2019 CIS Annual Meeting: Immune Deficiency & Dysregulation North American Conference

    Submission ID#551389

    Two Novel Mutations of Major Histocompatibility Class-II Associated Molecules

    Lauren Rigg, MD1, Neha Sanan, DO2, Devi Jhaveri, DO3, Haig Tcheurekdjian, MD3

    1Internal Medicine Resident, University Hospitals Cleveland Medical Center / Case Western Reserve University

    2Adult and Pediatric Allergy / Immunology Fellow, University Hospitals Cleveland Medical Center

    3Allergy / Immunology, Allergy Immunology Associates

    Introduction/Background: Major Histocompatibility Class II (MHC-II) molecules are transmembrane proteins that are essential to the development of the normal adaptive immune response. The genes that encode the MHCII include the Regulatory Factor X-Associated Ankyrin Containing Protein (RFXANK), Regulatory Factor X-Associated Protein (RFXAP), Regulatory Factor X, 5 (RFX5), and MHC-II transactivator (CIITA) proteins. Homozygous mutations in these genes lead to MHC-II Deficiency Syndrome and have been associated with early onset and severe respiratory and gastrointestinal infections, failure to thrive, and premature death. Herein we report two cases with significant clinical manifestations of immunodeficiency in patients with heterozygous mutations of the RFXANK proteins.

    Objectives: To describe two cases of novel RFXANK gene variants and their respective phenotypes.

    Methods: The patients were evaluated in the office for possible immune deficiency. A retrospective chart review was conducted examining medical history, diagnosis and response to treatments.

    Results/Case Description:

    Case 1: A 55-year-old female presented for recurrent mucocutaneous candida infections. Prior treatments included therapeutic and prophylactic fluconazole. Immunodeficiency workup showed a mannose binding lectin deficiency, low lymphocyte response to candida and tetanus antigen testing, and no response to candida skin testing. Genetic testing demonstrated a heterozygous variant in the RFXANK gene (c.612A>G/p.Arg167Cys).

    Case 2: An 18-year-old Caucasian female presented for lymphadenopathy, immune thrombocytopenic purpura and recurrent infections since early childhood. Prior treatments included antibiotics, subcutaneous and intravenous immunoglobulin (IVIG) therapy, and Rituximab. Immunodeficiency workup showed decreased immunoglobulin levels, B cells, and T cells. Genetic testing demonstrated a heterozygous variant in the RFXANK gene (c.726C>G/p.Ile242Met).

    Conclusions: Homozygous mutations of MHC-II associated molecules lead to a primary immunodeficiency known as MHC-II deficiency. Increasing genetic data is becoming available to physicians and patients including heterozygous mutations. While difficult to categorize, heterozygous mutations of MHC-II related proteins may still present with clinically significant immunodeficiency. As this data is further studied, it may assist in diagnosis and subsequent therapy.

    (2) Submission ID#551762

    The Effects of Adiantum Capillus Hydro Alcoholic Extract on Some Immunological Parameters in Mice

    Mehrdad Modaresi1, Masoomeh Pashaei2

    1Faculty Member, Isfahan (Khorasgan) Branch, Islamic Azad University, Isfahan, Iran

    2Laboratory employee, Department of Biology, Payam e Noor University, Isfahan Center, Isfahan, Iran

    The Adiantum capillus a known medicinal herb in traditional medicine which is widely used in traditional medicine to deal with infection by having chemical compounds that affect the immune system. The current study was carried out to investigate the effects of adiantum hydroalcoholic extract on plasma proteins and electrophoretic pattern of blood in small laboratory mice. Mature female mice (Balb/C) were divided into 5 groups including control, placebo, and 50, 100, and 200mg/kg of extract. The extract was injected intraperitoneal every other day for 20 days. At the end of the experiment, blood samples were taken and used to measure blood proteins and their electrophoretic pattern. Obtained data were analyzed using the SPSS program (p<0.05). According to the results, 100 and 200 mg/kg doses increased the amount of albumin, alpha-1 globulin, beta globulin, and A/G ratio. Therefore, it can be said that the extract has a positive effect on the blood system and plasma proteins and can increase the immune system without the presence of antigenic factors.

    (3) Submission ID#554014

    Unexpected Diagnosis in a Family with Autoimmune Multilineage Cytopenia and Hypogammaglobulinemia

    Yael Gernez, MD, PhD1, Jose Chavez, PhD2, James Bussel, MD3, Charlotte Cunningham-Rundles, MD, PhD4

    1Clinical Assistant Professor, Stanford School of Medicine

    2Post Doctoral, Division of Clinical Immunology, Icahn School of Medicine, Mount Sinai NY, NY

    3Professor in Pediatrics, Department of Hematology and Oncology, Weill Cornell Medicine, NY, USA

    4Professor in Medicine, Division of Clinical Immunology, Icahn School of Medicine, Mount Sinai, NY, NY, USA

    A 34 y.o. female was referred to our clinic with a history of multilineage cytopenias/Evans syndrome, a history of idiopathic thrombocytopenic purpura, hemolytic anemia, chronic neutropenia, lymphopenia, and hypogammaglobulinemia treated with IVIG.

    Our patient was healthy until she was 8 years old; at that time, she developed joint pain, rash, and bruising. She was found to have Evans syndrome with idiopathic thrombocytopenic purpura (ITP), neutropenia, and lymphopenia. She was initially diagnosed with lupus and was given steroids. Her bone marrow biopsy did not conclude myelokathesis. When she was 15 years old, she remained thrombopenic and was started on high dose of immunoglobulin replacement therapy. In 2012 (29 years old), she developed polyarthritis in her upper and lower extremities. In 2013 (30 years old), she had a severe nosebleed, for which she was admitted and treated with Amicar twice; her platelets were found to be 2,000 K/UL. She received rituximab weekly for 4 weeks resulting in an increase of platelet count to 90-100K/UL. She recently (March 2017) had a splenectomy to remove her large spleen, and since then, her platelets have rebounded to 400-500K/UL. In 2015, she was placed on long-term immunoglobulin replacement therapy after being hospitalized for bilateral pneumonia for 5 nights requiring IV antibiotics for treatment. In 2017, she developed and was treated for another pneumonia.

    Her family history is characterized by multiple members with autoimmune multilineage cytopenia as well as autoimmune diseases such as multiple sclerosis (mother), thyroiditis and enteropathy.

    On physical examination, she did not present with any warts and the remainder of her physical examination being unremarkable, except for her scar from her splenectomy and a cervical lymphadenopathy.

    Immunologic evaluations showed IgG 601 mg/dL, IgA <5 mg/dL, and IgM 208 mg/dL. CBC with differential and lymphocyte screen were as follows (cell/mm3): WBC 12.3 x103, Hemoglobin 10.2 g/dl, Platelets 503 x 103; 3 % neutrophils (ANC: 300), 82% lymphocytes, 10% monocytes, 0% eosinophils; absolute total T-cell number was 8884 (750-2500 cells/MCL), CD4+ T-cells 6554 (480-1700cells/MCL), CD8+ T-cells 2185 (180-1000cells/MCL), natural killer cells 206 (135-525 cells/MCL), and absolute number of B cells was 996 (75-375 cells/MCL).

    She came to our clinic with her sister, who also had multilineage cytopenia and hypogammaglobulinemia, treated with monthly IVIG; and her nephew whom had neutropenia. Based on this family presentation all three underwent Whole Exome Sequencing (WES). The patient, the patients sister and the patients nephew were all found to have a variant on CXCR4 (frameshift mutation on Chromosome 2, p.Val324fs; RefNt: TCA; AltNt: T). As an important note, the patient had a bone marrow biopsy, which did not conclude myelokathesis.

    In summary, our patient with trilineage cytopenia and hypogammaglobulinemia, without any warts or myelokathexis, had WHIM syndrome (Warts, Hypogammaglobulinemia, Immunodeficiency, and Myelokathexis), which was discovered by studying her WES. With the identification of her specific diagnosis, this allowed us to discuss the potential future indication of Plerifaxor (antagonist of the alpha chemokine receptor CXCR4). And equally important, we discussed family planning and future pregnancies given that the mutation is autosomal-dominant.

    (4) Submission ID#555017

    Risk of Bacterial Infections Among Patients with Secondary Complement Deficiency

    Taha Al-Shaikhly, MBChB1, Kathleen Mohan, ARNP2, Matthew Basiaga, DO, MSCE3, Eric Allenspach, MD, PhD4

    1Allergy & Immunology Fellow, Division of Allergy & infectious Diseases, University of Washington

    2Nurse Practitioner, Department of Immunology, Seattle Children's Hospital

    3Assistant Professor, Department of Rheumatology, Seattle Children's Hospital

    4Assistant Professor, Department of Immunology, Seattle Children's Hospital

    Introduction: Complement component-3 (C3) is shared by the classical, lectin and alternative complement activation pathways. C3, a major opsonin, facilitates phagocytosis of encapsulated microorganisms. Inherited C3 deficiency is rare and is associated with increased risk of bacterial infections. Subjects with connective tissue diseases (CTD) and C3 nephritic factors can have low and occasionally undetectable C3 levels, yet they are at an underappreciated infectious risk. We hypothesize that excessive C3 consumption in secondary complement deficiency disorders (SCD) is associated with higher risk of bacterial infections similar to primary complement deficiency disorders (PCD).

    Objectives: To compare the rate of bacterial infections between PCD and SCD patients and evaluate the association between C3 level and bacterial infection risk.

    Methods: We performed a retrospective cohort study. Subjects with an undetectable complement activity (CH50) or any of the complement components measured at Seattle Childrens hospital from 2002-2018 were included in our study. We recorded the number of infections, observation periods, diagnosis (PCD, SCD and its underlying etiology), lowest complement component levels, and the immunosuppressive agents used. The date of birth, and date of lowest C3 level were considered as start points to calculate the observation periods for PCD and SCD subjects respectively. Infections requiring hospitalization or parenteral antibiotics were categorized as serious bacterial infections (SBIs). Descriptive analyses were performed to determine medians and ranges for continuous variables. Differences in rates of bacterial infection were assessed using the chi-square and kruskal-wallis tests when appropriate. Among subjects with CTDs, we treated every C3 measurement as a single observation (n=1,197) and studied the association between C3 concentration and the 30-day odds of having a SBI. Multivariable logistic regression was performed to determine infection risk based on C3 level while controlling for contributing factors.

    Results: We identified 14 subjects with PCD, and 52 subjects with SCD. SCD consisted of three subgroups (CTD-related (n=44), nephritic factor-related (n=2), and infection-related (n=6)). Collectively, CTD subjects had a lower median rate of SBI compared to PCD subjects (P = 0.004). Subjects with CTD and C3 level <40 have higher rate of bacterial infection (of any severity) (P = 0.002) and of SBI (P = 0.004) when compared to CTD subjects with C3 >=40 at the beginning of observation period (Figure 1). While controlling for immunosuppression level and lupus nephritis diagnosis, C3 levels were predictive of SBI (P = 0.007, Figure 2). CTD subjects with a C3 level < 40 had a significantly higher risk for SBI compared to those > 40 (OR 3.63, 95% CI [1.03-12.7], P = 0.04). Subjects with infection-related hypocomplementemia had an undetectable CH50 lasted for at least 9 days.

    Conclusion: Among CTD patients, low C3 levels are predictive of more SBIs in the future. C3 level <40 is associated with higher 30-day odds of having a SBI. Therefore, CTD patients with very low C3 levels should be monitored for early signs of infections and should have a lower threshold for antibiotics initiation. SBI itself can result in an undetectable CH50 and re-testing is warranted before confirming a PCD diagnosis.

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    (5) Submission ID#556068

    Heterozygous TACI Mutation (TNFRSF13B: A181E) Causing Significant Infections in a Patient with Normal Immunoglobulins

    Shan Shan Wu, DO1, Jenny Lee, MD2, Michelle Sergi3, David P. McGarry, DO4, Robert Hostoffer, DO, LhD, FACOP, FACOI, FAAP, FCCP5

    1Allergy and Immunology Fellow, University Hospitals Cleveland Medical Center, Cleveland, Ohio

    2Internal Medicine/Pediatrics Resident, University Hospitals Cleveland Medical Center/Rainbow Babies and Childrens Hospital, Cleveland, Ohio

    3Medical Student, Ohio University Heritage College of Osteopathic Medicine, Warrensville Heights, Ohio

    4Allergy and Immunology Fellow, University Hospitals Cleveland Medical Center, Cleveland, Ohio

    5Allergy and Immunology Program Director, University Hospitals Cleveland Medical Center, Cleveland, Ohio; Allergy and Immunology Associates Inc. Mayfield Heights, Ohio

    Introduction: Common variable immunodeficiency (CVID) is a primary immune deficiency associated with loss of B-cell functions. Genetics of CVID are multifactorial, although both monogenic and polygenic forms have been described in the literature (1). Mutations (heterozygote and homozygote) in TNFRSF13B, the gene that encodes the transmembrane receptor, or TACI, are associated with 8-10% of CVID patients (2). TACI mutation with reduced TACI expression on marginal zone and CD27+ memory B-cells can impair B-cell differentiation, proliferation, and isotype switch (2), contributing to the pathogenicity of CVID. Asymptomatic individuals with normal immunoglobulin levels who have TACI mutation are also reported in the current literature (2). We aim to describe the significance of the heterozygous TNFRSF13B variant in a patient with recurrent sinopulmonary and skin infections without apparent B-cell dysfunction.

    Objective: To present a patient with the heterozygous TNFRSF13B variant with the clinical manifestations of those with CVID despite normal immunologic findings inconsistent with CVID.

    Method: Immunologic studies for the patient included serum immunoglobulins (IgG, IgA, IgM) and IgG subclasses, B-cell phenotyping, lymphocyte subset markers, mannose-binding lectin, mitogen and antigen stimulation, bacteriophage study, Streptococcus pneumoniae titers to 23 serotypes, and genetic sequence analysis with deletion/duplication testing of 207 genes. Genetic testing was also performed on the patients mother, father, and two sisters.

    Results: A 27-year-old male presented with a history of multiple infections since four months old, including recurrent episodes of acute otitis media, bronchitis, sinusitis and pneumonia, viral meningitis, mastoiditis, and cellulitis with abscesses of the axilla, thigh, and perianal region. The patient was found to have the TNFRSF13B, Exon 4.c.542C>A (p.Ala181Glu) heterozygous TACI variant associated with CVID. B-cell phenotyping showed an increase in naive B-cells (CD19+CD27-IgD+) and a decrease in both non-switched, memory B-cells (CD19+CD27+IGD+) and switched, memory B-cells (CD19+CD27+IgD-) with proper levels of transitional B-cells (CD19+CD24+CD38+) and plasmablasts (CD19+CD24-CD38+). This phenotype indicates a dysregulation in B-cell differentiation and proliferation into memory B-cells and impairment in isotype class-switching commonly found in individuals with CVID harboring TACI mutations. Yet the immunoglobulin levels and vaccine response were appropriate, excluding a diagnosis of CVID.

    The patients mother, who was asymptomatic, had the same TACI variant. Her immunoglobulins, lymphocyte subset markers, and B-cell phenotype were normal. Sister A has a history of multiple sinopulmonary infections with genetic results pending. The patients father and sister B did not have any immune issues and had no genetic mutations.

    Conclusion: CVID is a heterogeneous disease that may be associated with genetic defects. TACI mutations found in a small percentage of individuals with CVID, result in B-cell dysfunction and hypogammaglobinemia. We describe a patient with a TNFRSF13B exon Ala181Glu heterozygous mutation with recurrent infections and normal immunoglobulin levels and vaccine response.

    References:

    1 Bonilla FA, Barlan I, Chapel H, et al. International Consensus Document (ICON): Common Variable Immunodeficiency Disorders. J Allergy Clin Immunol Pract. 2016;4(1):38-59.

    2 Martinez-Gallo M, Radigana L, Belén Almejúne M, et. al. TACI Mutations and Impaired B-cell Function in Subjects with CVID and Healthy Heterozygotes. Allergy Clin Immunol. 2013;131(2):468476.

    (6) Submission ID#561686

    A Novel Mutation in Zap 70 Leading to an Infant with T+B+NK+ Severe Combined Immunodeficiency

    Kelsey Kaman, MD1, Alicia Johnston, MD2, Monique Abrams, MD1

    1Pediatric Resident, Baystate Medical Center

    2Faculty Advisor, Baystate Medical Center

    Introduction: ZAP70 codes for a 619-amino acid enzyme, ZAP70, a member of the Syk-protein tyrosine kinase family that plays an important role in T cell development and activation. ZAP70 is phosphorylated at tyrosine kinase residues upon T cell receptor (TCR) stimulation resulting in TCR-mediated signal transduction with Src family kinases. ZAP70 deficiency results in a rare T+B+NK+ Severe Combined Immunodeficiency (SCID). We report a novel compound heterozygous mutation in ZAP70 leading to presumed absent ZAP70 function in an infant with a normal TREC newborn screen and SCID.

    Case Description: The patient is a term, fully immunized female, born to non-consanguineous parents who was hospitalized for RSV bronchiolitis at 2 mo. At 4 mo she developed an erythematous, papular rash on her face and extremities, nonresponsive to topical antifungal therapy. At 6 mo she was re-hospitalized with RSV bronchiolitis and subsequently treated with multiple courses of antibiotics for presumed bacterial pneumonia followed by albuterol and oral steroids for possible reactive airways disease. During this course of treatment, her rash resolved. At 8 mo she presented with failure to thrive (wt <0.1% for age), multifocal pneumonia and respiratory failure requiring intubation. Bronchial alveolar lavage confirmed Pneumocystis jiroveci pneumonia prompting an immune evaluation. Total immunoglobulins were normal for age, however antibody titers to tetanus, diphtheria and Streptococcus pneumoniae were absent. Lymphocyte enumeration revealed elevated CD4 T cells and markedly diminished CD8 T cells, normal B and NK cells. T cell proliferation to mitogens (PHA, PWM) and antigens (Candida, tetanus) was absent, however T cells proliferated normally to stimulation with PMA and ionomycin. TREC number was normal by newborn screening, but was 2 std deviations below the mean and would have resulted in a positive screen upon repeat. Invitae 18 gene SCID panel revealed two variants of unknown significance, c.109C>G (p.Arg37Gly) leading to substitution of Arg with Gly and c.1529_1532dupGCAT (p.Ile511Metfs*65) resulting in a premature translational stop signal expected to disrupt the last 109 amino acids of ZAP70 protein. Parental sequencing revealed these variants to be on opposite chromosomes. The patient was successfully treated for PJP pneumonia and has since successfully engrafted a 9/10 matched unrelated donor stem cell transplant.

    Discussion: We report a novel compound heterozygous mutation in ZAP70 which we presume led to T+ B+ NK+ SCID. Our patients clinical presentation of failure to thrive, recurrent lower respiratory tract infections, dermatologic findings and PJP pneumonia are consistent with previously reported cases of ZAP70 SCID. Her paucity of CD8 T cells, abundance of CD4 T cells and absent proliferation to mitogens are also consistent with previously described cases of ZAP70. Normal proliferation of T cells when bypassing the TCR by stimulating cells with ionomycin and PMA confirms a defect in the TCR. We believe this is the second documented case of missed SCID by newborn screen in MA since the implementation of TREC screening in 2008.

    (7) Submission ID#564579

    A Case of Memory B-cell Dysfunction in a Child with Recurrent Otitis Media

    Arjola Cosper, DO MS1, Lisa Barisciano, MD2

    1Pediatric Resident (PGY III), Goryeb Children's Hospital

    2Attending Physician, Pediatric and Adult Asthma, Allergy and Immunology, LLC

    Introduction: Acute Otitis Media (AOM) is one of the most common reasons for antibiotic use in early childhood. We explored the challenges when AOM fails traditional therapies and immunologic evaluation does not identify a commonly described immunodeficiency.

    Case Description: An eighteen-month-old male presented with 12 episodes of AOM and recurrent purulent otorrhea requiring intravenous antibiotics. Laboratory evaluation revealed a normal CBC, normal immunoglobulins (IgG 588, IgA 76, IgM 63, IgE 12) and IgG subclasses. Lymphocyte subset panel was normal. Initial responses to DTaP and Prevnar boosters were normal, however, there was rapid decline to tetanus and pneumococcal antibody titers. A sub optimal response to Haemophilus influenza Type B vaccine was noted. Although vaccinated twice for MMR, he never mounted mumps specific IgG. Mitogen response to PHA was normal with decreased responses to ConA and pokeweed and no detectable tetanus nor candida responses. Further investigation revealed decreased non-class and class switched memory B-cells. The patient was recently vaccinated to PCV23 and at the present time has protective titers.

    Discussion: It has been previously suggested that decreased memory B cells may contribute to decreased antibody responses to select vaccine antigens resulting in recurrent AOM in children. Our case supports the need to investigate beyond typical immunologic screening for immunodeficiencies.

    (8) Submission ID#566756

    Coexistence of Lymphoproliferative Syndrome, Neurofibromatosis, Systemic Lupus Erythematosus and Hyper IgM Syndrome in a Patient with MSH6 Mutation

    Sukru Cekic1, Yasin Karali1, Sara Sebnem Kilic2

    1Fellow of Allergy and Clinical Immunology, Uludag University Faculty of Medicine

    2Professor of Allergy and Clinical Immunology, Uludag University Faculty of Medicine

    Introduction: DNA mismatch repair (MMR) system corrects replication errors in newly synthesized DNA, and prevent recombination between DNA sequences when they were not identical (1). MSH6 is a part of MMR genes, (2-4).

    Case: A ten-year-old girl presented with fever, brown spots on her skin, hair loss, recurrent pulmonary infections, arthritis on the left hand and right ankle. She has also been followed up with NF (Figure 1). There was a first-degree cousin marriage between her parents. Physical examination revealed findings of pneumonia and NF. Anti-nuclear antibody, anti-nDNA, anti-dsDNA, anti-histone, Anti Ro52 and anti-nucleosome antibodies were positive. In her immunologic assessment showed low IgG and IgA levels associated with high IgM level (Table 1). The coexistence of NF, hyper IgM syndrome, SLE, were considered in the patient. Intravenous Ig (400 mg/kg, every 3 weeks) treatment was started due to hypogammaglobinemia. The frame shift mutation in exon 2 of the MSH6 gene was detected in the Boztug's laboratory.

    In the follow up period, she admitted at 11 years old with back pain. A mass in the left paravertebral area, related to the spinal canal and neural foramina, was detected At the L4-L5 levels in spinal MRI. The lymphadenopathy around the liver and hilum and the left parietal bone lesions were developed within two months despite surgical excision of primary mass (Figure 2). As a result of PET examination; SUVmax was found to be around 6.5 in the mass lesion in the paravertebral region and SUVmax values did not exceed 2.5 in other lymphadenopathy and masses. Atypical cellular infiltration suggesting neoplastic events, which were including small-medium size atypical pleomorphic mononuclear cells and T cells. Since all these formations did not indicate definite cancer, chemotherapy was not started. Interestingly, although chemotherapy was not given, progression stopped, and partial spontaneous regression was observed.

    Discussion: The effect of MSH6 mutations on patients may significantly vary with the inheritance pattern (2). Leukemias or lymphomas are not common in heterozygote MMR gene defects (5,6). However, homozygote mutations in MMR genes show a different pattern. Wimmer and Etzler proposed the new term Constitutional mismatch repair-deficiency syndrome (CMMR-D) for patients who have a homozygous mutation in MMR (3). CMMR-D characterized by development of childhood cancers, mainly hematological malignancies and/or brain tumors, as well as early-onset colorectal cancers, and neurofibromatosis type 1 (3). Bi-allelic germline mutations in any of the MMR genes in which MSH6 is involved increases hematological malignancies by 15% (7,8). MSH6 mutation has been associated with many cancers since its identification. Leukemia, lymphoma, colorectal cancer, endometrial cancer, brain tumors are some of these cancer types (2-4,9).

    MSH6 deficiency is an important disease that can affect different systems at the same time. There is a high risk of malignancy in the cases and therefore they must be closely monitored. This case has also shown that atypical lymphoproliferation may occur in MSH6 homozygous mutant cases.

    Table 1. The immunologic assessment of patient
    IgG:213 mg/dl (normal rage: 842-1943) CD3: 83.1% (2717/mm3)
    IgA: 66,4 mg/dl (normal range: 62-390) CD4:%38,6 (1262/mm3)
    IgM: 334 mg/dl (normal range: 54-392) CD8: %41.5 (1357/mm3)
      CD19: %14.6 (477/mm3)
      HLA-DR+CD19:%14.5 (474/mm3)
      CD3-CD16+CD56+:%1.7 (55/mm3)

    Image 1. Cafe-au-lait spots on the extremities

    figurec

    Image 2. Paravertebral mass and parietal bone lesions in the magnetic resonance imaging

    figured

    References:

    1. Marinus MG. DNA Mismatch Repair. EcoSal Plus. 2012;5(1).

    2. Hegde MR, Chong B, Blazo ME, Chin LH, Ward PA, Chintagumpala MM, Kim JY, Plon SE, Richards CS. A homozygous mutation in MSH6 causes Turcot syndrome. Clin Cancer Res. 2005;11(13):4689-93.

    3. Wimmer K, Etzler J. Constitutional mismatch repair-deficiency syndrome: have we so far seen only the tip of an iceberg? Hum Genet. 2008;124(2):105-22.

    4. Ripperger T, Beger C, Rahner N, Sykora KW, Bockmeyer CL, Lehmann U, Kreipe HH, Schlegelberger B. Constitutional mismatch repair deficiency and childhood leukemia/lymphoma--report on a novel biallelic MSH6 mutation. Haematologica. 2010;95(5):841-4.

    5. Lynch HT, de la Chapelle A. Hereditary colorectal cancer. N Engl J Med. 2003;348(10):919-32.

    6. Lynch HT, Lynch JF, Lynch PM, Attard T. Hereditary colorectal cancer

    syndromes: molecular genetics, genetic counseling, diagnosis and management. Fam Cancer. 2008;7(1):27-39.

    7. Bakry D, Aronson M, Durno C, Rimawi H, Farah R, Alharbi QK, Alharbi M, Shamvil A, Ben-Shachar S, Mistry M, Constantini S, Dvir R, Qaddoumi I, Gallinger S, Lerner-Ellis J, Pollett A, Stephens D, Kelies S, Chao E, Malkin D, Bouffet E, Hawkins C, Tabori U. Genetic and clinical determinants of constitutional mismatch repair deficiency syndrome: report from the constitutional mismatch repair deficiency consortium. Eur J Cancer. 2014 Mar;50(5):987-96.

    8. Wimmer K, Kratz CP, Vasen HF, Caron O, Colas C, Entz-Werle N, Gerdes AM,

    Goldberg Y, Ilencikova D, Muleris M, Duval A, Lavoine N, Ruiz-Ponte C, Slavc I, Burkhardt B, Brugieres L; EU-Consortium Care for CMMRD (C4CMMRD). Diagnostic criteria for constitutional mismatch repair deficiency syndrome: suggestions of the European consortium 'care for CMMRD' (C4CMMRD). J Med Genet. 2014;51(6):355-65.

    9. Bougeard G, Charbonnier F, Moerman A, Martin C, Ruchoux MM, Drouot N, Frébourg T. Early onset brain tumor and lymphoma in MSH2-deficient children. Am J Hum Genet. 2003;72(1):213-6.

    (9) Submission ID#567651

    American Society of Pediatric Hematology and Oncology (ASPHO) Clinical Immunology Special Interest Group (SIG): Expanding Clinical Immunology Education, Research and Care

    David K. Buchbinder, MD, MSHS1, Sharat Chandra, MD, MRCPCH2, Blachy J. Davila Saldana, MD3, Rachael F. Grace, MD4, Kim E. Nichols, MD5, Süureyya Savasan, MD6, Nicola A. Wright, MD7, Roshini S. Abraham, PhD8, Shamuganathan Chandarkasan, MD9

    1Assistant Clinical Professor, Department of Hematology, Children's Hospital of Orange County, Orange, CA, Department of Pediatrics, University of California at Irvine, Orange, CA

    2Assistant Professor, UC Department of Pediatrics, Division of Bone Marrow Transplantation and Immune Deficiency, Cincinnati Childrens

    3Blood and Marrow Transplant Specialist, Division of Blood and Marrow Transplantation, Childrens National Medical Center, Department of Pediatrics, The George Washington University, Washington, DC

    4Director, Hematology Clinic, Assistant Professor of Pediatrics, Harvard Medical School, Pediatric, Hematology/Oncology, Dana-Farber/Boston Children's Cancer and Blood Disorders

    5Director, Cancer Predisposition Division, Division of Cancer Predisposition, St. Jude Children's Research Hospital, Memphis, TN

    6Director, Pediatric Blood and Marrow Transplantation Program, Professor of Pediatrics, Bone Marrow Transplant Program, Carman and Ann Adams Department of Pediatrics, Wayne State University School of Medicine, Children's Hospital of Michigan, Detroit, MI

    7Associate Professor, Department of Pediatrics, Alberta Children's Hospital, Calgary, Alberta, Canada

    8 Department of Pathology and Laboratory Medicine, Nationwide Childrens Hospital, Columbus, OH.

    9 Assistant Professor, Division of Bone Marrow Transplant, Aflac Cancer and Blood Disorders Center, Children's Healthcare of Atlanta, Emory University School of Medicine, Atlanta, GA

    Background: Advances in inborn errors of human immunity have supported the discovery of new syndromes that are marked by striking features of autoimmunity and immune dysregulation often associated with cytopenias, lymphoproliferation, and a predisposition to reticuloendothelial malignancies leading to evaluation with hematologists/oncologists. Moreover, hematologists/oncologists have also seen an increasing use of effector cell-based therapies, checkpoint inhibitors, immunomodulatory and targeted therapies resulting in autoimmunity and hyperinflammatory complications. A working knowledge of clinical immunology could help practicing hematologists/oncologists in the identification and management of these conditions.

    Objectives: To support the advancement of ASPHO members and the field by facilitating education regarding the best practices in diagnosis and management of immunological disorders. To create a platform for the development of collaborative clinical research in patients with hematological/oncological manifestations of immunological disorders or those requiring hematopoietic stem cell transplantation for a underlying immunological disorder.

    Design/Methods The ASPHO Clinical Immunology SIG was initiated based on collaboration with the Clinical Immunology Society (CIS). ASPHO members who are pediatric hematology/oncology clinicians, clinical researchers, and trainees are eligible to participate. We have established a steering committee with representatives from across the United States and Canada with diverse clinical and research expertise. Through regular teleconferences and annual in-person meetings, we have developed a platform to provide our members with a network of immunology resources to ensure a strong foundation of knowledge and tools to conduct clinical care and research pertaining to the diagnosis, evaluation, and treatment of patients with immunological disorders.

    Results: Prior to the inaugural meeting, we conducted a needs assessment of the ASPHO Clinical Immunology SIG Membership, which defined areas of priority pertaining to clinical immunology including education and research. At the inaugural meeting of the 2018 ASPHO Clinical Immunology Special Interest Group in May 2018, 43 ASPHO Members participated. We currently support over 50 members within our online community. Several educational initiatives have been successfully launched. We have submitted an invited review to Pediatric Blood and Cancer which provides a case-based review of primary immune regulatory disorders. We hosted the first Immunology for Hematology Oncology Practice (I-HOP) Cased-based Webinar Series. This series features case-based discussions of patients with primary immunodeficiency disorders presented by fellow trainees and mentored by senior clinicians. We will also be hosting an ASPHO Webinar focusing on the Laboratory Evaluation of Primary Immunodeficiencies and Immune Dysregulation Syndromes. We have also begun the process of laying the groundwork for clinical research initiatives.

    Conclusion: The ASPHO Clinical Immunology SIG seeks to serve as a collaborative resource for pediatric hematology/oncology clinicians and researchers. Through the development of educational and research initiatives, we envision improving the care of patients with immunological disorders that are often managed by pediatric hematologists/oncologists. Moreover, we hope to broaden our understanding and application of clinical immunology within pediatric hematology/oncology. We hope that this successful initiative will serve as a blueprint for the development of future collaborations with other specialty societies and patient groups.

    (10) Submission ID#569242

    Sepsis as a Sign of Immunodeficiency

    Katsiaryna Serhiyenka1, Oxana Romanova, PhD2

    1Assistant of professor, Belarussian State Medical University

    2Professor, Belarussian State Medical University

    Submission Text

    Background: T-cell immunity disorders among primary immunodeficiencies (PID) are 9% in the registry of the European Society of Immunodeficiency (ESID) and 10.5% in the United States . T-cell disorders are characterized by the absence or presence of T-lymphocytes. Because T cells are important for the normal functioning of B cells, most PID with a T-cell disorder lead to combined T- and B-cell disorders. Disturbances of the T-cell link of immunity are clinically manifested in early childhood. The most serious form of PID with violation of the T-cell link of immunity is a severe combined immunodeficiency (SCID), the first symptoms of which are already observed in infants and are characterized by the development of life-threatening infections.

    Results: Girl N. at the age of 3 months entered the Childrens Infectious Hospital with complaints of cough, high febrile temperature for 5 days, refusal to eat. From the anamnesis of life the girl from the 1st pregnancy, 1 birth, was born full term in 40 weeks gestation, birth weight 4640g. For 3 months of life, a bad increase in body weight was noted and at the time of admission, the weight in 3 months was 5400g. According to the parents, the child had atopic dermatitis. From the anamnesis of the disease on 08.01, the temperature rose to 38.2°C, there was a cough and a mucous discharge from the nose. Then the child refused to eat, the body temperature rose to 39.2°C. January 14 patient was hospitalized.

    According to the immunogram, a sharp decrease in CD3 + 26% (58-85%) was detected, activated T-lymphocytes (CD3 + HLA-DR +) were 19.9% (3-15%), T helper / inducers (CD4 + CD8 - 26.6% (30-56%) and T suppressors / cytotoxic (CD8 + CD4-) 0.5% (18-45%), a high ratio of Tx / Tc (CD4 + CD8 +) was detected 53.2% (0.6-2.3), cytotoxic non-T cells (CD3-CD8 +) -1,2, an increase in the number of B-lymphocytes (CD19 +) - 58.9% (7-20%), natural killers (CD16 + CD56 +) - 6.6% (5-25%), natural T-killers (CD3 + CD16 + CD56 +) - 0.3 (0-5%), leukocyte gates (CD45 + CD14-) - 99% (95-100 %). The absolute content of T-lymphocytes was 0.15 x 109/l, B - lymphocytes - 0.35 x 109/l. The number of thymic migrants (CD45 + CD45RA + CD31 +) was not detected (0%).

    01/17/2017 CT scan of the chest was diagnosed CT signs of a polysergic two-sided inflammatory process in the lungs.

    Blood for sterility - Staphylococcus epidermidis was isolated, CMV DNA was detected in an amount of 7.6 ×106copies/ml.

    Despite the therapy, the patient died.

    Posthumous diagnosis: Primary immunodeficiency (SCID, T0 B + Nk +). Complications: Sepsis. Septic shock. SPON: ARDS, renal failure, DIS, thrombocytopenia, anemia 3. Two-sided lower-lobe pneumonia. Generalized CMV infection.

    Conclusion: The peculiarity of the described clinical case was that the patient's first symptoms of SCID developed in the first months of life and were manifested by a bad weight gain, atopic dermatitis and the development of a life-threatening generalized cytomegalovirus infection and sepsis.

    (11) Submission ID#569933

    Two Siblings with Autoimmune Polyendocrinopathy-candidiasis-ectodermal Dystrophy-like Phenotype Demonstrating Classic and Atypical Symptoms

    Edith Schussler, MD1, Elise Ferre, PA-C, MPH2, Monica Schmitt, CRNP3, Michail Lionakis, MD, Sc.D4

    1Assistant Professor of Pediatrics, Division of Pulmonary, Allergy & Immunology, Weill Cornell Medicine

    2Physician Assistant, Fungal Pathogenesis Section, Laboratory of Clinical Immunology & Microbiology (LCIM) , National Institute of Allergy & Infectious Diseases (NIAID), NIH

    3Nurse Practitioner, Fungal Pathogenesis Section

    National Institute of Allergy and Infectious Diseases (NIAID) National Institutes of Health (NIH)

    4Chief, Fungal Pathogenesis Section, National Institute of Allergy and Infectious Diseases (NIAID) National Institutes of Health (NIH)

    Submission Text

    Autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) is a rare autosomal recessive disease caused by AIRE gene mutations. Clinical diagnosis is established by the presence of at least two components of the classic triad of chronic mucocutaneous candidiasis, hypoparathyroidism, and Addisons disease. In Europe, the classic presentation is widely recognized and nonendocrine autoimmune manifestations are rarely reported. A recent study of 35 American APECED patients demonstrated a more heterologous presentation, with many non-endocrine manifestations including urticarial eruption, hepatitis, gastritis, intestinal dysfunction, pneumonitis and Sjogrens-like syndrome, all uncommon in European reports. Within the American cohort, 80% of patients developed a mean of three non-triad manifestations before reaching the classic triad. Finding of AIRE mutations and high-titer antiIFN- autoantibodies is seen in both European and American cohorts.

    We present the case of two siblings, who demonstrate an APECED-like phenotype with both classical and atypical features. They share the same heterozygous c132+1_132+3delinsCT AIRE mutation.

    The older, an eight-year-old boy, with history of prematurity, bronchopulmonary dysplasia and onychomadesis in infancy, came to medical attention at 16 months of age due to failure to thrive (FTT), in addition to fevers and urticarial rash lasting months after his MMR vaccine. The fevers resolved with Anakinra, which was discontinued two years later due to pneumonia. From age 2-4 he developed an ALPs negative lymphadenopathy which self-resolved. Lung issues include chronic cough, initially treated as asthma but with poor bronchodilator response, and frequent lung infections, including 1-2 pneumonias per year. At age five evaluation for FTT revealed growth hormone deficiency. Two years later he was diagnosed with primary Addisons disease. Chronic abdominal discomfort, bloating, cyclical constipation/diarrhea, recurrent rashes, dystrophic nails, and SICCA symptoms are also present.

    His sister, age five, shows FTT, but no growth hormone deficiency. At age one, she too developed a fever and rash syndrome lasting 3 months. Severe GERD and constipation started in infancy and are ongoing. At age three she developed a transaminitis, initially diagnosed as EBV, but later thought to be autoimmune hepatitis. She has frequent viral respiratory infections, and pneumonia at age two. She has had a chronic cough, with poor bronchodilator response, for most of her life. Evaluation of seizure at age three showed normal brain activity. Brain MRI revealed partial agenesis of the corpus callosum and microgyria. Her brother has similar MRI findings. Both children have had developmental motor delay and poor tone. Brain dysgenesis and neurodevelopmental delay has not previously been described in APECED.

    Although there were both typical and atypical symptoms, the history in combination with genetic findings led to further investigation of an APECED-like syndrome. Autoantibody testing confirmed high-titer antiIFN- autoantibody typical of APECED in both children and high-titer BPIFB1 autoantibodies found almost exclusively in APECED pneumonitis in the brother. Whole exome sequencing and copy number variation analyses are underway to further evaluate the patients condition.

    This case demonstrates the importance of clinical presentation in the evaluation of genetic results and in the guidance of therapeutic management.

    (12) Submission ID#570047

    Different Clinical Manifestations in a Large Cohort of Predominantly Antibody Deficiency Patients with Monogenic Defects

    Reza Yazdani, PhD1, Hassan Abolhassani, MD, PhD2, Asghar Aghamohammadi, MD, PhD3

    1Fellow, Research Center for Immunodeficiencies, Childrens Medical Center, Tehran University of Medical Sciences, Tehran, Iran

    2Postdoctoral, Division of Clinical Immunology, Department of Laboratory Medicine, Karolinska Institute at Karolinska University Hospital Huddinge, Stockholm, Sweden

    3 Faculty member, Research Center for Immunodeficiencies, Pediatrics Center of Excellence, Children's Medical Center, Tehran University of Medical Science, Tehran, Iran

    BACKGROUND: Predominantly antibody deficiencies (PADs) are the most common primary immunodeficiencies, characterized by hypogammaglobulinemia and inability to generate effective antibody responses.

    OBJECTIVE: We intended to report most common monogenic PADs and to investigate how PAD patients who were primarily diagnosed as agammaglobulinemia, hyper IgM syndrome (HIgM) and common variable immunodeficiency (CVID) have different clinical and immunological findings.

    METHODS: Stepwise next generation sequencing and Sanger sequencing were performed for confirmation of the mutations in the patients clinically diagnosed as agammaglobulinemia, HIgM and CVID.

    RESULTS: Among 550 registered patients, the predominant genetic defects associated with agammaglobulinemia (48 BTK and 6 heavy chain deficiencies), HIgM (21 CD40L and 7 AID deficiencies) and CVID (17 LRBA deficiency and 12 atypical ICF syndromes) were identified. Clinical disease severity was significantly higher in patients with heavy chain and CD40L compared to patients with BTK (P = 0.003) and AICDA (P = 0.009) mutations. Paralysis following live polio vaccination was considerably higher in patients with heavy chain deficiency compared with BTK deficiency (P <0.001). We found a genotype-phenotype correlation among patients with BTK mutations regarding clinical manifestation of meningitis and chronic diarrhea. Surprisingly, we noticed that first presentations in the majority of ICF patients were respiratory complications (P = 0.008), while first presentations in LRBA patients were non-respiratory complications (P = 0.008).

    CONCLUSION: This study highlights similarities and differences in clinical and genetic spectrum of the most common PAD-associated gene defects. This comprehensive comparison will facilitate clinical decision making, and improve prognosis and targeted treatment.

    (13) Submission ID#574302

    Visualizing the Effect of Lymphatic Pump Techniques on Immune System in Normal Subjects “ Randomized Control Trial”

    Ahmed Abdelfattah, PhD1, Neveen Abdelraouf, PhD2, Samy Nasef, PhD2, Rania Ali, PhD2

    1Lecturer, Faculty of Physical Therapy - Cairo University

    2Professor of Physical Therapy - Cairo University Faculty of Physical Therapy

    Aim: this study was designed to investigate and compare the efficacy of selected osteopathic lymphatic techniques on the absolute CD4+ count in healthy subjects. Materials and Methods: Forty-five subjects (33 males and 12 female), age varies from 20 to 50 years old. They were allocated to three groups each one has 15 subjects: first one received sternal pump and sternal recoil techniques for 12 sessions, three sessions per week. Second one received thoracic lymphatic pump and splenic pump techniques for 12 sessions, three sessions per week. Third one (control group) didn't receive OMT. Absolute count of CD4 was used to evaluate participants before and after application of the osteopathic techniques. Results: analysis showed significant increase in CD4 count after treatment in the second group also there was no significance in the first and third groups. P-value was 0.05. Conclusion and discussion: thoracic lymphatic and splenic pump manipulative techniques are effective methods of enhancing the immune system in healthy subjects (TLPT & SPT).

    Key words: Osteopathy, Lymphatic techniques, Immune system

    (14) Submission ID#576532

    Infants with Idiopathic Transient and Persistent T Cell Lymphopenia Identified by Newborn Screening a Single Centers Experience from September 2010 December 2017

    Artemio M. Jongco, III, MD, PhD, MPH1, Omer Elshaigi2, Foysal Daian2, BS, Emily Bae2, Amanda Innamorato2, Brianne Navetta-Modrov, MD3, Robert Sporter, MD4, David Rosenthal, DO, PhD5, Vincent Bonagura, MD6

    1Assistant Professor of Medicine and Pediatrics, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell; Center for Health Innovations and Outcomes Research, Feinstein Institute for Medical Research, Manhasset, NY

    2Research Intern, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    3Fellow, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    4Allergist, ENT and Allergy Associates

    5Assistant Professor of Medicine and Pediatrics, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    6Professor of Medicine and Pediatrics, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    Rationale: Infants with low T cell receptor excision circles (TREC) born in Queens, Nassau, and Suffolk counties are referred to our center for further evaluation. This study elucidates the demographic and laboratory characteristics of referred infants with transient or persistent idiopathic T cell lymphopenia (TCL) without clearly identified genetic or acquired etiology.

    Methods: A retrospective analysis was performed from September 2010 (when TREC screening started) through the end of December 2017. Descriptive statistics were calculated for demographic and laboratory characteristics. T-test or Mann-Whitney tests were used to compare laboratory variables. Pearson or Spearman tests were used to determine correlation between initial TREC levels and T cell counts. By definition, the CD3+, CD4+, and CD8+ populations of transient TCL patients normalize by age 1 year.

    Results: Eighteen infants with transient and 17 with persistent TCL were identified. Males comprised 61.1% of the transient and 47.1% of the persistent TCL cohorts. Whites comprised 11.1% of the transient and 35.3% of the persistent TCL cohorts. The mean initial TREC levels did not differ between the transient and persistent cohorts (67.7 vs. 78.5 TRECs/L of blood, P = 0.56). Mean initial absolute counts of CD3+ (2149 vs. 1300 cells/L, P <0.0001), CD4+ (1462 vs. 922 cells/L, P <0.0001), and median initial absolute counts of CD8+ (524 vs. 309 cells/L, P = 0.0075), were higher for transient vs persistent cohorts. Initial TREC level did not correlate with initial CD3+, CD4+, or CD8+ absolute counts. The median age of resolution for the transient cohort was 121.5 days (range 23-244). The absolute CD3+, CD4+, or CD8+ counts rarely exceeded the reported median values for age, and remained closer or below the 5th percentile for age up to 1000 days of life. The majority of both transient and persistent TCL patients demonstrated unremarkable lymphocyte proliferation to mitogens.

    Conclusion: Our centers transient TCL cohort appears to be predominantly male and non-white, whereas the persistent TCL cohort is more evenly distributed by sex but still predominantly non-white. The transient cohort had lower initial TREC levels, but higher initial T cell counts. Both cohorts appear to have relatively intact in vitro function.

    (15) Submission ID#577904

    Primary Immune Deficiency Disease in Patients over Age 60: An Analysis from a Proprietary Immunology Patient Registry

    Roger H. Kobayashi, MD1, Daniel Suez, MD2, Ralph Shapiro, MD3, Donald L. McNeil, MD4, Mark R. Stein, MD5, Frank J. Rodino, MHS, PA6, Herbert Lewis, PhD7

    1Clinical Professor UCLA School of Medicine National Consultant, Immune Deficiency Foundation Executive Committee: CIIC, Consortium of Independent Immunology Clinics

    2President/Director of Daniel Suez, MD Allergy, Asthma & Immunology Clinic, PA, Past President CIIC Consortium of Independent Immunology Clinics

    3President/Director of Midwest Immunology Clinic; Past President: CIIC Consortium of Independent Immunology Clinics

    4President/Director of Optimed Research LTD Consortium of Independent Immunology Clinics

    5Physician, Allergy & immunology Allergy Section, Good Samaritan Medical Center, West Palm Beach, FL, USA

    6Founder and President Churchill Outcomes Research, LLC Clinical Assistant Professor Stony Brook University School of Health Technology and Management

    Consortium of Independent Immunology Clinics

    7Associate Professor - College of Business - Stony Brook University

    Consortium of Independent Immunology Clinics

    Introduction: Primary immune deficiency disease (PIDD) is typically considered a pediatric illness, although advances in treatment and diagnosis are changing this paradigm. Currently, data on PIDD in older patients are very limited.

    Objectives: To characterize the prevalence of PIDD among older individuals using a patient database maintained by the Consortium of Independent Immunology Clinics (CIIC), comprised of 17 specialty immunology outpatient practices in the US.

    Methods: Patients with PIDD were identified in the CIIC database using ICD-10 codes D80, D.80.3, D80.4, D80.5, D80.6, D81.1, D81.2, D82.0, D82.3, and D83.0. A total of 235 records from 11 geographically-diverse clinics were identified and characterized by age, gender, and PIDD diagnosis.

    Results: Of the 235 PIDD patients in the CIIC registry, 73 (31%) were between 60-87 years of age (see Figure). Within this age group, most patients were female (n=56, 77%). The most common diagnoses among patients >60 years of age included Common Variable Immunodeficiency with Predominant Abnormalities of B-Cell Numbers and Function (D83.0; n=41, 56%) and Antibody Deficiency with Near Normal Immunoglobulins (D80.6; n=14, 19%). In comparison, the registry included 36 (15%) patients aged 0-19 years; this age group was predominantly male (n=23; 64%). The most common ICD-10 codes within the younger cohort were relatively evenly distributed between Hereditary Hypogammaglobulinemia (D80.0), Antibody Deficiency with Near Normal Immunoglobulins (D80.6), and Common Variable Immunodeficiency with Predominant Abnormalities of B-Cell Numbers and Function (D83.0).

    Conclusions: Our data suggest that PIDD in patients over age 60 may be more prevalent than previously reported. Additional research is needed to corroborate these findings, further characterize the nature of PIDD in this population, and determine whether there are unique diagnostic and treatment considerations within this demographic.

    (16) Submission ID#579038

    A Case of C6 Complement Deficiency with a Novel Mutation

    Hassan A. Ahmad, MD1, Christopher D. Codispoti, MD, PhD2

    1Allergy/Immunology Fellow, Rush University Medical Center

    2Assistant Professor, Rush University Medical Center

    Introduction/Background: Increased susceptibility to invasive infections with Neisseria has been well documented in patients with deficiency of terminal complement proteins. The molecular attack complex is constructed with complement components C5 to C9. A deficiency in complement C6 has been described previously in both African American and South African populations. Complement C6 deficiency is inherited in a co-dominant pattern, with multiple known mutations. We present a case of a 19-year-old, previously healthy male, who presented with invasive N. Meningitides infection. He was found to have a novel mutation noted on genetic sequencing of the complement C6 gene.

    Objective: We present the case of a 19-year-old, previously healthy male, who presented with invasive N. Meningitides infection. On genetic sequencing, he was found to have three mutations of the complement C6 gene. Two of which have been described previously, and a third novel mutation.

    Methods: A 19-year-old male with no known history presented to us with a 3-hour history of emesis. He was found to be febrile, and quickly decompensated, developing septic shock. Blood cultures were drawn, and within 12 hours grew N. Meningitides. He was treated with broad spectrum antibiotics upon arrival, and subsequently narrowed to Ceftriaxone. His hospital course was complicated by disseminated intravascular coagulation, as well as acute tubular necrosis, leading to end-stage renal disease for which he is listed for kidney transplant.

    Results: On immunodeficiency evaluation, he was noted to have an undetectable CH50 (<13, reference range 31-60). Complement levels returned with C6 of 10.8 (reference range 28-69) and C1r of 41.5% (reference range 61-102%). Complement C6 function screen returned at 0% (reference range 40.7-169%). All other complement levels were within normal limits. Genetic sequencing showed the patient to be compound heterozygous for two of known four variants which have been reported to recur in African patients with complement C6 deficiency. This included c.821del and c.1879del, which are predicted to result in frameshift and premature protein termination. He was also found to be heterozygous for sequence c1202G>A, which results in amino acid substitution p.Arg401Lys. This variant is rare, with one large database reporting it in 6 of 276000 alleles, and not in a homozygous state. It has not been reported in a case of C6 complement deficiency previously.

    Conclusions: We present the case of a previously healthy 19-year-old male with invasive meningococcal disease. He is compound heterozygous for two mutations that have been associated with total complement C6 deficiency; however, he was found to have subtotal C6 deficiency. Furthermore, he has a third novel mutation of the complement C6 gene. Further investigation is warranted on the significance of this finding and impact on relevance to possible kidney transplant.

    (17) Submission ID#579501

    Assay Characteristics of an Automated, Liposome-based Assay for the Measurement of CH50 Complement Activity and Comparison with a Haemolytic Method

    Clare E. Tange, PhD1, Kattika Bootdee2, Kritraporn Deesin2, Leigh Williams, PhD3, Asada Leelahavanichkul4, Stephen Harding, PhD5

    1Medical Science Liaison, The Binding Site

    2Faculty of Medicine, Chulalongkorn University

    3Medical Science Liaison, The Binding Site

    4Asst Prof, Faculty of Medicine, Chulalongkorn University

    5Research and Development Director,The Binding Site

    Background: Measuring the function of the classical pathway of complement activation is useful in several disease states, including complement deficiency, autoimmune conditions such as systemic lupus erythematosus and certain forms of nephritis. The original method for assessing classical pathway activity was the haemolytic CH50 method, but this assay can be time consuming and has reagent stability issues due to the use of sheep red blood cells. There can also be high lab-to-lab variability due to differences in the protocols used. Here we report the assay characteristics of an automated, commercial, liposome-based assay to measure CH50 activity. We also compare the results obtained using the traditional haemolytic method with the automated, liposome-based method used on the SPAPLUS turbidimetric analyser.

    Methods: A linearity study was performed based on CLSI guideline EP06-A. The linear range of the SPAPLUS CH50 liposome assay was established by analysis of a series of sample dilutions and evaluation of results against pre-defined goals for recovery and %CV. Precision was assessed based on CLSI guideline EP05-A2 over 21 days. 4 samples with different CH50 activities (23.7-65.1 U/mL) were run in duplicate, with two runs per day using 3 reagent lots and 3 different analysers. Interference analysis was performed by spiking haemoglobin, bilirubin, chyle, ascorbic acid or saline (as a control) into samples before measuring the CH50 activity.

    For the assay comparison study, sera from 125 routine patient samples were used. Samples were collected from Chulalongkorn Hospital, Faculty of Medicine, Chulalongkorn University, Thailand. CH50 classical pathway activity was assessed using a haemolytic method and also using the liposome based CH50 assay for use on the SPAPLUS turbidimetric analyser (The Binding Site Ltd., Birmingham, UK). C3 protein concentrations were also available for 116 of these samples.

    Results: The liposome CH50 assay gives a linear response over the range 11.8-95.5 U/mL, covering the measuring range of the assay (12.0-95.0 U/mL) at the standard analyser dilution (neat). The within run, between run and between day %CVs were all 5.4%. The total %CV was 6.8% in all 4 samples. Minimal interference was observed with the four common interferents tested.

    A significant correlation was observed between the two CH50 methods (p<0.0001, r=0.66, y=1.1x±0.1), with 90.4% agreement between the methods in determining whether patients were above or below the lower limit of the assay normal range. The 12 individuals in disagreement had normal CH50 results using the haemolytic method, and low CH50 values in the liposome assay. Of these, C3 values were available for 10/12, and 5 had C3 concentrations below the lower limit of the assay normal range.

    Conclusion: The liposome CH50 assay for use on the SPAPLUS analyser has passed assay development guidelines based on those set out by the CLSI for linearity, precision and interference, and there is a strong correlation between this automated assay and the haemolytic CH50 method used here. Five additional patients with low C3 concentrations were defined as having a low CH50 using the SPAPLUS liposome method compared to the haemolytic method.

    (18) Submission ID#580179

    Frequency of Specific Antibody Deficiency (SAD) and Respiratory Allergy in Patients with Recurrent Sinusitis

    Charles Song, MD1, Dennys Estavez, Mr.2, Diana Cherinokova, MD3, Rie Sakai-Bizmark, MD4, Richard Stiehm, MD5

    1Chief of Pediatric Allergy and Immunology, Ronald Reagan UCLA Medical Center, UCLA Mattel Children's Hospital

    2Research statistician, Harbor-UCLA

    3Resident, Harbor-UCLA

    4Assistant Professor, Harbor-UCLA

    5Professor, Division of Allergy and Immunology, UCLA

    Submission Text

    Frequency of Specific Antibody Deficiency (SAD) and Respiratory Allergy in Patients with Recurrent Sinusitis

    Song CH1, Estavez D1, Chernikova D1, Sakai-Bizmark R1, Stiehm R2

    1Harbor UCLA Medical Center, Torrance, CA

    2UCLA Childrens Hospital , Los Angeles, CA

    Rational: Respiratory allergy and subtle immunodeficiency may lead to recurrent sinusitis. We sought to determine the frequency and relationship of allergic sensitization, allergic respiratory diseases, and specific antibody deficiency (SAD) among patients with recurrent sinusitis and respiratory infections (URI).

    Methods: The electronic medical records of 313 ambulatory patients from 6 to 70 years (median age 28 year) with recurrent respiratory infection (sinusitis >1x/y and URI>5x/yr) were screened for SAD, allergic sensitization (to mites, cockroach, cat, dog, and pollens), rhinitis (allergic and non-allergic), and asthma. Patients were divided into sinusitis and non-sinusitis classes; the pneumococcal antibody (PA) responses were categorized into three groups A, B, or C: A. Normal PA levels (defined as 70% of tested serotypes being above or equal to1.3 ug/mL for subjects, 6 years and older), B. Initially low with normal post-vaccination PA levels, and C. SAD with low PA levels even after Pneumovax).

    Results: Among the 213 sinusitis patients, 187(88%) had decreased initial protective PA serotypes (groups B & C). Of these, 45 subjects ( C, 21 % of total) had SAD. The prevalence of SAD among the sinusitis patients was significantly higher compared to non-sinusitis group (21% vs.12%, p<0.01). The SAD prevalence increased with age;10 % for 6-19 years olds, 24% for 20 -39 year olds, 21% for 40-59 year olds, and 26 % for 60- 70 year olds.( P <0.05 between 6-19 year olds vs. 20-39 year olds). The initial numbers of protective PA serotypes were highest among Group A and lowest in C (p<0.01). The allergy sensitization was equally high for the both groups (62% vs. 62%). Asthma was common for both groups (43% vs. 42%), and rhinitis was more prevalent among non-sinusitis group (74% vs. 84%, p=0.05).

    Discussion: SAD is a common and under-recognized cause of recurrent sinusitis. The prevalence rate increased with increasing age reflecting an aging immune response. Patients with SAD were more likely to present with initial very low number of protective PA serotypes compared to Group B representing a state that had experienced a greater difficulty mounting responses to polysaccharide antigens. The rates of allergy sensitization among both recurrent sinusitis and URI group were equal (62%) and significantly higher than the one reported by NHANES1 (45% among individuals 6 years and older), indicating an association between recurrent sinusitis /URI and allergy. Patients with recurrent sinusitis should be evaluated for both SAD and allergy.

    Reference:

    1. Arbes Jr SJ,Gergen PJ, Elliott L, Zeldin DC. Pfevalence of positive skin test response to 10 common allergens in the US population;results from the third National Health and Nutrition Examination Survey, J Allergy Clin Immunol 2009;124(3):522-7

    (19) Submission ID#583743

    Newborn Screening for SCID in Puerto Rico: A Three-year Experience

    Giannina Coppola-Fasick, MD1, Yanira M. Arce, MD1, Sonia Ramírez, MS, MT (ASCP)2, Ledith Resto, MS3, Sulay Rivera-Sanchez, MS, PhD4, Sylvette Nazario-Jimenez, MD5, Cristina Ramos-Romey, MD6

    1Allergy Immunology Fellow, University of Puerto Rico

    2Supervisor Molecular Genetics section of Puerto Rico Newborn Screening Program, University of Puerto Rico

    3NBS Follow Up Supervisor of Puerto Rico Newborn Screening Program, University of Puerto Rico

    4Associate Director of Puerto Rico Newborn Screening Program, University of Puerto Rico

    5Director of Allergy Immunology Program, University of Puerto Rico

    6Assistant Director of Allergy Immunology Program, University of Puerto Rico

    Background: Severe Combined Immune deficiency (SCID) is the most severe form of inborn immunodeficiencies, which are characterized in most cases by complete absence of T-cell-mediated immunity and by impaired B-cell-function. SCID is a pediatric emergency and is uniformly fatal without hematopoietic cell transplantation. Therefore, early diagnosis is important for prompt treatment. SCID can be detected using T cell receptor excision circle (TREC) assay. Newborn screening for SCID started in 2008 in Wisconsin and was added to the national recommended uniform panel for newborn screened disorders in 2010. There are currently 48 states performing mandatory screening for SCID in the United States. In August 2015 Puerto Rico (PR) added newborn screening for SCID to the mandatory newborn screening panel using dried blood spot specimen. A pilot program was done in 2011, in which one patient was diagnosed with SCID and successfully transplanted. The estimated incidence in PR is of 1:60,000 consistent with USA.

    Purpose/Objective: Report and describe the data of the first 3 years of newborn screening for SCID in PR.

    Method: We performed a retrospective record review of 15 positive newborn screening cases for SCID from patients born in PR during August 2015-October 2018. Patients received follow up at the Primary Immunodeficiency Clinic at the University of PR and NBS program.

    Results: A total of 81,600 infants were screened, identifying 15 cases with low TREC levels. Of these cases, five infants died (four of which were preterm babies), four were lost to follow up, and three had normal follow up TREC levels after surgery for gastroschisis and omphalocele. The other 3 infants were referred to our clinic for diagnostic and follow up evaluation which lead to the identification of one Di George syndrome, one Vici syndrome, and other non SCID lymphopenia who is still undergoing evaluation at our Immunology Clinic.

    Conclusion: We identified infants with abnormal TRECs that subsequently lead to diagnosis of non SCID lymphopenia, which may have not been recognized in the past and has enabled us to optimize management and outcomes of these infants. NBS has allowed for the early detection of infants with SCID (as found in our pilot study) and other lymphopenia disorders, which has permitted early diagnosis and management prior to developing symptoms or life-threatening complications.

    (20) Submission ID#584818

    Loss of Human ICOSL Results in Combined Immunodeficiency

    Lucie Roussel, PhD1, Marija Landekic, MSc2, Christina Gavino, MSc3, Alexis Blanchet-Cohen, PhD4, Ming-Chao Zhong, PhD5, Melanie Langelier, MSc6, Denis Faubert, PhD7, André Veillette, MD8, Don Vinh, MD9

    1Research Associate, Research Institute - McGill University Health Centre

    2Graduate student (PhD trainee), Research Institute - McGill University Health Centre

    3Research Assistant, Research Institute - McGill University Health Centre

    4Bioinformatics, Institute Recherche Clinique de Montreal

    5Research Associate, Institute Recherche Clinique de Montreal

    6Research Nurse - Clinical Program Manager, Research Institute - McGill University Health Centre

    7Proteomics Director, Institute Recherche Clinique de Montreal

    8Director, Molecular Oncology Research Unit, Institute Recherche Clinique de Montreal

    9Associate Professor, Clinician-Scientist, McGill University Health Centre

    Background: Primary Immunodeficiencies are inborn errors of immunity that represent naturally occurring experimental models to decipher human immunobiology. We present a patient with combined immunodeficiency, who suffered from recurrent respiratory tract and viral infections associated with hypogammaglobulinemia and panlymphopenia. He also had progressive moderate neutropenia, without evidence of bone marrow failure or associated severe prototypical infections.

    Methods: Identification of the causal gene was performed by whole exome sequencing, bioinformatics analyses, and Sanger sequencing. The impact of the variant on gene product was assessed by cDNA sequencing and protein detection (flow cytometry, Western blot, confocal microscopy) on various cell lines. The effect of the variant on protein function was assessed by co-culture experiments (Jurkat with lymphoblastoid cells derived from healthy controls or patient) and by transendothelial migration of cells across endothelial cell lines reconstituted with ICOSLG (wild-type vs. variant).

    Results: We identified a homozygous mutation in the Inducible T-Cell Costimulator Ligand gene (ICOSLG; c.657C>G; p.N219K). Whereas wild-type ICOSL is expressed at the cell surface, the ICOSL p.N219K mutant abolishes cell surface expression, due to retention of protein in the endoplasmic reticulum/Golgi apparatus. The mutant ICOSL was associated with diminished T cell costimulatory activity and with decreased transendothelial lymphocyte migration. Additionally, endothelial expression of mutant ICOSL compromised neutrophil transmigration, by reducing the proper localization of E-Selectin and ICAM-1 at the cell surface.

    Conclusions: Our work identifies human ICOSLG deficiency as a novel cause of a combined immunodeficiency syndrome. Moreover, findings from this natural experiment sheds light on the broad immunologic functions of ICOSLG in human immunobiology.

    (21) Submission ID#584884

    Safety of Administration of Rotavirus Vaccine in Infants Born to Mothers Receiving Biologic Therapy During Pregnancy: A Retrospective Case Series

    Christina Smith, MD1, Niraj C. Patel, MD, MS2, Richard Sigmon, MD3

    1Resident Physician, Department of Pediatrics, Levine Children's Hospital, Atrium Health

    2Physician, Department of Pediatrics, Division of Infectious Disease and Immunology, Levine Children's Hospital, Atrium Health

    3Physician, Department of Medicine, Division of Gastroenterology, Atrium Health

    Background/Aims: Rotavirus vaccine is a live viral vaccine that is part of the routine U.S. childhood immunization schedule. Live viral vaccines administered to infants of mothers who received biologic medications during pregnancy can potentially cause vaccine-associated disease. Infant death from disseminated mycobacterial infection after vaccination with bacille Calmette-Guerin (BCG) in infants whose mothers received infliximab during pregnancy has been reported. It is currently recommended that infants born to women who received biologic therapy during pregnancy not receive live viral vaccines, however there is a paucity of information regarding adverse events from live viral vaccines. We report two infants, born to mothers receiving infliximab during pregnancy, who tolerated the complete series of rotavirus vaccine.

    Methods: Two infants who received rotavirus vaccine and whose mothers received infliximab (monoclonal antibody against tumor necrosis factor alpha which blocks the inflammatory response) during pregnancy were identified and their charts were reviewed. Each mothers chart was assessed for timing of the biologic doses during pregnancy and concurrent immunosuppressant therapy.

    Results: The mother of the first infant had Crohn's Disease and received infliximab every 6 weeks throughout her pregnancy (final infusion at approximately 35 weeks estimated gestational age [EGA]). She did not take additional immunosuppressive drugs throughout her pregnancy. The infant was born at 39 weeks EGA. The infant received rotavirus vaccine at 2, 4, and 6 months of age. The infant did not have coexisting medical conditions or recorded hospitalizations during the first year of life. There were no side effects from rotavirus vaccine documented during well child examinations. The childs growth was normal during the first year of life.

    The mother of the second infant also had Crohn's disease and received infliximab infusions every six weeks during pregnancy until 27 weeks EGA. Additionally, she took mesalamine (anti-inflammatory) daily. The infant was born at 33 weeks EGA. The baby had a brief and uncomplicated neonatal intensive care unit stay. She did not have medical conditions diagnosed at the time of birth, or in the first year of life. The child received rotavirus vaccination at 2, 4, and 6 months of chronological age, and the infant did not experience documented adverse reactions. The child presented to the emergency department twice in the first year of life: once for thrush at 10 months of age and once for viral gastroenteritis at 11 months of age. The childs growth curve was unremarkable.

    Conclusions: We report two infants, whose mothers received infliximab during pregnancy, who safely tolerated the 3-dose series of rotavirus vaccination. Neither infant in this case series suffered from minor or severe adverse events as a direct consequence of receiving rotavirus vaccine. This suggests that administration of rotavirus vaccine may be safe in infants whose mothers received biologic therapy.

    (22) Submission ID#585141

    Combined Immune Deficiency in Association with a Single RAG1 Missense Variant in a 28-year-old Female

    Charles Song, MD1, Diana Cherinokova, MD2, Joseph A. Church, MD3, Henry Lin, MD4, Christin Deal, MD5, Manish Butte, MD, PhD6

    1Chief of Pediatric Allergy and Immunology, Ronald Reagan UCLA Medical Center, UCLA Mattel Children's Hospital

    2Resident, Harbor-UCLA

    3Professor, Pediatrics, Children's Hospital Los Angeles and Keck School of Medicine of U.S.C.

    4Chief, Pediatric Genetics, Harbor-UCLA

    5Allergy/Immunology Fellow, UCLA

    6Division of Allergy/Immunology Chair, Division of Immunology, Allergy, and Rheumatology, Dept. of Pediatrics and Jeffrey Modell Diagnos-tic and Research Center, University of California, Los Angeles

    Introduction: Combined immunodeficiencies (CIDs) can arise from partial loss of function variants in recognized SCID genes, which can lead to relative lymphopenia with poorly functioning and oligoclonal T cells. CIDs have been most commonly associated with variants of the RAG genes, but other genes are also implicated. Clinical symptoms may be less severe, and the onset generally is delayed, compared to typical SCID presentations.

    Case Report: A 28-year-old female presented with a history of recurrent and progressively worsening infections involving multiple microorganisms and organs, starting in infancy and requiring frequent hospitalizations. Bacterial or viral infections included rhinosinusitis, otitis media, herpetic stomatitis, dental abscesses, pneumonias, pulmonary mycobacterial abscesses, CMV hepatitis, urinary tract infections, dermal abscesses, and groin hidradenitis. Fungal and yeast infections included cryptococcal meningitis, oral thrush, dermatophytosis of the face, osteomyelitis of a finger, and onychomycosis. Laboratory tests in 2018 showed: mildly low T cell counts (791/uL) with a reversed ratio of CD4/CD8 T cells (0.22); almost absent B cells (2/uL); and low NK cell counts (19/uL). CD4+ T cells were mostly of the memory phenotype (87%). T cell development showed low counts of Th17 cells. T-cell stimulation tests demonstrated poor proliferation responses (<30%) to Concanavalin A, tetanus toxoid, and Candida albicans, with near-normal responses to pokeweed (>13%) and PHA (>84%). She had low Ig levels (IgA 72, IgM 23, IgE <2), except for IgG (872mg /mL; due to replacement since early childhood).

    Limited genetic evaluation at age 9 showed a heterozygous variant in the RAG1 gene (g.36595918T>C, c.1064T>C, p.Met355Thr; NM_000448.2).

    Discussion: Loss of function variants in RAG1 or RAG2 genes are known to cause a T- B- NK+ type SCID. More than 100 missense variants have been reported for RAG1, with disease-associated variants predominantly in zinc binding regions. The RAG1 missense variant in our patient also lies within the zinc binding region (amino acids 354-383). The variant is rare (mean allele frequency 0.0001521 in gnoMAD) and has been identified in at least one other individual with SCID (T-, B cell-, NK+). Although classified as a variant of unknown significance, occurrence in at least two individuals with deficiencies of T and B cells-- within a functionally important RAG1 domain -- supports an interpretation that the variant may be pathogenic. Most patients with CID with RAG variants are either homozygous for a poorly functional allele or have one nonunfucitonal and a second, poorly functional allele. We detected only a single potentially pathogenic allele. Our patient has decreased NK cells in addition to T and B cell defects. Further genetic studies including whole exome sequencing, are planned to identify further variants in RAG1 or other relevant genes.

    (23) Submission ID#586431

    Capturing Quality of Life in Patients with Common Variable Immunodeficiency (CVID) Using the Patient-Reported Outcomes Measurement Information System (PROMIS-29) Survey

    Shouling Zhang, MD1, Myriam Kline, PhD2, Ramsay Fuleihan, MD3, USIDNET Consortium4, Kathleen E. Sullivan, MD, PhD5, Artemio M. Jongco, III, MD, PhD, MPH6

    1Pediatrics Resident, Department of Pediatrics, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell, New Hyde Park, NY

    2Associate Research Statistician, Biostatistics Unit, Feinstein Institute for Medical Research, Manhasset, NY

    3Professor of Pediatrics, Division of Allergy and Immunology, Northwestern University Feinberg School of Medicine, Chicago, NY

    4United States Immunodeficiency Network, National Institute of Allergy and Infectious Diseases (NIAID), Towson, MD. The U.S. Immunodeficiency Network (USIDNET), a program of the Immune Deficiency Foundation (IDF), is supported by a cooperative agreement, U24AI86837, from the National Institute of Allergy and Infectious Diseases (NIAID).

    5 Professor, The Children's Hospital of Philadelphia

    6Assistant Professor of Medicine and Pediatrics, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell; Center for Health Innovations and Outcomes Research, Feinstein Institute for Medical Research, Manhasset, NY

    Introduction/Background: Common variable immune deficiency (CVID) is the most common antibody deficiency affecting both children and adults. Lifelong immunoglobulin replacement therapy (IGRT) is the mainstay of treatment. Information is limited about health-related quality of life (HRQOL) in patients with CVID receiving IGRT. The Patient Reported Outcomes Measurement Information System (PROMIS) is a validated self-report measure of physical, mental, and social health which can be used to assess HRQOL in patients with primary immunodeficiency diseases (PIDD).

    Objectives: The primary objective of this study was to compare patients with and without CVID on HRQOL domains using PROMIS-29 survey data from the United States Immunodeficiency Network (USIDNET) registry. USIDNET maintains a national registry of validated data from PIDD patients through the Immune Deficiency Foundation (IDF). The primary endpoint variables were scores in 7 HRQOL domains: 1) Depression, 2) Anxiety, 3) Physical Function, 4) Pain Interference, 5) Fatigue, 6) Sleep Disturbance, and 7) Social Participation. A secondary objective was to describe and compare patients with or without CVID with respect to IGRT.

    Methods: IDF prompts all its electronic personal health record users to answer PROMIS-29 biannually. Only those who have consented to participate in the USIDNET registry are included. Data from Fall 2015 to Spring 2018 were analyzed. Groups were compared using descriptive statistics and the Wilcoxon Mann-Whitney test. A mixed linear model approach compared groups with respect to the endpoint variables while adjusting for time, sex, age, and/or BMI. Simple models were tested followed by the addition of covariates. Interactions that were not significant were removed from models. All analyses use SAS, version 9.4.

    Results: Among the 222 PIDD patients in the registry, 173 patients (78%) were diagnosed with CVID. Humoral and cellular immunodeficiencies comprised the remaining non-CVID diagnoses (22%). Patients ranged from 18 to 83 years of age (mean age of 54). The study population was largely female (80%), Caucasian (96%), and with a mean BMI of 29. Twenty-five (11.5%) participants had a family history of PIDD. Results of the PROMIS-29 survey revealed that there was an effect of group (i.e., CVID/non-CVID) on the Fatigue subscale. The CVID group scored 3.05 points higher, on average, than the non-CVID group, after controlling for age and time (p=0.037). No other group differences were found among the remaining subscales. BMI was a significant predictor across all subscales (p<0.01), except for Anxiety (p=0.17). With regards to IGRT, the median IgG dose was less for CVID patients compared to non-CVID patients (17.8g vs. 25.0g, p=0.05), and the median number of days on IGRT was less for the CVID group compared to the non-CVID group (14 vs. 28, p=0.025).

    Conclusions: These data suggest that fatigue may be a key factor influencing the quality of life among PIDD patients with CVID. Future prospective longitudinal studies using PROMIS-29 will be needed to confirm this finding. Additional studies elucidating the role of BMI on HRQOL and IGRT dosing are recommended.

    (24) Submission ID#586449

    Describing Transient T Cell Lymphopenia in the United States Immunodeficiency Network (USIDNET) Following Infants with Low Lymphocytes (FILL) Program and a Single Referral Center from 2010-2017

    Shouling Zhang, MD1, Omer Elshaigi2, Foysal Daian, BS2, Emily Bae2, Amanda Innamorato2, Brianne Navetta-Modrov, MD3, Robert Sporter, MD4, David Rosenthal, DO, PhD5, Vincent Bonagura, MD6, Elizabeth A. Secord, MD7, Charlotte Cunningham-Rundles, MD, PhD8, John Routes, MD9, USIDNET Consortium10, Artemio M. Jongco, III, MD, PhD, MPH11

    1Pediatrics Resident, Department of Pediatrics, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell, New Hyde Park, NY

    2Research Intern, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    3Fellow, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    4Allergist, ENT and Allergy Associates, NY, NY

    5Assistant Professor of Medicine and Pediatrics, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    6Professor of Medicine and Pediatrics, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    7Professor of Pediatrics at Wayne State University, Children's Hospital of Michigan, Division of Allergy, Asthma and Immunology, Children's Hospital of Michigan, Detroit, MI

    8Professor in Medicine, Division of Clinical Immunology, Icahn School of Medicine, Mount Sinai, NY, NY, USA

    9Chief, Professor, Division of Allergy and Immunology, Children's Hospital of Wisconsin-Milwaukee, Milwaukee, WI

    10United States Immunodeficiency Network, National Institute of Allergy and Infectious Diseases (NIAID), Towson, MD.

    11Assistant Professor of Medicine and Pediatrics, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell; Center for Health Innovations and Outcomes Research, Feinstein Institute for Medical Research, Manhasset, NY

    Rationale: Infants with low T cell receptor excision circles (TREC) born in Queens, Nassau, and Suffolk counties in New York were referred to Northwell Health for further evaluation after abnormal newborn screens. The demographic and immune parameters of infants with transient T cell lymphopenia (tTCL) without clearly identified genetic or acquired etiology are described. TCL is considered transient if the lymphopenia resolves by 12 months of age. Similar data from the Following Infants with Low Lymphocytes (FILL) program of the United States Immunodeficiency Network (USIDNET) are presented.

    Methods: A retrospective analysis of two separate patient cohorts with tTCL are described. Cohorts include patients referred to a single center, Northwell Health, in NY from September 2010 to December 2017 and at USIDNET using data tracked by FILL from June 2011 to July 2018.

    Results: Out of 1,234 referrals at Northwell, 18 infants with tTCL were identified. Infants were predominantly male (61.1%) and non-Caucasian (89.9%). Out of 71 FILL participants, 9 infants with tTCL were identified. Infants were predominantly male (55.6%) and non-Caucasian (55.6%). Initial laboratory parameters for the Northwell versus FILL cohorts are summarized: a) median TREC levels: 54.5 vs. 47.0 TREC/L of blood; b) median absolute CD3+ count: 2135 vs. 1166 cells/L; c) median CD4+ count: 1460 vs. 777.0 cells/L; d) median absolute CD8+ count: 524.5 vs. 440.0 cells/L. Initial naïve CD4+ T cell information was available for 0 Northwell and 5 FILL infants (median 52%). Mitogen proliferation studies were performed in 10 (55.6%) Northwell and 6 (66.7%) FILL infants with 90% of these Northwell and 50% of these FILL infants demonstrating normal proliferation. Genetic testing, such as targeted genetic panels or chromosomal microarrays (CMA), was performed in 5 Northwell and 0 FILL infants. No genetic or chromosomal aberrations were identified. Whole exome sequencing (WES) was not performed in either cohort. 11 of 18 (61.1%) Northwell and 7 of 9 (77.8%) FILL infants did not receive the initial rotavirus vaccine. No FILL infants were vaccinated but no adverse effects were reported in 5 of 18 (27.8%) Northwell infants who received the first rotavirus dose. Of these, 3 of 5 (60.0%) had normal mitogen proliferation while 1 (20.0%) had decreased proliferation to phytohemagglutinin.

    Conclusions: Identifying biomarkers for tTCL and developing evidence-based guidelines for the diagnosis and management of tTCL are important knowledge gaps. This descriptive study is limited by small sample size and the constraints of registry-based research. Although there appear to be differences between these cohorts, our findings suggest that tTCL may disproportionately affect different segments of the population. tTCL infants with normal mitogen proliferation may be able to tolerate rotavirus vaccination. Thus, routinely checking proliferation studies in all tTCL infants may help risk stratify these patients and minimize vaccine-related adverse events. Currently, there is insufficient evidence to recommend more extensive genetic testing such as genetic panels, CMA, or WES. Systematically collecting information about patient characteristics and outcomes, as well as encouraging increased participation in registries such as FILL, may help address these shortcomings.

    (25) Submission ID#586903

    An Assay to Measure the Complement Binding Activities of Anti-dsDNA Antibodies in SLE

    Clare E. Tange, PhD1, David Taylor2, Marcos López-Hoyos, MD, PhD3, Victor Martínez-Taboada, MD, PhD4, Jaime Calvo-Alén, MD, PhD5, Leigh Williams, PhD1, Stephen Harding, PhDs6

    1Medical Science Liaison, The Binding Site

    2The Binding Site

    3Head of Service, Immunology Service., Hospital Universitario Marqués de Valdecilla

    4Staff, Rheumatology Service., Hospital Universitario Marqués de Valdecilla

    5Head of Service, Rheumatology Service., Hospital Universitario Araba

    6Research and Development Director, The Binding Site

    Background: Systemic lupus erythematosus (SLE) is a chronic, inflammatory disease that affects multiple organs. The measurement of anti-dsDNA antibodies (Abs) is a gold standard serological test used in the diagnosis and monitoring of SLE, with higher serum levels associated with worse prognosis. However, not all anti-dsDNA Abs are pathogenic, and some patients have consistently high levels with low disease activity. One mechanism suggested for the pathogenicity of these antibodies is complement activation. Here we describe an assay to measure the C1q binding activities of anti-dsDNA Abs in SLE patients.

    Materials & Methods: The concentration of anti-dsDNA Abs was determined using the QuantaLite dsDNA ELISA kit (INOVA) as per the manufacturers instructions. In order to determine the C1q binding capacity of bound Abs, samples were added to the pre-coated plate and incubated. Bound anti-dsDNA Ab/C1q complexes were then detected using a biotinylated anti-C1Q antibody (570 ng/mL) and streptavidin peroxidase (1 mg/mL). Normal reference ranges were developed in serum samples from healthy controls, and upper limits of these normal ranges were used as cut-offs. The dsDNA Abs and C1q binding capacity of bound Abs was then assessed in 49 SLE patients, and compared to other markers and the SLE Disease Activity Index (SLEDAI) score. Results are displayed as absorbance at 450nm (AU).

    Results and Conclusions: The 95th percentile ranges for anti-dsDNA Abs (0.068-0.137 AU) and C1Q binding activities (0.207-0.313 AU) were developed from the measurements generated in 17 healthy serum samples. SLE patients with an increased anti dsDNA Ab concentration (>0.137 AU) were then separated into those with low (<0.313 AU) and high (>0.313 AU) C1q binding activities. Patients whose dsDNA Abs had high C1q binding activity were found to have significantly higher SLEDAI scores (mean 6.70 vs 3.19). Serum C1q concentration, serum dsDNA Abs (measured by another method) and serum C3 and C4 concentrations were not significantly different between the two groups. This assay suggests that dsDNA Abs from SLE patients differ in their ability to bind complement, and that high complement binding activity of these antibodies may be linked to a more active form of disease.

    (26) Submission ID#587614

    Natural History of X-linked Lymphoproliferative Disease, Lessons Learned from a Long-term Survivor

    Tiphanie Vogel, MD, PhD1, Mihail Firan, MD2, Joud Hajjar, MD, MS1

    1Assistant Professor, Baylor College of Medicine, Texas Childrens Hospital Center for Human Immunobiology and Division of Immunology, Allergy and Rheumatology

    2Assistant Professor, Baylor College of Medicine, Departments of Pediatrics, Pathology and Immunology

    X-linked lymphoproliferative (XLP) is a primary immunodeficiency, caused by signaling lymphocyte activation molecule (SLAM)-associated protein (SAP) deficiency. Patients with XLP have severe immune dysregulation, usually triggered by EBV infection, leading to fulminant infectious mononucleosis, dysgammaglobulinemia and lymphoproliferation. Without hematopoietic stem cell transplant (HSCT) fatality is reportedly 100% by age 40. We report the natural history of XLP in a patient, and describe the lessons learned.

    Our patient was healthy and developed normally until 6-years of age, when he developed progressive respiratory symptoms. Lung biopsy revealed mature lymphoplasmacytic infiltrate in the alveolar septa, consistent with lymphoid interstitial pneumonia (LIP). He received corticosteroids and cyclophosphamide with significant improvement. At age 12, he developed severe infectious mononucleosis (fever, hepatosplenomegaly, lymphadenopathy, lymphocytosis). He had a protracted clinical course, but eventually recovered and seroconverted to a typical convalescent pattern. He subsequently developed hypogammaglobulinemia, and was started on intravenous immunoglobulin (IVIG). During the same year, his 10-year-old brother developed LIP, and subsequently hemophagocytic lymphohistocytosis (HLH) and died within 4 months from overwhelming candidiasis. Unfortunately, his youngest brother (age 7) then developed LIP and died 2 months later from a massive gastrointestinal bleed.

    Both siblings were treated with corticosteroids and cyclophosphamide; they did not have detectable EBV infection. At age 13 years, our patient experienced recurrent strokes and was found to have biopsy-proven CNS vasculitis. He was treated with interferon- and recovered with residual left sided weakness, but was lost to follow-up.

    He continued on IVIG, with no other immunomodulatory agents for several decades. He had progressive lung disease and recurrent seizures controlled with anti-epileptics. At age 43, he developed sudden vision change, headache and right-sided weakness, followed by a seizure. MRI of the brain revealed small bilateral areas of acute infarction suggestive of a central embolic event, however, no primary thrombus was identified. He did not receive any immunosuppression but was anti-coagulated. Eventually he was discharged home with resolution of weakness to his baseline.

    The patient was referred to our clinic after discharge and we re-evaluated him after 31 years. Immune profiles at the time showed therapeutic IgG troughs, low/undetectable IgM/A/E, normal T/B/NK-cell counts, normal spontaneous, but decreased antibody-dependent NK cytotoxicity, 0% SAP protein expression (on CD3+CD8+, CD3-CD56+ and CD3+CD56+ cells), and deletion on the X chromosome encompassing the SH2D1A gene which encodes SAP. His mother was a carrier of the same deletion. His functional status excluded the option of HSCT. A year later, he had rapid deterioration with recurrent lung infections, liver failure, and thrombocytopenia. Bone marrow biopsy revealed Hodgkins Lymphoma. He declined chemotherapy and died few days after diagnosis.

    Our case represents a rare patient with XLP surviving to the fifth decade without HSCT, particularly having experienced mononucleosis and non-EBV related CNS vasculitis. Our patient survived decades longer than his brothers (who most likely shared the same genetic defect) without evidence of somatic reversion (0% SAP expression in CD3+CD8+) to explain his milder clinical phenotype. This case may help in understanding the natural history of XLP, and confirms that prognosis remains poor without HSCT.

    (27) Submission ID#587907

    Abatacept for CTLA-4 Haploinsufficiency Presenting with Severe Bone Marrow Aplasia and Septic Shock - A Case Report

    Emilie Proulx, BSc, MD, FRCPC1, Antoine Morin-Coulombe, MD, FRCPC2, Jean-Philippe Drolet, MD, FRCPC3, Vincent Castonguay, MD, FRCPC4

    1Fellow-in-training, Clinical immunology, CHU de Québec, Université Laval

    2Fellow-in-training, Medical Oncology and Hematology, CHU de Québec, Université Laval

    3Allergy and Clinical Immunology, CHU de Québec, Université Laval

    4Haematology and Oncology, CHU de Québec

    CTLA-4 is a major negative regulator of immune responses, and CTLA-4 haploinsufficiency has been identified as a monogenic cause of primary immunodeficiency in patients presenting with a common variable immunodeficiency (CVID) phenotype with autoimmunity. Here we present the case of PB, a 40-year-old man who had been followed by the immunology service of our center for 17 years. A diagnosis of CVID had first been made when the patient presented with atypical transverse myelitis, low immunoglobulin levels, and lymphopenia. Over the years, his clinical picture was dominated by various forms of autoimmunity, namely inflammatory demyelinating disorder of the central nervous system, autoimmune haemolytic anemia, immune thrombocytopenia, cryptogenic organizing pneumonia, rheumatoid-like polyarthritis, chronic liver transaminitis with biopsy-proven moderate fibrosis, and lymphocytic colitis with malabsorption. Immunoglobulin replacement therapy was started at diagnosis, and autoimmunity was sequentially treated with methotrexate, interferon beta 1-a, cyclophosphamide, mycophenolate mofetil, rituximab, and finally a combination of low-dose prednisone and sirolimus, with stabilization of his neurological condition, the most debilitating complication of his immune dysregulation syndrome. Bone marrow transplant had been offered, but declined by the patient due to perceived good quality of life compared to transplant-associated risks.

    The patient was later referred to our hematology ward in July of 2018 for septic shock complicating febrile neutropenia, which was part of a two-month, gradual-onset pancytopenia. The diagnosis of immune-mediated aplastic anemia soon became apparent, as demonstrated by a bone marrow biopsy performed in a peripheral center two days prior to admission. The underlying pneumonia and thereafter biopsy-induced Staphylococcus aureus iliac osteomyelitis and soft-tissue abscess were treated with broad-spectrum antibiotics as well as multiple surgical interventions. The patient was started on eltrombopag, high-dose corticosteroids and cyclosporin A, the latter promptly switched to tacrolimus due to liver enzymes disturbances, all of which resulted in no significant hematologic response despite over seven weeks of treatment (with concurrent treatment of complicating infection, upper gastrointestinal bleeding, and intensive-care-unite myopathy). During that time, genetic confirmation of CTLA-4 haploinsufficiency was received, and the patient was thereafter started on abatacept on day 48 of current hospitalization. Administration of equine anti-thymocyte was initially foregone because of perceived infectious risk in the setting of poor iliac wound healing and superimposed adenovirus viremia; however, given the lack of response, it was given on days 52 through 54 of hospitalization. Haematologic response began on day 67 of hospitalization with a steady rise in all-lineage myelopoiesis up to a complete neutrophil response, platelet near-complete response as well as resolution of transfusion needs by day 101. While waiting for a well-matched bone marrow donor, isolated platelet decrease was observed and attributed to multiple factors, including low-grade thrombotic microangiopathy, inflammatory consumption and drug-related thrombocytopenia, but the patient remained well. To our knowledge, our patients presentation is one of the most severe manifestation of CTLA-4 haploinsufficiency to have responded to targeted therapy with abatacept, as a bridge to hematopoietic stem cell transplantation, with resolution of both immune and infectious complications, showing that genetic diagnosis is helpful in optimizing the management of presumed CVID patients.

    (28) Submission ID#588520

    Extreme Phenotypes, Identical Mutations: Two Patients with Same Nonsense XLF/Cernunnos Homozygous Mutation

    Luis I. Gonzalez-Granado, MD1, Nerea Dominguez-Pinilla, MD2, Melina Soledad Perrig, MD3, Carmen Rodriguez-Vigil, PhD4, Nerea Salmón-Rodriguez, MD5, Cristina Martinez Faci, MD4, Javier Blas-Espada, MD6, Marta López-Nevado, MD6, Raquel Ruiz-Garcia, PhD3, Rebeca Chaparro, MD3, Luis Allende, PhD6, Maria José Recio Hoyas, PhD3

    1Consultant. Head of the Primary Immunodeficiencies Unit. Pediatrics, Hospital 12 de Octubre

    2Consultant, Hospital 12 de Octubre Health Research Institute (i+12), Madrid, Spain. University Hospital Virgen de la Salud. Pediatric Hematology and Oncology Unit. Toledo. Spain.

    3Hospital 12 de Octubre Health Research Institute (i+12), Madrid, Spain, Dept. of Immunology, School of Medicine, Complutense University

    4Consultant, University Hospital Miguel Servet. Pediatric Hematology and Oncology Unit. Zaragoza. Spain

    5Consultant, Hospital 12 de Octubre Health Research Institute (i+12), Madrid, Spain. University Hospital 12 octubre. Madrid. Spain

    6Hospital 12 de Octubre Health Research Institute (i+12), Madrid, Spain, Dept. of Immunology, University Hospital 12 octubre. Madrid. Spain

    BACKGROUND: XLF/Cernnunos deficiency is a rare primary immunodeficiency classified within the DNA repair defects. These patients present severe growth retardation, microcephaly, lymphopenia and increased cellular sensitivity to ionizing radiation. Here, we describe two unrelated cases with the same nonsense mutation in the NHEJ1 gene showing significant differences in clinical presentation and immunological profile but a similar DNA repair defect.

    METHODS: Missense NHEJ1 mutation was identified by targeted next-generation sequencing with an in-house designed panel of 192 genes.

    For foci experiments, primary skin fibroblasts were irradiated with ionizing irradiation (137Cs) or treated with 20mM Etoposide for 1 hour. After irradiation, the cells were seeded at a density of 1x104 cells/mL in T75 flasks in triplicate. To evaluate cell sensitivity to gamma-IR (1 and 3 Gy),adherent cells were trypsinized and counted 11 days later. PBMCs from patient and healthy controls were irradiated with 10Gy, fixed and stained for CD3, CD19 and phospho-histone H2AX. Mean fluorescence intensities (MFI) of gamma-H2AX were evaluated on gated CD3+ lymphocytes.

    RESULTS:We report two patients harboring the same homozygous mutation in Cernunnos/XLF/NHEJ1 gene. Strikingly, their clinical phenotype ranges from severe combined immunodeficiency to isolated thrombocytopenia followed until escolar age (Table 1). They harbour the same c.169C>T mutation in NHEJ1 gene but different immunologic features (Table 2). P2 presented with mild T lymphopenia, hypersensitivity and NHEJ repair defect, typical for patients with XLF/NHEJ1 defects. On the other hand, P1 presented a more severe phenotype (T-B-), however hypersensitivity and NHEJ repair defect was similar to P2.Of note, P2 has survived into the first decade of live. Both patients are alive and well after HSCT.

    DISCUSSION: Usually the repair defect in these disorders is assessed by immunofluorescence assays of irradiation-induced gamma-H2AX foci using skin fibroblasts. A high throughput, sensitive and reliable assay to quantify gamma-H2AX foci in PBMCs isolated from blood samples would be a valuable tool to diagnose these patients and perform HSCT early. Flow cytometry (FC) can be applied as a rapid diagnostic tool for DNA repair disorders. Patients with the same homozygous mutation (p.R178X) in NHEJ1 gene have been previously reported. Two patients died at 1.5 and 4 years while another of the patients is already 8 years old and is alive (without HSCT). However,none of these patients presented severe T lymphopenia as it has been observed in our first patient.

    CONCLUSIONS: The assignment of a timely and accurate diagnosis is of paramount importance in the management of patients with defects in DNA repair. In the era of NBS an abnormal TREC assay should be followed by NGS approach as Cernunnos deficiency may present early in life as SCID,as other RS-SCID defects. Since genetic diagnosis takes time,functional radiosensitivity assays in peripheral blood may lead to the correct diagnosis and avoid exposure to alkylating agents during the conditioning regimen prior to genetic diagnosis. It would also be helpful in cancer patients to individualize and to guide the dosing of ionizing radiation (IR) and/or genotoxic agents to avoid accumulation of cells with genomic instability that could accelerate cancer development.

    Table 1. Clinical features of the patients with Cernunnos deficiency

       P1 P2
      Origin Caucasic Caucasic
      Consanguinity No No
    Age Onset 1m 9m
      Current 2y 7y
    Clinical features Microcephaly + +
      Growth retardation + +
      Facial dysmorphism - +
    Additional clinical features Neurological manifestations - -
      Bone malformation - -
      Autoimmunity - +
      Cytopenias - + (thrombocytopenia)
    Infections Respiratory tract infections - +
      Bacterial and opportunistic infection - -
      Urinary tract - -
    Outcome Status Alive and well (HSCT) Alive and well (HSCT)

    TABLE 2. Immunologic features of the patients

    Parameter RefValues
    (children)
    P1 P2
    Lymphocyte (n°/μL) 2500-6000 809 879
    T Cells    
    CD3+ n°/μL (%) 1400-4300(52-88) 60 (7) 661 (75)
    TCRαβ (%) 85-99 5 54
    TCRγδ (%) 2-15 1 16
    CD3+HLA-DR+ (%) 0-10 22 7
    DNT (%) 0-2.5 0.2 0.7
    CD4+ n°/μL (%) 1000-2500(33-55) 53 (7) 304 (35)
    CD4+CD45RA+CCR7+(Naïve) (%) 32-82 4.1 45.4
    CD4+CD45RA+CCR7- (CM) (%) 15-30 41.5 28.9
    CD4+CD45RA-CCR7- (EM) (%) 8-30 53.9 23.8
    CD4+CD45RA-CCR7+ (E) (%) 0.4-4 0.4 1.89
    CD4+CD45RA+CD31+ (%) 44-60 2  
    CD8+ n°/μL (%) 400-1400(17-34) 6 (1) 264 (30)
    CD8+CD45RA+CCR7+(Naïve) (%) 30-80 15.3 72.0
    CD8+CD45RA+CCR7- (CM) (%) 3-28 16.2 4.5
    CD8+CD45RA-CCR7- (EM) (%) 17-40 59.5 16.7
    CD4+CD45RA-CCR7+ (TEMRA) (%) 2-15 9 6.8
    TRECS (copies/punch) > 10 < 10 50
    NK Cells    
    CD56+CD3- n°/μL (%) 100-650(2-20) 671 (83) 191 (21.7)
    B Cells    
    CD19+ n°/μL (%) 400-1500(9-28) 49 (6) 22 (2.5)
    CD19+CD27+ (%) 7-19   32
    CD19+IgD-CD27+ (%Naïve) 75-89   63
    CD19+IgD+CD27+ (%MZ) 2.6-7.1   14.9
    CD19+IgD-CD27+ (%SW) 4.5-20   17.10
    CD19+CD38hiIgM+ (%Transitional) 3-10   13
    Plasmablasts 0.5-5   4.6
    KRECS (copies/punch) >10 <10 100
    Serum Immunoglobulins (mg/dl)    
    IgG (mg/dL) 600-1230 446 779
    IgA (mg/dL) 30-200 18 <6.67
    IgM (mg/dL) 50-200 40 109
    Specific antibodies    
    IgG vs Pneumococcus (mg/dL) >5.4   2.9
    IgG2 vs Pneumococcus (mg/dL) >2.4   0.36
    IgG vs Tetanus toxoid (IU/mL) >0.1   9.10

    (29) Submission ID#589566

    6 Month Old Female with Congenital Onset Indolent Systemic Mastocytosis Successfully Treated with Midostaurin

    Christin Deal, MD1, Manish Butte, MD, PhD2, Maria Garcia-Lloret, MD3

    1Allergy/Immunology Fellow, UCLA

    2Division of Allergy/Immunology Chair, Division of Immunology, Allergy, and Rheumatology, Dept. of Pediatrics and Jeffrey Modell Diagnos-tic and Research Center, University of California, Los Angeles

    3Allergy/Immunology Program Director, UCLA

    A 6-month-old female presented with symptoms of systemic mastocytosis including history of episodes of cyanosis and irritability, ALTE, chronic diarrhea (6-9 loose stools per day), hematochezia, diffuse skin lesions and bullae and a left axillary mass. Skin biopsy of left axilla was performed by her dermatologist showing presence of mast cells and she was subsequently started on cromolyn, cetirizine, ranitidine, prednisolone, hydroxyzine, diphenhydramine and montelukast without any improvement in symptoms. Axillary mass was initially believed to be abscess formation secondary to the skin biopsy but was found to have complex cystic structure on MRI and ultrasound. The lesion was initially improved by drainage but continued to recur over several weeks despite multiple antibiotic courses and surgical drainages. Cytology of drained fluid showed lymphocytosis.

    Bone marrow biopsy showed 5% mast cells and D816V variant of KIT gene. There was no associated hepatosplenomegaly. Germline sequencing of whole blood showed no mutation of KIT indicating congenital onset of acquired indolent systemic mastocytosis. Skin findings consistent with diffuse cutaneous mastocytosis with bullous eruptions and hemorrhage.

    She was subsequently hospitalized 2 weeks later for an unresponsive episode and had worsening of her skin lesions. During this hospitalization she started on Midostaurin at 30mg/m2 for systemic mastocytosis. Midostaurin is a novel protein kinase inhibitor that has shown efficacy treatment of patients with D816V c-kit positive advanced systemic mastocytosis (van Anrooij et al., 2018). Dose was increased to 45mg/m2 BID 3 weeks after initiation. She received premedication with ondansetron prior to each dose of Midostaurin. Tryptase levels responded well to Midostaruin and were 109 ug/L on the day of initiation of Midostaurin and were 54.1ug/L at her last check which was approximately 4 weeks after starting Midostaurin (Figure 1). Her skin lesions also significantly improved after starting the medication (Figure 2).

    Her hospitalizations were complicated by fluid overload and hypertension. Both fluid overload and hypertension resolved prior to discharge. She remains on 2mg prednisone daily, cetirizine, ranitidine, cromolyn and Benadryl and hydroxyzine PRN. To our knowledge, this is the youngest patient successfully treated with Midostaurin and she is doing very well on therapy with no apparent side effects. She has had resolution of many of her systemic mastocytosis symptoms including skin lesions, axillary mass and improvement in her diarrhea and growth as well as objective improvements in her tryptase levels.

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    (30) Submission ID#590027

    Two-year-old Male with Recurrent Cervical Lymphadenopathy Presenting with Rash

    Deborah Bloch, MD1, Meera Patrawala, MD2, Jennifer Shih, MD3, Whitney Sherry, MD4, Shelley Caltharp, MD5, Adina Alazraki, MD6, Jonathan Loewen, MD7, Christina Rostad, MD8,

    1Pediatric Infectious Diseases Fellow (PGY-6), Emory University School of Medicine

    2Allergy and Immunology Fellow, Emory University School of Medicine

    3Assistant Professor of Pediatrics and Internal Medicine, Department of Pediatrics, Division of Allergy and Immunology, Emory University School of Medicine

    4Assistant Professor of Pediatric Hospital Medicine, Department of Pediatrics, Emory University School of Medicine

    5Adjunct Assistant Professor, Department of Pathology and Laboratory Medicine, Emory University School of Medicine

    6Associate Professor, Departments of Pediatrics and Radiology, Division of Pediatric Radiology, Emory University School of Medicine

    7Assistant Professor, Departments of Pediatrics and Radiology, Division of Pediatric Radiology, Emory University School of Medicine

    8Assistant Professor, Department of Pediatrics, Division of Pediatric Infectious Diseases, Emory University School of Medicine

    Case report: A two-year-old male presented to the hospital with a painful, non-pruritic facial and groin rash. The rash started one week prior to presentation. He had no associated fevers. His history was remarkable for failure to thrive (FTT) and chronic bilateral leg pain with antalgic gait. Over the preceding months, he had been diagnosed with hand-foot-mouth disease and varicella. He had also had recurrent cervical lymphadenopathy (LAD) for greater than one year requiring incision and drainage. Gram stain and Gomori Methenamine-Silver Nitrate Stain (GMS) were negative and pathology showed only acute and chronic inflammation with areas of necrosis. His family history was negative for autoimmune disease or immunodeficiency. Infectious exposure history was significant for an incarcerated father with unknown tuberculosis status and history of living in a shelter.

    On physical examination, the patient was well appearing with multiple erythematous papules, with superficial erosions and scabbing on the face (Figure 1), lower abdomen, genital area, buttocks and proximal lower extremities. He had large, firm, non-tender submandibular lymph nodes. He also had small palpable axillary and inguinal lymph nodes bilaterally.

    His laboratory workup revealed normal white blood cell and platelet counts, but microcytic anemia, an erythrocyte sedimentation rate of 140 mm/hr, and C-reactive protein of 7.5 mg/dL. Full body magnetic resonance imaging (MRI) revealed bilateral cervical, supraclavicular, right hilar and inguinal lymphadenopathy and a patchy right upper lobe consolidation with at least one small area of cavitation (Figure 2) and an adjacent smaller area of ring enhancement. It also revealed three small nonspecific hypodense foci within the right lobe of the liver and borderline splenomegaly. Given these findings, there was concern for granulomatous diseases. The patient underwent a liver biopsy (Figure 3) which showed non-specific evidence of necrotizing granulomatous disease. Microbiological cultures and stains for bacteria, acid-fast bacilli and fungi were negative. His infectious work-up was negative for HSV, tuberculosis, HIV, syphilis, histoplasmosis, and toxoplasmosis. Superficial bacterial cultures from the face and groin grew mixed gram positive and negative organisms, including methicillin-susceptible Staphylococcus aureus (MSSA).

    His immunologic workup revealed borderline elevated IgA and IgG with normal IgM, normal T,B, NK-cell counts and pneumococcal and tetanus titers. A dihydrorhodamine (DHR) flow cytometric test was positive, consistent with a diagnosis of chronic granulomatous disease (CGD). Genetic testing confirmed X-linked disease.

    He was treated with acyclovir and ceftriaxone with resolution of his rash.

    Conclusion: We present a case of a two-year-old male with newly diagnosed X-linked CGD. Though he had been seen by multiple healthcare providers for recurrent lymphadenopathy over the preceding year, he had no other history of recurrent viral or bacterial infections or significant family history that might implicate a primary immunodeficiency. At time of presentation, he had diffuse rash which could have caused his palpable lymphadenopathy on exam. A high index of suspicion for CGD in the setting of recurrent LAD and FTT prompted sending the DHR, which led to the diagnosis.

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    Figure 1. Facial rash. Patient had similar rash in lower abdomen, genital area, buttocks and proximal posterior lower extremities. Bilateral submandibular fullness can also be appreciated.

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    Figure 2. Axial post gadolinium images showing abscess within a right submandibular lymph node (arrow) and consolidation in the right upper lobe with central cavitation (*)

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    Figure 3. Liver biopsy revealing palisading histiocytes surrounding central necrosis, consistent with necrotizing granulomas (400x).

    (31) Submission ID#590402

    Quality of Life in Adult Patients with Chronic Granulomatous Disease

    Samantha Kreuzburg, BA, RN1, Jennifer Treat, PA-C, MSHS2, Dawn Shaw, MBA, MN, RN3, Christa Zerbe, MD, MS4

    1Research Nurse Specialist, National Institutes of Health/National Institute of Allergy and Infectious Diseases

    2Physician Assistant, Medical Science & Computing

    3Protocol Nurse Coordinator II, Leidos Biomedical Research, INC

    4Director, Clinical Patient Services, National Institutes of Health/National Institute of Allergy and Infectious Diseases

    Chronic Granulomatous Disease (CGD) is an inherited primary immunodeficiency (PID) which results in both inflammatory response dysregulation and an increase in susceptibility to certain bacterial and fungal infections. Without curative treatment such as a bone marrow transplant, it remains a chronic disease with daily medication management, intermittent treatment and life-long surveillance. In general, chronic disease involves physical, psychological and social effects which can affect the patients quality of life. Although some research has been done on how PID affects quality of life, there is little research in the United States about how CGD affects patients quality of life.

    To examine the effect of CGD on patients quality of life, as a part of a voluntary research protocol examining the natural history of immune deficiencies, we administered the WHO QOL-BREF instrument to adult CGD patients enrolled on a NIH IRB approved protocol and seen in the Infectious Disease Clinic at the National Institutes of Health (NIH) over a five-month period. The WHO QOL-BREF is comprised of 26 items, which measure the following broad domains: physical health, psychological health, social relationships and environment. Each item is rated on 5-point Likert scale. It has been validated cross culturally and has been widely field tested. The survey was interview administered to 35 patients (23 males, 12 females) with genetically confirmed CGD. The age range was 18 - 60 years old (mean age 37.6 years) with a distribution of 57 % x-linked CGD and 43% autosomal recessive CGD.

    Results have been obtained and will be presented.

    (32) Submission ID#590834

    Health Disparities in CVID: a Report of 1,546 Patients from the USIDNET Registry

    Pragya Shrestha, MD1, Anthony Donato, MD, MHPE2, Avni Joshi, MD, MSc3

    1Resident Physician, Reading Hospital- Tower Health System

    2Associate Program Director, Reading Hospital- Tower Health System

    3Assistant Professor, Mayo Clinic, Rochester, MN

    Rationale: Common Variable immunodeficiency (CVID) is the most common primary immunodeficiency with an estimated prevalence of 1:25,000. We aimed to analyze the clinical presentations and their associated comorbidities amongst CVID patients in USA.

    Methods: Data on 1,546 CVID patients reported in the United States Immunodeficiency Network (USIDNET) from 1992 to 2018 were analyzed based on clinical, immunological and genetic factors. Univariate analysis with Spearman rank coefficients was done to analyze correlations between disease outcomes. Observed survival was estimated using the Kaplan-Meier method.

    Results: Among the 1546 patients, 908 (58.7%) were female and 638 (41.3%) were male. Median age at diagnosis was 29 years [mean (SD), 30.1 (20.2); range, 0-82; IQR, 12-47] with median age of onset of 14 years (mean (SD), 20.3 (19.2); range, 0-81; IQR, 3-33). Females showed a longer delay in diagnosis (9.5 vs. 6.6 years, P=0.006). Higher body mass index (BMI) linearly correlated with the age of diagnosis (r= 0.46). In survival analysis, a 5-year delay in age at diagnosis increased the risk of death by 7.4% (HR: 1.07, 95% CI: 0.98-1.18, p=0.14).

    Conclusions: Our study suggests a longer delay in diagnosis in female subjects and a strong association with diagnosis of CVID in patients with higher BMI. Females may have a longer period without symptoms leading to a diagnostic delay. Gender- based and disparities-based inquiry into these trends may need additional study.

    Demographics Number of patients % USIDNET CVID cohort 95%Confidence Interval
    Gender:    
    Male 638 41.3% 0.39-0.43
    Female 908 58.7% 0.56-0.60
    Age group:    
    <10 years 16 1% Median age 50 years
    10-20 years 188 12% Mean 47.6 years
    20-35 years 300 19% CI 46.6-48.74 for mean
    35-55 years 415 27%  
    >=55 years 627 41%  
    Race:    
    Caucasian 1283 96% 0.95-0.97
    Others (African-American, Asian, Hispanic) 34 3% 0.01-0.03
    Unknown   1% 0.05-0.01
    Living 1415 91.5% 0.95-0.97
    Deceased 55 3.5% 0.03-0.02
    Lost follow up 76 5%  
    Family history of PIDD    
    Yes 176 11.38% 0.20-0.26
    No 575 37.2% 0.73-0.79
    Unknown 795 51.4%  

    Table 1: Demographics of CVID patients in USIDNET registry including gender, age groups, race, living status, family history of Primary Immunodeficiency

      Female (N=908) Male (N=638) p- value
    BMI    0.00122
     N 567 364  
     Mean (SD) 26.2 (7.4) 24.5 (6.5)  
     Median 24.9 23.7  
     Q1, Q3 21.3, 29.8 19.3, 28.7  
     Range (11.6-60.2) (13.7-52.8)  
    Age of onset    <0.00012
     N 545 436  
     Mean (SD) 23.3 (19.7) 16.6 (17.8)  
     Median 19.0 10.0  
     Q1, Q3 5.0, 37.0 2.0, 27.5  
     Range (0.0-81.0) (0.0-79.0)  
    Years from onset to diagnosis    0.00662
     N 520 405  
     Mean (SD) 9.5 (12.9) 6.6 (9.8)  
     Median 4.0 3.0  
     Q1, Q3 1.0, 13.0 1.0, 8.0  
     Range (0.0-64.0) (0.0-72.0)  

    Table 2. Characteristics of Interest- BMI, Age of Onset and Years from onset to diagnosis based on gender among CVID patients in USIDNET registry

    (33) Submission ID#591345

    Indications of Depressive Disorders in Adults with Primary Immunodeficiency

    Christopher Scalchunes, MPA1, Tiffany S. Henderson, PhD2

    1Vice President of Research, Immune Deficiency Foundation

    2Survey Research Analyst, Immune Deficiency Foundation

    The physical well-being of those with primary immunodeficiency (PI) and the physical maladies of those with PI are well-documented. Since the 1950s, advances in identification and treatment of PI has for many led to lives where the physical infections of these groups of diseases are manageable. However, not as well understood are the emotional and mental health aspects of living with PI. As part of a larger survey project The IDF 2017 National Patient Survey, this study aims to quantify any potential mental health issues or challenges faced by adults with PI. Our hypothesis- those with PI, suffer from statistically higher rates of depression when compared to the U.S. general population.

    The 2017 IDF National Patient Survey was a nationally distributed, un-incentivized, mail-based survey of 4,500 persons in the IDF patient database identified as being either adults with PI or the parent/caretaker of a child with PI. The questionnaire comprised approximately 44 main questions about PI as well as the validated SF-12v2, Brief Fatigue Inventory and the Patient Health Questionnaire-2 (PHQ-2) instruments. Additional questions asked about current use of prescription medications for anxiety, depression, stress and pain. For the purpose of this study, only adult respondents with PI are included as the basis for analysis.

    The two-item Patient Health Questionnaire (PHQ-2) meets the criteria for general screening of depression suggested by the U.S. Preventive Services Task Force. Scored on a scale of 0-6, a score of three or higher is suggested as the cut-point for depressive screening. According to a 2014 AHRQ study that utilized MEPS data, 2,139 of the 23,770 (9%) respondents scored three or greater. In our survey 211 of the 925 (23%) adults scored three or greater (2 <.05.)

    Overall, those in our survey scored lower on the SF-12v2 MCS scale when compared to the U.S. population (44.3 v.50.0, p<.05). Further, adults with PI who scored three or higher on the PHQ-2 had an average MCS of 31.8.

    Those who met the PHQ threshold in our survey were also more likely to report moderate to severe limitations in normal activities as a result of emotional problems than those that fell below the threshold (74% versus 13%, p <.05).

    Not surprisingly, those that met the PHQ threshold reported much higher use of prescription medications for anxiety, depression, stress (69% versus 33% below threshold, p <.05) as well as a higher reported use of prescription pain medications (33% versus 17% below threshold, p <.05). Though moderate to severe fatigue was reported by 68% of those below threshold, 99% of those with PHQ scores at threshold reported experiencing moderate to severe fatigue (p <.05).

    Health care providers should consider including the PHQ-2 in the overall health assessments of their patients with PI. Those scoring three or higher should be referred to the appropriate professional for further evaluation.

    (34) Submission ID#592136

    DLCO Is a Reliable Noninvasive Approach for Pulmonary Monitoring in Patients with HIES

    Alyssa Kerber, MD1, Avni Joshi, MD2

    1Resident physician, Pediatric and Adolescent Medicine, Mayo Clinic

    2Assistant Professor, Allergy and Immunology, Mayo Clinic

    A 23-year-old female from Kuwait presented to our multi-disciplinary Primary Immunodeficiency Clinic, for a history of several years of eczema, multiple skin abscesses, recurrent sinusitis, and history of pneumonia complicated by pneumatocele. Based on patient history and outside records, she had been admitted multiple times for cutaneous abscesses on her abdomen, thigh, and gluteal regions, requiring multiple episodes of antibiotics and incision and drainages.

    The patients symptoms first started early on in her first year of life, with refractory eczema and recurrent cutaneous infections. During her second year of life, she had recurrent episodes of respiratory infections and was diagnosed with pneumonia complicated by pneumatocele formation, which required surgical intervention. Her medical history is also notable for fungal infections of the nails and multiple episodes of otitis media. There is no family history of recurrent infections or known immunodeficiency.

    In 2012 at the age of 17, she was diagnosed with clinical hyper IgE syndrome with eosinophilia and IgE level above 5000 KU/L.

    At the time of evaluation at our institution, examination did not reveal evidence of active infection. Her examination was significant for multiple scars to her upper and lower extremities and gluteal region from previous incision and drainages. She had a left thumb nail onychomycosis. She was also noted to have a broad nasal bridge and retained primary teeth.

    A CT scan of the chest was obtained, which showed fibrosis and volume loss with underlying bronchiectasis in the right lung. Pulmonary function testing revealed a mild decline in DLCO at 15.99 ml/(min*mmHg).

    The patients calculated NIH-HIES score was >60. Due to clinical diagnosis of Hyper-IgE, genetic testing was pursued, which revealed a likely pathogenic variant, p.Trp623Leu (W623L) (TGG>TTG): c.1868 G>T in exon 20 of the STAT3 gene. The W623L missense variant in the STAT3 gene has been previously observed in one individual in association with Hyper-IgE syndrome (Al-Mousa et al., 2016). This variant is not observed in large population cohorts (Lek et al., 2016). The W623L is a semi-conservative amino acid substitution, which may impact secondary protein structure. In-silico analyses supported a deleterious effect, located within the SH2 domain, which is a critical functional domain (Chandesris et al., 2012; Koskela et al., 2012). It was thus determined that this variant is likely pathogenic.

    The patients prophylactic treatment was optimized with TMP-SMX (800mg-160mg) twice daily for prevention of infections. She was also started on Hibiclens (chlorhexidine) baths once per week. She was referred to Pulmonology for optimization of pulmonary health in the setting of bronchiectasis and mild decline in DLCO. She was advised to follow-up on a yearly basis to the Primary Immunodeficiency Clinic to assess for recurrent infections and for changes in pulmonary health. Finally, targeted testing and clinical evaluation of both of the patients parents was recommended to determine if W623L was inherited or arose de novo. The pathogenic role of the W623L missense change would be further supported if it had occurred de novo or if it segregates with the disease in the family.

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    (35) Submission ID#592269

    The Effect of Hydroxychloroquine on CTLA4 Expression in Siblings with LRBA (Lipopolysaccharide-responsive and Beige-like Anchor Protein) Deficiency

    Nurcicek Padem, MD1, Melanie Makhija, MD2, John Routes, MD3, Jeffrey Woodliff, PhD4, Amer Khojah, MD2

    1Fellow, Northwestern University

    2Attending, Northwestern University

    3Chief, Professor, Division of Allergy and Immunology, Children's Hospital of Wisconsin-Milwaukee, Milwaukee, WI

    4Director, Flow Cytometry Core Facility, Medical College of Wisconsin

    Introduction: Lipopolysaccharide-responsive and beige-like anchor protein (LRBA) deficiency is a rare autosomal recessive disease of the immune systems characterized by hypogammaglobulinemia and decreased CTLA4 expression on T regulatory cell (T regs) due to defective intracellular trafficking of CTLA4. Previous in vitro study has shown a significant increase of CTLA4 expression on LRBA deficient T cells after overnight culture with chloroquine, an older anti-malarial agent. This effect is likely due to increasing lysosomal pH. However, there is no evidence of such effect in human subjects after administration of weight appropriate doses anti-malarial agents. We are presenting a set of siblings with LRBA deficiency who had CTLA4 expression measured before and four weeks after starting hydroxychloroquine.

    Case reports: Case 1 is a 14-year-old East-Indian boy with autoimmune thyroiditis, Type 1 diabetes mellitus (DM), short stature, autoimmune cytopenias, and lymphadenopathy. He was referred to immunology clinic at 9 years of age for suspicion of Autoimmune Lymphoproliferative Disorder. Primary Immunodeficiency Genetic Panel was sent which revealed a homozygous mutation in LRBA gene (c.6480_6481del). This novel variant resulted in a frameshift and created a premature stop codon 18 amino acids downstream from this location which may lead to absent or abnormal protein. Lung CT scan showed interstitial lung disease. Lung biopsy showed interstitial nodular and diffuse lymphoid proliferation. This diagnosis led to the testing of his sister (case 2) given her history of autoimmune illnesses and the family history of consanguinity. Case 2 is a now 13-year-old girl with type 1 DM, autoimmune thyroiditis, lymphadenopathy, psoriatic arthritis, and seizures. Her lung imaging showed pulmonary nodules without interstitial lung disease. Both cases received hydroxychloroquine while waiting for insurance approval of abatacept. CTLA4 expression on Tregs was measured prior to and four weeks after starting hydroxychloroquine treatment. At baseline, 8.6% of Case 1s CD4 cells were Treg (FOXP3+ve, CD25hi) and 51.4% of them expressed CTLA-4 (in contrast to 94.1% Tregs in the healthy control) with mean fluorescence intensity (MFI) of 335. This ratio and MFI did not change after 4 weeks of hydroxychloroquine treatment (6 mg/kg/day). Soluble Interleukin-2 receptor levels were measured: Case 1 had a baseline level of 8510 pg/mL, which decreased to 2228 pg/mL after 4 weeks of hydroxychloroquine treatment. For Case 2: 8.4% of her CD4+ T cells were found to be FOXP3+CD25hi and 36.1% of these Tregs expressed CTLA-4. This ratio increased by 7% after one month of hydroxychloroquine. Increase in MFI was also noted from 298 to 386. Case 2 had a drop in soluble Interleukin-2 receptor level from 1265 pg/mL to 950pg/mL after treatment.

    Conclusion: In contrast to the previous in vitro assays, we did not find a significant increase in CTLA4 expression on T regulatory cells in vivo after 4 weeks of 6mg/kg/day hydroxychloroquine. Interestingly, soluble IL-2 receptor levels improved dramatically with hydroxychloroquine.

    (36) Submission ID#592574

    Human NF-kappaB2 Defect Results in Defective Intrinsic B-cell Differentiation, Function and Class Switching

    Shancy Jacob, PhD1, Julie Feusier, MSc2, Krystin Krauel, PhD3, Li Guo, MD/PhD3, Jesse Rowley, PhD4, Robert Campbell, PhD5, Jacob Anderson, BS6, Michael D. Keller, MD7, Lynn Jorde, PhD8, Guy Zimmerman, MD/PhD9, Andrew Weyrich, PhD9, Karin Chen, MD10

    1Postdoctoral Research Fellow, Division of Allergy & Immunology, Department of Pediatrics, University of Utah

    2Graduate Student, Department of Human Genetics, University of Utah

    3Postdoctoral Research Fellow, Department of Internal Medicine, University of Utah

    4Assistant Professor, Division of Pulmonary Medicine, Department of Internal Medicine, University of Utah

    5Assistant Professor, Division of General Medicine, Department of Internal Medicine, University of Utah

    6MD Candidate, Pennsylvania State University College of Medicine

    7Assistant Professor, Center for Cancer and Immunology Research, Children's National Health System, Division of Allergy & Immunology, Children's National Health System, Washington, DC

    8Professor, Department of Human Genetics, University of Utah

    9Professor, Department of Internal Medicine, University of Utah

    10Assistant Professor, Department of Pediatrics, Division of Allergy and Immunology, University of Utah

    Introduction/Background: Autosomal dominant heterozygous mutations in NFKB2 (encoding for the protein NF-kB2) have been identified in the etiology of a form of primary immunodeficiency disorder that presents with hypogammaglobulinemia, defects in B-cell maturation, endocrinopathy, and autoimmune manifestations. In humans, the effects of altered NF-kB2 and mechanisms of immune system impairment have not been fully delineated.

    Objectives: To understand the mechanism of the antibody deficiency in patients with hypomorphic mutations in NFKB2 (c.2564delA; p.Lys855Serfs*7) by evaluating B-lymphocyte proliferation, differentiation, function, and gene expression.

    Methods: Immunophenotyping of primary B-cells from subjects with mutant NFKB2 was completed by flow cytometry. Proliferation of B-cells was assessed by CFSE stimulation of primary CD19+ B-cells from healthy and NFKB2 mutant subjects. Differentiation of healthy and affected naïve B-cells (CD27-CD38-) into plasmablasts (CD27+CD38+) following stimulation was assessed by flow cytometry. The supernatant from these cells were assayed for IgA, IgG and IgM production by ELISA. To study the defect in class-switch recombination, naïve B-cells and EBV-transformed B-cells from affected and healthy individuals were stimulated and expression of the AICDA gene was quantified by qPCR. In parallel experiments, EBV B-cells from wildtype and NFNB2 mutant individuals were stimulated and AID (Activation-induced cytidine deaminase) protein levels were determined by western blot.

    Results: Patients with hypomorphic mutations in NFKB2 (c.2564delA) had low memory B-cell (CD19+ CD27+ IgD- IgM+) and class-switched memory B-cell (CD19+ CD27+ IgD- IgM-) numbers. In vitro, primary B-cells from these patients demonstrated a 50% reduction in proliferation and cell division in response to CD40L and IL-10 (p =0.01). Compared to healthy naïve B-cells, mutant naïve B-cells had a significant reduction in plasmablast differentiation (p = 0.002) and secreted significantly lower levels of immunoglobulins in response to CD40L and IL-21 stimulation. Mutant naïve B-cells and mutant EBV B-cells failed to increase AICDA expression and AID protein levels in response to CD40L and IL-21 stimulation.

    Conclusions: Our studies demonstrate that a hypomorphic NFKB2 mutation in humans affects intrinsic B-cell proliferation and differentiation. The mutation impairs transcription of the AICDA gene that encodes AID, a key protein involved in B-cell class-switch recombination. The NFKB2 gene defect also impairs immunoglobulin production, as seen in common variable immunodeficiency-like cases. These studies provide unique translational insights into physiological activities of NF-kB2 in downstream immunologic outputs in humans, expanding those suggested by experimental observations in mice.

    (37) Submission ID#592712

    Provider Perceptions of Primary Immunodeficiency Disease Patients Quality of Life, Neurocognition, Physical Well-Being and Psychosocial Health

    Thomas F. Michniacki, MD1, Lauren E. Merz, BA2, Roshini S. Abraham, PhD3, Julie Sturza, MPH4, Kelly Walkovich, MD5

    1Pediatric Hematology/Oncology Fellow, University of Michigan

    2Medical Student, University of Michigan Medical School

    3Department of Pathology and Laboratory Medicine, Nationwide Childrens Hospital, Columbus, OH.

    4Statistician, University of Michigan Department of Statistics

    5Associate Professor, Pediatric Hematology/Oncology, University of Michigan Medical School

    Background: Few studies have evaluated the quality of life (QOL) and patient reported outcomes of primary immunodeficiency disease (PIDD) patients, and no studies have assessed medical provider perceptions of their PIDD patients QOL, neurocognition, physical well-being and psychosocial health. Understanding provider beliefs regarding patient reported outcomes is essential to improving clinical management of PIDDs. Here we report our PIDD medical provider survey results.

    Methods: Providers were contacted via email with the assistance of the Clinical Immunology Society. Participants completed adult and/or pediatric-based Likert scale survey questions via a secure online survey service. In addition to demographic information, survey questions assessed provider perceptions of patients overall QOL and their impression of the impact of disease or its associated treatment on mental health, physical well-being, neurocognition, social relationships and school/work performance. Clinicians were expected to make their assessments based on their PIDD patient cohort as a whole rather than on specific diagnoses or patients. Given the small sample size, a p-value < 0.1 was considered statistically significant; repeated measures ANOVA and paired t-test analyses were used.

    Results: Study participants (n=58) were primarily from the United States (64%), born between 1965-1979 (44%), and trained in allergy/immunology (77%). 85% of survey takers practiced within an academic center, 52% were female and 95% cared for children with 42% of providers concurrently caring for adults. There was a statistically significant difference (p=0.07) in the perceived overall QOL of pediatric versus adult PIDD patients with 41% of providers feeling as though their pediatric patients had a good QOL while only 25% believed their adult patients had a good QOL. Clinicians believed adult PIDD individuals had more difficulties related to associated co-morbidities rather than their actual PIDD compared to pediatric PIDD patients (p=0.046). Providers felt that the neurocognition and school performance of children were more often negatively affected by a PIDD than the neurocognition and work performance of immunodeficient adults (p=0.1). Clinicians believe children with PIDD more frequently had difficulties related to their concentration than memory (p<0.01). 96% of those who care for PIDD adults believe their patients work performance or daily mental functioning is at times negatively impacted. Anxiety symptoms and social relationships were viewed as being more negatively impacted by a PIDD diagnosis or treatment than anger or depressive symptoms in both children and adults (p<0.01). 38% of pediatric clinicians feel their PIDD patients experience anxiety symptoms often or almost always. Of physical health parameters, energy, rather than mobility or pain, was deemed to be more deleteriously influenced by an immunodeficiency in adult and pediatric patients (p<0.01).

    Conclusions: Our results show that medical providers perceive the overall QOL of pediatric PIDD patients to be superior to that of adults with PIDD, but most clinicians feel a diagnosis or associated treatment regimen for PIDD can negatively impact the physical well-being, psychosocial health, school/work performance and neurocognition of both children and adults.

    (38) Submission ID#592868

    Homozygosity for a Novel CARD11 Mutation Causes Severe Combined Immunodeficiency (SCID), Inflammatory Gastrointestinal Disease, and Complete Abrogation of MALT1 Activity

    Henry Y. Lu, BSc1, Sneha Suresh, MD, FRCPC2, Lyle McGonigle, MD, FRCPC3, Joanne Luider, BSc, ART, MLT4, Stuart E. Turvey, MBBS, DPhil, FRCPC5

    1PhD Candidate, BC Children's Hospital and University of British Columbia

    2Assistant Professor, University of Alberta and Alberta Health Services

    3Pediatrician, Alberta Health Services

    4Laboratory Scientist, Calgary Laboratory Services

    5Professor, BC Children's Hospital and University of British Columbia

    Introduction/Background: The caspase recruitment domain family member 11 (CARD11)B cell CLL/lymphoma 10 (BCL10)MALT1 paracaspase (MALT1) [CBM] complex is a critical signalling adaptor that regulates lymphocyte activation, proliferation, survival, and metabolism. Primary immunodeficiencies affecting each component (termed CBM-opathies) result in broad clinical manifestations ranging from severe combined immunodeficiency (SCID) to lymphoproliferation. We present the laboratory and clinical findings of two Canadian First Nations patients found to be homozygous for the same novel CARD11 mutation (c.2509C>T; p.R837*).

    Results: We have identified an 8-month-old boy who presented with a severe case of entero/rhinovirus bronchiolitis with interstitial lung disease and a 17-year-old boy with a history of severe pulmonary infections (including PJP), chronic sinusitis, candidiasis, invasive bacteremia, and severe ileo-colitis and oral ulceration requiring total colectomy. Both patients possessed absent Tregs, absent memory B cells, and hypogammaglobulinemia. However, only the 8-month-old had poor T cell proliferation to PHA, ConA, and CD3. Both patients were found to be homozygous for the same novel variant of CARD11 (c.2509C>T; p.R837*). The mutation rendered CARD11 protein expression unstable and it was undetectable by immunoblot. To confirm CARD11 deficiency, we stimulated patient B cells with phorbol 12-myristate 13 acetate (PMA) and ionomycin across a time-course and immunoblotted for various signalling proteins in both the NF-B (IKK/, IB, p65) and MAPK (MEK1/2, MKK4, JNK1/2, ERK1/2) pathways as well as various cleavage substrates of the MALT1 paracaspase (RelB, CYLD, BCL10, HOIL1). NF-B and JNK activation were completely absent and MALT paracapase activity was lost, but surprisingly, MKK4 (which acts upstream of JNK) was intact. Furthermore, co-immunoprecipitation experiments revealed that CARD11 was required for optimal MALT1 association with BCL10 in response to stimulation.

    Conclusions: These two cases highlight the crucial role of CARD11 in regulating lymphocyte development, function, and humoral responses. In addition, we have identified the oldest known living individual with CARD11 deficiency and he presented uniquely with inflammatory gastrointestinal disease in addition to SCID, further adding to the spectrum of phenotypes associated with CARD11-related primary immunodeficiencies.

    (39) Submission ID#593164

    NIH Participation to USIDNET Registry (Poster Submission)

    Elizabeth K. Garabedian, MSLS, RN1

    1Research Nurse, Principal Investigator, National Genome Research Institute, National Institutes of Health

    Abstract: The USIDNET Registry began in 1992 with an NIAID contract with the Immune Deficiency Foundation, which continues today. It aims to provide a resource for clinical and lab research through enrollment of known immunodeficiency patients into a national registry, the USIDNET. NIH is a major national and international referral center for clinical trials on inborn errors of immunity, or primary immunodeficiency diseases. It is a mechanism for depositing NIH data into USIDNET. A Registry of patient information may help us understand how many people have each disease. The information may improve how we diagnose and treat these conditions. The patient Registry is designed to obtain longitudinal data on a large number of patients with primary immunodeficiency diseases who come to NIH to participate in research. The data is collected from the NIH electronic medical record system, CRIS and is deposited into a secure registry with restricted and monitored access. All medical information is anonymized for patient privacy.

    (40) Submission ID#594012

    Heterozygous OAS1 Mutations Cause Spontaneous RNA Cleavage and Apoptosis with Resulting Multisystem Inflammation and Immunodeficiency

    Heimall, J1, Magg, T2, Sullivan, KE1, Albert, M2, Griese M2, Conway D3, Gray, PE4, Calderon, B5, Conn, G5, Klein, C2, Hauck, F2

    1Allergy Immunology, The Children’s Hospital of Philadelphia, Philadelphia, PA

    2Department of Pediatrics, Dr. von Hauner Children’s Hospital, University Hospital, LMU Munich, Germany

    3St. Christopher’s Hospital for Children, Philadelphia, PA

    4Dept. of Immunology and Infectious diseases, Sydney Children’s Hospital, Sydney, Australia

    5Department of Biochemistry, Emory University, Atlanta, GA

    OAS1 is an intracellular sensor for dsRNA that generates the second messenger 2'-5'-oligoadenylate to activate RNase-L as a means of antiviral defense. We describe four patients with a complex early-onset autoinflammatory and immunodeficiency disease caused by heterozygous de novo OAS1 mutations.

    Patients presented early in life with lung inflammation including pulmonary alveolar proteinosis and interstitial lung disease. They had febrile flares with dermatitis specifically with macular, pustule and bullous features often progressing to ulceration. Infants had episodes of bloody diarrhea in 3 patients (assoc. with villous blunting and cryptitis in two patients and oesophagitis in one patient). Immunoglobulin IgM, IgG, and IgA levels were low while T cell, B cell, and NK cell numbers were generally in the normal range. Exome sequencing identified de novo heterozygous OAS1 missense mutations in all patients.

    One patient had a heterozygous de novo OAS1 mutation p.Ala76Val, with mutant OAS1 protein being expressed in ex vivo generated T cell blasts. In sorted primary patient monocytes and B cells, OAS1 p.Ala76Val was associated with spontaneous RNA degradation and apoptosis as determined by RNA chip technology and flow cytometry, respectively, while T cells were not affected. Monocytes displayed disturbed terminal differentiation and functioning as indicated by reduced GM-CSF-R expression and signaling. B-cells display reduced class-switch-recombination. Proliferation of allogeneic T-cells was reduced in response to sorted OAS1 mutated monocytes and B-cells. Activation of interferon response genes in PBMCs was detected.

    Two further unrelated patients had a heterozygous de novo OAS1 mutation p.Cys109Tyr, which appeared to compromise protein stability in transformed patient fibroblasts and when transfected. Cells transfected with this mutant protein had reduced 2-5 oligoadenylate synthesis compared to wild type transfected cells. Immortalized fibroblast lines demonstrated higher levels of inflammatory cytokines and spontaneous cleavage of RNAs. A 4th patient with the clinical phenotype had a heterozygous de novo OAS1 variant p.Val121Gly, but has yet to have formal validation of the variant.

    Three patients underwent hematopoietic stem cell transplants in an effort to control their diarrhea and skin inflammation. One patient died with ongoing chronic graft versus host disease, while the two others (p.Ala76Val, Cys109Tyr) are alive and reasonably well with a follow-up of 0.5-7 years. The untransplanted patient died as a result of respiratory failure.

    In summary, patients with de novo heterozygous OAS1 mutations have chronic ongoing inflammation of multiple organs. This is at least in part due to spontaneous RNA cleavage, apoptosis and production of inflammatory cytokines and type I interferons. This defines a new category of autoinflammatory disorder.

    (41) Submission ID#594077

    Thinking Outside of Infection: Hemophagocytic Lymphohistiocytosis in a 5-week-old Male with Chronic Granulomatous Disease and Burkholderia Cepacia Sepsis

    Jacqueline Squire, MD1, Wil Chamizo, MD2, David M. Berman, DO3, Deepak Chellapandian, MD4, Beatriz Teppa, MD5, Laura Vose, DO6, Jennifer Leiding, MD7

    1Allergy and Immunology Fellow, USF - John Hopkin's All Children's Hospital

    2Medical director for Pathology and Laboratory medicine, Johns Hopkins All Childrens Hospital.

    3Pediatric Infectious Disease, John Hopkins All Children's Hospital

    4Bone Marrow Transplant, Johns Hopkins All Childrens Cancer & Blood Disorders Institute

    5Pediatric Intensive Care (PICU), John Hopkins All Children's Hospital

    6Pediatric Intensive Care (PICU). Co-chair of the Human Values and Ethics committee, John Hopkins All Childrens

    7Associate Professor, University of South Florida

    Introduction: Increased susceptibility to infections is the most common complication of chronic granulomatous disease (CGD). Hemophagocytic lymphohistiocytosis (HLH) is a severe disorder resulting from hyperinflammation and hypercytokinemia that can lead to multi-organ system dysfunction (1) characterized by certain criteria: fever, splenomegaly, cytopenias, hypofibrinogenemia or hypertriglyceridemia, hyperferritinemia, increased soluble CD25/IL-2Ra, evidence of hemophagocytosis, or decreased/absent NK cell cytotoxicity (2). Secondary HLH occurs infrequently but often is preceded by smoldering infection in CGD (3,4,5). We present a case of HLH in a 38-day old male, the youngest reported case with CGD.

    Case: A 38-day old male with previously diagnosed X-linked CGD, due to known family history, presented with fevers. Initial evaluation was unrevealing including chest x-ray, urinalysis, and blood and CSF cultures. He was admitted and treated empirically with cefepime. CT demonstrated multiple multifocal nodules of the lungs and spleen. After lung nodule biopsy was performed, antimicrobial therapy was broadened to IV meropenem, voriconazole, and micafungin. Despite this, he continued to have fever and developed new onset tachycardia, respiratory distress, and lactic acidosis. Further decompensation with vasoactive refractory shock was treated with vasopressors and stress dose hydrocortisone. Additional laboratory evaluation revealed rising liver enzymes (AST 1670u/L, ALT 307u/L), cytopenias (hemoglobin 7 g/dL, ANC 90/uL, platelets 96,000/uL), and coagulopathy (fibrinogen 93-135mg/dL). Splenomegaly was present on abdominal ultrasound. A diagnosis of evolving HLH was considered and dexamethasone was administered. Within 24 hours of clinical decompensation, the patient died of multiorgan failure. Subsequent blood cultures returned with gram-negative rods (and ultimately Burkholderia cepacia). Autopsy confirmed hemophagocytosis within the bone marrow. No mutations were found in genes associated with primary HLH.

    Discussion: Patients with CGD are susceptible to infectious complications and auto-inflammation most commonly involving the lungs, GI, and GU systems (6,7). Patients with CGD can be at increased risk of hyperinflammatory syndromes secondary to infections and chronic inflammation. As shown in the included case, HLH can present in infancy and can be deadly. Early consideration and directed treatment of HLH is imperative, even in the setting of sepsis.

    References:

    1. Janka GE, Lehmberg K. Hemophagocytic syndromes An update. Blood Rev. 2014 Jul;28(4):135-42

    2. Henter J, et al. HLH-2004: Diagnostic and Therapeutic Guidelines for Hemophagocytic Lymphohistiocytosis. Pediatr Blood Cancer. 2007 Feb;48(2):124-31.

    3. Bode et al. The syndrome of hemophagocytic lymphohistiocytosis in primary immunodeficiencies: implications for differential diagnosis and pathogenesis. Haematologica. 2015 Jul;100(7):978-88.

    4. Parekh C, Hofstra T, Church JA, Coates TD. Hemophagocytic lymphohistiocytosis in children with chronic granulomatous disease. Pediatr Blood Cancer. 2011 Mar;56(3):460-2

    5. Valentin G, Thomas TA, Nguyen T, Lai YC. Chronic granulomatous disease presenting as hemophagocytic lymphohistiocytosis: a case report. Pediatrics. 2014 Dec;134(6):e1727-30

    6. Henrickson SE, et al. Noninfectious Manifestations and Complications of Chronic Granulomatous Disease. J Pediatric Infect Dis Soc. 2018 May 9;7(suppl_1):S18-S24.

    7. Magnani A, et al. Inflammatory manifestions in a single-center cohort of patients with chronic granulomatous disease. J Allergy Clin Immunol. 2014 Sep;134(3):655-662.e8

    (42) Submission ID#594080

    Benign Reactive Gamma Delta T Cells Proliferation in Spleen The Mirage Effect

    Snegha Ananth, MD1, David J Haile, MD2, Andrea Yunes, MD3

    1Resident, UTHSCSA

    2Attending, Audie L Murphy VA Hospital, San Antonio

    3Pathologist, Audie L Murphy VA Hospital, San Antonio

    Malignant proliferation of Gamma-Delta T cells include hepato-splenic T-cell lymphoma (HSTL), primary cutaneous T-cell lymphoma and T-cell large granular lymphocytic leukemia (T-LGL). The former two have often been associated with splenomegaly and cytopenias. However, reactive proliferation of Gamma-Delta T cells in spleen mimicking malignancy has only been reported once and has a significant risk of misdiagnosis.

    A 30-year-old female presented with two years of unintentional weight loss, persistent leukopenia and thrombocytopenia, with leucocytes around 1-2 X 10^9/L and platelets around 100 X 10^9/L. She also had associated macrocytic anemia (Hemoglobin=10-11g/dl) with laboratory evidence of DAT (Direct Anti-globin Test) negative hemolysis. Physical examination and computed tomography (CT) imaging showed splenomegaly. There was no hepatomegaly or lymphadenopathy. Serum liver function test, auto-immune studies, hemolysis and hereditary diseases workup, viral and bacterial serologies were all normal or negative, except for mild hyperbilirubinemia and LDH elevation. Bone marrow examination performed four months prior to the splenectomy revealed mildly hypocellular marrow (50%) with trilineage hematopoiesis. Flow cytometric analysis and cytogenetics of the bone marrow aspirate and peripheral blood were normal except for small population of large granular lymphocyte and mild low absolute B cell counts in peripheral blood. A laparoscopic splenectomy was performed for diagnostic and therapeutic purposes due to patients worsening LUQ pain. There was no other treatment given prior to surgery. 24 hours post- splenectomy her leucocytes increased to 13.1 and platelets to 247. Her three-month post-splenectomy WBC count and platelet count was 8.9 and 391, respectively. Hemoglobin also improved to 14.9. Pathology showed red pulp expansion by small lymphocytes (Fig. 1) and subsequent IHC (Immunohistochemistry) was positive for CD3 (Fig. 2), CD2, CD7, TIA-1 and negative for CD8, CD5 and CD56. CD4 was difficult to interpret. EBER was negative. Flow cytometry (Fig. 3) showed increased gamma-delta T-cell population (20%) with positive CD3, CD2 and CD 7 and negative CD 5, CD4 and CD 8. Molecular studies by PCR didnt reveal any T-cell receptor gamma or beta gene rearrangement. Cytogenetics was negative for isochromosome 7q or any other abnormalities. She was symptom free at 6 months from her splenectomy.

    The morphology and immuno-phenotype of these Gamma-Delta T cells show significant overlap with the malignant cells seen in HSTL and T- LGL, such as loss or downregulation of CD5, CD4 and CD8. Awareness of this reactive condition is necessary to prevent making a wrong diagnosis of a malignant disease with a potentially benign, spontaneously resolving disease. Additional studies of similar cases is needed in order to establish more definitive criterion to separate benign from malignant processes and delineate the role of Gamma-Delta T cells.

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    (43) Submission ID#594181

    Sexual Dimorphism and AIRE-AIRE Interactors-miRNA Coexpression Networks in the Infant Human Thymus

    Silvia Y. Bando, PhD1, Fernanda B. Bertonha, PhD1, Lucila H. B. Oliveira, PhD2, Carlos Alberto Moreira-Filho, PhD3, Magda Carneiro-Sampaio, MD, PhD4

    1Scientific Researcher, Department of Pediatrics, Faculdade de Medicina da Universidade de Sao Paulo, Sao Paulo, Brazil.

    2Postdoctoral Fellow, Department of Pediatrics, Faculdade de Medicina da Universidade de Sao Paulo, Sao Paulo, Brazil.

    3Associate Professor, Department of Pediatrics, Faculdade de Medicina da Universidade de Sao Paulo, Sao Paulo, Brazil.

    4Full Professor, Department of Pediatrics, Faculdade de Medicina da Universidade de Sao Paulo, Sao Paulo, Brazil

    Background: Sex steroids in the human thymic environment influence AIRE expression as well as interactions with its partners, i.e. genes coding for AIRE interactors. Here we investigated the effects of sex steroids on these interactions during minipuberty the surge of sex hormones that occur along the first six months of life - and up to 18 months of life. We employed a network-based approach for investigating AIRE-interactors gene-gene relationships and how abundantly co-expressed thymic miRNAs covariate with those genes. AIRE-interactors networks allowed the measuring of gender-related differences in gene-gene expression correlation disclosing relevant differences between minipuberty groups.

    Methods: Total RNA was extracted from thymic surgical explants obtained from male (M) and female (F) infants - aged 0-6 months (groups MM and MF, for minipuberty) and 7-18 months (group NM and NF, for non-puberty) and used in DNA microarray assays. Gene coexpression network (GCN) analyses were performed for AIRE and its interactors and for miRNA-gene coexpression analysis. The set of genes coding for the AIRE-targeted proteins was previously identified in TECs by Abramson et al. (Cell 140:123-35, 2010). AIRE-interactors networks were obtained for all groups (link strength cut-off for gene-gene > |0.80| and for miRNA-gene < -0.80). AIRE expression in mTECs was quantified by immunohistochemistry. These methodologies are described in Moreira-Filho et al. (Sci Rep 8:13169, 2018).

    Results: The MM x MF networks comparison showed that 16 abundantly expressed miRNAs are interacting with the different AIRE interactor genes in both networks. It is interesting to note that network topology were more similar between NM and NF groups, although AIRE interacts with only one distinct miRNA in each network (miR-150-5p in the NM group or miR-7977 in the NF group). Conversely, in the non-puberty networks the sets of miRNAs and their interacting genes are distinct for each network. Immunohistochemistry analysis revealed a higher percentage of mTEC AIRE positive cells in the minipuberty groups: i.e. there is a significant difference between MM x NM (p = 0.0006) and between MF x NF (p = 0.0060).

    Conclusions Minipuberty and genomic mechanisms shape thymic sexual dimorphism along the first 6 months of life. This process does not involve changes in AIRE expression between genders, but differences in the interactions of AIRE with its partners that persist throughout the non-puberty period, probably regulated by miRNAs and also by genetic and epigenetic factors.

    FAPESP 2014/50489-9

    (44) Submission ID#595214

    Rescue Therapy with Granulocyte Transfusions (GT) as a Bridge to Hematopoietic Stem Cell Transplant (HSCT) in a Chronic Granulomatous Disease (CGD) Patient with Severe Disseminated Aspergillosis

    Sneha Suresh, MD1, Wendy Vaudry, MD2, Marta Rojas-Vasquez, MD3, Sunil Desai, MD4, Luis Murguia-Favela, MD5, Rashid Alobaidi, MD3, Victor Lewis, MD6, Jean Jacques De Bruycker, MD, FRCPC7

    1Assistant Professor, University of Alberta

    2Professor, University of Alberta

    3MD, University of Alberta

    4Clinical Professor, University of Alberta

    5Clinical Assistant Professor, University of Calgary

    6Associate Professor, University of Calgary

    7Clinical Assistant Professor, CHU Sainte-Justine, University of Montreal

    Introduction: Neutrophils are presumed to defend against Aspergillus species by releasing reactive oxygen species (ROS) and neutrophil extracellular traps (NETs) to degrade fungal hyphae. Triazole antifungals synergistically enhance neutrophil mediated hyphal degradation. Patients with CGD are particularly susceptible to Aspergillus species likely due to their inability to create ROS and NETs, and in severe cases may not be amenable to antifungal therapy alone.

    Objective: We present a case of severe disseminated aspergillosis in a patient with CGD in whom GT served as an important adjunct to antifungal therapy and bridge to transplant.

    Results: A 6-year-old boy with known CGD, lost to follow up and non-adherent to prophylaxis, presented acutely with right-sided hemiparesis. Neuroimaging revealed an embolic left middle cerebral artery infarction and cardiac magnetic resonance imaging showed extensive vegetations involving both right and left ventricles and atria, with an ejection fraction of 28%. The patient was admitted to intensive care, started on liposomal amphotericin B, meropenem and vancomycin, and underwent debulking of the intracardiac masses on post admission day (PAD) 1. Operative findings showed severe constrictive pericarditis with multiple abscesses and intracardiac vegetations. Thorough debridement of the vegetations was undertaken, however some deep seated abscesses in the myocardium were not amenable. Operative cultures were positive for Aspergillus fumigatus. Clinical status remained precarious, with ongoing requirement for inotropic and ventilator support. Antimicrobial therapy was refined to voriconazole, with amphotericin B remaining on board until therapeutic levels of voriconazole were achieved.

    As effective neutrophils are integral in the immune response against Aspergillus, the decision was made to start granulocyte transfusions to aid in clinical stabilization prior to HSCT. Interferon gamma infusions were not administered because of the risks of adverse effects and potentially increasing transplant rejection.

    GTs were started on PAD 6, at a dose of approximately 1X10^10 granulocytes, three times a week. The patient tolerated the infusions well, with no allergic or inflammatory response. Neutrophil oxidative burst measured one hour post infusion showed 23.9% mean fluorescent intensity, compared to a baseline of 0% (Figure 1). Clinical improvement was seen, with inotrope cessation on PAD 12 and extubation to BiPaP on PAD 41.

    Human leukocyte Antigen (HLA) allosensitizaton was tested on PAD 12, 6 days after the first GT, with no evidence of HLA antibodies. A total of 28 GTs were given over 3 months, prior to proceeding to a 10/10 HLA matched related donor transplant (PAD 69), with two transfusions given before neutrophil engraftment (ANC 500) on Day +14. The patient is now stable 13 months post transplant, with no evidence of graft rejection. He remains on chronic suppressive antifungal therapy, to continue until full lymphoid reconstitution.

    Conclusion: GT may be a useful adjunct to antifungal therapy in patients with impaired neutrophil function with severe invasive aspergillosis, and potentially provide a life sustaining bridge to HSCT.

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    (45) Submission ID#595387

    Investigational Cultured Thymus Tissue Transplantation (RVT-802) Following Failed Stem Cell Transplantation in Athymic Patients

    Tyler R. Yates, MD1, Jennifer Puck, MD2, Morna J. Dorsey, MD, MMSc3, Ziad Khatib, MD4, Karin Chen, MD5, Vivian Hernandez-Trujillo, MD6, William Blouin, CPNP7, Ralph Quinones, MD8, Erwin W. Gelfand, MD9, M. Louise. Markert, MD, PhD10

    1Immunology Fellow, Department of Pediatrics, Division of Allergy, Immunology, and Pulmonology, Duke University Medical Center, Durham, NC

    2Pediatric Immunologist, Department of Pediatrics, Division of Allergy, Immunology, and Bone Marrow Transplant, University of California San Francisco, San Francisco, CA

    3Pediatric Immunologist and Allergist, Department of Pediatrics, Division of Allergy, Immunology, and Bone Marrow Transplant, University of California San Francisco, San Francisco, CA

    4Pediatric Hematologist and Oncologist, Department of Oncology, Nicklaus Childrens Hospital/Miami Childrens Health System, Miami, FL

    5Assistant Professor, Department of Pediatrics, Division of Allergy and Immunology, University of Utah

    6Pediatric Immunologist and Allergist, Division of Pediatric Allergy and Immunology, Department of Pediatrics, Nicklaus Childrens Hospital, Miami, FL

    7Certified Nurse Practioner, Division of Pediatric Allergy and Immunology, Department of Pediatrics, Nicklaus Childrens Hospital, Miami, FL

    8Pediatric Hematologist and Oncologist, Department of Pediatrics, Section of Hematology, Oncology, and BMT, University of Colorado School of Medicine and Childrens Hospital of Colorado, Aurora, CO

    9Pediatric Immunologist and Allergist, National Jewish Health, Immunodeficiency Diagnosis and Treatment Program, Department of Pediatrics, Denver, CO

    10Professor of Pediatrics and Immunology, Department of Pediatrics, Division of Allergy, Immunology, and Pulmonology, Duke University Medical Center, Durham, NC

    Introduction: Transplantation of cultured thymus tissue (RVT-802) is an investigational therapy that has led to generation of naive T cells in 61/86 (71%) of subjects with complete DiGeorge anomaly (cDGA). Two children with cDGA and one with a FOXN1 mutation were treated with RVT-802 after prior failed hematopoietic transplants.

    Methods: Subjects were enrolled in IRB protocol 00051692 for RVT-802. RVT-802 was implanted into the quadriceps with immunosuppression.

    Results: Subject 1 was normal at 22q11.2 but had hypocalcemia, an ASD, PDA, and abnormal ears. The subject received a cord blood transplant mismatched at HLA-B and HLA-C alleles at age 3 months. Subsequently mild graft-versus-host disease (GVHD) developed and was treated with antithymocyte globulin, steroids and cyclosporine. Donor T cells developed in low numbers. Twelve years later, the subject developed Epstein Barr virus lymphoma and suffered two relapses. While in remission, subject 1 received unmatched RVT-802. Two weeks after RVT-802 implantation, the subject developed an adenovirus infection resulting in skin and gut GVHD, presumably from activation of the cord blood T cells. Subject 1 was treated with corticosteroids, cyclosporine, cidofovir and infliximab. Four years post RVT-802, subject 1 is healthy with 609 genetically recipient T cells/mm3 and 40% naïve CD4 T cells.

    Subject 2 was normal at 22q11.2 but had an ASD, PDA, hypoparathyroidism, and no T cells at birth. His genetic defect is unknown. Subject 2 was treated with a RIC myeloablative, allogenic, unrelated, 10/10 cord blood transplant, and a subsequent myeloablative, unrelated 9/10 cord blood transplant. Hematopoietic chimerism was established without T cell development. RVT-802 expressed the one allele in the recipient that was not expressed by the second cord donor. The post-thymic transplant course included immune thrombocytopenia requiring rituximab and splenectomy and generalized adenopathy for 3 years but no GVHD. He failed weaning of immunoglobulin replacement. Three years post RVT-802, he has 930 CD3, 750 CD4, and 105 CD8 T cells/mm3. He is active in school.

    Subject 3 had absent TRECs on newborn screening with 7 CD3+ T cells/mm.3 A single mutation in FOXN1 was identified; she has sparse scalp hair. Subject 3 received a 9/10 matched unrelated umbilical cord transplant. The post-transplant course was complicated by significant morbidity, and no naïve T cell development. RVT-802 expressed the one allele in the recipient that was not in the cord blood donor. The subject did not develop GVHD, is healthy and at 9 months has 98 naïve CD4+ T cells. She had resolution of longstanding norovirus and sapovirus gastroenteritis.

    Conclusion: RVT-802 can improve T cell immunity after poor or failed correction with allogeneic hematopoietic transplants. In subject 1, GVHD post RVT-802 was related to an acute viral infection; cord T cells attacked HLA mismatches in the recipient. Subjects 2 and 3 were given RVT-802 matched to recipient alleles that were not expressed in the hematopoietic donor. We hypothesize that thymocytes developing in RVT-802, if strongly reactive to the recipient-mismatched allele, are deleted by the bone-marrow-donor dendritic cells (that acquire recipient MHC from the recipient-allele-matched thymic epithelial cells) thereby preventing GVHD.

    (46) Submission ID#595739

    A Novel Mutation in the Cytotoxic T-lymphocyte Antigen4 (CTLA-4) Gene with Cytopenias, Interstitial Lung Disease, Hypogammaglobulinemia and Recurrent Bacterial Endocarditis

    Victoria Dimitriades, MD1, Samantha Swain, MD2

    1Associate Clinical Professor of Pediatrics, Division of Pediatric Allergy, Immunology & Rheumatology, University of California Davis Health

    2Assistant Professor of Medicine, Division of Allergy and Clinical Immunology, University of California Los Angeles

    Rationale: CTLA4 haploinsufficiency is an autosomal dominant immune dysregulation syndrome characterized by variable phenotypes. Here we present a young woman diagnosed with Evans Syndrome and lymphoproliferation as a child, found to have a novel CTLA4 variant as a young adult, and who developed hypogammaglobulinemia and a bacterial endocarditis while stabilized on CTLA-4 replacement therapy.

    Methods: Sequencing of 207 genes, including CTLA4, in Primary Immunodeficiency Panel.

    Results: Our patient was diagnosed with Evans Syndrome at age 2 with manifestations of anemia and thrombocytopenia recalcitrant to treatment over many years with steroids, cyclosporine, and vincristine. Bone marrow biopsy reportedly showed normal trilineage maturation and her symptoms responded for a short time to splenectomy at age 14. Symptoms recurred at age 16 when she was also found to have pulmonary reticular opacities, prominent lymph nodes, and elevated B cells. Repeat bone marrow and lymph node biopsies at that time were unrevealing. Minor responses to treatment with IVIg, rituximab, mycophenolate mofetil and GCSF were noted. At age 17, she developed varicella-related encephalitis shortly after vaccination. With a strong suspicion of an immune dysregulation syndrome, immune evaluation revealed normal immunoglobulins with good vaccine responses, elevated B cell numbers, normal T cell numbers, and normal mitogen proliferation. CTLA4 sequencing revealed a mutation in exon 2 [c.420C>A, p.Tyr140*] causing a premature translational stop signal, which was consistent with previously reported cases of CTLA4 haploinsufficiency. She was started on rapamycin initially for her cytopenias but was then transitioned successfully to abatacept with almost complete resolution of her anemia, neutropenia, and pulmonary opacities. After 6 months of stable control, she developed a precipitous drop in her platelets and was eventually diagnosed with Streptococcus viridans endocarditis of her native mitral valve. This responded to antimicrobial therapy, but eventually needed surgical intervention due to ongoing insufficiency. Around this time, she was also found to be newly hypogammaglobulinemic, necessitating ongoing IgG supplementation therapy. During successful replacement of her mitral valve with a biosynthetic prosthesis, it was noted that her aortic valve also had evidence of previous disease, implicating a prior endocarditis as part of her clinical syndrome as well.

    Conclusions: In this patient, the presentation of recalcitrant cytopenias, lymphadenopathy, elevated B cells, vaccine-induced viral infections and lung findings precipitated concern for immune dysregulation syndromes and allowed for identification of a novel deleterious CTLA4 mutation. In addition to previously reported clinical findings, our patient presents with the first reported case of repeated endocarditis in the setting of CTLA4 insufficiency disease. Given the finding in this patient of prior (unrecognized) disease, regularly screening patients with CTLA4 insufficiency for evidence of cardiac affectation may be prudent.

    (47) Submission ID#596393

    Alpha Fetoprotein Levels in Ataxia Telangiectasia as Related to Age, Disease Characteristics and Outcomes

    Ariela Agress, MD1, Howard M. Lederman, MD, PhD2, Rong Guo, MS3, Jennifer Wright, RN4

    1Pediatric Resident, Westchester Medical Center

    2Professor of Pediatrics, Medicine and Pathology, Division of Pediatric Allergy and Immunology at Johns Hopkins University School of Medicine

    3Statistician, Division of General Internal Medicine and Health Services Research, UCLA

    4Clinical Research Nurse, Johns Hopkins University

    Background: The relationship between elevated serum alpha fetoprotein (AFP) concentration and age, mortality, genotype and neurologic outcome in Ataxia Telangiectasia (A-T) patients has remained inconclusive over the past decades, leaving AFP as a useful marker for disease diagnosis without further clinical significance.

    Objective: To examine the relationship between AFP levels and age, mortality, genotype and neurologic outcome using a data set larger than any prior study.

    Methods: We retrospectively collected data on 280 A-T patients at Johns Hopkins Medical Center (0- 34 years of age) with both classical (predicted protein null) and variant A-T. This included 459 serum AFP measurements (179 serial levels in 50 A-T patients, max observations 9 per patient). Mixed model compound symmetry covariance was used for statistical analysis to examine the effect of age at visit on AFP levels. Subgroup analysis by mutation type, mortality, feeding/swallowing scores as a surrogate for neurologic function, x-ray induced in vitro chromosomal breakage and serum transaminase levels were similarly analyzed.

    Results: Significant association between age and AFP level was found such that for every 1 year increase in age, AFP level increases 20 ng/mL (p<0.0001). Subgroup analysis by mutation type found that the 12 patients with missense mutations showed a negative linear relationship between log AFP levels and age (r= -0.10, p=0.03). We found greater AFP levels in patients who subsequently died, after controlling for age (least square mean AFP level in log scale 0.67 greater in deceased patients versus living patients, p=0.002). We found a significant decline in feeding score by 0.18 units (score range 0-5) per 100 ng/mL AFP increase (p=0.05) after adjusting for age. There was no significant relationship between AFP levels and serum transaminase levels.

    Conclusion: AFP increases with age in A-T patients, though this may not apply to patients with missense mutations. There is a statistically significant increase in mortality and worsened swallowing scores with increasing AFP levels, but this remains to be proven clinically significant.

    figuren

    (48) Submission ID#596397

    Pill Endoscopy as a Diagnostic Tool for an Abdominal Exacerbation in a Pediatric Patient with Hereditary Angioedema: A Case Report

    Yatyng Chang, MD1, Melissa Cardenas-Morales, MD2, Luis Caicedo Oquendo, MD3, Jose Calderon, MD4, William Blouin, APRN5, Vivian Hernandez-Trujillo, MD4

    1Pediatric Resident, Nicklaus Children's Hospital

    2Allergy Immunology Fellow, Nicklaus Children's Hospital

    3Gastroenterology Attending, Nicklaus Children's Hospital

    4Allergy Immunology Attending, Nicklaus Children's Hospital

    5Nurse Practitioner, Nicklaus Children's Hospital

    Introduction: Hereditary Angioedema (HAE) is a chronic illness characterized by recurrent attacks of angioedema and results in frequent Emergency Department (ED) visits per year. Here we present a pediatric HAE patient who had recurrent abdominal attacks in which constipation, secondary to the ADHD medication dexmethylphenidate (Focalin), appears to be a trigger. Of importance, this is the first pediatric patient with HAE to be described as having safely undergone a capsule endoscopy for direct visualization of the gastrointestinal tract. This was done to decrease the risks associated with the more invasive procedure of traditional endoscopy and colonoscopy.

    Case Presentation: The patient was an 8-year-old male with Hereditary Angioedema who presented with 1 day history of diffuse abdominal pain and nausea. In the ED, patient was in no acute distress. Abdominal ultrasound showed severe circumferential thickening of the wall of multiple bowel loops and a large amount of simple ascites. X-ray revealed stool in the colon. He was admitted for pain control and hydration.

    In the next year, he visited the ED five more times for exacerbations of angioedema of his hand, penis, and bowel. Each time, he presented he had underlying abdominal pain and constipation. He was seen by Gastroenterology and had a workup that was negative for helicobacter pylori, parasites, and other gastrointestinal infections. To further evaluate his abdominal pain, capsule endoscopy was performed and well tolerated. During an admission in January 2016 he received a full inpatient bowel cleanout, after which, his angioedema finally improved. Of note, he was diagnosed with ADHD and started on dexmethylphenidate (Focalin) just prior to this period of recurrent angioedema attacks, and he did not have attacks during the summer months when he was off the medication.

    Discussion: Abdominal pain is a common complaint in pediatric hospitals, and further workup consists of endoscopy and colonoscopy. This may be easily accomplished in the general population, however, in patients with HAE, these procedures carry greater risk and may be avoided, leading to delayed diagnosis and treatment (2,4). A newer and less commonly used alternative for direct visualization of the gastrointestinal tract is capsule endoscopy. Some benefits are that it does not require sedation, is less invasive, and is less likely to be irritating to the mucosa (3). Additionally, since psychological stress may be a trigger for angioedema attacks, the decreased stress associated with a noninvasive procedure such as capsule endoscopy, makes it safer to use (1). Limitations of capsule endoscopy include dependence on battery life and its inability to biopsy or administer therapy if needed (3).

    Hereditary Angioedema treatment consists primarily of avoiding triggers and managing acute episodes. In this first case of HAE in a pediatric patient where capsule endoscopy was used, the procedure was well tolerated without any complications. Recognizing constipation as a trigger and capsule endoscopy as a safe method of direct visualization of the gastrointestinal tract will help others to control and decrease the severity of their HAE attacks as well.

    References

    (1) Aygoren-Pursun E, Saguer IM, Kreuz W, et al. Risk of angioedema following invasive or surgical procedures in HAE type I and II – the natural history. Allergy European Journal of Allergy and Clinical Immunology 2013; 68:1034-1039. (https://onlinelibrary.wiley.com/doi/epdf/10.1111/all.12186)

    (2) Nzeako U, Longhurst H. Many faces of angioedema: focus on the diagnosis and management of abdominal manifestations of hereditary angioedema. European Journal of Gastroenterology and Hepatology 2012; 24(4):353-359.

    (3) Robertson KD, Bhimji SS. Capsule Endoscopy. StatPearls Publishing LLC 2018; Last updated February 27, 2018. https://www.ncbi.nlm.nih.gov/books/NBK482306/.

    (4) Soni P, Kumar V, Alliu S, et al. Hereditary Angioedema (HAE): a cause for recurrent abdominal pain. BMJ Case Reports 2016; Published November 14 2016. https://www.ncbi.nlm.nih.gov/pubmed/27873761

    Capsule Endoscopy Images

    figureo

    (49) Submission ID#596915

    Hepatic Complications of CVID

    Kristine Vanijcharoenkarn, MD1, Merin Kuruvilla, MD2, Frances Lee, MD3, Ekemini Ogbu, MD4, Srihari Veeraraghavan, MD3, Jennifer Shih, MD5

    1Allergy and Immunology Fellow, Emory University School of Medicine

    2Assistant Professor of Medicine, Emory University School of Medicine

    3Associate Professor of Medicine, Emory University School of Medicine

    4Pediatric Rheumatology Fellow, Emory University School of Medicine

    5Assistant Professor of Pediatrics and Internal Medicine, Department of Pediatrics, Division of Allergy and Immunology, Emory University School of Medicine

    A 45 year old male with past medical history of common variable immune deficiency (CVID) and related autoimmune complications, including granulomatous-lymphocytic interstitial lung disease (GLILD), hepatosplenomegaly, leukopenia, and thrombocytopenia tolerated monthly subcutaneous immunoglobulin replacement as outpatient for several years with infrequent infectious complications. Four months ago, he was found to have elevated liver enzymes on routine chemistry. A liver biopsy two months later showed pathology consistent with nodular regenerative hyperplasia (NRH) without overt cirrhosis. A hepatic venous pressure gradient (HVPG) of 21 mmHg was found, consistent with portal hypertension. His hepatitis viral markers were negative, he did not drink, and portal venogram was negative for thrombosis. In early October, the patient was admitted to the hospital with anasarca and tense ascites. He underwent a diagnostic and therapeutic large volume paracentesis and was also found to have spontaneous bacterial peritonitis (SBP) and bacteremia with Group B streptococcus.

    The patients course was complicated by polymicrobial peritonitis, VRE bacteremia, fungemia, variceal hemorrhage, hepatic encephalopathy, and hepatorenal syndrome. His hepatic complications from portal hypertension were out of proportion to his liver parenchymal disease. Transjugular intrahepatic portosystemic shunt (TIPS) was considered to alleviate portal hypertension but was not feasible due to his degree of encephalopathy. Immunosuppressants such as high dose steroids were given while in the hospital with plans to start rituximab to treat patients GLILD after he had recovered from the acute infections. Unfortunately, after two months in the hospital, the patient succumbed to sepsis and progressive liver failure.

    This case emphasizes the importance of systematic screening and continued vigilance for hepatic complications in patients of CVID as studies have shown that NRH of the liver is present in more than 80% of CVID patients who undergo a liver biopsy (PMID: 23219764). A cross-sectional study of patients with primary hypogammaglobulinemia and hepatic dysfunction found that histological findings of NRH were present in 84% of CVID patients and was associated with portal hypertension in 75% of cases (PMID: 17998147). Another study estimated the minimal prevalence of NRH in CVID patients as 12% (PMID: 18647320), stating that this was likely a gross underestimate as NRH may also be present in patients with normal liver function tests that are not routinely biopsied. Therefore, liver enzyme levels may not anticipate the severity of liver involvement. There is currently no treatment for CVID-related liver disease. Other causes of non-cirrhotic portal hypertension, including hepatic veno-occlusive disease and Budd-Chiari Syndrome should be ruled out or treated in CVID patients presenting with hepatic disease. In the case of hepatic NRH in CVID patients, early detection could lead to earlier interventions (such as TIPS prior to hepatic encephalopathy), to mitigate complications.

    (50) Submission ID#597549

    Epigenetic Immune Cell Quantification for Diagnosis and Monitoring of Patients with Primary Immune Deficiencies and Immune Regulatory Disorders

    Janika Schulze, MSc1, Jeannette Werner, PhD2, Konstantin Schildknecht, MSc3, Uma Lakshmanan4, Andreas Grützkau, PhD5, Julia Chu6, Yael Gernez, MD7, Carsten Speckmann, MD8, Katja G. Weinacht, MD9, Alice Bertaina, MD10, Udo Baron, PhD11, Stephan Borte, MD, PhD12, Sven Olek, PhD13, Rosa Bacchetta, MD14

    1Research Scientist, Epimune GmbH

    2VP Research & Development, Epimune GmbH

    3Statistician, Epiontis GmbH, part of Precision for Medicine

    4Staff, Stanford University

    5Head Immunomonitoring, Deutsches Rheumaforschungszentrum (DRFZ)

    6Instructor, Pediatrics - Stem Cell Transplantation, Stanford University

    7Clinical Assistant Professor, Pediatrics - Immunology And Allergy, Stanford University

    8Assistant Medical Director - Pediatric Immunology, University Hospital Freiburg

    9Assistant Professor Of Pediatrics (Stem Cell Transplantation And Regenerative Medicine), Stanford University

    10Associate Professor Of Pediatrics (Stem Cell Transplantation), Lucile Salter Packard Children's Hospital

    11 Principal Scientist, Epiontis GmbH - part of Precision for Medicine

    12Clinical Research Director, ImmunoDeficiencyCenter Leipzig, Municipal Hospital St. Georg Leipzig

    13Managing Director, Epiontis GmbH -a Precision for Medicine Company

    14Associate Professor, Department of Pediatrics, Stem Cell Transplantation, at the Lucile Salter Packard Childrens Hospital, Stanford school of medicine, Stanford, CA, USA

    We describe the application of epigenetic quantification of T regulatory (Treg) cells in addition to CD3+, CD4+, CD8+ T cells, B cells, NK cells, monocytes and neutrophils from as little as 50 μl of fresh, frozen or dried blood. The method yields identical results to flow cytometry from fresh blood samples of a healthy donor cohort, with the advantage of being more sensitive and precise with limited amount of blood and minimal sample preparation (Sci Transl Med 2018). We have used this method 1) to immunophenotype patients with early onset immune regulatory disorders (PIRD) and primary immune deficiency (PID), and 2) to evaluate cell subsets reconstitution early after hematopoietic stem cell transplantation (HSCT).

    Patients with Immune Dysregulation, Polyendocrinopathy, Enteropathy, X-Linked (IPEX) and IPEX-like PIRD were evaluated by analyzing the Treg-Specific Demethylated Region (TSDR) of the FOXP3 locus in the total of CD3+ T-cells. Despite the dysfunctional FOXP3 mutated protein, IPEX patients exhibited elevated Treg/CD3+ cell ratios which seemed to correlate with disease severity. In contrast, most of the patients with IPEX-like symptoms without FOXP3 mutations exhibited decreased Treg/CD3+ cell ratios - in line with the possible central pathogenic role of Treg function and number in PIRD.

    Using epigenetic quantification of CD3+/B- and NK cells, 23 out of 24 confirmed SCID and XLA cases were correctly identified within a cohort of 250 newborn dried blood spot (DBS) samples (96% sensitivity, 100% specificity). The method identified one delayed onset SCID as well as a XLA case that were missed by combined TREC/KREC testing. Epigenetic immune cell quantification missed one SCID case with maternal engraftment that was identified by combined TREC/KREC testing. Abnormally elevated Treg/CD3+ ratio was also detected in a DBS from a newborn who was subsequently confirmed to be affected with IPEX Syndrome.

    When applied to serial blood samples during engraftment and reconstitution post-HSCT, the epigenetic method allowed identification of the different blood cell subsets, including Treg cells, at earlier time points than flow cytometry according to current clinical practice. This opens the way to a better understanding of the correlation between early immune reconstitution events and Graft vs. Host Disease or viral reactivation, earlier than with the current methods, in different types of HSCT.

    These studies underscore the suitability of epigenetic immune cell quantification for accurately measuring multiple immune cell types from limited blood sample sources. We propose this method as uniquely suitable for novel molecular diagnostic applications in settings with limited fresh blood sample or limited cell number, at the point of care as well as for newborn screening.

    (51) Submission ID#597700

    Patient with Hypohidrotic Ectodermal Dysplasia and Recurrent Infections Mimicking NEMO-Deficiency Syndrome

    Roman Deniskin, MSc, MSc, MD, PhD1, Tara Rosenberg, MD2, Nicholas Rider, DO3

    1Resident, Baylor College of Medicine/Texas Children's Hospital

    2Surgical Director, Vascular Anomalies Center, Baylor College of Medicine/Texas Children's Hospital

    3Clinic Chief of Allergy and Immunology, Baylor College of Medicine/Texas Children's Hospital

    We evaluated a 5-year-old male with hyperpyrexia, hypertrichosis, conical hypodontia, and a history of illnesses concerning for NEMO-deficiency syndrome. Starting at six months of age, he suffered recurrent episodes of acute otitis media (non-typeable Hib and Actinobacter Iwolffli), pneumonia, and RSV bronchiolitis. Whole exome sequencing demonstrated a de novo heterozygous c.1259G>A (p.R420Q) mutation in the EDA-receptor (EDAR) gene not present in the parental DNA. His physical exam findings and mutation were consistent with hypohidrotic ectodermal dysplasia (HED), a rare genetic condition characterized by abnormal development of skin, teeth, hair, and sweat glands. HED is caused by defects in the ectodysplasin-A (EDA)-NFkB signaling pathway but is not typically associated with immune deficiency. Consistent with this, immunophenotyping showed normal sub-populations of T-, B-, and NK-cells. Immunoglobulin and complement levels were quantitatively appropriate. He had normal mitogen-induced lymphocyte proliferation and normal antibody response to pneumococcal vaccination. NK-cell studies demonstrated robust cytotoxicity. However, nasal mucosa biopsy showed diffuse squamous metaplasia and the absence of ciliated epithelial cells. We hypothesize that recurrent infections in our patient arose from impaired mucociliary clearance due to a ciliary defect. This case raises the possible association between EDAR variants and ciliary dysfunction. It also underscores the importance of evaluating the immune status of HED patients with recurrent infections which could mimic NEMO-deficiency and have broad implications about clinical management.

    (52) Submission ID#597704

    Show Me the Phenotype: The Ordering Clinicians Role in Genetic Variant Interpretation for Primary Immunodeficiency Diseases

    Jennifer Holle, MS, CGC1, Rebecca Truty, PhD2, Shiloh Martin, MD, PhD3, Hui Yu, PhD4, Michael Anderson, PhD5, Britt Johnson, PhD, FACMG6

    1Genetic Counselor, Invitae

    2Scientist, Invitae

    3Scientist, Invitae

    4Scientist, Invitae

    5Scientist, Invitae

    6Lab Director, Invitae

    The rapid pace of new gene discovery and phenotype expansion for Primary Immunodeficiency Diseases (PIDDs) creates challenges for genetic testing and variant interpretation. Whereas well-described clinical case reports in published literature have traditionally served as the source of phenotypic data used for variant interpretation, for PIDDs the causal variants are often private to the patients family and thus the sole source of phenotypic information for a novel genetic variant is frequently the history provided by the clinician on the test requisition form. Taking into account such heterogeneous information during variant interpretation requires establishing objective criteria for its inclusion as part of the variant interpretation process. To this end, we adapted our laboratorys pre-existing, evidence-based variant classification framework, called Sherloc, by developing point-based criteria for the inclusion of clinical information such as a patients phenotype, familial segregation patterns, and whether the variant is inherited or de novo in the patient. As part of this process, we defined clinical criteria for 154 PIDD genes. Here, we illustrate the application of this method and the importance of integrating clinical information into variant interpretation.

    Between April 2017 and October 2018, our commercial diagnostic laboratory performed 4057 immunological genetic tests, and information about the patients clinical history was provided in 2849 (70%) of these orders. Restricting our analysis to just the 154 genes for which case report information is currently used in variant interpretation, these tests revealed 3868 variants, 370 (10%) of which were classified as pathogenic or likely pathogenic (P/LP). Information from case report descriptions, segregation patterns, and de novo status were applied for 32%,15% and 4% of P/LP variants, respectively. In 37 (10%) cases, the clinical information provided by the clinician on the test requisition form was used as evidence in the classification of the patients variant as P/LP. Ten variants were initially classified as being of uncertain significance and reclassified following receipt of further clinical information or testing of additional relatives. In addition, 35 suspicious variants of uncertain significance were identified in which one or two additional patient case reports would allow for reclassification from uncertain significance to P/LP. These data illustrate the importance of providing good quality clinical information to the genetic testing laboratory both at the time of sample submission and following the receipt of genetic test results.

    (53) Submission ID#597776

    A 30-year Prospective Study Reveals Risk Factors for Malignancies and Early Death in Cartilage-hair Hypoplasia

    Svetlana Vakkilainen, MD1, Mervi Taskinen, MD, PhD2, Paula Klemetti, MD, PhD3, Outi Mäkitie, MD, PhD4

    1Fellow in Pediatric Infectious Diseases, Children's Hospital, Pediatric Research Center, University of Helsinki and HUS Helsinki University Hospital, and Folkhälsan Institute of Genetics, Helsinki, Finland

    2Senior Consultant, Children's Hospital, Pediatric Research Center, University of Helsinki and HUS Helsinki University Hospital

    3Consultant, Children's Hospital, Pediatric Research Center, University of Helsinki and HUS Helsinki University Hospital

    4Professor, Children's Hospital, HUSLAB, University of Helsinki and HUS Helsinki University Hospital, and Folkhälsan Institute of Genetics, Helsinki, Finland, and Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden

    Background: Cartilage-hair hypoplasia (CHH) is a skeletal dysplasia with combined immunodeficiency, variable clinical course and increased risk of malignancy, mostly non-Hodgkin lymphoma and basal cell carcinoma. There is a paucity of long-term follow-up data, as well as knowledge on prognostic factors in CHH.

    Objective: We conducted a prospective cohort study in Finnish patients with CHH to describe clinical course and analyze risk factors for adverse outcomes.

    Methods: We recruited 80 Finnish patients with CHH in 1985-1991 and performed clinical follow-up in 2011-2015. We obtained health information from Finnish National Medical Databases (covering time period of 1969-2016), the Finnish Cancer Registry (1953-2016) and the Cause-of-Death Registry of the Statistics Finland (1971-2016) and analyzed all patients’ health records. Standardized mortality ratios (SMRs) were calculated based on the population data. Primary outcomes included immunodeficiency-related death (from infections, respiratory diseases or malignancies), the development of lymphoma and the development of skin cancer.

    Results: The study cohort included 35 males and 45 females. Median age at recruitment was 14.6 yrs (range 2 weeks – 49.6 yrs) and median duration of follow-up for the surviving patients was 29.2 yrs (range 25.6 – 31.0 yrs). Half of the patients (46/80, 57%) had no symptoms of immunodeficiency, while 15 (19%) and 19 (24%) patients manifested symptoms of humoral or combined immunodeficiency respectively, including six cases of late-onset immunodeficiency. In a significant proportion of patients (17/79, 22%), clinical features of immunodeficiency progressed over time. Of the 15 patients with non-skin cancer, eight had no preceding symptoms of immunodeficiency. Altogether 20 patients had deceased (SMR=7.0, 95% confidence interval (CI)=4.3-11) including deaths due to pneumonia (n=4), malignancy (n=7, SMR=10, 95%CI=4.1-21) and lung disease (n=4, SMR=46, 95%CI=9.5-130). Malignancy was diagnosed in 21/80 (31%) patients, mostly lymphoma (n=9) and skin cancer (n=15). Severe short stature at birth (compared to normal, SMR/SMR ratio=5.4, 95%CI=1.5-20), symptoms of combined immunodeficiency (compared to asymptomatic, SMR=19 (95%CI=8.0-36) vs SMR=4.8 (95%CI=2.3-8.9), Hirschsprung disease (odds ratio (OR) 7.2, 95%CI=1.04-55), pneumonia in the first year of life or recurrently in adulthood (OR=7.6/19, 95%CI=1.3-43/2.6-140), and autoimmunity (OR=39, 95%CI=3.5-430) in adulthood associated with early mortality. In addition, recurrent pneumonia in childhood was associated with the development of lymphoma, while warts and actinic keratosis were associated with the development of skin cancer. Birth length standard deviation score correlated significantly with the age at the diagnosis of first malignancy (p=0.0029), lymphoma (p=0.011) and skin cancer (p=0.014), demonstrating that patients with shorter birth length developed malignancies at an earlier age.

    Conclusions: Patients with CHH have high mortality due to infections and malignancies, but also from lung disease. Some subjects present with late-onset immunodeficiency or malignancy without preceding symptoms of immune defect, warranting careful follow-up and screening for cancer even in asymptomatic patients. We provide clinicians with the risk factors for adverse outcomes to assist in management decisions.

    (54) Submission ID#599129

    A Novel Genetic Etiology for FAS-associated Protein with Death Domain Deficiency

    Lisa A. Kohn, MD, PhD1, Caroline Y. Kuo, MD2

    1Fellow, Pediatrics in the Division of Allergy, Immunology, and Rheumatology at UCLA

    2Assistant Clinical Professor of Pediatrics, University of California, Los Angeles

    Autoimmune Lymphoproliferative Syndromes (ALPS and related disorders) are characterized by insufficient apoptosis due to defects in the FAS apoptosis pathway. FADD deficiency (OMIM 602457) is an autosomal recessive disorder resulting from a mutation in FAS-associated protein with death domain (FADD), the adaptor protein involved in Fas signaling to Caspases 8 and 10. We present a case of FADD deficiency identified by whole exome sequencing with a novel genetic mutation

    We describe two brothers with recurrent febrile episodes accompanied by seizures and respiratory compromise. The older sibling initially presented with status epilepticus following the Measles Mumps Rubella vaccination later experiencing similar episodes until his demise at 18 months of age.

    The younger sibling, who is unvaccinated, presented at 14 months with fever, rash, vomiting, and diarrhea. He developed status epilepticus with respiratory depression that required intubation. He also had enlarged cervical lymph nodes that regressed with antibiotics and steroids. He recovered from that episode but subsequently had a series of similar illnesses with fevers, altered mental status and seizures. With the exception of elevated HHV6 IgG, extensive infectious workup up in all instances was negative.

    Previously described FADD deficiency patients demonstrate an ALPS like phenotype with increased circulating double negative T cells, lymphocyte apoptosis defects, elevated Fas ligand and IL10, encephalopathy, functional asplenism but no splenomegaly or lymphadenopathy. Our patients clinical and laboratory findings were similar. He had normal IgG and IgA, decreased IgM, and lack of isohemagglutinins. Absolute CD3+ count is elevated, with elevated percent of CD3+ TCR+ CD4- CD8-. Normal mitogen and antigen T lymphocyte stimulation, but with defect in pokeweed induced B cell proliferation. Fas ligand and IL10 level are increased (See Table 1). No hepatosplenomegaly, but Howell Jolly bodies were detected in peripheral blood indicating functional hyposplenism.

    Whole-exome sequencing revealed two different genetic alterations in the FADD gene: a maternally inherited nonsense mutation predicted to severely truncate the protein and a paternally inherited missense mutation in codon 105. Although this paternal mutation has not been described as pathogenic, a different variant in same nucleotide of FADD has been associated with FADD deficiency (Reference1).

    There are very few cases in the literature of FADD deficiency patients and the overall prognosis is poor compared to classical ALPS patients, as these patients are at significant risk of deadly sepsis from encapsulated organisms or death from neurologic complications. Of the FADD deficiency patients described in the literature, several died prior to 5 years old. While pneumococcal prophylaxis may reduce the risk of sepsis, hematopoietic stem cell transplant has been reported for patients with FADD deficiency (Reference2), and is being considered for our patient.

    References:

    1. Bolze A et al. Whole-exome-sequencing-based discovery of Human FADD Deficiency. Am J Hum Genet. 2010

    2. Savic S, et al. A new case of Fas-associated death domain protein deficiency and update on treatment outcomes. JACI. 2015.

      6 years old Reference Range
    Total Lymphocytes (cells/μl) 8740 1500-7000
    CD3+ (T cells) absolute 5947 1253-2216
    CD16+CD56+ (NK cells) absolute Not done  
    CD19+ (B cells) absolute 2186 214-624
    Phenotype of CD3+
    (% T cells)
      
    TCR αβ+ T cells 64% 59-81%
    TCRαβ+
    CD4-CD8-
    4.7% 0.3-1.7%
    TCR γδ+ T cells 3.6% 1.2-12.7%
    TCR γδ+
    CD4-CD8-
    2.5% 0.7-7.3%
    HLADR+ activated T cells 14% 3-25%
    Phenotype of B cells   
    Total B cells CD19+ 25% 8-22%
    CD5+ 29% 6-33%
    CD27+ B cells 6% 11-51%
    Serum immunoglobulins (g/L)   
    IgG 1,068 397-1,652
    IgA 204 50-240
    IgM 27 (low) 40-140
    Isohemmaglutins (Blood type A+)   
    A1 <1:2 <1:2
    A2 <1:2 <1:2
    B <1:2 <1:2
    Other studies   
    IL10 level 9 pg/ml <=6 pg/ml
    Soluble FasL 758 70-308
    Vitamin B12 level 1415 254-1060

    (55) Submission ID#599209

    Infusion Parameters and Key Characteristics of Pediatric and Adolescent Patients with Primary Immunodeficiency Initiated on Ig20Gly in a Patient Program

    Lisa Meckley, PhD1, Yanyu Wu, PhD2, Spiros Tzivelekis, MSc3, Andre Gladiator, PhD4

    1Director, GHEORE, Shire

    2Lead, Health Economics and Outcomes Analytics, Shire

    3ORE Lead, ORE Immunology & Opthalmolgy, Shire

    4Global Medical Lead Immunology - Global Medical Affairs, Shire

    Rationale: HCUVP is a patient product-introduction program that provides Cuvitru® (immune globulin subcutaneous [human], 20% solution [Ig20Gly]) free of charge for the first 4 infusions to eligible patients with primary immunodeficiency disease (PID). Using patient data from this ongoing program, our analysis described the clinical characteristics and infusion parameters of pediatric and adolescent patients who were initiated on Ig20Gly through HCUVP.

    Methods: HCUVP eligibility criteria were: patients aged 2 years old, with a primary ICD-10-CM code verifying diagnosis of PID, and no current or prior use of Ig20Gly at program initiation. Data from patients who received the first Ig20Gly infusion between January 1, 2017, and September 1, 2017 were included. Data from patients receiving infusions after October 31, 2017 were censored. Descriptive statistics were calculated for patients demographic and clinical characteristics and prescribed and actual infusion characteristics by age group (<18 years and 18 years).

    Results: In total, 817 patients who completed all 4 infusions were included in the analysis, of whom 97 were aged <18 years. Among those who previously received immunoglobulin (IG) therapy, a greater percentage of patients aged <18 years were treated with intravenous IG therapy (n=46; 73%) compared with adult patients (n=222; 62%) before initiating Ig20Gly. Nine patients aged <18 years were treatment naïve. The mean infusion volume per site was lower among patients aged <18 years (25 years: 17.9 mL; 611 years: 26.4 mL; and 1217 years: 34.6 mL) than among patients aged 18 years (1864 years: 38.5 mL and 65 years: 38.9 mL). However, the mean infusion rate per site was similar between patients aged <18 years (25 years: 45.9 mL/h; 611 years: 47.3 mL/h; and 1217 years: 40.7 mL/h) and patients aged 18 years (1864 years: 43.3 mL/h and 65 years: 44.0 mL/h). In addition, by the final infusion, fewer patients aged <18 years were infused weekly (n=18 [19%] patients) compared with patients aged 18 years (n=232 [32%] patients). Conversely, a greater percentage of patients aged <18 years were infused biweekly (n=35 [36%] patients) compared with patients 18 years (n=168 [23%] patients).

    Conclusion: The results provide insights into the clinical and infusion characteristics of pediatric and adolescent patients who have received Ig20Gly and clinical use of Ig20Gly outside of a controlled clinical trial setting.

    Funding: This research was sponsored by Shire.

    (56) Submission ID#599435

    XMEN: MAGT1 Mutation Associated Immunodeficency. Case Report of an Atypical Presentation

    Carlos A. Verdugo, Medical Doctor1, Alejandra V. King, MD2

    1Immunology Resident, Universidad de Chile

    2Staff, Immunology Unit Luis Calvo Mackenna Children's Hospital, Clínica Alemana de Santiago

    XMEN disease (X-linked Immunodeficency with Magnesium defect, Epstein-Barr virus infection and Neoplasia) is a primary immune deficiency caused by mutations in MAGT1 and characterized by chronic infection with Epstein-Barr virus (EBV), EBV-driven lymphoma, CD4 T-cell lymphopenia, and dysgammaglobulinemia. MAGT1 gene codifies to MagT1 protein, a Mg2+- selective transporter, expressed in the human immune system, specifically in the spleen and the thymus. Functional studies have established the key role of MAGT1 in T cells and natural killer (NK) cell activation. Upon CD4+ T-cell receptor stimulation, MAGT1 mediates a transient Mg2+ influx that is necessary for phospholipase C gamma 1 (PLCy1) activation, which drives Ca2+ rise and downstream signaling. This Mg2+ influx also regulates cytotoxic functions of NK and CD8 T cells through NKGD2, reason why these patients have impaired cytolytic responses against EBV. Eleven male XMEN patients have been described. We present the case of a 1-year old Hispanic infant with a pathogenic variant in MAGT1 gene that clinically manifested with early Pneumocystis jirovecii and cytomegalovirus (CMV) interstitial pneumonia, and EBV chronic infection with good response to intravenous immunoglobulins supplementation without hematopoietic stem cell transplantation or gene therapy. Laboratory study highlights low levels of NKG2D ligands. The objective of this case report is to broaden the spectrum of clinical presentation of XMEN disease, that manifests initially as a Combined Immune Deficiency (CID) and evolved with a favorable course of the disease with intravenous immunoglobulins supplementation therapy and chemoprophylaxis with trimethoprim-sulfamethoxazole.

    (57) Submission ID#599449

    Smoldering Hemophagocytic Lymphohistiocytosis Secondary to Compound Heterozygous Variants in SLCA7 Treated with Anakinra

    Nicholas L. Hartog, MD1, Beth Kurt, MD2, Stacie Adams, MD3, Johanna Zea-Hernandez, MD4, Surender Rajasekaran, MD5

    1Allergy and Immunology, Helen DeVos Children's Hospital and Michigan State University

    2Hematology and Oncology, Helen DeVos Children's Hospital and Michigan State University

    3Biochemical Genetics, Helen DeVos Children's Hospital and Michigan State University

    4Pediatric Pulmonary, Helen DeVos Children's Hospital and Michigan State University

    5Pediatric Intensive Care Unit, Helen DeVos Children's Hospital and Michigan State University

    Introduction: Lysinuric protein intolerance (LPI) is a recessively inherited disorder of the cationic amino acids transporter subunit y+LAT1 caused by variants in the SLC7A7 gene. The disease is characterized by protein-rich food intolerance has a heterogeneous presentation. The clinical findings are a result of depletion of lysine, ornithine, and arginine. Symptoms can include hyperammonemia, failure to thrive, protein aversion, neurologic disease, and lung disease. There is also evidence that inflammatory manifestations are mediated through upregulation of NFB, IL1, and TNF that occur independent of intracellular arginine levels and can lead to life-threatening episodes of hemophagocytic lymphohistiocytosis (HLH).

    Case Presentation: A 17-year-old male presented with history of anxiety, depression, eating disorder, delayed puberty and complex partial seizures. Due to poor nutrition and failure to thrive, a gastrostomy tube was placed. Following commencement of enteral feeds, he presented with altered mental status, bilateral mydriasis, hyperreflexia, and agitation which lead to a PICU admission. Ammonia peaked as high as 181 μmol/L and episodes ceased with cessation of enteral feedings. Prior to enteral feeds, he had been self-restricting protein in his diet. Biochemical testing was consistent with LPI and Illumina next-generation sequencing revealed compound heterozygous variants in SLC7A7 (p.S396Lfs*122 and p.E465Dfs*54). Hyperammonemia resolved quickly with cessation of protein intake and high rate dextrose infusion without the need for ammonia scavenging agents. He was subsequently started on protein-restricted enteral feeds.

    At diagnosis he did not have any respiratory symptoms, CT scan of chest showed patchy areas of groundglass opacification that was suggestive of early pulmonary alveolar proteinosis (PAP). Bronchoalveolar lavage demonstrated foamy, cloudy pink fluid and elevated bronchioalveolar macrophages on cell differential.

    His clinical course and SLC7A7 genotype led to suspicion for smoldering HLH. The findings of elevated ferritin, hypertriglyceridemia, decreased fibrinogen, splenomegaly, elevated IL-2 receptor, decreased NK cell function, along with hemophagocytosis on bone marrow biopsy confirmed the diagnosis. Because of his PAP and HLH, in addition to dietary modifications, a trial of IL-1 beta inhibition (anakinra) at 3 mg/kg/day was initiated.

    Follow up CT scan of chest 2 months after initiation of anakinra showed complete resolution of pulmonary groundglass opacifications and PAP. Bone marrow evaluation showed continued hemophagocytosis in spite of the normalization in ferritin, soluble IL-2 receptor, NK function, and triglycerides levels. Overall, he is significantly improved on daily anakinra and no longer meets criteria for HLH or PAP.

    Discussion: Recent data has shown in y+LAT1 models that THP-1 macrophages and A549 airway epithelial cells upregulate IL1 and TNF regardless of intracellular arginine content. This suggests that inflammatory manifestations may continue independent of dietary modifications. We present a 17 year old patient with newly diagnosed LPI who was treated dietary modification and anti-IL1 therapy resulting in resolution of HLH and PAP. More research is needed to see if long-term IL1 blockade that can consistently control both the immunologic and pulmonary manifestations of LPI and positively impact morbidity and mortality.

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    (58) Submission ID#599526

    Bartonella Endocarditis in a Child with Probable ALPS

    Keerti Dantuluri, MD1, James A. Connelly, MD2, Donna Hummell, MD3, Leigh Howard, MD, MPH4, Yasmin Khan, MD5, Daniel Dulek, MD6

    1Pediatric Infectious Diseases Clinical Fellow, Vanderbilt University Medical Center

    2Assistant Professor Hematology/Oncology/Bone Marrow Transplant, Vanderbilt University Medical Center

    3Professor Allergy and Immunology, Vanderbilt University Medical Center

    4Assistant Professor Pediatric Infectious Diseases, Vanderbilt University Medical Center

    5Assistant Professor Pediatric Allergy/Immunology, Vanderbilt Children's Hospital

    6Assistant Professor Pediatric Infectious Diseases, Vanderbilt University Medical Center

    Learning Objective: Recognize that symptoms of Bartonella endocarditis and associated complications can share features of certain immunocompromising conditions.

    Case Description: An 8-year-old Caucasian boy with history of repaired pulmonary atresia and aortic root dilation was diagnosed with pancytopenia and splenomegaly during a brief hospitalization for atypical pneumonia. Pancytopenia persisted, splenomegaly worsened, and five months after presentation, he developed hypertension and renal insufficiency. He was diagnosed with hypocomplementemic, diffuse sclerosing and crescentic glomerulonephritis and was started on mycophenolate mofetil with improvement in kidney function and stabilization of cytopenias. As part of a comprehensive immune work-up, ALPS (autoimmune lymphoproliferative syndrome) panel was sent and demonstrated elevated double-negative T (DNT) cells with 3 out of 4 positive immunologic criteria for ALPS. Neither targeted sequencing for ALPS and ALPS-like disorders nor whole exome sequencing revealed pathogenic mutations.

    By age 10, the patient remained on mycophenolate, but developed failure to thrive, with weight dropping from 37th percentile to less than 3rd percentile. He was hospitalized again for low-grade fever, increased work of breathing, left shoulder pain and fatigue and was found to have right lower lobe pneumonia. Pancytopenia worsened, and he was started on cefepime and azithromycin without improvement in symptoms. Echocardiogram revealed vegetations in his pulmonary conduit and bilateral branch pulmonary arteries, but multiple blood cultures were negative. Upon further history, the patient reported contact with kittens. Bartonella henselae titers and polymerase chain reaction (PCR) from blood were sent and were both positive. He completed a 2-week course of gentamicin, 1-month course of ceftriaxone, and was transitioned to doxycycline and rifabutin. After initiating antimicrobial therapy, his weight and energy significantly improved, his blood Bartonella PCR became negative, and his splenomegaly resolved. Approximately one year later, the patient underwent pulmonary artery conduit replacement and Bartonella PCR testing of the tissue specimen was positive. He has had sustained weight increase, resolution of hypocomplementemia and splenomegaly, decrease in DNT cell frequency from >2% to 0.9%, and improvement though not resolution of cytopenias. He currently remains on doxycycline and rifabutin and continues treatment with mycophenolate.

    Discussion: ALPS is characterized by defective lymphocyte apoptosis and clinical features such as lymphadenopathy, splenomegaly, hepatomegaly, cytopenias, and glomerulonephritis. The hallmark laboratory finding is expansion of DNTs. Our patient met criteria for a probable ALPS diagnosis based on the presence of both required criteria (chronic splenomegaly and elevated DNT cells) and secondary additional criteria (typical immunologic findings noted on ALPS panel). Pediatric cases of Bartonella henselae endocarditis have been associated with splenomegaly, cytopenias, and glomerulonephritis which mimic many features of monogenic immune dysregulatory disorders. The diagnosis of Bartonella endocarditis in our patient therefore raises the question of whether his immunosuppression predisposed him to infection or if his entire clinical presentation can be explained by Bartonella endocarditis. Physicians taking care of patients with immune dysregulatory disorders should consider Bartonella endocarditis in the differential diagnosis of onset or exacerbations of immune dysregulation.

    (59) Submission ID#599570

    Body Temperature in Patients with Primary Immunodeficiency

    Shouling Zhang, MD1, Tiffany S. Henderson, PhD2, Christopher Scalchunes, MPA3, Kathleen E. Sullivan, MD, PhD4, Artemio M. Jongco, III, MD, PhD, MPH5

    1Pediatrics Resident, Department of Pediatrics, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell, New Hyde Park, NY

    2Survey Research Analyst, Immune Deficiency Foundation

    3Vice President of Research, Immune Deficiency Foundation

    4Professor, The Children's Hospital of Philadelphia

    5Assistant Professor of Medicine and Pediatrics, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell; Center for Health Innovations and Outcomes Research, Feinstein Institute for Medical Research, Manhasset, NY

    Rationale: While fever is considered a sign of infection, many individuals with primary immunodeficiency (PI) anecdotally report a lower than normal average body temperature. On Immune Deficiency Foundation (IDF) Friends and IDF PI CONNECT Research Forum online, PI patients report a diminished fever response even when other signs of infection are present. There is limited knowledge about the average body temperature in persons with PI. However, the implications of missing an infection in those with PI is well established.

    Methods: Study investigators partnered with patient investigators to design a prospective cohort study to determine whether body temperature differed between persons living with and without PI. Three hundred fifty adults with PI were recruited from IDF and one adult household member without PI was also recruited. McKesson digital oral thermometers (Model 01-413BGM) were provided and used to record temperatures in all participants three times a day for five consecutive days. Descriptive statistics were calculated. Median body temperatures were compared between the two cohorts at each time point using Mann-Whitney test.

    Results: Data from 254 households were used for analysis (72.6% participation rate). The PI population was largely female (85.8%) with a median age of 49 years and largely Caucasian population (97.6%). The non-PI population was largely male (66.9%) with a median age of 53 years and largely Caucasian population (92.9%). PI diagnoses included CVID (74.8%), hypogammaglobulinemia (12.6%), IgG subclass deficiency (4.7%), selective IgA deficiency (3.1%), specific antibody deficiency (3.1%), agammaglobulinemia (0.4%), chronic granulomatous disease (0.4%), combined immunodeficiency (0.4%), and complement deficiency (0.4%). A total of 123 individuals with PI (48.4%) reported a lower than normal non-sick body temperature, while 108 individuals with PI (42.5%) reported a normal (between 97°F - 99°F) non-sick body temperature. A total of 172 individuals with PI (67.7%) reported absence of fever with infection, while 50 individuals (19.7%) reported a normal fever response with infection. The median body temperature was significantly higher for the PI patients in the morning, but not evening or bedtime, reading in 4 of the 5 days (Monday: PI = 97.5°F vs. non-PI = 97.2°F, p = 0.0291; Tuesday: PI = 97.4°F vs. non-PI = 97.2°F, p = 0.0020; Wednesday: PI = 97.5°F vs. non-PI = 97.2°F, p = 0.0009; Thursday: PI = 97.4°F vs. non-PI = 97.2°F, p = 0.0575; Friday: PI = 97.4°F vs. non-PI = 97.2°F, p= 0.0008).

    Conclusions: Despite the limitations of this non-clinical study, individuals with PI are knowledgeable about their conditions and can offer unique insights and direction to researchers. This study demonstrates that collaboration with patient advocacy groups may facilitate patient-centered and patient-driven research with high participation among the target population.

    (60) Submission ID#599598

    A 2-year-old Male with Compound Heterozygous Familial Mediterranean Fever (FMF)

    Shouling Zhang, MD1, Zoya Treyster, MD2, Vincent Bonagura, MD3

    1Pediatrics Resident, Department of Pediatrics, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    2 Fellow, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    3Professor of Medicine and Pediatrics, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    Introduction: Familial Mediterranean Fever (FMF) is a hereditary condition characterized by recurrent episodes of painful inflammation caused by mutations in the pyrin (MEFV) gene. Alterations in the MEFV gene affect pyrin production leading to recurrent fevers and painful inflammation in the peritoneum, synovium, and pleura. Amyloidosis may also develop as a complication. Arabic, Turkish, Armenian, and Sephardic Jewish populations are most commonly affected. Homozygosity for MEFV mutations are associated with a more severe course. There is a paucity of information regarding pediatric FMF in the literature.

    Case: We present a case of a 2-year-old male with minor speech delay diagnosed with compound heterozygous FMF. Patient was initially referred due to recurrent fevers and infections. At 4 months of age, he was hospitalized with septic shock requiring intubation secondary to adenovirus. At 5 months of age, the patient began to have recurrent fevers every 3 to 4 weeks, leading to multiple blood draws and courses of antibiotics prior to referral. At 11, 12, and 22 months of age, he developed three separate episodes of febrile seizures. A total of 10-15 lifetime episodes of acute otitis media occurred prior to bilateral myringotomy tube placement. Four episodes of Streptococcus pyogenes pharyngitis confirmed by throat culture preceded tonsillectomy. No oral ulcers, joint pain, or abdominal pain were reported. No other infections such as pneumonia, sinusitis, UTI, non-viral gastroenteritis, fungal infections, or skin infections were reported. Both parents are Ashkenazi Jewish and a maternal history of early miscarriage was noted. Family history was negative for immunodeficiency, malignancy, and autoimmunity.

    The patients vital signs and physical exam were unremarkable. Serology indicated leukocytosis of 18.53 K/L with elevated monocytes of 1390 cells/L, elevated eosinophils of 1200 cells/L, and slightly elevated CD8 T cell count of 2653 cells/L. Neutrophil, CD4 T cell, B cell, NK cell enumeration, and immunoglobulin panel were normal for age. Tetanus, diphtheria, rubella, Streptococcus pneumoniae, and Haemophilus influenzae B titers were protective. Genetic analysis identified that the patient was compound heterozygous for the E148Q and V726 mutations in the MEFV gene.

    Family was instructed to keep a fever diary. Colchicine 0.6mg once a day was given initially, then increased to 1.2mg once a day for inadequate response. Loose stools were observed while patient was maintaining a lactose free diet so he was switched to colchicine 0.6mg BID with resolution of loose stools. Apart from two occasions when his colchicine dose was missed, the patient remained afebrile at his follow up visits.

    Conclusion: We present a pediatric case of compound heterozygous FMF (E148Q and V726 MEFV mutations) in an otherwise healthy 2-year-old male of Ashkenazi Jewish background, initially symptomatic at 5 months of age. Individuals who are compound heterozygous for the E148Q and a second MEVF mutation are generally symptomatic, although severity cannot be predicted. Additional pediatric research on symptomatic heterozygous and compound heterozygous FMF is recommended.

    (61) Submission ID#599700

    CD27 Deficiency Causes Human NK Cell Deficiency with Specific Loss of the CD56(bright) Subset: A Single Case Report

    Natalia Chaimowitz, MD, PhD1, Sarah Nicholas, MD2, Leonora Noroski, MD, MPH3, Lisa R. Forbes, MD4, Sara Nandiwada, Ph.D., D(ABMLI)5, Nicholas Rider, DO6

    1Fellow, Texas Childrens Hospital/Baylor College of Medicine

    2Assistant Professor, Baylor College of Medicine/Texas Children's Hospital

    3Associate Professor of Pediatrics, Baylor College of Medicine/Texas Children's Hospital

    4Assistant Professor, Department of Pediatrics, Baylor College of Medicine, Houston, TX, USA

    5Assistant Professor of Pediatrics, Baylor College of Medicine/ Texas Children's Hospital

    6Associate Professor of Pediatrics, Texas Childrens Hospital/Baylor College of Medicine

    Natural killer (NK) cells are innate lymphocytes that play a key role in defense against virally-infected cells and in tumor surveillance. NK cells can be divided in two subsets. The majority of NK cells in peripheral blood expressed intermediate levels of CD56 and are referred to as CD56(dim). These NK cells are responsible for NK cell cytotoxicity. A minor population of NK cells express very high expression of CD56 and are referred to as CD56(bright). These NK cells are responsible for cytokine production and are precursors to CD56(dim) NK cells. A few immunodeficiencies have been described in which there are abnormal NK cell subsets, such as autosomal dominant GATA2 deficiency where CD56(bright) NK cells are absent and IRF8 where there is a paucity of CD56(dim) NK cells and relative expansion of CD56(bright) NK cells.

    Here we present a patient with an absence in CD56(bright) NK cells secondary to CD27 deficiency. Our patient is a 6-year-old African American female born to non-consanguineous parents. The patients past medical history is significant for chronic lung disease secondary to prematurity, recurrent acute otitis media, failure to thrive and congenital hypothyroidism. Family history is significant for an older sister that presented at age 3 with EBV-associated Hodgkin lymphoma whose treatment was complicated by chronic activated EBV infection and who ultimately underwent hematopoietic stem cell transplantation (HSCT). Our patient presented with pancytopenia, fever, lymphadenopathy and splenomegaly. She was found to have EBV viremia with greater than 550,000 copies in whole blood by PCR. She was treated with two doses of rituximab followed by etoposide and dexamethasone as a bridge to HSCT. Whole exome sequencing demonstrated a homozygous mutation in CD27. CD27 is a member of the tumor necrosis factor receptor family and influences the function of T cells, B cells and NK cells. In NK cells, CD27 is primarily expressed in CD56(bright) NK cells. CD27 deficiency is an autosomal recessive disorder associated with persistent symptomatic EBV viremia, including EBV-driven hemophagocytosis and lymphoma, hypogammaglobulonemia and specific antibody deficiency. Our patients immune evaluation prior to initiation of chemotherapy and immunosuppression was notable for very elevated IgG, IgA and IgM. Despite hypergammaglobulonemia patient had only 3 out of 11 protective titers against streptococcus pneumoniae. The patient had pan-lymphopenia with appropriate percentages of lymphocyte subsets. Assessment of her B cell subsets showed a slight increase in the percentage of transitional B cells/plasmablast and a nearly complete absence of CD27-expressing B cells. Her NK cell phenotyping demonstrated a complete loss of CD56(bright) NK cells with reduced NK cell cytotoxicity, comparable to what has been previously reported in patients with GATA2 deficiency. Previous reports of patients with CD27 deficiency denote normal NK cell numbers with normal to moderately reduced NK cell cytotoxicity, however, CD27 deficiency causing a specific loss of the CD56(bright) NK cell subset has not been previously reported. CD27 deficiency should be consider in patients with EBV driven disease and abnormal NK cell studies.

    (62) Submission ID#599897

    Allogeneic Hematopoietic Stem Cell Transplant Outcomes for Patients with Dominant-Negative IKFZ1/IKAROS Mutations

    Erinn S. Kellner, MD1, Christa Krupski, MD2, Hye Sun Kuehn, PhD3, Sergio D. Rosenzweig, MD/PhD4, Nao Yoshida, MD/PhD5, Seiji Kojima, MD/PhD6, David Boutbol, MD7, Sylvain Latour, PhD8, Vincent Barlogis, MD/PhD9, Claire Galambrun, MD10, Asbjørg Stray-Pedersen, MD/PhD11, Hans Christian Erichsen, MD/PhD12, Rebecca A. Marsh, MD13

    1Clinical Immunodeficiency Fellow, Division of Bone Marrow Transplantation and Immune Deficiency, Cincinnati Childrens Hospital Medical Center

    2Instructor, Division of Bone Marrow Transplantation and Immune Deficiency, Cincinnati Childrens Hospital Medical Center

    3unknown, Immunology Service, Department of Laboratory Medicine, Clinical Center

    4Chief, Immunology Service, Department of Laboratory Medicine, NIH Clinical Center, Bethesda, MD, USA

    5unknown, Department of Hematology and Oncology, Childrens Medical Center, Japanese Red Cross Nagoya First Hospital

    6Professor, Department of Pediatrics, Nagoya University Graduate School of Medicine, Nagoya, Japan

    7unknown, Laboratory of Lymphocyte Activation and Susceptibility to EBV Infection, Inserm UMR 1163 and Clinical Immunology Department, Hôpital Saint Louis, Assistance Publique Hôpitaux de Paris (APHP) Université Paris Diderot

    8unknown, Laboratory of Lymphocyte Activation and Susceptibility to EBV Infection, Inserm UMR 1163 and University Paris Descartes Sorbonne Paris Cité, Imagine Institut

    9unknown, Department of Pediatric Immunology, La Timone Hospital

    10unknown, Department of Pediatric Hematology and Oncology, Timone Enfants Hospital and Aix-Marseille University

    11unknown, Norwegian National Unit for Newborn Screening, Oslo University Hospital

    12unknown, Section of Specialized Pediatric Medicine, Oslo University Hospital

    13Associate Professor, Division of Bone Marrow Transplantation and Immune Deficiency, Cincinnati Childrens Hospital Medical Center

    Introduction/Background: The transcription factor IKAROS is encoded by the IKZF1 gene and plays a crucial role in lymphopoiesis. Somatic, and more recently also germline mutations of IKZF1 are associated with a hematologic malignancies, most notably B-cell precursor acute lymphoblastic leukemia. Germline mutation in IKZF1 was first reported as a monogenic cause of human disease characterized by marrow failure and immune deficiency in a single neonate in 2012. Subsequently, mutations leading to haploinsufficiency were discovered to underlie a proportion of patients with CVID and low B cell numbers, and dominant-negative mutations have been observed to cause more severe combined immune deficiency phenotypes. At this time, there is very little known regarding allogeneic hematopoietic cell transplantation (HCT) outcomes for patients with severe dominant-negative IKZF1 mutations. Concerningly, IKAROS deficiency has been observed to have a negative impact on graft versus host disease in mouse models.

    Objective: To describe allogeneic stem cell transplant outcomes in patients with the dominant-negative IKAROS mutation.

    Methods: We collected transplant data from 4 patients who underwent allogeneic HCT at transplant centers around the world.

    Results: Patients underwent allogeneic HCT using a variety of conditioning regimens. Patients received bone marrow (N=3) or cord blood (N=1) grafts from an HLA-matched sibling donor (N=1) or single allele HLA-mismatched unrelated donor (N=3). Neutrophil engraftment occurred between Day +12 and +51 post-transplant. Platelet engraftment occurred between Day +8 and +167 except in one patient who did not have return of normal platelet counts due to underlying liver dysfunction. All patients were documented to have greater than 99% whole blood donor chimerism at a median of 28 days (range 12-51 days) following transplant and maintained >95% donor chimerism until last follow-up. Only one patient developed grade II acute GVHD. No patients developed chronic GVHD. One patient died approximately 1 year post transplant related to cryptosporidium cholangitis which existed prior to HCT. At the most recent follow up of the 3 surviving patients (range: 0.99-7.2y), IVIG had been discontinued, antimicrobial prophylaxis had been stopped, and patients had received routine vaccinations. They all had excellent performance status.

    Conclusions: Allogeneic HCT may be a safe option to consider for patients with dominant-negative IKAROS mutation as there does not appear to be an increased risk of death or GVHD. Moreover, 3-out-of-4 of the transplanted patients are alive and well and show no features of the disease. However, because of the limited number of patients evaluated and the retrospective nature of this analysis, our data do not allow firm conclusions to be made, and further studies will be needed to evaluate outcomes in larger cohorts.

    (63) Submission ID#600169

    Plastic Bronchitis and Secondary T-cell Lymphopenia

    Saara Kaviany, DO1, Yasmin Khan, MD2, Dan Dulek, MD3, Michael O'Connor, MD4, Stacy Dorris, MD4, John Newman, MD5, Rizwan Hamid, MD6, John Phillips, MD6, James A. Connelly, MD7

    1Clinical Fellow, Vanderbilt Children's Hospital

    2Assistant Professor Pediatric Allergy/Immunology, Vanderbilt Children's Hospital

    3Assistant Professor Pediatric Infectious Disease, Vanderbilt Children's Hospital

    4Assistant Professor Pediatric Pulmonology, Vanderbilt Children's Hospital

    5Professor Allergy/Immunology, Vanderbilt University Medical Center

    6Professor Genetics, Vanderbilt University Medical Center

    7Assistant Professor Hematology/Oncology/Bone Marrow Transplant, Vanderbilt University Medical Center

    Introduction: When evaluating patients with T- cell lymphopenia, we often are concerned about defects in lymphocyte production and function, especially in the setting of frequent infections. Here we outline a case demonstrating T-cell lymphopenia due to increased loss, which should be considered in the differential diagnosis.

    Case Report: We report a 13-year-old male who initially presented with recurrent, right-sided pneumonias requiring frequent hospital admissions including severe episodes necessitating intensive care unit admission. His work up for the pneumonias included a bronchoscopy revealing normal anatomy with minimal inflammation, and a chest CT with mild peribronchial wall thickening.

    As his pulmonary disease progressed, he developed a persistent, productive cough with expectorated mucous plugs that were plastic-like in appearance. While his pulmonary symptoms responded to steroids, his mucous plug production persisted. Sputum cultures were intermittently positive, isolating Cryptococcus neoformans and Aspergillus niger. He underwent VATS and wedge biopsy, concerning for recurrent aspiration. An immunologic evaluation initially demonstrated normal T- and B-cell counts, but serial evaluation of his lymphocyte population demonstrated low CD4+ cells (ranging 151-367 cells/cumm), and low normal CD8 cells (ranging 101-177 cells/cumm) with normal B- and NK-cell numbers.

    Further T-cell evaluation revealed normal ratios of naive and memory populations (CD4CD45RA+ 61%, CD4CD45RO+ 39%, CD8CD45RA+ 74%, CD8CD45RO 33%), normal TREC (7768 copies per 10^6 CD3 cells) and normal thymic emigrants (CD4CD31CD45RA+: 158, normal 150-1500), indicative of sufficient thymopoiesis. Mitogen and antigen stimulation assays demonstrated normal responses to phytohemagglutin, concanavalin A, and pokeweed mitogen, with a low lymphocyte response to Candida. He had normal quantitative immunologlobulins, normal diphtheria, tetanus and streptococcus pneumonia titers. His dihydrorhodamine flow cytometry and FISH for chromosome 21q11.2 deletion were negative.

    Given normal function and thymic output, his immunologic profile was concerning for T-cell loss. Our patient was registered with the Undiagnosed Disease Network, and had a second review of his lung biopsy, concerning for plastic bronchitis. Subsequent lymphatic imaging demonstrated abnormal lymphatics within the bilateral clavicular space, right greater than left, with questionable partial thoracic duct, explaining his unilateral symptoms. He was diagnosed with plastic bronchitis secondary to abnormal lymphatic drainage, with lymphatic fluid filling his airways and secondary T-cell loss.

    Discussion: Plastic bronchitis is a rare and potentially fatal disorder, seen commonly after the Fontan procedure for congenital heart disease. This process has resulted in T-cell loss into the airway and subsequent T-cell lymphopenia.

    In patients with Fontan-related protein losing enteropathy, multiple immune abnormalities have been described including reduced immunoglobulins, lymphopenia, and selective CD4 lymphocyte deficiency. Similar findings have been reported in patients with lymphatic malformations. Although the impact of T-cell loss on adaptive immunity is not entirely known, there is no indication of increased risk for atypical infections.

    Given his normal mitogen assay, our patient did not start prophylactic antibiotics. He continues to have symptomatic episodes with lymphopenia, but has had no opportunistic infections, and remains stable with an aggressive pulmonary regimen. We conclude by reiterating the importance of considering T-cell loss in patients presenting with lymphopenia, particularly with evidence of normal thymopoeisis and T-cell function.

    (64) Submission ID#600267

    Granulomatous Disease and Lymphoma in a Cohort of 1395 Patients with CVID in the USIDNET Registry

    Joao Pedro Matias Lopes, MD1, Nicole Ramsey, MD, Phd1, Ramsay Fuleihan, MD2, Kathleen E. Sullivan, MD, PhD3, Avni Joshi, MD4, Daniel Suez, MD5, Patricia Lugar, MD, MS6, John Routes, MD7, Charlotte Cunningham-Rundles, MD, PhD8

    1Allergy and Immunology Fellow, Icahn School of Medicine at Mount Sinai

    2Professor of Pediatrics, Division of Allergy and Immunology, Northwestern University Feinberg School of Medicine, Chicago, NY

    3Professor, The Children's Hospital of Philadelphia

    4Assistant Professor, Allergy and Immunology, Mayo Clinic

    5President, Allergy, Asthma & Immunology Clinic, PA

    6Assistant Professor, Allergy and Immunology, Duke Health

    7Chief, Professor, Division of Allergy and Immunology, Children's Hospital of Wisconsin-Milwaukee, Milwaukee, WI

    8Professor in Medicine, Division of Clinical Immunology, Icahn School of Medicine, Mount Sinai, NY, NY, USA

    Introduction: Granulomatous disease (GD) has been described with a variable incidence (8.0-22.0%) in patients with common variable immunodeficiency (CVID). An increase in malignancies has been reported in CVID patient cohorts, particularly for lymphoma, reported in 1.6-8.2% of the CVID patients depending on the cohorts. Prior analysis of a cohort of 436 CVID patients included 59 patients with GD (GD+). In these, there was a suggestion of more cases of lymphoma (12.5%) when compared to cases without (GD-) (5.0%) although the difference was not statistically significant (p=.07).

    Objectives: Compare the frequency of lymphoma in GD+ and GD- patients in the CVID patient cohort from the USIDNET Registry.

    Methods: We submitted a query to the USIDNET registry requesting de-identified data for patients with the diagnosis of CVID, through August 2018. Statistical analysis was performed on SPSS, with comparisons done with Pearson chi-square or Fisher's exact test, depending on the sample sizes, using an alpha level of .05.

    Results: A cohort of 1395 CVID patients from the USIDNET registry was analyzed. Ninety-one patients (6.5%) were GD+. Overall, 152 patients (10.9%) had a malignancy diagnosis, 47 of these (3.4%) with lymphoma. Lymphoma was present in 6/91 GD+ patients (6.6%) versus 41/1304 GD- patients (3.1%) (p=.12). Overall malignancy was present in 15/91 GD+ (16.5%) versus 137/1304 (10.5%) (p=.08).

    Discussion: In the cohort of 1395 CVID patients from the USIDNET registry, we found a frequency of lymphoma of 3.4%, which is in the range of previously described cohorts. The frequency of lymphoma was 6.6% in patients with GD, higher than the 3.1% frequency for GD- patients, but these differences were not statistically significant. Our identified frequency of lymphoma in GD+ patients was lower than the one previously identified in the 436 CVID patient cohort, but with similar proportional differences between GD+ and GD- patients. Despite no statistical significance, the frequency of lymphoma, as shown here and elsewhere, was higher in CVID patients GD+ than GD- in both studies, with no full understanding of this increased risk of lymphoma. Expanding this analysis to larger groups of CVID patients may help to confirm, or deny a more robust association, which may have a meaningful impact in the outcomes of this particular population.

    (65) Submission ID#600312

    Four Patients with Refractory Pericarditis Treated with Concurrent Hyaluronidase-facilitated Subcutaneous Immunoglobulin and Anti-interleukin 1 Therapy

    Melissa D. Gans, MD1, Rachel Eisenberg, MD2, Arye Rubinstein, MD, Ph.D.2

    1Fellow in Allergy & Immunology, Montefiore Medical Center

    2Attending in Allergy & Immunology, Montefiore Medical Center

    Introduction: Patients with refractory pericarditis have been treated with intravenous immunoglobulin (IVIG) or interleukin 1 receptor antagonist (Anakinra) with limited and transient benefit. Separate or combined therapy with subcutaneous immunoglobulin (SCIG) and interleukin (IL) 1 inhibitor (Rilonacept) for refractory pericarditis in a cohort of patients has not been previously described.

    Case Descriptions: 4 patients were referred for recurrent pericarditis refractory to traditional therapies at ages ranging from 16 to 54 years. They all had multiple serious sequelae of their pericarditis and abnormal immune parameters including hypogammaglobulinemia, poor responses to vaccines, poor mitogen induced lymphocyte proliferation, and/or B cell lymphopenia. The patients had varied past medical histories and associated conditions. Patients were started on IG, with some initiated on IVIG, though all were transitioned to hyaluronidase-facilitated SCIG (HYQVIA). Patients were then started on either Anakinra or Rilonacept with 3 patients continuing on Rilonacept and 1 remaining on Anakinra. All patients had complete or near complete resolution of their pericarditis on dual therapy for greater than 1 year. The markedly elevated IL1 prior to therapy seen in all of the patients normalized post-therapy. Some patients had elevated IL6 prior to therapy that also improved post-therapy. 1 patient who has also been diagnosed with Familial Mediterranean Fever (FMF) has stopped both therapies for greater than 1 year with no further episodes of her pericarditis.

    Discussion: 4 patients with recurrent refractory pericarditis and signs of immunodeficiency and autoinflammatory disease on laboratory testing responded to dual therapy with HYQVIA and Rilonacept or Anakinra resulting in resolution of pericarditis. Inflammasome and immune abnormalities may be implicated or associated with recurrent pericarditis and may respond to targeted therapies.

      Patient #1 Patient #2 Patient #3 Patient #4 Reference Range
    Age at first pericarditis episode (years) 16 54 57 21 N/A
    Pericarditis pre-therapy 6 acute episodes over 6 years Constrictive pericarditis for 6 years 2 episodes over 1 year Constrictive pericarditis for 12 years, signs of pericarditis on imaging starting at 5 years old N/A
    Pericarditis complications Pulmonary edema, ventricular tachycardia, myocarditis, depressed ejection fracture Empyema, congestive heart failure Cardiac arrest, pacemaker placed for complete heart block Pericardiocentesis N/A
    Therapies failed NSAIDs, steroids, azathioprine NSAIDs, steroids, colchicine NSAIDs, steroids, methotrexate Adalimumab, Mycophenolic acid, rituximab NSAIDs, steroids, colchicine N/A
    Age at starting therapy (years) 22 (IG,)
    22 (Rilonacept)
    60 (IG)
    63 (Anakinra)
    54 (IG)
    64 (Rilonacept)
    33 (IG)
    33 (Anakinra)
    34 (Rilonacept)
    N/A
    Pericarditis post-therapy No subsequent episodes Resolved on imaging No subsequent episodes 1 episode on Anakinra, none on Rilonacept; stopped IG and Rilonacept for past 18 months with no episodes N/A
    Time on therapy (years) 2 (IG)
    2 (Rilonacept)
    4 (IG)
    1 (Anakinra)
    11 (IG)
    1 (Rilonacept)
    2 (IG)
    1 (Rilonacept)
    N/A
    Associated conditions None Protein loosing enteropathy Rheumatoid arthritis, ITP, thyroid disease, CIDP Familial Mediterranean Fever N/A
    Immunodeficiency diagnosis Selective antibody deficiency with B cell lymphopenia Hypogammaglobulinemia with poor T cell function CVID with poor T cell function Selective antibody deficiency N/A
    CD3+ cells
    (cells/μL, %)
    1009, 89% 226, 53% 1588, 93% 1897, 80% 1087-2198, 65-80%
    CD4+ cells
    (cells/μL, %)
    657, 56% 66, 15% 1105, 65% 946, 39% 677-1401, 38-55%
    CD8+ cells
    (cells/μL, %)
    342, 29% 139, 32% 472, 28% 878, 37% 212-1007, 15-38%
    CD19+ cells
    (cells/μL, %)
    29, 3% 148, 37% 31, 2% 280, 12% 180-492,
    8-23%
    CD16+56+ cells
    (cells/μL, %)
    73, 7% 34, 8% 60, 3% 181, 8% 97-421,
    5-17%
    IgG (mg/dL) 995 423 492 1000 844-1912
    IgA (mg/dL) 193 124 225 137 68-423
    IgM (mg/dL) 83.2 25.5 15.3 72 50-196
    Streptococcal pneumonia titers post-vaccination poor adequate poor poor adequate
    Mitogen induced lymphocyte proliferation adequate poor poor adequate adequate
    IL1β (pg/mL) 39.8 (pre)
    <3.9 (post)
    >250 (pre)
    <3.9 (post)
    150.4 (pre)
    <3.9 (post)
    69.6 (pre)
    <3.9 (post)
    <3.9
    IL6 (pg/mL) 0.92 (pre) 373.76 (pre)
    4.8 (post)
    3547 (pre) 85 (off tx) 0.31-5
    Tumor necrosis factor-α (pg/mL) <1 (pre) 2 (pre) 1.8 (pre)
    2.84 (post)
    1.9 (pre)
    <5 (off tx)
    1.2-15.3
    Interferon-Υ (pg/mL) <5 (pre) N/A N/A <5 (off tx) <=5
    Genetic testing negative negative not performed MEFV V726A heterozygous negative

    Table 1. Clinical characteristics and immune evaluation for 4 patients.

    (66) Submission ID#600335

    Characterization of Gut Inflammation and Autoimmunity in Mice Carrying Rag1 Hypomorphic Mutations

    Riccardo Castagnoli, MD1, Marita Bosticardo, PhD2, Rosita Rigoni, PhD3, Elena Fontana, PhD4, Ottavia M Delmonte, MD, PhD5, Lisa M Ott de Bruin, MD6, John P Manis, MD7, Cristina Corsino8, Yu Han, PhD9, Emilia Falcone, MD10, Anna Villa, MD11, Luigi D. Notarangelo, MD, PhD12

    11 Laboratory of Clinical Immunology and Microbiology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD. 2 Department of Pediatrics, University of Pavia, Pavia, Italy

    2Staff Scientist, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    33 San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), Division of Regenerative Medicine, Stem Cells and Gene Therapy, San Raffaele Scientific Institute, Milan, Italy

    44 Humanitas Clinical and Research Institute, Rozzano, Italy. 8 Milan Unit, Istituto di Ricerca Genetica e Biomedica, Consiglio Nazionale delle Ricerche, Milan, Italy

    5Staff Clinician, 1 Laboratory of Clinical Immunology and Microbiology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD

    65 Department of Pediatric Immunology, Wilhelmina Children's Hospital, Utrecht University Medical Center, Utrecht, The Netherlands

    76 Department of Laboratory Medicine, Boston Children's Hospital, Harvard Medical School, Boston, MA

    81 Laboratory of Clinical Immunology and Microbiology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD

    9Research Associate, Immunopathogenesis Section, Laboratory of Clinical Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, USA

    107 Immunopathogenesis Section, Laboratory of Clinical Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, USA

    11Clinician, Head of Unit, San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), Division of Regenerative Medicine, Stem Cells and Gene Therapy, San Raffaele Scientific Institute, Milan, Italy. 8 Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, Milan, Italy

    12Chief, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    Hypomorphic Recombination Activating Gene 1 (RAG1) mutations result in residual T- and B-cell development in both humans and mice and have been found in patients presenting with delayed-onset combined immune deficiency with granulomas and/or autoimmunity (CID-G/AI). Recent studies have shed light on how hypomorphic RAG1 mutations alter the primary repertoire of T and B cells, but less is known about their effect on immune dysregulation in targeted organs. In order to investigate the role of these mutations in determining intestinal disease, we set out to evaluate gut immunity and microbiota interplay in Rag1 mutant hypomorphic mice.

    We evaluated two mouse models carrying homozygous Rag1 mutations (R972Q and R972W), corresponding to human mutations (R975Q and R975W, respectively) described in patients with CID-G/AI. Both mutations fall in the coding flanksensitive region of the RAG1 C-terminal domain. On the basis of aminoacid properties and in vitro studies, the R972Q mutation has demonstrated a moderate effect on Rag1 protein stability while the R972W mutation resulted highly disruptive.

    Analysis of intestinal pathology in Rag1 mutant mice (NIAID animal protocol LCIM 6E) revealed different degrees of spontaneous colitis, with the most severe inflammatory infiltrate observed in mice carrying the most disruptive mutation, R972W. Colonic inflammation was characterized by crypt elongation, epithelial hyperplasia, and an abundant inflammatory infiltrate extending to the colonic lamina propria, with occasional crypt abscesses. A significant increase in activated CD44hiCD62LCD4+ T cells expressing the gut homing receptor 47 was observed in mesenteric lymph nodes (MLNs) of both mutant strains, and was especially prominent in R972W mutant mice. Additionally, the proportion of MLN CD4+ T regulatory (Treg) cells was increased in both mouse models. Finally, MLN of mutant mice contained a high number of myeloid cells (CD11b+) along with a decreased number of B220+ B cells, and these abnormalities were also more prominent in R972W than in R972Q mice.

    In summary, we have shown that Rag1 mutant hypomorphic mice present with different degrees of inflammatory bowel disease, with the mouse model carrying the most disruptive mutation presenting with the most severe phenotype. We are currently performing studies to evaluate the impact of Rag1 mutations on microbiome composition and diversity in these mouse models of CID-G/AI.

    (67) Submission ID#600337

    Immunomodulatory Effects of Immunoglobulin Replacement Therapy on T-cells in Patients with Hypogammaglobulinemia

    Tri M. Dinh, BSc1, Jun Oh, MSc2, Bill Cameron, MD, FRCPC, FACP3, Seung-Hwan Lee, PhD4, Juthaporn Cowan, MD, PhD, FRCPC5

    1Honour's Research Student Associate, University of Ottawa

    2PhD Candidate, University of Ottawa

    3Medical Director for Clinical Research, Ottawa Health Research Institute

    4Associate Professor, University of Ottawa

    5Associate Scientist and Assistant Professor, University of Ottawa, Ottawa Health Research Institute

    Background: Hypogammaglobulinemia or low serum immunoglobulin G (IgG) levels either inherited (primary) or acquired (secondary) is associated with increased infection rates. Primary (1°) hypogammaglobluinemia can be caused by many primary immune deficiencies (PID) including combined variable immune deficiency (CVID), while secondary (2°) hypogammaglobluinemia can be caused by many acquired conditions such as lymphomas, leukemias, or chemotherapies and other immunosuppressive drugs. Immunoglobulin replacement therapy (IRT) has been the mainstay of treatment in patients with hypogammaglobulinemia by reducing infection through replenishing the quantitative IgG. There are other applications of Ig therapy such as in autoimmune diseases, where the mechanism of action is thought to be Ig mediated immunomodulation. Innate immune cells have shown to be involved in such mechanism, but whether IRT modulates adaptive immune cells in patients with hypogammaglobulinemia is not well known.

    Hypothesis: IRT has an immunomodulatory effect on T-cell function and proliferation in patients with hypogammaglobulinemia.

    Methods: Blood from thirty patients with 1°(n=12) or 2° (n=18) hypogammaglobulinemia recruited from the Immunodeficiency Clinic at the Ottawa Hospital was drawn for peripheral blood mononuclear cell (PBMC) isolation, before starting IRT and minimum 8 weeks after starting IRT. Data regarding IgG level, number and type of infections after receiving IRT was collected. PBMCs were analyzed using flow cytometry for quantitation of T-cell subset. Cultured and anti-CD3/CD28 stimulated PBMC were also analyzed for extracellular and intracellular cytokine production, measured by ELISA and flow cytometry, respectively. Combined Cytomegalovirus, Epstein-Barr Virus and Influenza virus (CEF) peptides were used to study specific T-cell responses. Anti-CD3/CD28 stimulated PBMC were used for CellTrace T-cell proliferation assays. Data was grouped based on nature of hypogammaglobulinemia i.e. 1° or 2°. Results were compared between before and after IRT using Wilcoxon matched-pairs signed rank test.

    Results: IRT was not found to significantly alter proportion of Treg, CD4+, or CD8+ T-cell populations or activation state as measured by CD45RA/R0 expression. However, IRT was found to significantly increase expression of intracellular IFN-y in CD4+ and CD8+ T-cells post-CD3/CD28 stimulation in 2° (p = 0.007), but not in 1° hypogammaglobulinemia patients. There was no change in extracellular IL-10 and IL-17 cytokine production in both groups. In contrast, CD8+ T-cells in 1° hypogammaglobulinemia patients showed significantly higher expression of intracellular IFN-y and TNF-a post-CEF viral peptide stimulation (p = 0.027). CD3+ and CD8+ T-cell proliferation after CD3/CD28 stimulation was found to be decreased after IRT for both groups (p = 0.025 & p = 0.049).

    Conclusions: Our results suggest that IRT can alter CD4+ and CD8+ T-cell function with differential effect in patients with 1° or 2° hypogammaglobulinemia in addition to replenishing serum IgG level. More experiments assessing cytotoxicity of T-cells will be conducted to further study T-cell subset function as well as B-cell function. These laboratory results will be analyzed for association with clinical outcomes.

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    (68) Submission ID#600349

    Severe Congenital Neutropenia Caused by ELANE Gene Mutation in a Malaysian Girl

    Siti Mardhiana Binti. Mohamad, MD, PhD1, Intan Juliana Abdul Hamid, MD, MMed, PhD2, Asrul Abdul Wahab, MD, MPath3, Adli Ali, MD,MMed4, Choo Chong Ming, MBBS, MMed5, Florence Bakon, MD, MMed6, Amir Hamzah Abdul Latiff, MBBS, MMed, MRCP, FACAAI, FAAAA7, Lokman Mohd Noh, MBBS, DCH,MRCP,FRCPE,Cert. Fellowship Immunology8

    1Clinical Scientist, Primary Immunodeficiency Diseases Group, Regenerative Medicine Cluster, Institut Perubatan & Pergigian Termaju, Universiti Sains Malaysia

    2Paediatric Immunologist, Primary Immunodeficiency Diseases Group, Regenerative Medicine Cluster, Institut Perubatan & Pergigian Termaju, Universiti Sains Malaysia

    3Immunopathologist, Universiti Kebangsaan Malaysia, Kuala Lumpur, Malaysia

    4Paediatrician, Universiti Kebangsaan Malaysia, Kuala Lumpur, Malaysia

    5Paediatric Infectious Diseases, Hospital Sultan Abdul Halim, 08000 Sungai Petani, Kedah

    6Paediatrician, Kuching Specialist Hospital, Kuching, Sarawak

    7Clinical Immunologist and Allergy, Allergy and Immunology Centre, Pantai Hospital, Kuala Lumpur, Malaysia

    8Paediatric Immunologist Consultant, Hospital Kuala Lumpur, Jalan Pahang, 50586 Kuala Lumpur, Wilayah Persekutuan

    Background: Severe congenital neutropenia (SCN) is a rare immunodeficiency disorder characterised by the extremely low absolute neutrophils count (ANC) less than 0.5x109/L. The clinical feature of SCN is recurrent bacterial infections and the patients the risk of leukemia development. The incidence of SCN is estimated to be 1 in 200 000 individuals. Mutations in more than 20 genes have been described causing SCN and it is either recessive, dominant or X-linked inheritance.

    Case presentation: We described an 11 years old Malaysian girl who presented with recurrent abscesses over the whole part of the body, recurrent oral candidiasis, growth failure and recurrent pneumonias since 4 months old. She also had history of a few episodes of acute tonsillitis, chronic suppurative otitis media and herpes zoster infections. Throughout her age, she had persistent neutropenia less than 0.5x109/L but in few occasions, her ANC elevated up to more than 1.0x109/L . She was treated as autoimmune neutropenia, respectively due to few positive results of autoimmunity workout such as antinuclear antibodies (ANA) and double stranded DNA (dsDNA) but eventually later to be negative. Later at the age 9 years old, whole exome sequencing was performed and confirmed by Sanger sequencing, found a heterozygous variant in ELANE gene(c.640G>T; p.Gly214Ter), an autosomal dominant which was described to cause SCN. Both parents do not carry this mutation, hence, it is a de novo mutation. Currently, she had few on and off recurrent infections. Despite that, she is relatively well and on prophylaxis antibiotic.

    Conclusion: To our knowledge, we report for the first time a Malaysian girl with SCN, with confirmed mutational analysis of the ELANE gene. The delayed diagnosis might be due to the insufficient awareness of the phenotypic presentation of this rare disease. Moreover, the genetic analysis is not available in Malaysia and need to be done outside of the country. This case demonstrates the importance of the genetic analysis which may help in improving the diagnosis and management of the patient.

    (69) Submission ID#600360

    Autologous Ex Vivo Lentiviral Gene Therapy for the Treatment of Severe Combined Immune Deficiency Due to Adenosine Deaminase Deficiency

    Donald B. Kohn, MD, MS, BS1, Kit L. Shaw, PhD2, Elizabeth K. Garabedian, MSLS, RN3, Denise A. Carbonaro-Sarracino, PhD4, Theodore B. Moore, MD5, Satiro De Oliveira, MD6, Gay M. Crooks, MBBS7, John Tse, PharmD8, Sally Shupien, BA9, Dayna Terrazas, RN10, Alejandra Davila, BS11, Amalia Icreverzi, PhD12, Allen Yu, BS11, Provaboti Barman, PhD13, Maritess Coronel, MS14, Beatriz Campo Fernandez, PhD, MSc, BSc15, Ruixue Zhang, Master16, Roger Hollis, PhD17, Chilenwa Uzowuru, MSc, BSc18, Hilory Ricketts19, Jinhua Xu-Bayford, Degree (graduate diploma)20, Valentina Trevisan, MD21, Serena Arduini, PhD22, Frances Lynn, MSc23, Mahesh Kudari, MBBS, MA24, Andrea Spezzi, MD, MBBS25, Lilith Reeves, MS, MT(ASCP)26, Kenneth Cornetta, MD27, Robert Sokolic, MD, FACP28, Roberta Parrott, BS29, Rebecca Buckley, MD30, Claire Booth, MBBS, PhD, MSc31, Fabio Candotti, MD, PhD32, Harry L. Malech, MD33, Adrian J. Thrasher, MBBS, PhD34, H Bobby Gaspar, MD, PhD35

    1Professor of Microbiology, Immunology and Molecular Genetics (MIMG) and Pediatrics, University of California, Los Angeles

    2Study Manager for Gene Therapy Clinical Trials, University of California, Los Angeles

    3Research Nurse, Principal Investigator, National Genome Research Institute, National Institutes of Health

    4Senior Scientist, University of California, Los Angeles; Orchard Therapeutics, Boston MA

    5Professor of Pediatrics, Chief of Pediatric Hematology/Oncology and Director of the Pediatric Blood and Marrow Transplant Program at UCLA, University of California, Los Angeles

    6Assistant Professor, Pediatrician and Cancer Immunotherapy Researcher, University of California, Los Angeles

    7Professor, Pathology & Laboratory Medicine; Paediatric oncologist, Division of Stem Cell Transplantation and Regenerative Medicine, Department of Pediatrics, Stanford School of Medicine, Stanford, CA

    8Clinical Pharmacist, University of California, Los Angeles

    9Clinical Trials staff, University of California, Los Angeles

    10Clinical Research Nurse Coordinator, University of California, Los Angeles

    11SRAII, University of California, Los Angeles

    12 Head of Clinical Manufacturing and Gene Therapy, University of California, Los Angeles

    13Lead Manufacturing and QA at UCLA GMP (Human Gene and Cell Therapy Facility), University of California, Los Angeles

    14Regulatory and data manager, University of California, Los Angeles

    15Associate Project Scientist, University of California, Los Angeles

    16SRA, University of California, Los Angeles

    17Project Scientist VII, University of California, Los Angeles

    18Clinical Trial coordinator, University College London; Great Ormond Street Hospital NHS Trust

    19Data Manager, University College London; Great Ormond Street Hospital NHS Trust

    20Gene Therapy and Immunology CNS Team lead, University College London; Great Ormond Street Hospital NHS Trust

    21Clinical Research Fellow, University College London; Great Ormond Street Hospital NHS Trust

    22Clinical Development Scientist, Orchard Therapeutics, London, UK

    23Biostatistician, Orchard Therapeutics, London, UK

    24Senior Director, Clinical Development, Orchard Therapeutics, London, UK

    25Chief Medical Officer, Orchard Therapeutics, London, UK

    26Assistant Professor; Translational Core Director, Cincinnati Childrens Hospital Medical Center

    27Professor of Clinical Medical & Molecular Genetics, Indiana University School of Medicine

    28Hematologist/Oncologist at Lifespan Cancer Institute, National Genome Research Institute, National Institutes of Health; Comprehensive Cancer Center at Rhode Island Hospital

    29Research Associate, Duke University

    30Sidbury Professor of Pediatrics, in the School of Medicine; Professor of Immunology, Duke University

    31Consultant Paediatric Immunologist and senior clinical lecturer at Great Ormond Street Hospital for Children NHS Foundation Trust, University College London; Great Ormond Street Hospital NHS Trust

    32Associate Professor of Medicine and Head Physician, Division of Immunology and Allergy; Director, Vaccine and Immunotherapy Center, University Hospital of Lausanne, Switzerland, National Genome Research Institute, National Institutes of Health; Current Address: Division of Immunology and Allergy, University Hospital of Lausanne, Lausanne, Switzerland

    33Chief, Genetic Immunotherapy Section, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    34Professor of Paediatric Immunology and Wellcome Trust Principal Research Fellow, University College London; Great Ormond Street Hospital NHS Trust

    35Professor of Paediatrics and Immunology, University College London; Great Ormond Street Hospital NHS Trust; Orchard Therapeutics, London, UK

    Background: ADA-SCID is a rare genetic disorder which causes severe combined immunodeficiency. Historically, ADA-SCID has been treated using enzyme replacement therapy (ERT) followed by allogeneic hematopoietic stem cell (HSC) transplant (HSCT) from a matched related donor (MRD) or, if none is identified, a non-MRD (matched/mismatched unrelated or mismatched related donor). We developed a self-inactivating lentiviral vector (LV), in which a codon optimized human ADA cDNA is driven by the short form of the elongation factor-1alpha (EFS) promoter (EFS-ADA LV). The drug product (OTL-101), composed of autologous HSCs transduced ex vivo with the EFS-ADA LV, was evaluated in a prospective, historically-controlled Phase I/II clinical trial in ADA-SCID pediatric subjects. We report safety and efficacy at 24 months in 20 ADA-SCID subjects treated with lentiviral gene therapy (GT) compared to a historical cohort of 26 ADA-SCID patients treated with HSCT.

    Methods: Twenty subjects (9 male, 11 female; 4 mo 4.3 yrs) were treated with GT. Autologous CD34+ HSCs were isolated from bone marrow and pre-stimulated with cytokines before transduction with EFS-ADA LV. Busulfan was administered at a single dose (4 mg/kg) prior to infusion of OTL-101. The control group included 26 patients (0.2 mo 9.8 yrs) treated with allogeneic HSCT (MRDs n=12, non-MRDs n=14) at Great Ormond Street Hospital, UK (n=16) or Duke University Childrens Hospital, USA (n=10) between 20002016.

    Results: At 24 months, overall survival (OS) and event-free survival (EvFS), defined as survival in the absence of ERT reinstitution or rescue allogeneic HSCT) were statistically significantly higher in the GT group compared with the HSCT group (Table). Successful engraftment of genetically modified HSC was observed in all GT subjects at 6 months, which persisted over 24 months, based on vector gene marking in granulocytes (median 0.085 copies/cell [range 0.04-2.50] at 24 months) and peripheral blood mononuclear cells (median 0.843 copies/cell [range 0.13-1.86] at 24 months), and was associated with increased red blood cell ADA enzyme activity and metabolic detoxification from deoxyadenosine nucleotides.

    Over 24 months, none of the GT subjects required PEG-ADA ERT reinstitution and 90% were able to stop receiving immunoglobin replacement therapy (IgRT), whereas 38% HSCT patients required rescue HSCT or reinstitution of PEG-ADA ERT, and 52% were able to stop receiving IgRT (Table). Nine subjects in the GT group experienced a serious adverse event (SAE), most frequently infections and gastrointestinal events; only one was considered treatment-related. In the GT group, there were no events of autoimmunity during the study. Due to the autologous nature of the product, there was no incidence of graft vs host disease (GvHD) in the GT group; whereas 5 patients in the HSCT group experienced acute GvHD and 3 experienced chronic GvHD events, one of whom died.

    Conclusions: Treatment with lentiviral GT for ADA-SCID is well tolerated and has a favorable benefit-risk profile at 24 months based on sustained gene correction and restoration of immune function, as well as improved OS and EvFS compared with HSCT (MRD or non-MRD) at 24 months.

    Grant Support:

    Supported by a research grant from the NIAID, NIH (U01 AI100801), the National Gene Vector Biorepository (5P40HL116242), the California Institute for Regenerative Medicine (CL1-00505-1.2, FA1-00613-1), Medical Research Council (MR/K015427/1), and the National Institute for Health Research Biomedical Research Centre at Great Ormond Street Hospital for Children NHS Foundation Trust and University College London.

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    (70) Submission ID#600370

    Autologous ex vivo lentiviral gene therapy for the treatment of Severe Combined Immune Deficiency due to Adenosine Deaminase Deficiency (ADA-SCID) Improves B Cell Function

    Donald B. Kohn1, Kit L. Shaw1, Elizabeth Garabedian2, Denise A. Carbonaro-Sarracino1,3, Theodore B. Moore1, Satiro De Oliveira1, Gay M. Crooks1, John Tse4, Sally Shupien1, Dayna Terrazas1, Alejandra Davila1, Amalia Icreverzi1, Allen Yu1, Provaboti Barman1, Maritess Coronel1, Beatriz Campo Fernandez1, Ruixue Zhang1, Roger Hollis1, Chilenwa Uzowuru5, Hilory Ricketts5, Jinhua Xu Bayford5, Valentina Trevisan5, Serena Arduini3, Frances Lynn3, Mahesh Kudari3, Andrea Spezzi3, Lilith Reeves6, Kenneth Cornetta7, Robert Sokolic2,*, Roberta Parrott8, Rebecca Buckley8, Claire Booth5, Fabio Candotti2,**, Harry Malech9, Adrian J. Thrasher5, and H. Bobby Gaspar3,5

    1University of California, Los Angeles

    2National Genome Research Institute, National Institutes of Health

    3Orchard Therapeutics Limited

    4Department of Pharmaceutical Services, Ronald Reagan Medical Center, UCLA

    5University College London/Great Ormond Street Hospital

    6Cincinnati Children’s Hospital Medical Center

    7Indiana University School of Medicine

    8Duke University

    9National Institute of Allergy and Infectious Disease, National Institutes of Health

    *Current Address: Comprehensive Cancer Center at Rhode Island Hospital

    **Current Address: Division of Immunology and Allergy, University Hospital of Lausanne, Lausanne, Switzerland

    Grant Support:

    Supported by a research grant from the NIAID, NIH (U01 AI100801), the National Gene Vector Biorepository (5P40HL116242), the California Institute for Regenerative Medicine (CL1-00505-1.2, FA1-00613-1), Medical Research Council (MR/K015427/1), and the National Institute for Health Research Biomedical Research Centre at Great Ormond Street Hospital for Children NHS Foundation Trust and University College London.

    Background: ADA-SCID is a rare genetic disorder that causes severe combined immunodeficiency, with minimal or absent B cell function. Prior to, and often after, treatment with allogeneic hematopoietic stem cell (HSC) transplant (HSCT) or autologous ex vivo HSC gene therapy (GT), patients are managed with enzyme replacement therapy (ERT) and immunoglobulin (Ig) replacement therapy (IgRT). We evaluated a GT treatment with autologous HSCs transduced ex vivo with a self-inactivating lentiviral vector (LV), in which a codon optimized human ADA cDNA is driven by an internal short form of the elongation factor-1alpha (EFS) promoter (“EFS-ADA LV”). At 24 months follow-up, 20 pediatric ADA-SCID subjects treated with GT were compared to a historical cohort of 26 ADA-SCID patients treated with HSCT. Here, we report on B cell reconstitution in these cohorts.

    Methods: Twenty subjects (9 male, 11 female) aged 4 mo – 4.3 yrs received GT. Autologous CD34+ HSCs were isolated from bone marrow and pre-stimulated with cytokines before transduction with EFS-ADA LV. Genetically modified cells were administered after conditioning with single dose busulfan (4 mg/kg). The control group included 26 patients aged 0.2 mo to 9.8 yrs treated with HSCT at Great Ormond Street Hospital (UK) (n=16) or Duke University Children’s Hospital (US) (n=10) between 2000 - 2016. The HSCT patients received an allogeneic transplant from matched related donors (MRDs) (n=12) or non-MRDs (n=14). Subjects continued to receive IgRT post-GT until a clinical decision was made to stop, factoring in B cell reconstitution, general medical condition and seasonal infections.

    Results: By Month 12, in the GT group, 45% had stopped treatment with IgRT compared to 38% in the HSCT group overall. By Months 18 and 24, higher proportions of GT-treated subjects had stopped IgRT (70% and 90%, respectively) compared with MRD HSCT patients (55% and 70%, respectively) and non-MRD HSCT patients (42% at both timepoints) (Table).

    In the GT group, vector gene marking was detectable in peripheral blood mononuclear cells within 3 months and persisted at 24 months post-infusion (median 0.843 copies/cell [range 0.13-1.86]), suggesting successful gene modification. As evidence of B cell reconstitution, IgA and IgM levels in peripheral blood sera more than doubled by 18 months, from 18.5 mg/dL (range 8 to 95) to 48.0 mg/dL (range 20 to 110) and 32.5 mg/dL (range 16 to 107) to 69.0 mg/dL (range 20 to 180), respectively. Additionally, antibody response following tetanus vaccination, was evaluated in 3 subjects. All 3 subjects mounted a protective response to the vaccine (median antibody response 3.2 IU/mL [range 0.1 to 3.5]), based on a normal threshold of 0.01 IU/mL (Hammarlund Clin Infect Dis 2016) and a laboratory reference range (0.10 to 2.9 IU/mL).

    Conclusions: GT with autologous HSCs transduced ex vivo with EFS-ADA LV resulted in B cell reconstitution, as evidenced by doubled IgA and IgM production at 18 months, cessation of IgRT in 90% of patients by 24 months, and protective specific antibody responses to tetanus vaccine in patients that were evaluated.

    (71) Submission ID#600374

    Lentiviral Vector Gene Therapy for X-linked Chronic Granulomatous Disease Corrects Neutrophil Function

    Harry L. Malech, MD1, Claire Booth, MBBS, PhD, MSc2, Elizabeth M. Kang, MD3, Sung-Yun Pai, MD4, Kit L. Shaw, PhD5, Giorgia Santilli, PhD6, Myriam Armant, PhD7, Karen F. Buckland, PhD, BSc8, Uimook Choi, PhD, BSc9, Suk See De Ravin, MD, PhD10, Morna J. Dorsey, MD, MMSc11, Caroline Y. Kuo, MD12, Diego Leon-Rico, PhD, MS, BSc13, Christine Rivat, PhD8, Katie Snell, Dip (children's nursing), BSc14, Jinhua Xu-Bayford, Degree (graduate diploma)15, Emma C. Morris, MB BChir, MA, PhD, MRCP, FRCPath16, Dayna Terrazas, RN17, Leo D. Wang, MD, PhD18, Geraldine Honnet, MD19, Peter Newburger, MD, BA20, Frederic D. Bushman, PhD, BA21, Manuel Grez, PhD, BSc22, H Bobby Gaspar, MD, PhD23, David A. Williams, MD24, Anne Galy, PhD25, Donald B. Kohn, MD, MS, BS26, Adrian J. Thrasher, MBBS, PhD27

    1Chief, Genetic Immunotherapy Section, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    2Consultant Paediatric Immunologist and senior clinical lecturer at Great Ormond Street Hospital for Children NHS Foundation Trust, University College London; Great Ormond Street Hospital NHS Trust

    3Staff Clinician and Chief of the Hematotherapeutics Unit of Genetic Immunotherapy Section, National Institute of Allergy and Infectious Diseases, National Institutes of Health

    4Associate Professor of Pediatrics, Harvard Medical School, Boston Childrens Hospital, Harvard Medical School

    5Study Manager for Gene Therapy Clinical Trials, University of California, Los Angeles

    6Non-clinical lecturer in gene therapy, University College London; Great Ormond Street Hospital NHS Trust

    7Instructor in Pediatrics, Boston Childrens Hospital, Harvard Medical School

    8Healthcare Scientist, University College London; Great Ormond Street Hospital NHS Trust

    9Staff Scientist, National Institute of Allergy and Infectious Diseases, National Institutes of Health

    10Clinician, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    11Pediatric Immunologist and Allergist, Department of Pediatrics, Division of Allergy, Immunology, and Bone Marrow Transplant, University of California San Francisco, San Francisco, CA

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    12Assistant Clinical Professor of Pediatrics, University of California, Los Angeles

    13Senior Research Associate, University College London; Great Ormond Street Hospital NHS Trust

    14Lead gene therapy research nurse, University College London; Great Ormond Street Hospital NHS Trust

    15Gene Therapy and Immunology CNS Team lead, University College London; Great Ormond Street Hospital NHS Trust

    16Haematologist and Professor of Haematology, University College London; Great Ormond Street Hospital NHS Trust

    17Clinical Research Nurse Coordinator, University of California, Los Angeles

    18Assistant Professor, Department of Immuno-Oncology, City of Hope National Medical Center

    19Director of Development, Genethon, Evry, France

    20Professor of Hematology, University of Massachusetts, Worcester

    21Professor of Microbiology, University of Pennsylvania

    22Research Group Leader, Georg-Speyer Haus, Frankfurt, Germany

    23Professor of Paediatrics and Immunology, University College London; Great Ormond Street Hospital NHS Trust; Orchard Therapeutics, London, UK

    24President, Dana-Farber/Boston Children's Cancer and Blood Disorders Center, Boston Childrens Hospital, Harvard Medical School

    25Head of Inserm Unit 951, Genethon, Evry, France

    26Professor of Microbiology, Immunology and Molecular Genetics (MIMG) and Pediatrics, University of California, Los Angeles

    27Professor of Paediatric Immunology and Wellcome Trust Principal Research Fellow, University College London; Great Ormond Street Hospital NHS Trust

    Background: X-linked Chronic Granulomatous Disease (XCGD) results from mutations in CYBB encoding the gp91phox subunit of phagocyte NADPH-oxidase. Attempts to treat XCGD with gene therapy (GT) using transduced autologous hematopoietic stem cells (HSC) transduced ex vivo with a gammaretroviral vector have met with limited efficacy due to transient engraftment of gene corrected HSCs, gene silencing, and vector insertion-mediated activation of oncogenes leading to myelodysplasia. We developed a novel self-inactivating (SIN) lentiviral vector (G1XCGD LV) with a chimeric cathepsin G/cFes myeloid-specific promoter driving gp91phox expression from a codon optimized cDNA. Following transplant of G1XCGD LV ex vivo transduced autologous HSCs into busulfan-conditioned XCGD patients, there was long-term restoration of oxidase activity in peripheral blood polymorphonuclear neutrophils (PMN) at 12 months in 6 of 9 severely affected XCGD patients without evidence of genotoxicity. Here we present data about the multiple assays used to assess quality and quantity of restoration of PMN oxidase activity.

    Methods: Similar trials of GT with G1XCGD LV were initiated in the UK (n=3, plus 1 compassionate use patient) and USA (n=5). All patients had histories of inflammatory disease and severe, persistent infections (some non-responsive to conventional therapy at time of GT). G-CSF plus Plerixafor-mobilized CD34+ HSCs were transduced with ex vivo G1XCGDLV. Subjects received myeloablative conditioning with single-agent busulfan, targeted to net area-under-the-curve of 70,000 ng/mL*hr. Freshly prepared or cryopreserved quality-tested genetically-modified HSC, manufactured on-site, were administered intravenously. PMN oxidase activity post-GT was assessed by p-nitroblue tetrazolium (NBT) reduction, dihydrorhodamine (DHR) flow cytometry assay, and quantitative Ferricytochrome C Assay (FerriC) measurement of superoxide generation.

    Results: We report results for 7 patients (aged 2-27 years) with 1-2.5 years of follow-up; two additional patients were treated but died within three months of GT from complications deemed related to pre-existing disease-related co-morbidities (severe pulmonary disease and anti-platelet antibodies). Within 1 month post-GT, oxidase (+) PMN were present in peripheral blood based on NBT testing and DHR flow cytometry. Expression of the corrective transgene was confirmed by flow cytometry using antibody detection of gp91phox. Quantitative biochemical measurements of oxidase activity were also confirmed in some samples using the FerriC assay, demonstrating quantitative levels of superoxide production per corrected cell that were within the normal range. Functional testing of oxidase burst activity using DHR fluorescent assays was applied serially to follow levels of corrected PMN where oxidase activity per corrected cell also were in the normal range. All patients had >15% PMN DHR+ within one month, which remained stable for most patients over the follow-up period (Figure). Follow-up demonstrated sustained stable persistence of 12-46% oxidase burst positive neutrophils in 6 of 7 surviving subjects at 12 months, with restoration to clinically beneficial levels (defined as 10% of PMN being DHR+) in these patients as of December 2018.

    Conclusion: These results demonstrate corrected PMN function within 1 month in X-CGD patients treated with autologous GT. PMN oxidase activity was sustained at levels which restore biochemical function and provide clinically beneficial levels of immunity for 12 months in 6/7 patients.

    Grant Support:

    Supported by research grants from the: California Institute of Regenerative Medicine (CLIN2-08231; FA1-00613-1), the Gene Therapy Resource Program from NHLBI, NIH (CRB-SSS-S-15-004351 1840), the

    NIAID Intramural Program, NET4CGD (FP7 EU grant agreement no. 305011), the Wellcome Trust (104807/Z/14/Z), and the National Institute for Health Research Biomedical Research Centre at Great Ormond Street Hospital for Children NHS Foundation Trust and University College London.

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    (72) Submission ID#600426

    A New High Concentration Immunoglobulin Product for Subcutaneous Administration (IGSC 20%)

    William Alonso1, John Lang2, Pete Vandeberg3

    1Principe Process Development Scientist II, Grifols Bioscience Research Group

    2Principle Research Scientist I, Grifols Bioscience Research Group

    3R&D Program Director, Grifols Bioscience Research Group

    Introduction/Background: Grifols has developed a new 20% immunoglobulin liquid product for subcutaneous administration (IGSC 20%). The formulation for IGSC 20% was developed based on the knowledge acquired from the formulation of Grifols currently licensed 10% Immune Globulin (Human), Gamunex®-C; however, the protein concentration was increased from 10% to 20% to facilitate efficient subcutaneous administration. Gamunex-C has an extensive record of safety and tolerability when administered intravenously and subcutaneously for greater than 15 years in diverse patient populations. The IGSC 20% manufacturing process employs the same purification steps as Gamunex-C and was demonstrated to be robust and to provide an IgG product with the required potency, purity, and quality. The formulation excipient characteristics and compatibility with the drug product have been well established. Glycine has been an excipient of Intramuscular Immune Globulin (Human) for fifty years and Intravenous Immune Globulin (IGIV) for over twenty years. The IGSC 20% formulation has low buffering capacity, and a low pH was selected to achieve a product with low aggregates, low fragments and viscosity suitable for subcutaneous administration. To improve visual clarity, the IGSC 20% formulation contains a small amount of polysorbate 80 (PS80), which is widely used in biopharmaceutical products. Subcutaneous administration of the IGSC 20% formulation has been well tolerated in clinical studies.

    Objectives: The goal was to provide the PID population with a new 20% immunoglobulin liquid product for subcutaneous administration (IGSC 20%).

    Methods: IGSC 20% is manufactured using the current manufacturing process for Gamunex-C, followed by an additional concentration step so that the product can be formulated at a higher protein concentration. IGSC 20% and Gamunex-C batches were produced at full industrial scale and then subjected to a series of analytical testing including assessment of purity, composition and neutralizing activity.

    Results: The IGSC 20% and Gamunex-C manufacturing processes and formulations have preserved the IgG integrity, molecular characteristics and potency. The manufacturing processes have eliminated lipids, alcohols, and acetate and coagulation factor impurities, including FXIa, which were undetectable by either specific or global methods. The IGSC 20% and Gamunex-C batches were 100% gamma globulin by agarose membrane electrophoresis, and have a subclass distribution similar to normal plasma and acceptable specific antibody content. IGSC 20% was shown to be primarily monomer plus dimer IgG (99±1%) with minimal aggregate or fragment, which confirms that appropriately gentle processing conditions were used during the concentration of 10% IgG solutions to 20% IgG.

    Conclusions: IGSC 20% is a highly concentrated IgG solution with characteristics comparable to Gamunex-C, but with twice the IgG concentration in order to facilitate subcutaneous administration with reduced volumes and shorter infusion times. Analytical testing demonstrates suitable potency, purity, and neutralizing activity for a number of specific antigens.

    Funding: This study was funded and conducted by Grifols, a manufacturer of 20% immunoglobulin for subcutaneous administration.

    Disclosure: All authors are employees of Grifols.

    (73) Submission ID#600434

    Miller-Dieker Syndrome May Be Another Syndromic Primary Immunodeficiency

    Erika Tsutsui, MD1, Deepti Deshpande, MD, MPH2, Yesim Demirdag, MD3

    1Resident, The University of Tokyo Hospital

    2Fellow, Division of Allergy, Immunology and Rheumatology, Department of Pediatrics, Columbia University Medical Center

    3Faculty, Division of Allergy, Immunology and Rheumatology, Department of Pediatrics, Columbia University Medical Center

    INTRODUCTION: Miller-Dieker Syndrome (MDS) is a contiguous gene deletion on chromosome 17p13.3, characterized by lissencephaly, distinctive facial features and severe intellectual disability and seizures. Frequent respiratory tract infections and seizures cause recurrent hospitalizations in these children and are typically considered a result of neurological impairment and poor airway clearance. Evaluation of these patients for immunodeficiency is not a common clinical practice. Here we report combined immune deficiency in 2 patients with MDS and recurrent respiratory tract infections.

    CASE PRESENTATION

    Case 1: A boy with MDS was initially referred at age 2 months for an abnormal newborn screen with low T cell receptor excision circles (TREC) for severe combined immunodeficiency (SCID). Initial evaluation revealed moderate CD3+ and CD4+ T cell lymphopenia (figure 1). Initial immunoglobulins levels were normal. He was placed on anti-seizure medications. He later developed recurrent and severe respiratory tract infections starting in infancy. At 12 months of age, he developed hypogammaglobulinemia (figure 2). In addition, T cell counts progressively decreased and stayed around 600 cells/ul. Immunoglobulin replacement therapy started at 18 months of age. Hospitalizations due to respiratory tract infections significantly decreased.

    Case 2: A 3-year-old boy with MDS had recurrent bacterial and viral respiratory infections which required numerous hospitalizations including intensive care unit stays. Newborn screening for SCID was negative. He had been on anti-seizure medications. Immunologic evaluation at 3 years of age revealed low total CD3+ cells and CD8+ T cells (CD3+: 1284cells/uL[normal range 1400-3700cells/uL], CD8+:278cells/uL[normal range 490-1300cells/uL]), hypogammaglobinemia (IgG: 252mg/dL[normal range 453-916mg/dL]), and non-protective IgG levels to tetanus, varicella and pneumococcus serotypes. Immunoglobulin replacement therapy started at 3 years of age which resulted in reduced frequency and severity of respiratory infections, and improved quality of life.

    DISCUSSIONS: T cell lymphopenia and hypogammaglobulinemia were seen in both our cases of Miller-Dieker Syndrome. To our knowledge, immune deficiency has never been reported in MDS. One of our cases suggests that low T cell counts may start as early as at birth and may be detected by newborn screening. Hypogammaglobulinemia may be primary or secondary due to antiepileptics. Both children had reduced frequency and severity of respiratory infections and improved quality of life after immunoglobulin replacement highlighting the importance of screening and early management of immunodeficiency.

    CONCLUSION: Miller-Dieker Syndrome is likely another syndromic primary immune deficiency disorder. A high index of suspicion with early screening and management of immunodeficiency may be beneficial for children with Miller-Dieker Syndrome.

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    (74) Submission ID#600442

    Safety and Pharmacokinetics of IGSC 20% in Subjects with Primary Immunodeficiency in an Open-label, Multicenter, Phase 3 Study

    John Sleasman, MD1, Amy Darter, MD2, William Lumry, MD3, Iftikhar Hussain, MD4, H. James Wedner, MD5, James Harris, III, MD6, Kecia Courtney7, Elsa Mondou, MD8, Jiang Lin, PhD9, Mark R. Stein, MD10

    1Professor of Pediatrics, Division of Allergy, Immunology, and Pulmonary Medicine, Duke University School of Medicine

    2Physician, Oklahoma Institute of Allergy & Asthma Clinical Research, LLC

    3Physician, Allergy and Asthma Specialists

    4President and Principal Investigator, Vital Prospects Clinical Research Institute, P.C., and Allergy, Asthma and Immunology Center, P.C.

    5Chief, Division of Allergy and Immunology, Washington University Physicians, Washington University School of Medicine in St. Louis

    6Physician, Allergy & Immunology, The South Bend Clinic Center for Research

    7Director, Clinical Development, Grifols Bioscience Research Group

    8Medical Director II, Grifols Bioscience Research Group

    9Biostatistician III, Grifols Bioscience Research Group

    10Physician, Allergy & immunology, Allergy Section, Good Samaritan Medical Center, West Palm Beach, FL, USA

    This prospective, multi-center, open-label study assessed the pharmacokinetic (PK), safety, and tolerability of Immune Globulin Subcutaneous (Human), 20% Caprylate/Chromatography Purified (IGSC 20%) in subjects with primary immunodeficiency (PI). The objectives were to determine a weekly subcutaneous (SC) dose of IGSC 20% that is noninferior to the intravenous (IV) dose of Immune Globulin Injection (Human), 10% Caprylate/Chromatography Purified (IGIV-C 10%) and to determine the steady state trough IgG levels after IGSC 20% and IGIV-C 10% infusions. There were 3 possible phases. If not on a qualifying IgG regimen at enrollment, subjects (n=44) were required to enter the Run-In Phase, receiving IGIV-C 10% to achieve steady-state before entering the IV Phase to determine steady-state area-under-the-curve (AUC) of IV infusions. Subjects with a qualifying IGIV-C 10% regimen (300-800 mg/kg) (n=9) directly entered the IV Phase for steady-state IV PK assessments. Upon completion of the IV PK assessments subjects entered the SC Phase, receiving weekly doses of IGSC 20% for up to24 weeks, with steady-state AUC determined at the 13th dose. IGSC 20% was not associated with any reports of serious local infusion site reactions (ISRs). The majority of local ISRs were mild-to-moderate. IGSC 20% (at a dose conversion factor of 1.37) provided equivalent exposure to IGIV-C 10% as assessed by steady-state AUC0-7 days, with 33% higher mean IgG trough values, lower fluctuations in IgG concentrations and the flexibility of at home administration. IGSC 20% was well tolerated with a safety profile comparable to IGIV-C 10%.

    ClinicalTrials.gov Identifier: NCT02604810

    Disclosure: Kecia Courtney, Elsa Mondou, and Jiang Lin are employees of Grifols, a manufacturer of IGSC 20%. Grifols is the sponsor of this study.

    (75) Submission ID#600556

    Deficiency of Adenosine Deaminase 2: An Expanding Spectrum of Disease

    Jenna Bergerson, MD/MPH1, Karyl Barron, MD2, Deborah Stone, MD3, Patrycja Hoffmann, MSN, FNP4, Natalia Sampaio Moura, BS5, Oskar Schnappauf, PhD6, Ivona Aksentijevich, MD7, Daniel Kastner, MD, PhD8, Amanda Ombrello, MD3

    1Staff Clinician, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    2Deputy Director, DIR, NIAID

    3Staff Clinician, NIH/NHGRI/Inflammatory Disease Section

    4Commissioned Corps, NIH/NHGRI/Inflammatory Disease Section

    5Post Baccalaureate IRTA, NIH/NHGRI/Inflammatory Disease Section

    6Postdoctoral Fellow, NIH/NHGRI/Inflammatory Disease Section

    7Staff Scientist, NIH/NHGRI/Inflammatory Disease Section

    8Scientific Director, NHGRI, NIH/NHGRI/Division of Intramural Research

    Background: In 2014 two reports described the deficiency of adenosine deaminase 2 (DADA2) as early-onset lacunar strokes, intermittent fevers, livedoid rash, and early onset polyarteritis nodosa (PAN). Since these first reports, the clinical spectrum has dramatically expanded to include antibody deficiency, liver disease, vasculopathy, pure red cell aplasia, cytopenias, and lymphoproliferative disease.

    Methods: Forty-two patients were enrolled in an IRB approved study at the NIH. Sequencing of ADA2, the gene encoding adenosine deaminase 2 (ADA2), was performed in all patients. Information was obtained by chart review of all clinical, serologic, and radiographic testing.

    Results: All 42 patients had germline biallelic loss of function mutations in ADA2, leading to absent or significantly decreased protein expression and function of ADA2. The cohort comprises 20 females (48%) and 22 males (52%). There were 6 sibling pairs and 2 families with 3 affected individuals. Twenty-seven patients had a history of at least one ischemic stroke and 6 experienced a hemorrhagic stroke. The average age at the time of first stroke is 5.6 years (range 4 months - 24 years), and the average number of strokes is 3 (range 1-11). No new strokes have occurred in patients on anti-TNF therapy. Skin manifestations occurred in 86% of patients and include livedo (74%), cutaneous vasculitis resembling PAN (64%), and Raynauds (19%). Hepatomegaly (43%) and splenomegaly (55%) were also notable. Portal hypertension was observed in 6 (14%) patients, with 1 patient requiring a spleno-renal shunt for a massive variceal bleed. Abdominal MRA revealed arteritis and aneurysm in 7/13 patients evaluated; 3 patients developed bowel necrosis. Peripheral vasculopathy was seen in 3 patients, with one requiring amputation of gangrenous digits.

    The most common immune abnormality seen in this cohort is hypogammaglobulinemia (62%); 20 patients have low IgG, 20 patients have low IgM, and 14 patients have low IgA. Ten of these patients are on immunoglobulin replacement. Specific antibody responses to vaccines were inadequate in 5/16 patients challenged. Lymphocyte phenotyping revealed decreased class-switched memory B cells in 23/32 patients (72%) tested. However, there was no relationship between absolute number of class switched memory B cells and hypogammaglobulinemia or infection frequency. Hematologic abnormalities include transfusion depended anemia (7%), neutropenia (7%), lymphopenia (5%), and thrombocytopenia (2%). Seven patients developed pancytopenia, 1 presented with pure red cell aplasia, and 1 developed aplastic anemia. Three patients have undergone bone marrow transplant, with two of those patients requiring a second transplant for graft failure.

    Conclusions: The spectrum of DADA2 has expanded from strokes, intermittent fever, and cutaneous manifestations to include portal and systemic hypertension, immune deficiency, cytopenias, vascular abnormalities, and bone marrow failure. While initiation of anti-TNF therapy improves inflammatory markers, and no new strokes have occurred while on therapy, cytopenias do not seem to improve. Bone marrow transplantation should be considered in patients with findings of bone marrow failure, although transplant of our patients has been complicated by immune mediated neutropenia. Disease manifestations are heterogenous, making a comprehensive evaluation critical to our understanding of this disease.

    (76) Submission ID#600606

    Guidance for the Care of Patients Undergoing Cultured Thymus Tissue Transplantation (RVT-802)

    Stephanie E. Gupton, MSN, CPNP1, Elizabeth A. McCarthy, RN, MSN2, Mary Louise. Markert, MD, PhD3

    1Nurse Practitioner, Department of Pediatrics, Division of Allergy, Immunology and Pulmonary, Duke University Medical Center

    2Research Program Leader, Sr., Department of Pediatrics, Division of Allergy, Immunology and Pulmonary, Duke University Medical Center

    3Professor of Pediatrics and Immunology, Department of Pediatrics, Division of Allergy, Immunology and Pulmonary, Duke University Medical Center

    Cultured thymus tissue transplantation (RVT-802) is an investigational therapy used to treat athymia or other conditions with severely diminished thymic function. Since 1993, 97 transplants of RVT-802 have been performed under the direction of Dr. M. Louise Markert. The overall survival rate after RVT-802 is 71% with most deaths secondary to pre-existing infections, cardiac defects and/or respiratory conditions. With the advent of widespread newborn screening for primary immunodeficiency, the average number of patients referred for RVT-802 implantation is 18 per year. Given the increase in neonatal diagnosis of athymia, clinical care is provided by the referring medical centers prior to RVT-802 implantation and patients return to the referring centers earlier after RVT-802. This creates the need for clear, concise guidelines for the care of these patients.

    Primary goals of pre-transplantation clinical care are (1) management of pre-existing medical needs such as feeding difficulties, airway obstruction, congenital cardiac defects and developmental disabilities; (2) management of symptoms related to oligoclonal recipient T cell expansion (autologous GVHD/atypical complete DiGeorge anomaly) and (3) prevention of infections. Most deaths in the pre and early post-transplantation period are secondary to pre-existing infections. Necessary surgical and medical procedures (ie cardiac surgery, hearing aids) should not be delayed.

    For the first 6 to 9 months after RVT802, patients have profoundly low naïve T cell numbers and may require immunosuppression to prevent rejection of RVT-802 by oligoclonal recipient T cells. Immunosuppression needs to be closely monitored and titrated for desired effect while minimizing side effects such as renal toxicity, electrolyte abnormalities and hypertension. T cell counts should be performed every 3 months and are used to guide weaning of immunosuppression. Most patients with successful transplants develop greater than 100/mm3 naïve T cells by 12 months post RVT-802. Infection prevention, clinical stability and optimal nutrition are critical for lasting engraftment. Clinical guidelines have been developed to address immunosuppression, management of autologous GVHD symptoms (gut, skin and liver), preservation of renal function, and developmental considerations.

    After the development of naïve T cells, patients should continue to be monitored regularly by an immunologist. Patients may develop autoimmune complications such as thyroid disease and transient cytopenias. While risk of complications related to viral infections is greatly decreased after development of naïve T cells, patients with comorbidities (central venous access device dependence, tracheostomy, chronic lung disease) continue to require complex care from multidisciplinary teams. Medical conditions associated with athymia but not alleviated by thymus transplantation, such as hypoparathyroidism or cardiac defects, may require lifelong medical care. Lastly, patients must be evaluated for readiness for killed and live vaccines.

    Transplant outcomes are influenced by the clinical condition at the time of RVT-802 implantation and optimization of immunosuppression, nutrition and clinical stability in the first 9 months following RVT-802. Clinical care that maintains a well-nourished, clinically stable, infection free patient yields the best chance for successful T cell development. Guidance documents supporting these goals ensure patients are best prepared to receive RVT-802 and develop long lasting thymic function.

    (77) Submission ID#600641

    Severe Inflammatory Episodes Associated with COG4-Congenital Disorder of Glycosylation (CDG-IIj) Presenting as Hemophagocytic Lymphohistiocystis (HLH)

    Jeffrey Lo, MD1, William J. Brucker, MD, PhD2, John Prensner, MD, PhD3, Anita Pai, MD4, Mary Beth Son, MD5, Christine K. Lee, MD6, Matthew M. Heeney, MD7, Olaf Bodamer, MD, PhD8, Christina S.K. Yee, MD, PhD9

    1Pediatric Fellow, Division of Immunology, Boston Childrens Hospital/Harvard Medical School

    2Pediatric Fellow, Division of Genetics and Genomics, Boston Childrens Hospital/Harvard Medical School

    3Pediatric Fellow, Division of Hematology/Oncology, Dana-Farber/Boston Children's Cancer and Blood Disorders Center/Harvard Medical School

    4Pediatric Fellow, Division of Gastroenterology, Hepatology, and Nutrition, Boston Childrens Hospital/Harvard Medical School

    5Assistant Professor of Pediatrics, Program Director Pediatric Rheumatology, Division of Immunology Boston Childrens Hospital/Harvard Medical School

    6Instructor of Pediatrics, Division of Gastroenterology, Hepatology, and Nutrition, Boston Childrens Hospital/Harvard Medical School

    7Assistant Professor of Pediatrics, Associate Chief Hematology, Dana-Farber/Boston Children's Cancer and Blood Disorders Center/Harvard Medical School

    8Associate Professor of Pediatrics, Associate Chief of Genetics and Genomics, Division of Genetics and Genomics, Boston Childrens Hospital/Harvard Medical School

    9Instructor of Pediatrics, Division of Immunology, Boston Childrens Hospital/Harvard Medical School

    Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening disease of immune dysregulation characterized by unchecked inflammatory responses leading to end-organ dysfunction. Primary HLH results from inherited mutations that impair capacity for immune regulation whereas secondary HLH arises from inappropriate response to an immune stimulus such as infection, malignancy or autoimmunity. We report a 9-month-old male who presented with symptoms of HLH as an initial manifestation of congenital disorder of glycosylation (CDG) due to mutations in the gene Component of Oligomeric Golgi Complex 4 (COG4) resulting in COG4-CDG (CDG-IIj).

    A 9-month-old male with history of mild motor delay presented with 3 days of fever, vomiting, and diarrhea. Initial evaluation identified highly elevated ferritin and triglycerides, transaminitis, coagulopathy, and hyperammonemia. He subsequently developed generalized seizures. Liver and bone marrow biopsies demonstrated erythrophagocytosis consistent with HLH. Immunologic evaluation was notable for mild hypogammaglobulinemia, neutropenia, thrombocytopenia, and anemia. Serum CD25 levels and NK functional studies were later found to be normal.

    The patient was initially treated with ammonia-scavenger therapy and fresh frozen plasma (FFP) for coagulopathy with subsequent intravenous immunoglobulin and dexamethasone several days later. Within 24 hours after starting FFP, the patients ferritin level declined sharply. Hyperammonemia and transaminitis also resolved, and his fever curve improved. Additional immunosuppression was considered, but not initiated due to the patients ongoing clinical improvement.

    Over the next 3 months, the patient experienced two further acute episodes of fever, liver dysfunction, coagulopathy, and sepsis physiology. The second episode was successfully treated with FFP, though no clear infectious trigger was identified. The third episode occurred 4 days after routine vaccinations. The patient had prolonged hypotension requiring ionotropic support that resolved after receiving daily FFP, and hypoxia with pleural effusions that resolved after a single treatment with protein C concentrate.

    As the patient had met 5/8 clinical diagnostic criteria for HLH, but also had a history of hyperammonemia, he underwent concurrent biochemical and genetic evaluation for both primary HLH and inborn errors of metabolism. Whole exome sequencing identified compound heterozygous mutations in COG4, part of an oligomeric protein complex involved in Golgi apparatus structure and function. COG4 mutations have previously been reported in two patients with autosomal recessive COG4-CDG (CDG-IIj), who were described to have similar clinical symptoms of hypotonia, seizures, coagulopathy, and liver dysfunction, as well as recurrent infections. Subsequent immune phenotyping while the patient was healthy was notable for slightly low numbers of NK cells, but normal CD107a mobilization and perforin/granzyme B expression in vitro.

    Our patient represents a novel presentation of CDG due to COG4 defect with associated immune dysfunction manifesting as recurrent episodes of inflammatory crisis with features of HLH. CDG and inborn errors of metabolism should be considered during diagnostic evaluation for patients with HLH symptoms, as CDG patients may develop acute episodes of severe inflammation, in the absence of cellular regulatory defects, for which FFP and protein C concentrate may have therapeutic value.

    (78) Submission ID#600705

    Natural History of Anti-Interferon-gamma Autoantibody-associated Immunodeficiency Syndrome in Thailand

    Gloria H. Hong, BA1, Ploenchan Chetchotisakd, MD2, Siriluck Anunnatsiri, MD2, Piroon Mootsikapun, MD2, Lindsey B. Rosen, BS3, Christa S. Zerbe, MD, MS4, Steven M. Holland, MD5

    1NIH Medical Research Scholar, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health

    2Faculty of Medicine, Division of Infectious Diseases and Tropical Medicine, Department of Medicine, Khon Kaen University, Thailand

    3NIH-Oxford Scholar, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health

    4Senior Research Physician, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH

    5Director, Division of Intramural Research, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health

    Introduction/Background: Autoantibodies to interferon-gamma (IFN-g) are associated with disseminated nontuberculous mycobacterial (NTM) and other opportunistic infections in previously healthy adults, predominantly in or from Southeast Asia. Although the clinical manifestations of this acquired immunodeficiency syndrome have been reported, its natural history is not well understood.

    Objectives: To characterize demographic data, recurrence of infections, clinical outcomes, and autoimmunity-related complications in patients with anti-IFN-g autoantibodies.

    Methods: Eighty-one HIV uninfected voluntary participants (40 with disseminated NTM infection and 41 with another opportunistic infection with or without NTM infection) at Srinagarind Hospital in eastern Thailand were enrolled in an Institutional Review Board-approved protocol (09-I-N060) beginning in 2010 and followed annually until November 2018. Demographic information and clinical histories were recorded on standard forms at each visit and plasma samples were obtained. Serial plasma samples are being analyzed for anti-IFN-g antibody levels.

    Results: Seventy-four out of 81 patients (91%) had anti-IFN-g autoantibodies. The median [interquartile range, IQR] age of patients with anti-IFN-g autoantibodies was 50 [46,56] years. Forty-seven patients (64%) were female. At the time of diagnosis, 36 patients (49%) with anti-IFN-g autoantibodies had disseminated NTM infection, 35 patients (47%) had another opportunistic infection with NTM infection, and 3 patients (4%) had another opportunistic infection without NTM infection. Mycobacterium abscessus was the most commonly isolated organism and lymph nodes (69 patients, 93%) were the most commonly involved site.

    During the follow-up period, 25 patients (34%) with anti-IFN-g autoantibodies had at least one recurrence of culture-proven infection. After a median [IQR] follow-up time of 85 [42,96] months, 41 patients (55%) with anti-IFN-g autoantibodies had inactive disease after prolonged antibiotic treatment, 6 patients (8%) had active/progressive disease, and 18 patients (24%) had died. Of the 14 deaths with identifiable causes, 10 (71%) were related to infections. The rate of death per person-year was 0.044. The most common autoimmunity-related complication was Sweets syndrome, seen in 29 patients (39%) with anti-IFN-g autoantibodies. Sixteen of those patients (55%) had recurring Sweets syndrome. Additionally, 14 patients (19%) developed lymphatic obstruction, which continued to recur in 12 patients (86%).

    Seven patients (9%) in this study did not have anti-IFN-g autoantibodies. The median [IQR] age of autoantibody-negative patients was 38 [27,54] years and 3 patients (43%) were female. None of the autoantibody-negative patients developed new infections during follow-up. At the end of the follow-up period, none of the patients had active/progressive disease and 2 patients (29%) had died.

    Conclusions: Ninety-one percent of HIV uninfected Thai patients with disseminated NTM infection with or without other opportunistic infections had detectable anti-IFN-g autoantibodies. About one third of patients with autoantibodies to IFN-g had recurrent infections during follow-up. After approximately 7 years of follow-up, 55% of patients with anti-IFN-g autoantibodies had inactive disease following multi-drug antibiotic therapy while 8% had active/progressive disease and 24% had died. Patients with anti-IFN-g autoantibodies are at risk for recurrent infections and autoimmunity-related complications. Therefore, long-term follow-up is recommended. Life-long secondary antibiotic prophylaxis may be required to prevent recurrence of infection in the setting of persistent anti-IFN-g autoantibodies.

    (79) Submission ID#600727

    Artificial Thymic Organoids Represent a Reliable and Quick Tool to Study T Cell Differentiation in Human Bone Marrow Samples from Patients with Severe T Cell Immunodeficiency

    Marita Bosticardo, PhD1, Francesca Pala, PhD2, Enrica Calzoni, MD3, Cameron Gardner, BSc4, Kerry Dobbs, BSc5, Suk See De Ravin, MD, PhD6, Nicholas Hartog, MD7, M. Louise. Markert, MD, PhD8, Katja G. Weinacht, MD, PhD9, Harry L. Malech, MD10, Christopher Seet, MD, PhD11, Amelie Montel-Hagen, PhD12, Gay M. Crooks, MBBS13, Luigi D. Notarangelo, MD, PhD14

    1Staff Scientist, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    2Post doctoral Fellow, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    3Graduate Student, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    4Graduate Student, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA; Department of Medicine, University of Oxford, Oxford, UK

    5Biologist, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    6Clinician, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    7Assistant Professor, Michigan State University, College of Human Medicine, Grand Rapids, MI

    8Professor of Pediatrics and Immunology, Department of Pediatrics, Division of Allergy, Immunology, and Pulmonology, Duke University Medical Center, Durham, NC

    9Assistant Professor, Division of Stem Cell Transplantation and Regenerative Medicine, Department of Pediatrics, Stanford School of Medicine, Stanford, CA

    10Chief, Genetic Immunotherapy Section, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    11Fellow, Department of Pathology and Laboratory Medicine, DGSOM, UCLA, Los Angeles, CA

    12Associate Project Scientist, Department of Pathology and Laboratory Medicine, DGSOM, UCLA, Los Angeles, CA

    13Professor, Pathology & Laboratory Medicine; Paediatric oncologist, Division of Stem Cell Transplantation and Regenerative Medicine, Department of Pediatrics, Stanford School of Medicine, Stanford, CA

    14Chief, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    The study of early T cell development in patients with severe T cell immunodeficiencies is challenging because of the rarity of these diseases, the difficulty to obtain hematopoietic stem cells (HSCs), and limitations in the assays to assess in vitro differentiation of HSCs to mature T cells. We recently developed a serum-free system that allows faithful analysis of sequential steps of T cell differentiation. In this system, artificial thymic organoids (ATOs) are generated, based on the 3D aggregation and culture of a delta-like canonical Notch ligand 4 (DLL4)-expressing stromal cell line (Ms5-Dll4) with CD34+ cells isolated from bone marrow (BM) samples of normal donors (ND). In this project, we set out to evaluate the possibility of using the ATO system to study T cell differentiation in patients carrying T cell defects, in order to define the exact steps of T cell development affected by different genetic defects. Using the ATO system, we studied in vitro T cell differentiation from CD34+ cells obtained from patients carrying defects that are intrinsic to hematopoietic cells (RAG1, RAG2, AK2, IL2RG) or that affect thymus development (DiGeorge syndrome, DGS). The AK2-deficient patient showed a markedly decreased viability in CD34+ cells and a very early defect in T cell development, already at the pro-T cell stage. This defect was very similar to that observed in a patient carrying a null IL2RG mutation who was reported to show autologous reconstitution after unconditioned haploidentical HSC transplantation. In contrast, CD34+ cells from a patient carrying a missense IL2RG mutation and with a leaky SCID phenotype were capable of differentiating into mature T cells in vitro, although with 100-fold decreased efficiency as compared to normal donors (ND). Interestingly, in the patient carrying the null IL2RG mutation, we noticed very few cells that could reach full maturation, with an absolute number of CD3+TCRab+ cells around 1000-times less than in ND. At variance with pro-T cells (that failed to express the gc protein), these mature T cells did express normal levels of gc, suggesting that they may have derived from residual CD34+ cells from the BM donor. In addition, CD34+ cells from the patients carrying RAG1 and RAG2 hypomorphic mutations were able to differentiate to CD4+CD8+ double positive cells, but not to CD3+TCRab+ cells. Finally, the DGS patient showed a completely normal in vitro T cell differentiation, confirming that T cell deficiency reflected thymic abnormalities. In summary, our data show that the ATO system could be extremely useful in determining whether the lack of T cells in patients with unknown gene defects reflect hematopoietic or thymic intrinsic problems, and may therefore provide critical evidence in deciding whether HSC or thymus transplantation is warranted, even without knowing the actual gene defect.

    Supported by the Intramural Research Program, DIR, NIAID, NIH

    Protocol 18-I-N128

    (80) Submission ID#600761

    Ataxia Telangiectasia with Chronic Skin Granulomas Preventable with SCID Newborn Screening?

    Sara Seghezzo, MD1, Dana Feigenbaum, MD2, Sonal D. Shah, MD3, Erin F. Mathes, MD4, Morna J. Dorsey, MD, MMSc5, Jennifer Puck, MD6,

    1Clinical Fellow, Department of Pediatrics, Division of Allergy, Immunology, and Bone Marrow Transplant, University of California, San Francisco

    2Resident, Department of Dermatology, University of California San Francisco, San Francisco, CA

    3Assistant Professor, Department of Dermatology, University of California San Francisco, San Francisco, CA

    4Associate Professor, Department of Dermatology, University of California San Francisco, San Francisco, CA

    5Pediatric Immunologist and Allergist, Department of Pediatrics, Division of Allergy, Immunology, and Bone Marrow Transplant, University of California San Francisco, San Francisco, CA

    6Pediatric Immunologist, Department of Pediatrics, Division of Allergy, Immunology, and Bone Marrow Transplant, University of California San Francisco, San Francisco, CA

    Introduction: Ataxia-Telangiectasia (AT) is an autosomal recessive disorder caused by mutations in the Ataxia Telangiectasia Mutated (ATM) gene, which aids in detection and repair of DNA damage. AT is characterized by progressive cerebellar ataxia, oculomotor apraxia, choreoathetosis, conjunctival telangiectasias, variable degrees of T-cell lymphopenia (TCL) and immune compromise. Patients are at an increased risk for malignancy, particularly leukemia and lymphoma, and are unusually sensitive to ionizing radiation. With the advent of TREC-based newborn screening (NBS) for SCID, AT patients are being recognized with asymptomatic TCL in early infancy.

    Objectives: We present an older child with AT and chronic granulomatous lesions and discuss how this may be avoided in individuals with AT diagnosed following abnormal NBS.

    Case Report: A 12 y/o male was born at term following an uncomplicated twin pregnancy and delivery, prior to institution of SCID NBS. He demonstrated mild gross motor and speech delay as an infant and was diagnosed with AT at age 3. He had received all routine immunizations, including live vaccinations. He developed granulomatous skin lesions at age 1, initially small papules on his cheeks and ears, which subsequently formed large disfiguring plaques on sun-exposed areascheeks, arms and hands (Fig 1). Following an extensive workup, his lesions were found to be secondary to a mutated vaccine-strain Rubella (RA27/3) based on 739bp genotyping, previously described in other immunocompromised individuals [Perelygina/Sullivan et al. JACI 2016]. His lesions have been refractory to multiple treatments including nitazoxanide. He is currently on daily oral and topical steroids, TMP/SMX and IVIG. Retrieval of his NBS for TREC determination revealed that he would have screened positive [Mallot/Puck et al. J Clin Immunol 2013]. When first measured at age 3, CD3 T-cells were low, 443/ul, with CD4 227/ul and CD8 140/ul. B and NK cell numbers were normal.

    Since April 2017, 4 cases of AT were seen at UCSF in infants with non-SCID TCL on NBS. These 3 males and 1 female were all born at term and discharged from well-infant nurseries. AT was diagnosed at 2-7 months of age. Their initial TRECs ranged from 5-12/ul (normal with PerkinElmer Enlite kit >18), and all had low T-cells on initial flow cytometry (242-1612 CD3/ul, ref range>2500) with decreased CD4 (146-1178/ul) and CD8 (87-403/ul) T-cells; however naïve T-cells were present, ruling out typical SCID and raising concern for non-SCID TCL. Three infants also demonstrated low B-cells (<20-77/ul), while NK cells were normal in all. Two are currently receiving IVIG, one of whom is also on TMP/SMX. All have avoided not only rotavirus but also MMR and varicella live vaccinations.

    Conclusions: AT is now often diagnosed in infants with low TRECs on SCID NBS, prior to neurologic manifestations. Benefits of early diagnosis include avoidance of live vaccines, including MMR, which led to the debilitating granulomas in our older patient. Additionally, patients receive prompt immunologic monitoring and treatment, avoidance of unnecessary radiation, specialty referrals and family genetic counseling. While there is no cure for AT, ongoing research may bring neuroprotective treatments in the future.

    figurex

    (81) Submission ID#600763

    Comorbidities, Concomitant Medications, Infusion Parameters, and Tolerability in Advanced Age Patients with Primary Immunodeficiency Diseases Treated with Ig20Gly

    Mark R. Stein, MD1, Daniel Suez, MD2, Iftikhar Hussain, MD3, Sudhir Gupta, MD4, Amy Darter, MD5, Ping Wang, PhD6, Barbara McCoy, PhD7, Leman Yel, MD8

    1Physician, Allergy & immunology, Allergy Section, Good Samaritan Medical Center, West Palm Beach, FL, USA

    2President, Allergy, Asthma & Immunology Clinic, PA

    3President and Principal Investigator, Vital Prospects Clinical Research Institute, P.C., and Allergy, Asthma and Immunology Center, P.C.

    4Professor, University of California at Irvine, Irvine, CA, USA

    5Medical Director, Oklahoma Institute of Allergy & Asthma Clinical Research

    6Project Lead, Biostatistics & Statistical Programming, Shire

    7Clinical Scientist Lead – Immunology, Shire

    8Sr Medical Director, Global Development Leader, IG, Clinical Research Immunology, Shire, Cambridge, MA, USA

    Introduction: Subcutaneous immune globulin 20%, Ig20Gly, was well tolerated in the phase 2/3 North American study in patients with primary immunodeficiency diseases (PIDD). Here we assess comorbidities, use of concomitant medications, infusion parameters, and tolerability in advanced age patients (60 y) treated with Ig20Gly in the North American study.

    Methods: Patients aged 2 years with PIDD received weekly Ig20Gly infusions at volumes 60 mL/site and rates 60 mL/h/site for ~1.3 years in the North American study (NCT01218438). The medical history at baseline, medical conditions that were ongoing (defined as comorbid events), use of concomitant medications, adverse events (AEs), tolerability, and infusion parameters were assessed by age: in advanced age patients (60 y; n=14), adult (16<60 y; n=39), and pediatric/adolescent patients (<16 y; n=21).

    Results: The mean number of medical history events at baseline was higher in advanced age patients (28.7 events/patient; 402 events in 14 patients) versus adult (16.8 events/patient; 657 events in 39 patients), and pediatric/adolescent patients (6.5 events/patient; 137 events in 21 patients). Of these, the medical conditions that were ongoing at baseline (comorbid events) were also higher in the advanced age patients (20.9 events/patient; 292 events in 14 patients) versus adult (12.4 events/patient; 482 events in 39 patients), and pediatric/adolescent patients (3.4 events/patient; 71 events in 21 patients). In the advanced age patients, neurological comorbidities (51 events) were the most common, followed by those related to eyes, ears, nose, and throat (49 events), gastrointestinal (43 events), and musculoskeletal comorbidities (43 events). Concomitant medications were given to treat a preexisting condition in all patients in the advanced age group (225 medications in 14 patients). Despite the higher mean number of comorbid conditions, infusion parameters in the advanced age patients were comparable to those in the adult age group. Median maximum infusion rates and infusion volumes/site were comparable in the advanced age patients (60 mL/h/site; 47.5 mL/site) and adults (60 mL/h/site; 44 mL/site); lower infusion rates and volumes/site were reported in the pediatric/adolescent patients (30 mL/h/site; 26.8 mL/site). Infusions were well tolerated in all patients. Percentages of infusions associated with causally related AEs were low in advanced age patients (all [1.3 %], local [0.4%], systemic [0.9%]), adults (all [3.5%], local [1.1%], systemic [2.7%]) and pediatric/adolescent patients (all [2.8%], local [2.7%], systemic [0.4%]). Larger infusion volumes and faster infusion rates were not associated with increases in causally related local AEs in the advanced age group, consistent with the trends seen in the pediatric/adolescent and adult patients.

    Conclusions: Despite the higher mean number of comorbidities in advanced age patients with PIDD, Ig20Gly was infused at relatively high rates and volumes and was well tolerated.

    (82) Submission ID#600823

    Interim Analysis of Infusion Characteristics and Adverse Events During Facilitated Subcutaneous Immunoglobulin Treatment for Primary Immunodeficiency Diseases: Global Post Authorization Safety Study

    Arye Rubinstein, MD, Ph.D.1, Tracy Bridges, MD2, H. James Wedner, MD3, Donald McNeil, MD4, Richard L. Wasserman, MD, PhD5, Raffi Tachdjian, MD6, Katharina Fielhauer, MA7, Heinz Leibl, PhD8, Leman Yel, MD9

    1Attending in Allergy & Immunology, Montefiore Medical Center

    2Allergy / Immunology, Allergy & Asthma Clinics-Ga

    3Chief, Division of Allergy and Immunology, Washington University Physicians, Washington University School of Medicine in St. Louis

    4President, Founder and Principal Investigator, Optimed Research, LTD

    5Allergist/immunologist, Allergy Partners of North Texas Research, Dallas, TX, USA

    6Assistant Clinical Professor of Medicine and Pediatrics in the Division of Allergy and Clinical Immunology, Ronald Reagan Medical Center, UCLA School of Medicine

    7Clinical Scientist, Shire

    8Sr Medical Director, Global Development Leader, IG, Clinical Research Immunology, Shire, Vienna, Austria

    9Sr Medical Director, Global Development Leader, IG, Clinical Research Immunology, Shire, Cambridge, MA, USA

    Introduction: HyQvia (IGHy; immunoglobulin infusion 10% with recombinant human hyaluronidase [rHuPH20]) is an immunoglobulin (IG) replacement therapy approved for patients with primary immunodeficiency diseases (PIDD) that allows larger infusion volumes, up to 600 mL/site, and has improved IG bioavailability compared with conventional subcutaneous IG products. A post-authorization safety study is being conducted in the United States to acquire long-term safety data on IGHy and to assess prescribed administration regimens in routine clinical practice. Infusion characteristics and treatment-related adverse events from an interim analysis are reported here.

    Methods: Patients aged 16 years with PIDD receiving IGHy were included in this ongoing, prospective, non-interventional, open-label, uncontrolled, multicenter study. As a part of routine clinical practice, patients are treated with IGHy according to standard medical care and their treatment regimen is at the discretion of the treating physician. Adverse events (AEs) are collected from enrollment to study completion/discontinuation using a subject diary and assessed at every study visit (every 3 months or standard practice). AEs are assessed based on seriousness, severity, and causal relatedness to IGHy. The presence of anti-rHuPH20 antibody is evaluated on a voluntary basis. Treatment preferences for various attributes of IG therapy were assessed annually using a treatment preference questionnaire.

    Results: A total of 175 patients were enrolled at 26 US study sites (data cut-off date: August 21, 2017). Infusions were self-administered at home (56%) or at the clinical site (44%) most commonly using 4-week infusion intervals (56.6%). The mean maximum IG infusion rate was 302.8 mL/h and the mean IG dose was 418 mg/kg bodyweight/4weeks. The mean number of infusion sites used for administration was 1.9 and mean infusion duration was 2.8 hours. Most infusions (97.3%) were administered without a rate reduction, interruption, or discontinuation due to AEs. There were no serious AEs (SAEs) related to IGHy. Sixteen patients experienced a causally related non-serious local AE (9.1%; 0.43 events/patient-year, 0.07 events per infusion) and 25 patients experienced a causally related non-serious systemic AE (14.3%, 0.88 events/patient year, 0.14 events per infusion). Seven of 113 patients who were tested for anti-rHuPH20 antibody had 1 positive binding antibody test to rHuPH20 (titer 1:160; maximum titer 1:10240 at enrollment, 1:5120 during the study); no neutralizing rHuPH20 antibodies were detected. Of the patients who responded to the treatment preference questionnaire at the end of year 1, the majority (38/52 [73.1%]) preferred to receive their IG therapy at home; 21.2% (11/52) preferred the doctors office; 3 patients preferred treatment at the hospital, had no preference, or indicated other. Almost all patients (51/52 [98.1%]) indicated a preference to continue treatment with IGHy.

    Conclusion: This interim analysis of 175 patients with PIDD treated with IGHy in routine clinical practice supports previous observations that IGHy is a well-tolerated and preferred therapy with no reports of treatment-related SAEs or neutralizing anti-rHuPH20 antibodies.

    (83) Submission ID#600846

    Lymphocyte Radiosensitivity in Cartilage Hair Hypoplasia

    Jennifer R. Yonkof, MD1, Sharat Chandra, MD, MRCPCH2, Matthew J. Smith, MS3, Roshini S. Abraham, PhD4

    1Fellow, Department of Pediatrics, Division of Allergy and Immunology, Nationwide Children's Hospital

    2Assistant Professor, UC Department of Pediatrics, Division of Bone Marrow Transplantation and Immune Deficiency, Cincinnati Childrens

    3Research Technologist, Division of Hematology, Department of Medicine, Mayo Clinic

    4Department of Pathology and Laboratory Medicine, Nationwide Childrens Hospital, Columbus, OH.

    Background: Cartilage hair hypoplasia (CHH) is an autosomal recessive chondrodysplasia associated with variable immunodeficiency. Pathogenic defects in RMRP, encoding the untranslated RNA subunit of ribonucleoprotein endoribonuclease complex (RMRP), result in reduced mRNA and rRNA cleavage. RMRP c.70A>G is the most common variant, increased in Finnish and Amish populations. While cellular immunodeficiency is associated with increased morbidity and mortality, there is no established correlation between clinical and immunological phenotype. Lymphocyte radiosensitivity has not been described.

    Case: A full-term Amish female infant had low TREC copies on newborn SCID screen. Flow cytometry at 3 months-old demonstrated severe T and B cell lymphopenia (CD3+T-cells 413 cells/mcL, range: 2,300-6,500 cells/mcl; CD19+B-cells 214 cells/mcL, range: 600-3,000 cells/mcL) with normal NK quantitation (CD16/56+ 340 cells/mcl, range: 100-1,300 cells/mcL) and CD4+ memory T-cell expansion (33.2%) relative to the naïve subset (67.0%). T-cell functional mitogen responses were normal. She was diagnosed with CHH with homozygous RMRP c.70A>G mutation. Lymphocyte subset (T, B and NK cells) radiosensitivity was evaluated by flow cytometric analysis of phosphorylated (p) ATM, SMC1 and gamma-H2AX after low-dose (2Gy) irradiation. An increase in gamma-H2AX level was observed in a subset of non-irradiated T cells (17.66% v. 1.36% gamma-H2AX+) and NK cells (23.07% v. 1.04% gamma-H2AX+) in the patient, suggestive of a constitutive defect in DNA repair. The relative distribution of T, B and NK cells expressing pATM, pSMC1 and gamma-H2AX at 1 hour post-irradiation (IR) was not significantly different from the experimental healthy control (EHC) or pediatric reference range (pRR). However, the kinetics of dephosphorylation at 24 hours post-IR was altered with residual gamma-H2AX expression in a subset of the patients T cells (delta 3.84%, mode ratio mean fluorescence intensity (MFI)=2.58; EHC: delta 0.10%, mode ratio MFI=1.39; pRR: delta 2.16%, mode ratio MFI=2.42). A similar finding was observed in a subset of patient B-cells for gamma-H2AX (delta 11.35%, mode ratio MFI=1.48; EHC: delta 0.82%, mode ratio MFI=0.86; pRR: delta 1.95%, mode ratio MFI=1.19). The frequency of the patient's lymphocytes with residual gamma-H2AX persistence at 24h post-IR was prominent, with 8.29% T-cells demonstrating persistence of gamma-H2AX (compared to 0.82% in the EHC, and 2.60% in the pRR), and 18.02% B-cells gamma-H2AX+ (compared to 1.80% in the EHC, and 2.96% in the pRR). There has been lack of follow-up, but verbal report suggests no significant immunological or infectious concerns at 1 year of age.

    Discussion: Lymphocyte radiosensitivity is a novel finding in CHH with T and B cell lymphopenia. The ability of RMRP to associate with telomerase reverse transcriptase (TERT) and function as an RNA-dependent RNA polymerase, yielding distinct silencing RNA sequences, may underlie radiosensitivity in RMRP mutants. Systematic characterization of lymphocyte radiosensitivity and immunological phenotype could provide useful information on whether this could serve as a biomarker for the magnitude or complexity of immunodeficiency. Assessment of radiosensitivity has implications in conditioning regimen selection for patients requiring allogeneic hematopoietic cell transplantation. We recommend lymphocyte radiosensitivity assessment in CHH infants identified by NBS SCID and CHH patients with significant immunodeficiency and/or malignancy.

    (84) Submission ID#600887

    Novel Primary Immunodeficiency with Lymphoproliferative Disease Due to Biallelic Defects in NCKAP1L

    William A. Comrie, PhD1, M. Cecilia Poli, MD, PhD2, Douglas B. Kuhns, PhD3, Jason W. Caldwell, DO4, Morgan Similuk, ScM5, Alexandre F. Carisey, PhD6, Lisa R. Forbes, MD2, Emily M. Mace, PhD7, Tram N. Cao, MS8, Zeynep H. Coban-Akdemir, PhD9, Shalini N. Jhangiani, PhD10, Donna M. Muzny, MSc11, Richard A. Gibbs, PhD12, James R. Lupski, MD, PhD12, V. Koneti Rao, MD, FRCPA13, Jordan S. Orange, MD, PhD14, Ivan K. Chinn, MD2, Michael J. Lenardo, MD15

    1Postdoctoral Research Fellow, Molecular Development of the Immune System Section, Laboratory of Immune System Biology, NIAID, National Institutes of Health, Bethesda, MD, USA

    2Assistant Professor, Department of Pediatrics, Baylor College of Medicine, Houston, TX, USA

    3Principal Scientist, Neutrophil Monitoring Lab, National Cancer Institute-Frederick, Frederick National Laboratory for Cancer Research, Frederick, MD, USA

    4Assistant Professor, Section of Pulmonary, Critical Care, Allergic and Immunological Diseases, Wake Forest University School of Medicine, Winston-Salem, NC, USA

    5Genetic Counselor, Molecular Development of the Immune System Section, Laboratory of Immune System Biology, NIAID, National Institutes of Health, Bethesda, MD, USA

    6Postdoctoral Research Fellow, Department of Pediatrics, Baylor College of Medicine, Houston, TX, USA

    7Assistant Professor, Department of Pediatrics, Columbia University Irving Medical Center, New York, NY

    8Research Coordinator, Department of Pediatrics, Baylor College of Medicine, Houston, TX, USA

    9Postdoctoral Research Fellow, Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX

    10Project Manager, Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX

    11Assistant Professor, Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas, USA

    12Professor, Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas, USA

    13Staff Physician, Molecular Development of the Immune System Section, Laboratory of Immune System Biology, NIAID, National Institutes of Health, Bethesda, MD, USA

    14Professor and Chair, Department of Pediatrics, Columbia University Irving Medical Center, New York, NY

    15Senior Investigator, Molecular Development of the Immune System Section, Laboratory of Immune System Biology, NIAID, National Institutes of Health, Bethesda, MD, USA

    BACKGROUND: Three children from 2 non-consanguineous families and different ethnic backgrounds developed lymphoproliferative disease by 2 years of age. They also had recurrent infections, including pneumonia and bronchiectasis, otitis media, and skin pustules. Immune phenotyping revealed low CD4+ T cell percentages, an accumulation of memory-like CD8+ T cells, impaired T cell proliferation, and low total NK cell numbers.

    METHODS: The affected individuals, unaffected parents, and other unaffected family members underwent exome sequencing.

    RESULTS: All 3 affected cases had rare and bioinformatically damaging biallelic variants, with appropriate familial segregation, in NCKAP1L, which encodes Hem1. Hem1 is an essential component of the WAVE2 Regulatory Complex (WRC). Immunoblotting confirmed destabilization of the WRC in all patients. Immunofluorescence microscopy demonstrated defective F-actin and WAVE2 localization to immune synapses in NK cells. Significant abnormalities were identified in patient lymphocyte and neutrophil migration and morphology, consistent with altered WRC-mediated cytoskeletal dynamics. All patients exhibited impaired inside-out integrin activation. Knockdown of Hem1 produced deficient proliferative responses and mTORC2-mediated AKT activation in control T cells.

    CONCLUSIONS: The immunologic and clinical phenotype in the affected individuals recapitulates the phenotype observed in Hem1-deficient mice. Biallelic defects in NCKAP1L therefore result in a novel human primary immunodeficiency disease characterized by lymphoproliferation and susceptibility to infections.

    (85) Submission ID#600899

    Prevalence of Hypogammaglobulinemia in Newly Diagnosed Lymphoma

    Namrata Singh, MD, MSCI, FACP1, Sarah Mott, MS2, Ashley McCarthy, MPH3, Aaron Knaack, BA4, James Cerhan, MD, PhD5, Zuhair Ballas, MD6, Brian Link, MD6

    1Clinical Assistant Professor, University of Iowa Hospitals and Clinics

    2 Biostatistician, College of Public Health

    3Manager, Holden Comprehensive Cancer Center

    4Division Coordinator, University of IOwa Hospitals and CLinics

    5Professor, Mayo Clinic

    6Professor, University of Iowa Hospitals and Clinics

    Background: Concurrent existence/significance of immunodeficiency with new onset lymphoproliferative disease remains understudied. Just two studies to date have evaluated the prevalence of hypogammaglobulinemia in chronic lymphocytic leukemia (CLL) and neither studied prevalence and impact of IgE deficiency on outcomes in CLL [1, 2]. Therefore, the objective of this study was to examine the prevalence of hypogammaglobulinemia, examining all isotypes, in newly diagnosed CLL patients and to test the hypothesis that patients with hypogammaglobulinemia have a distinct clinical profile and outcome.

    Methods: Using the banked sera of 150 newly diagnosed, treatment-naïve, CLL adult patients from the Lymphoma Molecular Epidemiology Resource (L-MER), Ig (IgG, IgA, IgM and IgE) levels were measured. The L-MER was initiated as an observational cohort study of prospectively enrolled newly diagnosed lymphoma patients evaluated at the Mayo Clinic (Rochester, MN) and the University of Iowa (Iowa City, IA) [3]. IgG/A/M levels were measured using immunoturbidimetric assay whereas the IgE level was determined using electrochemiluminescence immunoassay. The associations between Ig deficiencies and clinical factors were evaluated with Wilcoxon rank sum and chi-squared (Fishers exact, where appropriate) tests. Cox regression models were used to assess the effects of clinical variables on overall survival (OS). Time was calculated from biopsy to death due to any cause; patients still alive were censored at last contact. All tests were two-sided and assessed for significance at the 5% level using SAS v9.4 (SAS Institute, Cary, NC).

    Results: The mean age (SD) of the selected CLL cohort was 63.8 (11.0) years with a male predominance (69.3%). 96.2% of the patients were white. With a median follow-up of five years, there were 50 deaths. Hypogammaglobulinemia in newly diagnosed, treatment-naïve CLL was common in our cohort with 88 (58.7%) patients having a measurable isotype deficiency. The most common Ig deficiency was IgM (44.0%, 95% CI 35.9-52.3%), followed by IgG (34.7%, 95% CI 27.1-42.9%), IgE (16.7%, 95% CI 11.1-23.6%) and IgA (12.0%, 95% CI 7.3-18.3%). Multiple deficiencies in the same patient were common (Figure 1). IgA and IgE deficiency were associated with higher Rai stages (grading system for CLL) at presentation (p<0.01 and 0.04 respectively) as well as with higher white blood cell counts at presentation (p=0.02 and 0.01 respectively). A higher proportion of IgA deficient patients needed second treatment during follow-up (61% compared to 36%, p=0.04). When comparing predictors of overall survival, higher Rai stage [3-4 vs 0, Hazard ratio (HR) 2.43, 95% CI 1.08-5.46, p=0.03] and age (HR 1.08, 95% CI 1.05-1.12, p<0.01) correlated with worse overall survival. Individual immunoglobulin deficiencies did not correlate with overall survival.

    Conclusions: A significant proportion of treatment-naïve patients with CLL have underlying Ig deficiencies- both in isolation and a combination of different isotypes. A deficiency of IgA or IgE was associated with severe disease at presentation. The underlying relationship between these two immunologic disorders deserves further study.

    Figure 1. Distribution of immunoglobulin deficiencies among patients with chronic lymphocytic leukemia (CLL).

    figurey

    (86) Submission ID#600905

    Clinical Features and Management of Patients with Rheumatoid Arthritis and a Coexisting Immunodeficiency Disorder

    Ruth Fernandez, MD1, Scott Vogelgesang, MD2, Bharat Kumar, MD, MME3, Zuhair Ballas, MD2, Namrata Singh, MD, MSCI, FACP4

    1Fellow, NYU Langone Health System

    2Professor, University of IOwa Hospitals and Clinics

    3Clinical Assistant Professor, UIHC

    4Clinical Assistant Professor, University of Iowa Hospitals and Clinics

    Background: Patients with primary immunodeficiency (PID) have an increased risk of developing autoimmune diseases, including rheumatoid arthritis (RA). Management of these patients is challenging as immunomodulators can further increase their risk for infections. Additionally, patients with RA that undergo therapy with drug modifying anti-rheumatic drugs (DMARDs) may develop a secondary immunodeficiency. There are few studies reviewing the characteristics of patients with a PID who later develop RA, and no studies have been reported comparing these patients to those who develop an immunodeficiency after starting DMARD therapy for RA.

    Methods: 65 patients were identified as having inflammatory arthritis and a concomitant immunodeficiency (ID) at our institution between 1/1/2000-10/03/2017 using ICD-9 and 10 codes. Manual chart review was performed to confirm and identify the timing of diagnosis of these disorders. Patients were excluded if either there was no definitive diagnosis of ID or RA (clinically diagnosed by a practicing allergist/immunologist and meeting ACR 2010 criteria for RA with a score of 6 or higher, respectively), or rituximab was administered prior to diagnosis of ID . Clinical symptoms, treatment, and laboratory data were extracted. Fishers exact test was used to compare the categorical variables between the groups; t-test was used to compare the continuous variables.

    Results: 10 patients met the inclusion criteria. 5 patients were diagnosed with an ID and developed RA later in life (group 1), and 5 patients were diagnosed with RA and subsequently developed a clinically significant ID (group 2). The mean ages of diagnosis of ID and RA in group 1 patients were 32.0 years (SD ± 26.9) and 42.6 years (SD ± 19.0), respectively. In group 2, the mean age of diagnosis of RA was 37.8 (SD ± 14.2), compared to 54.8 years (SD ± 12.7) for the diagnosis of ID. Most patients in both groups were female (60% in group 1 and 80% in group 2). All patients in both groups had a humoral ID, including common variable immunodeficiency (CVID) (40% of group 1 patients), specific antibody deficiency (SAD) (20% of group 1 and 60% of group 2 patients), and hypogammaglobulinemia (20% of group 1 and 40% of group 2 patients). All patients in group 2 were seropositive for rheumatoid factor (RF) or anti-cyclic citrullinated peptide (anti-CCP), whereas only 20% of patients in group 1 were positive for RF or anti-CCP (Table 1). Most patients in both groups were treated with immunoglobulin replacement therapy. Treatment of RA in both groups was similar, but combination DMARD therapy was not used in group 1 patients in contrast to group 2 patients.

    Conclusions: Our study indicates that even though clinical characteristics and management are similar in patients with coexisting ID and RA, RF and anti-CCP are usually negative in patients who develop RA after ID, possibly due to impaired antibody production in immunodeficient patients.

    Table 1. Comparison of the clinical and laboratory features of patients with an immunodeficiency disorder diagnosed prior to development of rheumatoid arthritis (Group 1) to those diagnosed with an immunodeficiency disoder after diagnosis of rheumatoid arthritis (Group 2)

      Group 1
    n (%)
    Group 2
    n (%)
    P-value
    Sample Size
    Demographics
    5 (100%) 5 (100%)  
    Race/Ethnicity    
     White, not Hispanic 5 (100.0) 5 (100.0) 1.0000
    Age at the time of diagnosis of RA (years) 42.6 ± 19.0 37.8 ± 14.2 0.6964
    Age at the time of diagnosis of ID (years) 32.0 ± 26.9 54.8 ± 12.7 0.1638
    Gender    1.0000
     Male 2 (40.0) 1 (20.0)  
     Female 3 (60.0) 4 (80.0)  
    Deceased 2 (40.0) 1 (20.0) 1.0000
    Positive RF 1 (20.0) 4 (80.0) 0.2063
    Positive anti-CCP 1 (33.3) 2 (66.7)* 1.0000
    Positive RF or anti-CCP 1 (20.0) 4 (80.0) 0.0476

    RA: Rheumatoid arthritis; ID: Immunodeficiency disorder; RF: Rheumatoid factor; anti-CCP: Cyclic citrullinated peptide antibody

    (87) Submission ID#600937

    A Case of Complement Factor D Deficiency with Streptococcus Pneumoniae Pneumonia with Associated Lung Abscess and Empyema

    Ashleah Courtney, MD, MSPH1, Matthew Bell, MD2, Diana K. Bayer, DO3, Sheva Chervinskiy, DO4

    1Pediatric Resident, Arkansas Children's Hospital, University of Arkansas for Medical Sciences

    2Assistant Professor of Allergy and Immunology, Arkansas Children's Hospital, University of Arkansas for Medical Sciences

    3Clinical Assistant Professor of Allergy/Immunology, Division of Allergy/Immunology and Pulmonary, Department of Pediatrics, University of Iowa Stead Family Childrens Hospital

    4Assistant Professor of Allergy and Immunology, Arkansas Children's Hospital, University of Arkansas Medical Sciences

    Introduction/Background: Complement deficiencies are relatively rare, comprising less than 1% of primary immunodeficiencies. They are associated with increased risk for infections with encapsulated organisms and autoimmunity. Of all complement deficiencies, the rarest are defects in the alternative complement pathway. Properdin deficiency is the most commonly described alternative pathway deficiency, with Factor B and Factor D deficiency more rarely described. Fewer than 5 patients with factor D deficiency have been reported with all reported cases being children of consanguineous parents who succumbed to meningococcal sepsis.

    Objectives: To describe a case of Factor D deficiency associated with recurrent respiratory infections with Streptococcus pneumoniae pneumonia with associated lung abscess and empyema.

    Methods: Retrospective chart review was conducted. Laboratory investigations included lymphocyte immunophenotyping by flow cytometry, lymphocyte proliferation to mitogen, quantitative serum immunoglobulins, vaccine titers, complement assays and functional evaluation, and genetic evaluation by next generation sequencing.

    Results: A 2 year old Marshallese male was transferred from an outside hospital to our facility for further evaluation of worsening pneumonia and was found to have right-sided pleural effusion and pulmonary abscess in the right lower lobe. The abscess was drained and was found to be positive for Streptococcus pneumoniae via polymerase chain reaction. He improved after chest tube placement and treatment with intravenous antibiotics. His medical history was significant for recurrent acute otitis media and prior hospitalization out-of-state for pneumonia with empyema secondary to Streptococcus pneumoniae, which required chest tube placement and admission to the pediatric intensive care unit at 18 months of age. Immunologic work up revealed age-appropriate lymphocyte subpopulations, lymphocyte proliferative responses to mitogens, quantitative immunoglobulin levels, pneumococcal/tetanus/diphtheria titers, and CH50 complement assay. AH50 complement assay was decreased to 44 units/mL. Complement testing was repeated - with normal CH50 and AH50 of 0 units/mL. Further evaluation revealed normal levels of Factors B, H, I and properdin. Factor D level was 0.12 mcg/mL, and Factor D function was decreased to 2 units/mL, indicating a diagnosis of Factor D deficiency. Sequencing of the CFD gene revealed a previously undescribed homozygous deletion (c.721_723del and p.Lys241del). The parents were not agreeable to personally undergoing genetic evaluation to determine if this was a de novo mutation. The patient was managed with pneumococcal and meningococcal immunizations, prophylactic amoxicillin and intravenous gamma globulin (IVIG) without any further infections. Unfortunately, after two IVIG infusions, he was lost to follow up.

    Conclusion: Factor D deficiency is an extremely rare alternative complement pathway deficiency, described in less than 5 patients. All infections described thus far have been secondary to Neisseria meningitidis. This case represents not only a novel mutation in the CFD gene leading to Factor D deficiency, but also the first description of a patient with Factor D deficiency developing invasive infection secondary to Streptococcus pneumoniae.

    (88) Submission ID#600938

    Hexaviral-Specific T-cells for Treatment and Prevention of Viral Infections Post Hematopoietic Stem Cell Transplant

    Michael D. Keller, MD1, Katherine Harris, MD2, Patrick Hanley, PhD3, Blachy J. Davila Saldana, MD4, Allistair Abraham, MD5, Nan Zhang, PhD6, Gelina Sani, BS7, Haili Lang, MS8, Richard Childs, MD9, Richard Jones, MD10, Catherine Bollard, MD11,

    1Assistant Professor, Center for Cancer and Immunology Research, Children's National Health System, Division of Allergy & Immunology, Children's National Health System, Washington, DC

    2Clinical Fellow, Children's National Health System

    3Director, Stem cell Therapy Lab, Children's National Health System

    4Blood and Marrow Transplant Specialist, Division of Blood and Marrow Transplantation, Childrens National Medical Center, Department of Pediatrics, The George Washington University, Washington, DC

    5Assistant Professor, Children's National Health System

    6Lead Cell Therapy Technician, Children's National Health System

    7Technician, Children's National Health System

    8Staff Scientist, Children's National Health System

    9Clinical Director, NHLBI

    10Professor, Johns Hopkins

    11Director, Center for Cancer and Immunology Research, Children's National Health System

    Background: Viral infections are a significant cause of morbidity and mortality in patients with primary immunodeficiency disorders and following hematopoietic stem cell transplantation. Adoptive immunotherapy using virus specific T-cells (VSTs) has been shown to prevent and treat viral infections in immunocompromised hosts. Human Parainfluenza Virus-3 (HPIV3) is a common cause of severe respiratory illness in immunocompromised patients and has no approved antiviral therapies and has not previously been used as a target for T cell therapeutics.

    Objective: The primary aim was to determine whether donor derived hexaviral specific T-cells are effective in preventing and treating CMV, EBV, AdV, BK virus, HHV-6, and HPIV3.

    Patients and Methods: This was a first in man study where we studied the antiviral effect in 8 patients who received hexa-valent VSTs after stem cell transplant on a Phase I trial. Assessment of virus-specific immunity was measured by IFN-g ELIspot. Viral loads for the primary targeted viruses were measured at specific time points post VST infusion.

    Results: Three patients were treated for active CMV and had resolution of viremia. Two patients treated for active BK virus had complete resolution of symptoms and viremia, while one had resolution of hemorrhagic cystitis but fluctuating viral loads in the blood and urine. Two patients were treated prophylactically. One patient did not develop any infections, while the other developed EBV viremia requiring rituximab. Two patients received VSTs under expanded access for emergency treatment 1 patient was treated disseminated adenoviremia and the second patient was treated for HPIV3 pneumonia. These critically ill patients demonstrated partial clinical improvements, but VST persistence was likely hindered by concomitant steroid use which resulted in incomplete antiviral responses. ELISpot showed evidence of antiviral T-cell activity in 3 of 4 evaluable patients by 3 months post-infusion, with in vivo VST expansion detectable in 2 patients.

    Conclusions: Preliminary results show that hexaviral specific VSTs are safe and may be effective in preventing and treating multiple viral infections. Further studies are warranted to determine if VSTs are effective against active HPIV3 infections.

    Table 1: Demographics and Outcomes for Recipients of Hexaviral Specific T-cells

    Patient # Infections Pre-VSTs Prior antiviral therapy CMV EBV Adv BKV HHV6 HPIV3 Outcome
    1 - Cidofovir (CMV cleared prior to VSTs) - - - - - - Free of viral infections;
    Alive and well
    2 CMV, BKV Cidofovir CR - CR CR - - Transient Adv detection post-infusion; cleared. Alive and well
    3 CMV, BKV Ganciclovir, Cidofovir CR - - PR - - BK hemorrhagic cystitis resolved post-infusion,with fluctuating BK viremia/viruria.
    Alive and well
    4 CMV, BKV Cidofovir CR - - CR - - Alive and well
    5 (EIND) AdV Cidofovir - - PR - - - Died, veno-occlusive disease
    6 - Valacyclovir - NR - - - - EBV reactivation without PTLD, treated with rituximab. Alive and well.
    7 (EIND) HPIV3 - - - - - - PR Transient radiographic improvement by 1 month.

    CR: complete response (resolution of viral infection); PR: partial response; NR: no response

    (89) Submission ID#600976

    Predictors of Fatigue in Common Variable Immunodeficiency

    Joud Hajjar, MD, MS1, Carleigh Kutac, MPH2, Tiffany S. Henderson, PhD3, Christopher Scalchunes, MPA4

    1Assistant Professor, Baylor College of Medicine, 1Texas Childrens Hospital Center for Human Immunobiology and Division of Immunology, Allergy and Rheumatology

    2Biostatistician, Baylor College of Medicine

    3Survey Research Analyst, Immune Deficiency Foundation

    4Vice President of Research, Immune Deficiency Foundation

    Introduction: We previously reported that fatigue is increased in common variable immunodeficiency (CVID). However, in previous studies, fatigue was not defined using validated tools. Our aim from this study is to identify the prevalence of patient-reported fatigue, using validated questionnaires, and determine the factors predisposing to fatigue in CVID

    Methods: Data from CVID who responded to the IDF 2017 patient national survey a were analyzed. Fatigue was measured using the Brief Fatigue Inventory (BFI) questionnaire, which includes seven items to identify fatigue, and measure fatigue severity. A total of 555 patients with CVID and responses to BFI were enrolled. Demographics, co-morbidities, immunoglobulin replacement therapy (IgGRT) route and dose, co-morbidities, infections, depression, quality of life (QOL) (using the SF-12v2) and disability were compared between fatigued and non-fatigued. Logistic regression was used to identify the significant variables.

    Results: The overall prevalence of fatigue was 83.24% (437/555), 69.55% of fatigued patients reported having moderate/severe fatigue. Significant predictors of fatigue were: Asthma (p=0.014), Patients Current Health status (p<0.001), history of Bronchitis (p=0.002), Sinus Infections (p<0.001), Pneumonia (p=0.028), Female sex (p<0.001), Employment Status (employed vs disabled; p<0.001), Household Income (p<0.001), BMI (p=0.024), abnormal digestive function (p<0.001), Permanent lung impairment (p<0.002), Recurrent Diarrhea (p<0.001), Inflammatory Bowel Disease (p=0.022), Depression (p<0.001), Noticeable wear-off of immunoglobulins (p<0.001), SF-12 Physical Component Score (PCS) (p<0.001), and SF-12 Mental Component Score (MCS) (p<0.001).

    Increased fatigue severity corresponded with significantly lower PCS and MCS scores. Using a univariate linear regression model for PCS and MCS score prediction, and the BFI variable (fatigue= none, mild, moderate, severe) as the independent variable, there was a significant increase in both PCS and MCS predicted scores as the severity of fatigue decreased (p<0.001). Improvement of fatigue scores from severe to moderate/mild predicted improvement in QoL scores, PSC scores by (5.3 and 12.7 points respectively) and MSC scores (4.9 and 9.9, respectively).

    Conclusion: Moderate to severe fatigue is significantly increased in CVID patients, increased fatigue severity predicts lower PCS and MCS, and improving fatigue score could lead to improvement in QoL and patient-reported health outcomes in CVID patients.

    (90) Submission ID#600981

    Rapid Response of CVID Skin Granulomatous Disease to Infliximab

    Maria Gabriela Torre, MD1, Eloisa Malbran, MD2, Maria Cecilia Juri, MD3, Alejandro Malbran, Phd4

    1Staff, Unidad de Alergia, Asma e Inmunologia Clinica; British Hospital; Army Hospital

    2Staff, Unidad de Alergia, Asma e Inmunologia Clinica; Sanatorio Mater Dei

    3Staff, Unidad de Alergia, Asma e Inmunologia Clinica; British Hospital; Hospital de Clinicas Jose de San Martin

    4Director; Head of Department, Unidad de Asma, Alergia e Inmunologia Clinica; British Hospital

    Granulomas are the most significant day-to-day problem for CVID patient management. Currently, there are limited options for their treatment and the optimal therapy is unknown. In case reports and small series, Infliximab has been reported effective while others found it useless.

    We here describe a 26yo white male referred for monthly IVIG in august 2016. At age 1, he developed large areas of erythematous polymorphic plaques in his cheeks, arms and legs. A skin biopsy showed tuberculoid granulomas negative for bacteria, BAAR and fungi, with infiltrating CD4+ lymphocytes. A prolonged course of steroids did not improve his skin. He also had multiple pneumonias and bronchiectasis, and oral candidiasis. He received all vaccines, including BCG with no complications. With low immunoglobulins and a poor response to pneumococcal polysaccharides and tetanus toxoid he was diagnosed as CVID and placed on IVIG at 7yo with excellent infectious control since then. At age 8, his skin lesions persisted and deepened to the bone on his left leg. Broad spectrum antibiotics for 3 months were unsuccessful. At 16yo to 18yo, skin grafts were performed on his arms, legs and both cheeks. Two ulcers persisted on his left leg until August 2018 that increased in size, deepened and became erythematous and extremely painful (Fig. 1). In September, two new ulcers appeared on his right cheek and right gluteus, respectively. One week later a third ulcer was found on his left calf. On September 28th, Infliximab 5mg/kg (300mg) was administered. On the second Infliximab dose, October 12th, the pain was completely gone and all ulcers were shrinking, and those ones in the cheek, gluteus and calf almost completely resolved. By the third dose, on November 23rd the ulcers in his right leg were almost closed (Fig. 2). Infliximab 300mg treatment continues every 8 weeks. Lab test remained unchanged from 2016 till 2018, when his wounds got worsened. (Table 1)

    Granulomatous disease in CVID is a challenge. Both B and T cell directed therapies are encouraged. We add a new case of an Infliximab responsive patient to others already reported.

    Fig 1. Before Infliximab After second dose Infliximab

    figurez

    Fig. 2

    figureaa

    Table 1

      08/2016 08/2017 12/09/2018 20/09/2018 28/09/2018
    IgG (600-1600mg/dl) 868 885    928
    IgA (70-400mg/dl) 41 31    30
    IgM (50-300mg/dl) 9 12    11
    IgE (1-100mg/dl) 2 3    4
    CD3 % (67-75) 96 98    98
    CD4 % (36-46) 47 51    50
    CD8 % (31-40) 43 45    44
    CD56/CD16 % (10-19) 3 1    
    CD19 % (11-16) 0 1    1
    CD20 % (11-16) 0 1    1
    Leucocytes (3700-9500/ul) 4000 3900 4100 3500 3600
    Beta2 micro globulin (0.8-2.3mg/l)   4.1 4.3   
    Sedimentation ((0-15mm) 12 14 28 21 13
    C3 (80-160mg/dl) 147 134 177 171 162
    C4 (15-45mg/dl) 52 51 50 51 51

    (91) Submission ID#601004

    A Novel Form of Partial Recessive IFN-gamma R2 Deficiency Caused by a Mutation of the Initiation Codon Presenting with a Severe Phenotype

    Ayse Metin, MD, PhD1, Saliha Knk Yüksel, MD2, Belgin Gulhan, MD2, Aslnur Ozkaya Parlakay, MD2, Carmen Oleaga Quintas, MSci3, Jacinta Bustamante, MD, PhD4

    1Prof. of Pediatric Immunology, MD, PhD, SBU, Ankara Children's Health and Diseases Hematology Oncology Training and Research Hospital, Ankara, Turkey

    2Pediatric Infectious diseases Unit, Ankara Children's Health and Diseases Hematology Oncology Training and Research Hospital, Ankara Turkey

    3M. Sci, Laboratory of Human Genetics of Infectious diseases, Necker Branch, INSERM UMR 1163, Imagine Institute, Necker hospital for Sick children, Paris, France

    4Senior Investigator, Laboratory of Human Genetics of Infectious diseases, Necker Branch, INSERM UMR 1163, Imagine Institute, Necker hospital for Sick children, Paris, France

    Introduction: Mendelian susceptibility of mycobacterial disease (MSMD) is characterized by infections caused by weakly virulent mycobacteria in otherwise healthy individuals. Known genetic etiologies disrupt IFN-gamma (IFN-g) immunity like IFNGR2. Germline bi-allelic mutations of IFNGR2 can underlie partial or complete deficiency of IFN-g receptor2 (IFN-gR2). Patients with partial IFN-gR2 deficiencies express dysfunctional molecule on the cell surface. We describe here a novel mutation within the start of IFNGR2 in two siblings born to a consanguineous parents and their clinical presentation.

    Methods: We studied 2 siblings fron one kindred from Turkey by whole- exom sequencing (WES). Candidate mutation of IFNGR2 was analzed experimentally.

    Conclusion: We describe a novel mutation in both siblings at the first codon of IFNGR2 that define a new form of partial recessive IFN-gR2 deficiency. The low amounts of full length IFN-gR2 confer a more severe clinical phenotype in these siblings than that of patients with other forms of partial recessive IFN-gR2 deficiency due to surface expressed dysfunctional receptors.

    (92) Submission ID#601013

    Maternal Diabetes Causing Atypical, Complete DiGeorge Syndrome

    Richa Panara, MD1, Kiran Patel, MD2, Lisa Kobrynski, MD, MPH3, Gerald Lee, MD2, Kristine Vanijcharoenkarn, MD4, Meera Patrawala, MD4, Jennifer Shih, MD5

    1Resident Physician, Department of Internal Medicine, Emory University School of Medicine

    2Assistant Professor of Allergy, Department of Pediatrics, Emory University School of Medicine

    3Associate Professor of Allergy, Department of Pediatrics, Emory University of School of Medicine

    4Allergy and Immunology Fellow, Emory University School of Medicine

    5Assistant Professor of Pediatrics and Internal Medicine, Department of Pediatrics, Division of Allergy and Immunology, Emory University School of Medicine

    INTRODUCTION: DiGeorge Syndrome (DGS) is a primary immunodeficiency characterized by thymic hypoplasia, cardiac defects and hypoparathyroidism. Approximately 90% of cases of DGS are due to a chromosomal microdeletion at 22q11.2, however non-genetic etiologies that have been described in the literature include maternal diabetes, fetal alcohol syndrome, and prenatal exposure to isotretinoins.

    CASE PRESENTATION: We describe an 8 month-old male born at 38 weeks to a woman with insulin dependent diabetes mellitus. At delivery, examination revealed microcephaly, small upper lips, small nares and pyriform aperture stenosis. Chest imaging revealed no thymic tissue, right-sided rib and spinal anomalies. Renal ultrasound was normal.

    Newborn Screen showed critically low levels of TREC. Flow cytometry confirmed low T-cells: CD4 count 1 cell/ uL, CD8 count 7 cell/uL, CD3 count 8 cells/uL/ <1%, CD19 at 1398 cells/uL/79%. Helper and suppressor t-cells <1%. CD4/CD8 ratio 0.6. Lymphocyte proliferation to mitogens was absent to phytohemagglutinin (PHA) and decreased to pokeweed mitogen (PWM). Calcium and parathyroid hormone were low. Quantitative immunoglobulins: IGG 488mg/dL, IgA <8mg/dL, IgM 40mg/dL.

    DNA microarray for 22q deletion, TBX1 testing, and CHD7 sequencing was negative. FISH for trisomy 13, 18, 21 was normal. Given the negative genetics, the cause of DGS in this case is likely maternal diabetes.

    Patient has had a complicated course with recent admission for hypocalcemia requiring IV calcium infusions complicated by acute liver dysfunction with coagulopathy, MRSA and pseudomonas bacteremia, and tube feeding intolerance with abdominal distention and diarrhea. He was noted to have lymphadenopathy, generalized rash and eosinophilia, concerning for oligoclonal expansion of T-cells. Repeat testing showed increased total T cells from 43 cells/uL to 342 cells/uL with an increase in CD4 cells from 21 cells/uL to 293 cells/uL that were 96% memory. Cytokine panel IL2 receptor 1059 pg/ml suggested T-cell activation. EGD and colonoscopy biopsies showed descending colon and rectum colitis.

    DISCUSSION: Maternal diabetes mellitus has been described as a possible etiology of DGS in several case reports. Infants born to insulin-dependent diabetic mothers are eight times more likely to have major malformations than infants of non-diabetic mothers. The cause of these defects is hypothesized to be from alteration of neural crest migration and development. Since 10% of DGS are not due to a chromosomal microdeletion at 22q11.2, it is prudent to consider maternal diabetes mellitus as an etiology for DGS, especially when the microdeletion is not found, but clinical presentation is suggestive of DGS.

    (93) Submission ID#601038

    Novel Mutation in the WAS Gene Causing a Phenotypic Presentation of Wiskott- Aldrich Syndrome

    Elisa Ochfeld, MD1, Melanie Makhija, MD2

    1Allergy/ Immunology Fellow, Northwestern University

    2Attending, Northwestern University

    Introduction/ Background: Wiskott- Aldrich Syndrome (WAS) is a rare X- linked disorder characterized by immunodeficiency, thrombocytopenia and atopic dermatitis. The phenotype is variable and depends on the type and location of the mutation in the WAS gene (located on Xp11.23). The treatment of choice for WAS is hematopoietic stem cell transplant (HSCT).

    Objective: To present a clinical case with a novel genetic mutation causing WAS.

    Results: We describe a now 5 month-old infant who presented at birth with petechiae and bruising of his legs and abdomen after an elective cesarean section at 38 weeks gestation. He was found to have severe neonatal thrombocytopenia based on complete blood count (CBC). He received a platelet transfusion on day of life (DOL) 1, was discharged home with stable platelets on DOL 5 but had worsening thrombocytopenia on DOL 8 requiring re-admission for further work up (platelet count 40,000). Platelet count reached a nadir of 26,000 on DOL 11. He was initially thought to have neonatal alloimmune thrombocytopenia. Parental testing showed no platelet abnormalities or platelet alloantibodies. He re-presented for acute lower GI bleeding on DOL 19. His platelet morphology at that time showed small platelets, though previously his platelet size was reportedly normal. Genetic testing for WAS revealed a variant of unknown significance (VUS) in the WAS gene (WAS NM_000377.2. Exon 2. C.151GT. P.VAL51Phe. Hemizygous. VUS. Chr X: g: 48542690G>T). This mutation, WAS c.151G > T (p. Val51Phe) is a missense variant in exon 2 which changes an amino acid valine at codon 51 to phenylalanine. This is a region essential for interaction with the WASP interacting protein (Rajmohan, 2009). This variant has not been reported in the literature as pathogenic and has not been reported as a known variant in the general population. In silico computational evidence predict the variant to be damaging (Polyphen2, Mutation Taster, SIFT, HSF). Further immune evaluation was performed. Flow cytometry revealed a low absolute CD3 count, low absolute CD4 count, low absolute CD8 count, CD4/CD8 ratio of 5:1, with normal NK cell count and low absolute CD19 count. His immunoglobulins were abnormal, with low IgM and elevated IgA with normal IgG and slightly elevated IgE. WAS protein expression testing revealed decreased expression of WAS protein on lymphocytes (0.24 ratio, normal range 0.71-1.31). At the time of his initial presentation to immunology at age 3 months, he had not developed atopic dermatitis nor any significant infections. However, at 4 months he was admitted for otitis externa with adjacent facial cellulitis, which responded to IV antibiotics. During that admission he was given IVIG and started on Bactrim prophylaxis. He has since developed significant seborrheic dermatitis skin lesions. Family history was not significant for immunodeficiency. No known WAS in family, no consanguinity. Maternal genetic testing was positive for the same missense mutation in the WAS gene. The patient is currently undergoing stem cell transplantation with a matched sibling donor.

    Conclusion: We report a new mutation in the WAS gene that causes a phenotypic presentation of Wiskott-Aldrich Syndrome.

    (94) Submission ID#601045

    Common Variable Immunodeficiency (CVID) Associated with Variants in Neuroblastoma Amplified Sequence (NBAS) Gene

    Joseph A. Church, MD1, Dominic Lenz, MD2, Christian Staufner, MD3, Georg Hoffmann, MD4

    1Professor, Pediatrics, Children's Hospital Los Angeles and Keck School of Medicine of U.S.C.

    2Resident, Department of General Pediatrics, University Hospital Heidelberg, Heidelberg Germany

    3Consultant, Department of General Pediatrics, University Hospital Heidelberg, Heidelberg, Germany

    4Professor of Pediatrics, Department of General Pediatrics, University Hospital Heidelberg, Heidelberg, Germany

    Over 20 genes have been reported to cause monogenic CVID.

    A 4 year old girl presented with recurrent pneumonias and a diagnosis of CVID. The parents sought a second opinion. Born at 33 weeks gestational age, she was "always smaller and sicker than her friends," and in the prior 8 months she had 3 episodes of pneumonia with fever to 104F requiring emergency department treatment. Two of these were associated with RSV and metapneumovirus, respectively.

    Laboratory evaluation confirmed low levels of IgG (326 mg/dL) IgA (7) and IgM (6), and poor antibody responses to tetanus, Hib, MMR and VZV antigens. Her response to Pneumovax was marginal (+12 of 23 serotypes tested) and transient (#2 of 23 after 4 months). CBC including smear and chemistry panel were normal. Results of routine lymphocyte subset analysis were normal except for relatively low NK cells (3%). On examination she appeared well. Ht and wt were at the 20th%iles. There was no evidence of active infection, hepatosplenomegaly, lymphadenopathy or skin rash. She had a small VSD noted since birth, and bilateral optic atrophy identified during routine ophthalmology follow-up for prematurity. Ig replacement was initiated and continues as weekly infusions of SCIG, maintaining IgG levels over 1000 mg/dL.

    At 4.5 years she was hospitalized for pneumonia, and transiently elevated transaminases (AST 117 U/L, ALT 552) were noted for the first and only time.

    Exome sequencing revealed compound heterozygous, likely pathogenic, variants limited to the C-terminal domain of the NBAS gene. The product of this gene is a component of the syntaxin 18 complex that is implicated in Golgi to ER retrograde transport.

    Mutations in NBAS are associated with Infantile Liver Failure Syndrome 2 (ILFS2, OMIM#616483) and Short Stature, Optic Atrophy and Pelger-Huet Anomaly in neutrophils (SOPH syndrome). Hyopgammaglobulinemia and low NK cell numbers have been reported in affected patients, and NBAS deficiency is listed in the IUIS 2017 PID Committee report, but under "Defects in intrinsic and innate immunity."

    Clinical features reported in NBAS deficiency include familial consanguinity, fever-related recurrent liver crises, short stature (<3%ile), skeletal dysplasia, cervical instability (small C1-C2) and retinal dystrophy. To the present the patient has not exhibited these findings.

    Our patient's NBAS variants are being studied for functional validity. A skin biopsy was performed and fibroblasts cultured for analysis. In contrast to our patient's variants that are both in the C-terminal domain, most reported cases have at least 1 mutation affecting domains further toward the N-terminus. To the present our patient's fibroblasts do not show a clear reduction in NBAS protein, and protein interaction partners were not diminished. Additional studies planned include challenging the fibroblasts with elevated temperature to measure any reduction in variant NBAS expression and function.

    (95) Submission ID#601049

    An Unexpected Diagnosis in a Premature Infant with Persistent Fever, Respiratory Distress and Significant Neutrophilia: Congenital Tuberculosis

    Idil Ezhuthachan, MD1, Sara Sussman, MD2, Renata Schillizzi, NP3, Shanika Uduwana, MD1, Lily Glater, MD4, Blanka Kaplan, MD5, Todd Sweberg, MD6, Maria Santiago, MD7, Stefan Hagmann, MD7, Vincent Bonagura, MD8, Gina Coscia, MD5

    1Fellow, Northwell Health

    2Fellow, Department of Pediatrics, Zucker School of Medicine at Hofstra Northwell School of Medicine

    3Nurse Practitioner, Northwell Health

    4Pediatric Intensivist, Northwell Health Cohen Children's Medical Center

    5Assistant Professor of Medicine and Pediatrics, Northwell Health

    6Assistant Professor of Pediatrics, Northwell Health

    7Associate Professor of Pediatrics, Northwell Health

    8Professor of Medicine and Pediatrics, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    Congenital tuberculosis (CTB) is a rare disease most often associated with maternal genitourinary (GU) tuberculosis (TB) or disseminated TB. Due to infertility caused by GU TB, CTB is rarely reported even in endemic countries. Infants can acquire TB hematogenously via the placenta or umbilical vein or by fetal aspiration of infected amniotic fluid. Presenting symptoms include respiratory distress, fever, hepatosplenomegaly, poor feeding, lethargy, and low birth weight.

    We report a premature female infant conceived via in vitro fertilization (IVF), who was born to Indian immigrant parents at 29 weeks of gestation due to preterm premature rupture of membranes. Maternal history was significant for pulmonary TB at 9 years of age. She denied abdominal or GU symptoms. Infants NICU course was complicated by opacifications in the right lung and leukocytosis with neutrophil predominance, identified during evaluation of frequent apnea and bradycardia episodes at 1 month of age. Clinical improvement was noted after treatment with vancomycin, amikacin and piperacillin-tazobactam; however, leukocytosis of unknown etiology persisted. At 1.5 months of age she was discharged to inpatient rehabilitation.

    At 3 months of age, she was readmitted for fever and respiratory distress. During this admission, an immune evaluation was undertaken due to persistence of symptoms along with unresolved leukocytosis with a peak of 58,000 cells/l with neutrophilia to 42,850 cells/l, and chest CT evidence of progressive multifocal lung disease worse in the right upper lobe despite empiric treatment with broad-spectrum antibiotics. Infectious work-up was negative, including acid-fast bacilli testing from bronchoalveolar lavage. Due to the pronounced and persistent leukocytosis and neutrophilia, a primary immune defect was suspected. Immune evaluation included: normal immunoglobulins (Ig) G, A, and E, elevated IgM, vaccine-specific antibody titers protective to diphtheria and 9 of 13 Streptococcus pneumonia strains, mildly elevated T and B cells, a normal flow cytometry for dihydrorhodamine, myeloperoxidase stain and glucose-6-phosphate dehydrogenase level, as well as a peripheral smear with no giant azurophilic granules. Her primary immunodeficiency genetic panel was unrevealing. She underwent lung biopsy via video-assisted thoracoscopic surgery, which showed non-caseating granulomas and eventual growth of multi-drug-resistant Mycobacterium tuberculosis (MTB). Upon treatment with an appropriately adjusted anti-tuberculosis regimen, she showed rapid clinical and laboratory improvement. Endometrial samples obtained from mother showed GU TB, confirming the diagnosis of CTB.

    The slow-growing nature of MTB that resulted in delayed diagnosis, along with the presence of non-caseating granulomas and persistent neutrophilia, prompted an immune work up that was completely normal. This case demonstrates the importance of considering CTB in the differential diagnosis of an infant presenting with severe lung infection, persistent neutrophilia, suboptimal response to broad-spectrum antibiotics and relevant epidemiologic risk factors. Furthermore, in the setting of appropriate parental exposures and infertility prompting the use of IVF, maintaining a high level of suspicion of CTB can aid in earlier diagnosis of affected neonates.

    (96) Submission ID#601056

    Herpes Simplex Virus Whole Genome Sequencing for Antiviral Resistance in a Child with DOCK8 Deficiency and Chronic Infection

    Sean Stout, MD1, Alexander Greninger, MD, PhD, MS, M.Phil.2, Rangaraj Selvarangan, PhD3, Alexandra F. Freeman, MD4, Brandon Newell, MD5, Erin Stahl, MD6, Dwight Yin, MD, MPH7

    1Pediatric Resident, Department of Pediatrics, Children's Mercy Hospital

    2Department of Laboratory Medicine, University of Washington School of Medicine

    3Department of Pathology and Laboratory Medicine, Children's Mercy Hospital; School of Medicine, University of Missouri-Kansas City

    4Director, Primary Immune Deficiency Clinic, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    5Division of Dermatology, Department of Pediatrics, Children's Mercy Hospital; School of Medicine, University of Missouri-Kansas City

    6Section of Ophthalmology, Department of Surgery, Children's Mercy Hospital; School of Medicine, University of Missouri-Kansas City

    7Division of Infectious Diseases, Department of Pediatrics, Children's Mercy Hospital; School of Medicine, University of Missouri-Kansas City

    Background: Patients with dedicator of cytokinesis 8 (DOCK8) deficiency are prone to severe, recurrent or chronic mucocutaneous herpes simplex virus (HSV) infections that may develop antiviral resistance. We present the case of a child with DOCK8 deficiency and chronic, resistant HSV-1 mucocutaneous infections to illustrate the potential clinical utility of an investigational viral whole genome sequencing approach to detecting active and latent HSV resistance mutations longitudinally.

    Methods: We abstracted clinical and laboratory data in a 14 year old boy with DOCK8 deficiency with repeated viral culture growth of HSV-1. HSV-1 DNA from seven positive viral culture specimens collected during 2015-2018 were sequenced on an Illumina® MiSeq to >150X depth. Consensus genomes were called using an established HSV genome pipeline, and reads were mapped to the HSV-1 strain reference genome (NC_001806). Sequence variants were checked against an online database of UL23 (thymidine kinase) and UL30 (DNA polymerase) variants associated with antiviral resistance.

    Results: The patient had low CD4+ T cells (initial 248 cells/mm3), eosinophilia, elevated IgE (initial 9660 kU/L), severe eczematous dermatitis, chronic obstructive and interstitial lung disease, growth delay, and presented with recurrent infections including Staphylococcus aureus, Candida albicans, JC virus, and HSV-1. He was receiving prophylaxis with subcutaneous immunoglobulin G, trimethoprim-sulfamethoxazole, and acyclovir. Family declined hematopoietic stem cell transplantation. Mucocutaneous HSV-1 infections extensively involved his face, trunk, and extremities with more severe infections involving his cornea, lips, perineum, scalp, and periorbital regions. Scalp and periorbital lesions were proliferative and edematous papules and plaques consistent with HSV vegetans (Figures 1-2) and confirmed by scalp biopsy. Although early HSV-1 infections responded to oral or IV acyclovir, clinical response decreased over time, requiring advancement of therapies to high-dose acyclovir IV, foscarnet IV, cidofovir IV, topical cidofovir cream, and/or interferon-alpha with variable clinical response.

    Phenotypic testing detected acyclovir resistance in HSV isolated from four samples while the patient was on acyclovir and no resistance in a sample while not on acyclovir. Phenotypic foscarnet resistance was detected in one sample without prior patient exposure to foscarnet. Viral whole genome sequencing detected the UL23 variant R176Q (associated with acyclovir resistance) on all specimens, whether on acyclovir or not, and the UL30 variant T821M (associated with acyclovir and cidofovir resistance) only when on cidofovir. When phenotypic testing and genome sequencing were discordant, clinical response appeared to be more consistent with genome sequencing results.

    Conclusions: This patient with DOCK8 deficiency illustrates the potential severity of chronic, resistant mucocutaneous HSV-1 infection. Viral genome sequencing for antiviral resistance mutations may provide additional information about the presence of clinically significant variants, which may result from detecting smaller or latent HSV-1 sub-populations.

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    (97) Submission ID#601066

    Variable Phenotypes in Three Patients with Two Novel STAT3 Gain of Function Mutations

    Musa G. Bolkent1, Melissa Elder, MD, PhD2, Tiphanie Vogel, MD, PhD3, Akaluck Thatayatikom, MD, RhMSUS4

    1Fellow, University of Florida

    2Division Chief, University of Florida

    3Assistant Professor, Baylor College of Medicine, 1Texas Childrens Hospital Center for Human Immunobiology and Division of Immunology, Allergy and Rheumatology

    4Associate Professor, University of Florida

    Introduction: Gain-of-function (GOF) mutations in signal transducer and activator of transcription 3 (STAT3) cause autosomal dominant, early-onset autoimmune disease. STAT3 GOF syndrome is characterized by lymphadenopathy (LAD), hepatosplenomegaly (HSM), autoimmune cytopenias, hypogammaglobulinemia and other solid organ autoimmunities, including interstitial lung disease, enteropathy, endocrinopathy, arthritis, and scleroderma-like skin changes. We report three children with two different novel missense STAT3 GOF mutations.

    Objectives: To describe novel GOF mutations of STAT3 that expand our current understanding of the clinical manifestations of the disease.

    Methods: Molecular investigation was performed in suspected patients using whole exome sequencing (WES). The STAT3 mutations caused GOF were tested using a luciferase assay.

    Results: Results of WES identified three children with novel STAT3 mutations at the same amino acid (Table 1). Standardized luciferase assay confirmed the mutations caused GOF in the STAT3 protein. Case#1 was a 15-year-old Caucasian male who initially presented with recurrent otitis media, persistent HSM, LAD, and hypogammaglobinemia (IgG <170 mg/dL) at 2 years of age. He was diagnosed with common variable immunodeficiency (CVID) and chronic arthritis when he was 6 and 9 years of age, respectively. Subsequently, he developed hepatitis and recurrent pneumonia with Mycobacterium avium complex (MAC). His arthritis partially responded to anti-tumor necrosis factor (TNF) agents and tofacitinib, but did not respond to anti-interleukin-6 treatment. A combination of anti-TNF inhibitor, tofacitinib, and low dose prednisone was required to control his arthritis. Case#2 was an 18-year-old Caucasian male who initially developed thrombocytopenia, hypogammaglobulinemia (IgG <110 mg/dL), recurrent otitis media, pneumonia, Crohn's disease, celiac disease, LAD and failure to thrive at 2 years of age with more recent development of HSM. He required only immunoglobulin replacement therapy. Case#3 is a 9-year-old Caucasian male, the half-brother of case#2, who initially presented with recurrent pleural effusion and bilateral pulmonary infiltrates, HSM, LAD, abdominal distension and ascites at 7 years of age. A transbronchial lung biopsy revealed chronic eosinophilic pneumonitis. Liver biopsy showed increased eosinophils in the sinusoids with diffuse enlargement of hepatocytes, but without hepatitis. Colon biopsy revealed minimal colonic eosinophilia. His pulmonary infiltrates and pleural effusion responded to prednisone, and he has not required additional treatment for past 1.5 years.

    Conclusions: The clinical manifestations of the same genetic variant may be variable and unpredictable even in the same family. STAT3 GOF syndrome should be considered in children with multisystem autoimmune diseases, LAD, HSM and low switched memory B cells regardless of presence of hypogammaglobulinemia or history of recurrent infections.

    Table.1 Patient Characteristics

    Pt Age at onset (years) STAT3 GOF mutation Class switched memory B cells Infections GI MSK* Lympho-proliferation Other
    #1 2 c.1165 A>T (p.T389S) <1% Sinusitis, otitis media, pneumonia Hepatitis Chronic arthritis LAD, HSM Uveitis
    #2 2 c.1165 A>G (p.T389A) 1% Pneumonia Crohn’s disease None LAD, HSM *ITP, Portal
    HTN
    #3 7 c.1165 A>G (p.T389A) <1% none Abdominal distensionwith colonic eosinophilia None LAD, HSM Chronic eosinophilic pneumonitis

    (98) Submission ID#601069

    Immunophenotype and Metabolic Characteristics of EBV-Specific T Cells Generated Using Different Manufacture Approaches

    Danielle E. Arnold, MD1, David M. Barrett, MD, PhD2

    1Fellow In Training, Children's Hospital of Philadelphia

    2Attending Physician, Children's Hospital of Philadelphia

    Background: Patients with primary immune deficiencies characterized by severe T lymphopenia and/or poor T cell function and patients post-hematopoietic cell transplantation are at high risk of severe viral infections. Antiviral medications are expensive, not always effective and associated with significant toxicity and/or long-term side effects. As such, there has been increasing interest in the use of donor-derived or third-party virus-specific T cells (VSTs), and several studies have demonstrated efficacy of VSTs generated using various manufacture strategies. However, in depth immunologic and metabolic characterization of VSTs has not been reported, limiting correlative investigations into efficacy.

    Methods: EBV-VSTs were generated from apheresis T cells collected from healthy donors using three methods: (1) stimulation and expansion with HLA-matched EBV-lymphoblastoid cell lines (LCLs) purchased from Astarte Biologics or Sigma-Aldrich over a period of 4 weeks, (2) stimulation with EBV PepTivator from Miltenyi followed by expansion over 9-12 days with different cytokines, and (3) stimulation with EBV PepTivator followed by isolation of activated cells using the IFN-gamma capture system from Miltenyi. Immunophenotyping by flow cytometry was performed using the Miltneyi MACSQuant Analyzer. The NanoString nCounter system was used to measure gene expression for metabolic pathway analysis, and the Agilent Seahorse XF cell mito stress test system was used to measure mitochondrial respiration.

    Results: EBV-VSTs generated using LCLs or PepTivator plus IL-15 both resulted in a high percentage of CD8 T cells skewed to the effector memory and terminal effector memory phenotype with high expression of the exhaustion markers PD-1, TIM-3, and LAG-3. Conversely, EBV-VSTs generated using PepTivator plus IL-4 and IL-7 and the IFN-gamma capture system resulted in a mixed CD4 and CD8 T cell population with a high number of central memory T cells and lower percentage of cells positive for PD-1, TIM-3, and LAG-3. Stimulation with PepTivator followed by expansion with IL-2 resulted in an intermediate immunophenotype. NanoString results demonstrated upregulation of the glycolytic pathway in EBV-VSTs stimulated with PepTivator followed by expansion with IL-2 or IL-15 compared to EBV-VSTs generated using the other manufacture approaches. The Seahorse mito stress test demonstrated that the PepTivator plus IL-2 EBV-VSTs had a significantly lower spare respiratory capacity than other EBV-VSTs and a low extracellular acidification rate despite upregulation of the glycolytic pathway. The Peptivator plus IL-4 and IL-7 EBV-VSTs had the highest basal oxygen consumption rate, ATP-linked respiration, and extracellular acidification rate.

    Conclusions: Manufacture of EBV-VSTs using the various approaches currently employed clinically results in T cell pools with different immunophenotypes and different metabolic profiles. EBV-VSTs stimulated with PepTivator followed by expansion in IL-4 and IL-7 and EBV-VSTs isolated using the IFN-gamma capture system have immunophenotypes and metabolic phenotypes suggestive of potential greater in vivo persistence, whereas EBV-VSTs expanded in IL-2 and IL-15 have characteristics correlated with increased effector function. However, these VSTs are more likely to be short-lived and to have impaired metabolic fitness. These phenotypes will enable better correlation with clinical results and suggest combinatorial approaches depending on clinical indication.

    (99) Submission ID#601079

    Efficacy, Tolerability and Safety of Cutaquig®, a New Subcutaneous Human Immunglobulin 16.5% in Adult Patients with Primary Immunodeficiencies

    Latysheva E.1, Rodina Y.2, Sizyakina L.3, Totolian A.4, Tuzankina I.5

    1National Research Center Institute of Immunology FMBA, Russia

    2Dmitry Rogachev National Research Center of Pediatric Hematology, Oncology and Immunology, Moscow

    3State Medical University, Rostov

    4Pasteur Institute, Saint-Petersburg

    5Institute of Immunology and Physiology of the Ural Branch of the Russian Academy of Sciences, Yekaterinburg

    Introduction: Majority of patients with primary immunodeficiencies (PID) require life-long replacement therapy with immunoglobulins (Ig) to prevent severe infections and irreversible complications. In addition to safety and efficacy, tolerability and convenience of administration of Ig products are essential factors for patients. A new 16.5% Ig preparation octanorm (Octapharma, Lachen; tradename cutaquig® in North America) has been developed for subcutaneous administration (SCIG) derived from the established manufacturing process of Octapharmas intravenous Ig (IVIG) brand octagam®.

    Objectives: Primary outcome was assessment of the efficacy of octanorm in preventing serious bacterial infections. Main secondary endpoints included (among others) evaluation of tolerability and safety of octanorm, the number and rate of other infections, number of days missed at work, and use of antibiotics.

    Methods: A prospective, open-label, non-controlled, single-arm phase 3 study involving 25 adult patients with PID was conducted at 5 Russian centers. Patients treated with at least 4 infusions of IVIG prior to enrollment and with IgG trough levels 5.0 g/L underwent an 8-week wash-in/wash-out period followed by a 24-week efficacy period. During the study, patients received weekly administrations of octanorm at the same monthly dose as during previous IVIG treatment (monthly IVIG dose divided by 4 for weekly dose). In total, each patient received 32 SCIG infusions.

    Results: Twenty-four patients completed the study. One patient terminated early (after infusion 7, during wash-in/wash-out phase; personal reasons). Mean age was 35.24 years (range 18-64 years). Fifteen patients (60%) were female and 10 patients (40%) male.

    No serious bacterial infections were recorded. During the efficacy period a total of 26 non-serious infections was observed in 14 patients. Seventeen infections in 11 patients were of mild and 9 infections in 5 patients of moderate intensity. The infection rate per person-year was 2.37.

    In total 25 patients received 775 infusions of study drug. The average dose of cutaquig® was 0.11 g/kg/week. During the entire study, 59 systemic adverse events were reported (including 34 infections). Three of these systemic adverse events were rated as related to study drug, all were non-serious. There was no serious or significant adverse event nor was there an adverse event leading to withdrawal. Infusion site reactions were reported for 15% of infusions.

    Serum IgG trough levels were nearly constant during the efficacy period. Median IgG trough levels were 8.15 g/L at Screening, 9.52 g/L at the end of wash-in/wash-out period and 10.71 g/L at the Termination Visit. One patient had a trough level 5g/L at 2 visits during the efficacy period and the dosing was subsequently adjusted for this patient.

    During the primary treatment period 10 patients (41.7%) used antibiotics in 19 treatment episodes (total of 229 treatment days; range 5-76 days) and 3 patients had 4 absences from work or school due to infections (total of 14 days of absence).

    Conclusion: This study demonstrated that the new subcutaneous human normal immunoglobulin 16.5% is well tolerated, safe and effective in adult patients with PID.

    (100) Submission ID#601082

    Infective Endocarditis, Osteomyelitis of Skull and Invasive Aspergillosis in a Child with Chronic Granulomatous Disease

    Gummadi Anjani, MBBS;MD Pediatrics1, Amit Rawat, MD (Pathology) PDCC (Laboratory Immunology) PDCC (Nephropathology) MAMS2, Ankur Jindal, MD;DM (pediatric clinical immunology and rheumatology)3, Pandiarajan Vignesh, MD;DM (pediatric clinical immunology and rheumatology)3, Ankita Singh, MD4, Surjit Singh, MD; DCH (Lon.); FRCP (Lon.); FRCPCH (Lon.); FAMS5

    1Fellow in pediatric clinical immunology and rheumatology, Postgraduate institute of medical education and research

    2Professor of Pediatric Allergy and Immunology, Paediatric Allergy Immunology Unit, Department of Paediatrics, Advanced Paediatric Centre, Postgraduate Institute of Medical Education & Research

    3Assisstant professor, department of pediatrics, Postgraduate Institute of Medical Education & Research

    4fellow in pediatric clinical immunology and rheumatology, Postgraduate Institute of Medical Education & Research

    5Head, Department of Pediatrics and Chief, Allergy Immunology Unit, Advanced Pediatrics Centre,Principal Investigator, Indian Council of Medical Research (ICMR) Centre for Advanced Research in Primary Immunodeficiency DiseasesVice-President, Indian Rh, Postgraduate Institute of Medical Education & Research

    Background: Children with chronic granulomatous disease (CGD) are at high risk for fungal infections (especially with Aspergillus species) and these infections usually have contiguous site involvement. Most patients have pulmonary presentation. Infective endocarditis and fungal osteomyelitis of skull are distinctly unusual. We report one such case.

    Case: A 6-year-old boy, born out of a non-consanguineous marriage, presented with soft tissue swellings of skull for 2 months. His past history was significant with an episode of pneumonia at 1 year and recurrent soft tissue swellings all over the body since 1½ years of age. On examination he was wasted, had signs of micronutrient deficiency, rickets, pallor, cervical lymphadenopathy and two abscesses, 12x4 cm on right temporo-parietal region and 4x3 cm over left frontal region. He was also found to have hyperdynamic precordium with an ejection systolic murmur. Investigations revealed hemoglobin 85g/L; platelet count 7.34x109/L; total leukocyte count 13x109/L(N60/L23/M13/E1); elevated C-reactive protein( 244 mg/L) and a raised erythrocyte sedimentation rate(104 mm 1sthr). Chest x ray revealed cardiomegaly (cardiothoracic ratio 67%) and 2D echocardiography showed vegetation of 6x3 mm over the anterior mitral leaflet suggestive of infective endocarditis. Blood and urine cultures were sterile. Culture from pus over the temporo-parietal abscess showed growth of Aspergillus fumigatus. Human immunodeficiency virus serology was non-reactive. Immunoglobulin profile revealed elevated IgG 21.20g/L (5.40-16.10g/L) and IgA 5.66 g/L(0.5-2.4g/L); IgM was 1.63 g/L(0.50-1.8g/L). In view of strong suspicion of CGD, nitroblue tetrazolium dye reduction test (NBT) was carried out- it revealed no reduction and Dihydrorhodamine (DHR) assay showed a low stimulation index (4.34). Flow cytometry for gp 47 phox and gp 67 phox was normal and DHR of mother did not reveal X linked carrier state. Contrast enhanced computerized tomography (CECT) of head showed osteomyelitis of the calvarial bones. Contrast enhanced magnetic resonance imaging (CEMRI) brain showed heterogeneously enchancing soft tissue lesion in the scalp at right fronto-parietal region and left frontal region with underlying bony destruction suggestive of osteomyelitis. He was given intravenous antimicrobials (ceftriaxone, gentamycin, cloxacillin, voriconazole). After 6 weeks of therapy, he showed resolution of findings on MRI brain and a repeat 2D echocardiography showed significant decrease in size of mitral leaflet vegetation.

    Conclusion: This case highlights a rare presentation of CGD with infective endocarditis and skull osteomyelitis due to Aspergillus fumigatus. To the best of our knowledge, this has not been reported previously.

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    (101) Submission ID#601094

    Enigmas of IL-12R1 Deficiency: Contemporary of Two Disease, Mendelian Susceptibility to Mycobacterial Disease and Crohn Disease

    Roya Sherkat, MD 1, Razieh Khoshnevisan, MD2, Nioosha Nekoei, PhD3, Christoph Klein, MD, PhD4, Daniel Kotlarz, MD , PhD5, Mahdieh Behnam, MSc6, Soodabeh Rostami, PhD7, Majid Yaran, PhD8, Hamid Tavakoli, MD9, Abbas Rezaei, PhD10

    1Associate Prof. ,Head of Acquired Immunodeficiency Research Center, Acquired Immunodeficiency Research Center, Isfahan University of Medical Sciences , Isfahan , Iran

    2Phd, Acquired Immunodeficiency Research Center, Isfahan University of Medical Sciences , Isfahan , Iran

    3Research Assistant, Acquired Immunodeficiency Research Center, Isfahan University of Medical Sciences , Isfahan , Iran

    4Director, Dr. von Hauner Children's Hospital, University Hospital, LMU, Munich Germany

    5Senior Scientist, Department of Pediatrics Dr. von Hauner Childrens Hospital, Ludwig- Maximilians-University, Munich, Germany

    6Research Assistant, Medical Genetics Laboratory, Alzahra University Hospital, Isfahan University of Medical Sciences, Isfahan, Iran

    7Assistant Prof., Nosocomial Infection Research Center, Isfahan University of Medical Sciences, Isfahan, Iran.

    8Scientist, Acquired Immunodeficiency Research Center, Isfahan University of Medical Sciences , Isfahan , Iran

    9Associate prof., Integrative Functional Gastroenterology Research Center, Isfahan University of Medical Sciences, Isfahan, Iran

    10Prof., Immunology Department, Isfahan University of Medical Sciences, Isfahan, Iran.

    Background: Genetic defect in IL12R1 affect cellular immunity, underlie Mendelian Susceptibility to Mycobacterial Disease (MSMD) and Inflammatory bowel disease (IBD) through different pathways.

    We present for the first time a patient with IL-12R1 deficiency from a consanguine family with two different phenotypes. Initially diagnosed as Crohn's disease prior to the MSMD diagnosis.

    Method and Material:Patient was referred to the clinical Immunology and Allergy clinic at the at Alzahra University Hospital for immunological and genetic evaluation . Blood samples from patient, his family and healthy donor controls were collected upon informed consent.

    In this study, we investigated effect of IL12R1 mutation in IL-12/IFN- axis by evaluation of patients whole blood cell response to IL-12 and IFN-, IL-12R1 expression in PBMCs and T cell blasts. Also Whole-exome sequencing has been performed.

    Result and Discussion: A 26 years old male from consanguine family , with history of right sub-axillary BCG lymphadenitis, recurrent mouth ulcers , chronic diarrhea in childhood and appendectomy at age of 5 was investigated. Based on his clinical presentation abdominal pain, significant weight loss, chronic and bloody diarrhea , endoscopic and pathological findings treatment for Crohn's disease (CD) was started at the age of seven . Unfortunately, protracted patient's symptoms ends up to resection of his colon and colostomy two years later. He was presented with multi focal osteomyelitis at the age of 13 . Although no bacteria was detected in PCR and tissue culture of the bone biopsy and the patient was not responded to antibacterials , he had a dramatic response to empirical anti mycobacterial treatment and his severe bone pain and lesions were healed. Even though the bone manifestations were completely controlled, he continuously was under treatment for his gastrointestinal symptoms. Genetic analysis was confirmed segregation of homozygous mutation in 3splice site of exon 15 in IL-12R1. Expression of gene was completely abolished in PBMCs of patient and the surface expression of IL12RB1 was not detectable in T cell derived PBMCs of the patient compared to healthy control. Furthermore, did not response to IL12 stimulation since we could not detect increase of INF- after stimulation with Il12 and BCG.

    Our patient received BCG vaccination at birth and had BCG lymphadenitis as an infant, CD and mycobacterial multifocal osteomyelitis as a child. Furthermore there are some evidences which indicate the role of atypical mycobacterial infections as a trigger for CD.

    Conclusion: We reported for the first time contemporary MSMD and IBD in 26 years old patient, who had impaired IL-12 signaling and abolished IL12 R1 expression in PBMCs and T cell blast. However, mycobacterial osteomyelitis is a typical phenotype of MSMD patients with deficiency in IFN-R1 or STAT, there were no mycobacterial osteomyelitis reported in IL-12R1 deficient patients.

    (103) Submission ID#601130

    NBAS Compound Heterozygous Variants as a Cause of Recurrent Acute Liver Failure Triggered by Common Childhood Infections

    Suthida Kankirawatana, MD1, Anna Hurst, MD2, Janaina Nogueira, MD3, Julie Jones, PhD4, Prescott Atkinson, MD, PhD5

    1Assistant Professor, Division of Allergy and Immunology, Department of Pediatrics, Children's of Alabama, UAB

    2Assistant Professor, Division of Genetics, Department of Pediatrics, Children's of Alabama, UAB

    3Associate Professor, Division of Gastroenterology, Department of Pediatrics, Children's of Alabama, UAB

    4 Director, Clinical Genomic Sequencing Program, Greenwood Genetic Center

    5Professor, Division of Allergy, Immunology, Department of Pediatrics, Children's of Alabama, UAB

    Background: Advanced genetic studies help explain the occurrence of many undiagnosed, rare conditions. Recently, NBAS variants were identified as a causative basis of recurrent liver failure in infants (Infantile Liver Failure Syndrome 2, ILFS2). The NBAS (Neuroblastoma Amplified Sequence) gene encodes a protein involved in Golgi to ER retrograde transport. NBAS functions seem to be broad and loss of function variants in NBAS have been associated with multisystem manifestations.

    Case report: A 5y 9m old Chilean male presented to the ER with a three day history of vomiting, diarrhea and one day of fever (38.3° F). On examination he was pale, lethargic, and tachycardic. A chemistry profile revealed markedly elevated liver enzymes, increased bilirubin, and coagulopathy, consistent with the acute hepatic failure (ALT 6291, AST >4000, total bilirubin 3.49 (2.82 DB), GGT 52, and INR of 2.1). He was hospitalized, given Vitamin K, and kept on intravenous fluids, ursodiol, and anti-pyretics. His liver function improved significantly within 5 days of admission (ALT was down to 980, AST 45, total bilirubin 1.62). Work-up of possible etiologies including autoimmunity and infectious hepatitis was negative. Liver sonogram was normal, but liver biopsy was consistent with acute hepatitis with some necrosis. Urine organic acid and plasma amino acid screens were not consistent with any inherited metabolic disorders. His parents recalled two previous episodes of liver failure at ages 3 and 4 years. Both were preceded with a mild febrile illness and non-specific symptoms including fever, coughing, vomiting, diarrhea, lethargy, and decreased PO intake. These subsequently were followed by jaundice and marked elevation of liver enzymes. Flu A and adenovirus were identified as causes of febrile illnesses of the two previous episodes. For this admission, adenovirus was found in the respiratory secretions and a mild EBV viremia was also detected. Genetic evaluation in Chile was reportedly normal. After a literature review we obtained sequencing of NBAS which revealed two variants: c.2827G>T,p.Glu943* and NBAS c.2951T>G, p.IIe984Ser. Both variants have been previously reported in patients with an infantile onset, recurrent liver failure syndrome. His other clinical features include developmental and speech delays, failure to thrive, and facial dysmorphism. He also has a history of recurrent ear infections and has had 3 sets of tympanostomy tubes. Further testing was limited due to the lack of insurance coverage.

    Conclusion: NBAS deficiency is a newly described syndrome of recurrent acute liver failure that occurs early in life. Once individuals have survived to adulthood they do not seem to develop liver failure with illness. Typically, liver crisis is triggered by a common childhood febrile illness. The mechanism of disease is thought to be thermal instability of hepatocytes which improves over time in most cases. However, although spontaneous recovery can occur following the crises, each episode can be fatal or result in permanent liver damage required liver transplantation. Increased awareness of this disease will lead to the early establishment of the diagnosis. Appropriate and timely management of fever at the onset of illness can significantly improve outcome in this potentially fatal disease.

    (104) Submission ID#601139

    Molecular Study in Children with Chronic Granulomatous Disease (CGD) at a Tertiary Care Center in North India

    Dharmagat Bhattarai, MD, DM Fellow1, Pandiarajan Vignesh, MD;DM (pediatric clinical immunology and rheumatology)2, Madhubala Sharma, PHD scholar3, Jitendra Shandilya, PHD scholar3, Amit Rawat, MD, PDCC4, Kohsuke Imai, MD, PHD5, Shigeaki Nonoyama, MD, PHD6, Osamu Ohara, PHD7, Yu-lung Lau, MBChB, MD (Hon), FRCPCH, FRCPS8, Surjit Singh, MD; DCH (Lon.); FRCP (Lon.); FRCPCH (Lon.); FAMS9

    1DM Fellow, Postgraduate Institute of Medical Education and Research, Chandigarh

    2Assisstant professor, department of pediatrics, Postgraduate Institute of Medical Education & Research

    3PHD Scholar, Postgraduate Institute of Medical Education and Research, Chandigarh

    4Professor, Postgraduate Institute of Medical Education and Research, Chandigarh

    5Faculty, Tokyo Medical and Dental University

    6Faculty, Tokyo Medical and Dental University

    7Immunologist, Kazusa DNA Research Institute

    8Chair Professor of Paediatrics Doris Zimmern Professor in Community Child Health, Department of Paediatrics and Adolescent Medicine, The University of Hong Kong

    9Head, Department of Pediatrics and Chief, Allergy Immunology Unit, Advanced Pediatrics Centre,Principal Investigator, Indian Council of Medical Research (ICMR) Centre for Advanced Research in Primary Immunodeficiency DiseasesVice-President, Indian Rh, Postgraduate Institute of Medical Education & Research

    Background: Chronic granulomatous disease (CGD) results from an inherited, genetically heterogeneous functional defect of phagocytes. CGD results from defects in different components of NADPH oxidase enzyme complex. Mutations in the seven structural genes of this complex (CYBB, CYBA, NCF1, NCF2, NCF4 and Rac1/Rac2 GTPase binding protein gene) cause CGD.

    Methods: All children diagnosed to have CGD between 1998 and 2018 on the basis of abnormal result of nitroblue tetrazolium dye reduction test (NBT) and Dihydrorhodamine 123 (DHR) assay were enrolled in the study. Clinical findings, diagnostic tests and outcomes were recorded from scrutiny of case notes. Assessment of carrier state of mother and level of gp91phox, p47phox, p67phox , p22phox by flow cytometry were done as guiding tools for possible genetic defects. Mutation analysis was done by gene scan, Sanger sequencing and next-generation sequencing.

    Results: Among 62 patients with CGD, 51 (40 boys; 11 girls) had been tested proven for mutation. Male female ratio was 3.63:1. Twenty five (49.1%) patients had X-linked CGD and 26 (50.9%) had autosomal recessive (AR) forms of CGD. Mean age at initial presentation was 2 years (range 15 days - 10 years), while mean age of diagnosis was 3.5 years (range 15 days - 20 years). Forty nine percent (25 among 51) children were diagnosed to have CYBB gene mutation. Sixteen patients (31%) had NCF1 gene mutation. NCF2 gene defect was detected in 10 patients (19.6%). There was history of consanguineous marriage in 15% of the patients.

    All children were receiving cotrimoxazole and itraconazole prophylaxis after being diagnosed with CGD. Thirty three (64.7%) patients recovered completely from their infections. Two patients needed rib resection due to locally invasive Aspergillus pneumonia while two patients lost follow up. Sixteen (32%) children succumbed to their illness despite therapy.

    Conclusion: A high proportion of heterogeneity was detected in our cohort of CGD. Assessment of mothers carrier status and flow cytometric evaluation of membrane bound and cytosolic protein component of NADPH oxidase complex can be assessed to get help for determining possible mutation. Genetic testing has diagnostic and prognostic importance for the children with CGD.

    References:

    1. Gennery A. Recent advances in understanding and treating chronic granulomatous disease. F1000Res. 2017 Aug 11;6:1427.

    2. Wolach B, Gavrieli R, de Boer M, van Leeuwen K, Berger-Achituv S, Stauber T, et al. Chronic granulomatous disease: clinical, functional, molecular, and genetic studies. The Israeli experience with 84 patients: research article. Am J Hematol. 2017;92(1):28–36.

    (105) Submission ID#601150

    Cytomegalovirus Specific Cell-mediated Immunity Status in Women with Pre-eclampsia: A Case-control Study

    Roya Sherkat, MD1, Zahra Shahshahan, MD2, Sahar Memar Montazerin, MD3, Maryam Kalateh Jari, MD4, Majid Yaran, PhD5, Maryam Nasirian, PhD6, Somaye Najafi, MSc3

    1Associate Prof. ,Head of Acquired Immunodeficiency Research Center, Acquired Immunodeficiency Research Center, Isfahan University of Medical Sciences , Isfahan , Iran

    2Associate prof., Obstetrics and Gynecology department, Isfahan University of Medical Sciences, Isfahan, Iran.

    3Research Assistant, Acquired Immunodeficiency Research Center, Isfahan University of Medical Sciences , Isfahan , Iran

    4Obstetrics and Gynecology Assistant, Obstetrics and Gynecology department, Isfahan University of Medical Sciences, Isfahan, Iran.

    5Scientist, Acquired Immunodeficiency Research Center, Isfahan University of Medical Sciences , Isfahan , Iran

    6Associate prof., Infectious Diseases and Tropical medicine research center, Isfahan University of Medical Sciences, Isfahan, Iran

    Background: Pre-eclampsia, a pregnancy-specific complication, has been shown to be associated with Cytomegalovirus (CMV) infection. CMV specific T-cell response plays the major role in CMV infection or disease .We explored whether a change in CMV-specific cell-mediated immunity (CMI) Is related to the development of pre-eclampsia.

    Method: CMV-specific CMI was assessed using CMV-QuantiFERON (QF-CMV) assay in serum from 35 women with pre-eclampsia as well as 35 normal pregnancy controls retrospectively. Participants were matched for gestational age individually. Proportion of reactive results, mean value of Interferon- level produced in mitogen and antigen tubes were compared between the cases and controls via Chi-Square, Wilcoxon rank-sum tests, respectively. Odds ratio (OR) and confidence interval (CI) were calculated as well.

    Result: No significant differences observed between demographic characteristics of the case and control groups. The QF-CMV assay turned reactive (QF-CMV [+]) in 22 of 35 of patients (63%) VS. 32 of 35 controls (91.4%) (P = 0.004). Women with pre-eclampsia had lower mean IFN- levels in antigen tube (1.57 ± 1.79) compared with normal pregnancy controls (2.40 ± 2.21) (P = 0.028). There was no statistically significant differences in this value of mitogen tube between cases (3.53 ± 1.67) and controls (3.53 ± 1.67) (P = 0.209). Women with suppressed CMV-CMI were 6.3 times more likely to manifest pre-eclampsia (OR= 6.30, 95% CI: 1.60-24.7). This result even strengthened after adjustment for age, gestational age and gravidity (OR = 12.86, 95% CI: 2.68-61.6).

    Conclusion: Our finding support an association between suppressed CMV specific CMI and pre-eclampsia.

    (106) Submission ID#601152

    Auto-inflammation and Immunodeficiency 2 Genes One Presentation

    Amarilla B. Mandola, MD1, Chaim M. Roifman, MD2

    1Fellow in Training, Canadian Centre for Primary Immunodeficiency, The Roifman Laboratory, Research Institute Division of Immunology and Allergy, Department of Paediatrics, The Hospital for Sick Children, University of Toronto

    2Head of Canadian Centre for Primary Immunodeficiency, Head of the Roifman Laboratory, Canadian Centre for Primary Immunodeficiency, The Roifman Laboratory, Research Institute Division of Immunology and Allergy, Department of Paediatrics, The Hospital for Sick Children, University of Toronto

    Introduction: The triad of susceptibility to infections, auto-inflammation, and cancer in a patients personal and family history are always suggestive of an underlying primary immunodeficiency; however, in some cases the diagnosis might be delayed for years. Furthermore, the results of immunological and inflammatory evaluation can also be affected by ongoing immunomodulatory therapy initiated by different specialists upon clinical diagnosis.

    Objective: To describe a unique presentation of auto-inflammatory disease with combined immunodeficiency in an adult patient.

    Case presentation: We report here the case of a 64 year old male, who had a long history of infections including recurrent sino-pulmonary bacterial infections starting during childhood, osteomyelitis at 7 years of age, recurrent tonsillitis requiring tonsillectomy at 21 years of age, recurrent cellulitis, an episode of prostatitis with septicaemia, as well as recurrent varicella zoster and warts. The patient was also diagnosed with sclerosing mesentheritis, and Reynauds phenomenon, recurrent oral ulcers, arthritis, uveitis, autoimmune thyroiditis, lung fibrosis and suffered repeated episodes of abdominal pain. Furthermore, there is a family history of early childhood death, multiple soft tissue cancers, Crohns disease, and autoimmune thyroiditis.

    Upon physical examination, the patient had multiple telangiectasia, baseline erythroderma, and flushing. Immunological evaluation showed lymphopenia with significant reduction in both circulating B and T cells, however, assessment of humoral immunity revealed low IgG and decreased IgM with normal IgA levels. At the time of the evaluation he had been on low dose daily prednisone (7.5mg), colchicine, and methotrexate as immuno-modifying therapy.

    Genetic evaluation revealed a heterozygous mutation in NOD2 as well as compound heterozygous mutations in the MEFV gene.

    Discussion: Mutations in NOD2 have been described in association with Blau syndrome a multisystem auto-inflammatory syndrome which may explain many of the features experienced by our patient. To our surprise next generation sequencing revealed a second aberration in the MEFV gene which causes Familiar Mediterranean Fever, another multisystem auto-inflammatory disease, which might lead to the phenotype observed in the patient.

    Conclusion: This is the first report of genetic lesions in two different genes leading to a severe course of auto inflammation.

    (107) Submission ID#601164

    Novel CDC42 Mutation Causes Severe Autoinflammatory Syndrome Responsive to IL-1 Inhibition

    Yael Gernez, MD, PhD1, Matthew Kirbey, PhD2, Jay M. Balagas, MD3, Claudia I. Macaubas, PhD4, Scott Canna, MD, PhD5, David B. Lewis, MD6, Elizabeth Mellins, MD, PhD7, Rosa Bacchetta, MD8, Katja G. Weinacht, MD, PhD9

    1Clinical Assistant Professor, Stanford School of Medicine

    2Scientist, Department of Pediatrics, Stem Cell Transplantation, at the Lucile Salter Packard Childrens Hospital, Stanford school of medicine, Stanford, CA, USA

    3Physician, Pediatric Hematology/Oncology, 2018 John Muir Health Physician Network Member

    4Basic Life Res Scientist, Department of Pediatrics Rheumatology/RK Mellon Institute, Children's Hospital of Pittsburgh of UPMC, Pittsburgh, Pa, USA

    5Assistant Professor, Department of Pediatrics Rheumatology/RK Mellon Institute, Children's Hospital of Pittsburgh of UPMC, Pittsburgh, Pa, USA

    6Professor, Department of Pediatrics, Allergy, Immunology and Rheumatology at the Lucile Salter Packard Childrens Hospital, Stanford School of Medicine

    7MD, PhD, Department of Pediatrics, Human Gene Therapy at the Lucile Salter Packard Childrens Hospital, Stanford school of medicine, Stanford, CA, USA

    8ASSOCIATE PROFESSOR, Department of Pediatrics, Stem Cell Transplantation, at the Lucile Salter Packard Childrens Hospital, Stanford school of medicine, Stanford, CA, USA

    9Assistant Professor, Division of Stem Cell Transplantation and Regenerative Medicine, Department of Pediatrics, Stanford School of Medicine, Stanford, CA

    Monogenic autoinflammatory syndromes (MAIS) are a diverse group of disorders characterized by primary over-activation of the innate immune system. Induction of the inflammasome complex by innate immune sensors and increased production of IL-1b are implicated in the pathogenesis of MAIS. Macrophage activation syndrome (MAS) is a life-threatening illness defined by acute hyper-inflammation and unopposed cytokine release. It is considered an acquired condition secondary to infection, rheumatoid disease or malignancy. The early therapeutic use of IL-1b inhibition has profoundly improved the prognosis MAS. It has recently been shown that increased free IL-18 levels in the blood are causatively linked to the development of MAS. Significant overlap in clinical presentation and laboratory markers between patients with MAIS and MAS led us to explore the role of free IL-18 and therapeutic use of IL-1b inhibition in a patient with CDC42 mutation.

    Here, we report the case of an 18 months-old female who presented with hydrops fetalis in utero, and later developed failure-to-thrive, splenomegaly, anemia, thrombocytopenia, arthralgias, rashes, frequent febrile episodes and mild facial dysmorphism along with massive increase in CRP, ESR and ferritin. Whole Exome Sequencing (WES) identified a heterogenous likely pathogenic de novo variant in cell division control protein 42 homolog (CDC42) c.563G>A (p.C188Y). CDC42 encodes a small RHO family GTPase that regulates multiple signaling pathways controlling cell polarity, migration, endocytosis and cell cycle progression. Single allele mutations in the CDC42 gene were recently reported to cause Takenouchi-Kosaki syndrome manifesting with growth retardation, developmental delay, facial dysmorphism, and thrombocytopenia however systemic autoinflammation has not been described. CDC42 closely interacts with the Wiskott-Aldrich Syndrome Protein but little is known about the mechanism underlying immune abnormalities associated with CDC42 mutations.

    Our patient had an inflammamosopathy-like syndrome. Because of significant clinical overlap to MAS, we measured IL-6, IL-18, free IL-18 and IL-18 binding protein, all of which were significantly increased. This increase in free IL-18 heightened her risk of developing MAS. Her IL1-b level was normal, but an increase in IL-1b is hardly ever detectable in the serum despite playing a critical role in this type of inflammation. Indeed, chronic IL-1b excess in the tissues promotes systemic inflammation and is associated with chronically elevated CRP and ESR. With this rationale we started the IL-1 receptor antagonist anakinra. Within 48 hours from starting anakinra, the parents observed an increase in appetite, resolution of arthralgias and improved mobility. Over the course of the following weeks, fever, anemia, thrombocytopenia and rash disappeared, the spleen massively decreased in size and the patient started to meet developmental milestones. CRP, ESR eventually normalized while ferritin and free IL-18 are still trending down.

    Conclusions: Significant increase in free IL-18 and extremely encouraging clinical response to therapy with anakinra in a patient with novel CDC42 mutation suggests a link between MAS and defects in CDC42. Elucidating the mechanism of inflammasome activation and the drivers of IL-18 increase in MAS and MAIS more broadly may shed light on novel therapeutic targets like the use of human recombinant IL-18 binding protein.

    (108) Submission ID#601190

    First Three Years Experience in the Immunology Outpatient Clinic of a University Hospital in Cali,Colombia

    Andres F. Zea-Vera, MD, PhD1, Vanessa Montoya-Lozano2, Mario A. Chacon-Acevedo, MD3

    1Assistant Professor, Universidad del Valle. Hospital Universitario del Valle.

    2Nursing Student, Universidad del Valle

    3MSc Student, Universidad del Valle. Hospital Universitario del Valle.

    Introduction: In August 2015 the Clinical Immunology outpatient clinic was established in the Hospital Universitario del Valle (HUV) in Cali, Colombia. The clinic evaluate an average of 8 to 10 patients weekly (new and follow up patients). Most of the cases are referred in the context of Recurrent Infection syndrome, disseminated mycobacterial disease, hypogammaglobulinemia and severe autoimmune or allergic disease.

    Results: The Clinical Immunology outpatient clinic has evaluated 261 patients classified as: Primary Immunodeficiencies (PID)= 64 patients (25%), Secondary Immunodeficiencies= 14 patients (5%), Autoimmunity/Rheumatic disease= 49 patients (19%), Severe/Refractory Allergy= 61 patients (23%) and Infectious diseases with high suspicious of PID in follow up= 24 patients (9%).

    According to the IUIS-2017 classification, 64 patients with confirm Inborn Errors of Immunity (PID) were diagnosed: I. Immunodeficiencies affecting cellular and humoral immunity 4(6%), II. CID with associated or syndromic features 10(16%), III. Predominantly Antibody deficiencies 26(41%), V. Congenital defects of phagocyte number, function or both 7(11%), VI. Defects in intrinsic and innate immunity 6(9%), VII. Auto-inflammatory disorders 7(11%), VIII. Complement deficiencies 2(3%), IX. Phenocopies of PID 2(3%).The mean age was 16 years with a Male:Female ratio 34:30. Molecular tests have been done in 14 cases with 8 gene mutation confirmation.

    Conclusion: Our Clinical Immunology service constitutes an opportunity for low income people with public health care insurance in the southwest of Colombia. The Universidad del Valle and Hospital Universitario del Valle combine effort has contribute to improve the suspicious, diagnosis and treatment of patients living with Inborn Errors of Immunityr (PID) as well as patients with other immune disorders.

    (109) Submission ID#601193

    Telomeres in Schimke Immuno-Osseous Dysplasia: Comparing Telomere Length in Individuals with Homozygous and Heterozygous SMARCAL1 Mutations

    Elizabeth A. Lippner, MD1, Geraldine Aubert, PhD2, Vasavi Ramachandran, MS3, David B. Lewis, MD4

    1Postdoctoral Scholar and Clinical Instructor, Division of Allergy, Immunology, and Rheumatology, Department of Pediatrics, Stanford University School of Medicine

    2Director of Clinical Development & Research, Repeat Diagnostics, Inc

    3Life Science Research Professional, Division of Allergy, Immunology, and Rheumatology, Department of Pediatrics, Stanford University School of Medicine

    4Professor, Department of Pediatrics, Allergy, Immunology and Rheumatology at the Lucile Salter Packard Childrens Hospital, Stanford School of Medicine

    BACKGROUND: Schimke Immuno-Osseous Dysplasia (SIOD) is a rare, autosomal recessive disease characterized by spondyloepiphyseal dysplasia, vasculopathy, T-cell immunodeficiency, progressive nephrotic disease, and increased neoplastic risk. SIOD is caused by homozygous mutations in the SMARCAL1 gene. SMARCAL1 encodes a DNA-annealing helicase that functions in gene expression modulation and maintaining genome integrity at stalled DNA replication forks. Recent in vitro studies implicate a role for SMARCAL1 in telomere maintenance; SMARCAL1 is enriched in cells that maintain telomeres via the alternative lengthening of telomeres pathway and SMARCAL1-decifient cells demonstrate telomere instability with replication fork collapse and increased telomere-associated DNA damage.[1,2] Telomere analysis of 4 SIOD patients, including one patient who received a hematopoietic stem cell transplant (HSCT) 20 years prior, as well as 5 heterozygous family members revealed significantly shorter telomeres in SIOD patients compared to heterozygous family members and compared to age-matched, healthy controls.

    METHODS: Peripheral blood mononuclear cells were isolated using a Ficoll-Hypaque density gradient, cryopreserved, then sent to Repeat Diagnostics in North Vancouver, BC. Telomere length measurements were performed at a single-cell level using flow-fluorescence in situ hybridization as previously described.[3] Telomere length was measured in total lymphocytes, naive and memory enriched T cells, B cells, and NK cells and compared to reference samples from age-matched, healthy individuals.

    RESULTS: Compared to age-matched healthy controls, three SIOD individuals had mean telomere lengths (MTLs) less than the 1st percentile for age across all lymphocyte subsets (total lymphocytes, B cells, NK cells, naïve and memory T cells). In comparison, three unaffected family members had normal MTLs (10th percentile< x <90th percentile) across all subsets, and two unaffected family members had low MTLs (1st< x <10th percentile) in some subsets. The SIOD individual who received a matched-sibling HSCT 20 years prior, had normal MTL in NK cells (10th < x <90th percentile) but low MTLs (1st< x <10th percentile) for all other subsets.

    CONCLUSIONS: These data show that SIOD patients have significantly impaired telomere lengths across multiple lymphocyte lineages and support a limiting role for SMARCAL1 deficiency in telomere maintenance. In comparison, unaffected family members, heterozygous for SMARCAL1 mutations, have mean telomere lengths that are normal or slightly low for age. This suggests that abnormally short telomeres are seen in individuals with homozygous but not heterozygous SMARCAL1 mutations. For the individual who received a HSCT, we do not have pre and post-HSCT telomere data, but these results support obtaining pre and post-HSCT telomere length analysis in future cases.

    Abnormally short telomeres have been linked to widespread perturbation of gene expression.[4] We hypothesize that SMARCAL1 deficiency, by the effect of stalled forks and shortened telomeres, leads to perturbation in the transcriptome of affected tissues. Shortened telomeres may explain the reduced hematopoietic bone marrow production in SIOD, as bone marrow failure is a cardinal feature of dyskeratosis congenita, a disorder of impaired telomere maintenance. Future studies to investigate the role of telomere maintenance in SIOD include measurement of telomerase activity in polyclonally activated T cells and transcriptome analysis using RNA-Seq.

    REFERENCES:

    1. Cox KE, Maréchal A, Flynn RL. SMARCAL1 resolves replication stress at ALT telomeres. Cell Reports. 2016; 14:1032-1040.

    2. Poole, LA et al. SMARCAL1 maintains telomere integrity during DNA replication. Proc Natl Acad Sci U S A. 2015; 112:14864-14869.

    3. Baerlocher GM, Vulto I, de Jong G, Lansdorp PM. Flow Cytometry and FISH to measure the average length of telomeres (flow FISH). Nat Protocols. 2006; 1:2365-2376.

    4. Robin JD, Ludlow AT, et al. SORBS2 transcription is activated by telomere position effect-over long distance upon telomere shortening in muscle cells from patients with facioscapulohumeral dystrophy. Genome Res. 2015; 25:1781-1790.

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    (110) Submission ID#601200

    Yellow Fever: Is It Possible to Vaccinate Patients with IgA Deficiency?

    Magda Carneiro-Sampaio, MD, PhD1, Mariana Castiglioni, MD2, Nathalia Souza, MD2, Bruna Aquilante, MD3, Mayra Dorna, MD3, Ana Paula Castro, MD3, Antonio Carlos Pastorino, MD, PhD4,

    1Full Professor, Department of Pediatrics, Faculdade de Medicina da Universidade de Sao Paulo, Sao Paulo, Brazil

    2Allergy Immunology Fellow, Allergy Immunology Department from Child´s Institute from Medicine Faculty from Uiversity of Sao Paulo (Instituto da Criança do Hospital das Clinicas da Faculdade de Medicina da Universidade de São Paulo)

    3Allergy Immunologist, Allergy Immunology Department from Child´s Institute from Medicine Faculty from Uiversity of Sao Paulo (Instituto da Criança do Hospital das Clinicas da Faculdade de Medicina da Universidade de São Paulo)

    4Allergy Immunologist Professor, Allergy Immunology Department from Child´s Institute from Medicine Faculty from Uiversity of Sao Paulo (Instituto da Criança do Hospital das Clinicas da Faculdade de Medicina da Universidade de São Paulo)

    Background: Yellow fever is a potentially fatal disease for which only supportive treatment is available. Vaccination is the primary strategy for prevention of this disease And the vaccine is extremely effective, but there are a few specific populations where it is contraindicated. Regarding IgA deficiency (the most frequent primary immunodeficiency), current recommendations in the literature are controversial. There are no specific studies in this disease, so case series addressing the safety or possible adverse events after vaccination are essential for decision-making during epidemic scenarios, as experienced in Brazil in the last years. In this context, this study aimed to describe adverse events after the use of the yellow fever vaccine in IgA deficient patients.

    Method: a retrospective cross-sectional study was conducted including IgA deficient patients followed at a specialized pediatric outpatient clinic between 2017 and 2018. All patients had at least one year of follow-up. Immunoglobulin levels, antibody response to vaccines and lymphocyte subset count were evaluated to exclude other immunodeficiencies or the presence of abnormalities that could contraindicate vaccination. Demographic data, the presence of infections and comorbidities, use of immunosuppressive medication and adverse events after vaccine administration of the vaccine were described.

    Results: Thirty-eight patients with IgA deficiency were included in the study and 18 received the vaccine. Vaccinated patients had a mean age at the time of the study of 13.7 years (SD ± 3.5y). Six out of the 18 presented comorbidities: thyroiditis (n=3), type 1 diabetes mellitus (n=1), celiac disease (n=1) and juvenile rheumatoid arthritis (n=1). All patients were atopic and only one had recurrent infections in the last year despite the use of antibiotic prophylaxis. All 18 patients had normal IgG and IgM levels for their age, positive vaccine responses for measles, rubella and mumps, and age-appropriate lymphocyte subset count. After 6 months of observation, no immediate or late adverse events were reported. Among the 20 non-vaccinated patients, only one had a formal contraindication (systemic erythematosus lupus using immunosuppressive therapy). Five out of the 20 non-vaccinated patients reported being afraid of receiving the vaccine, 7 still intended to receive it and for other 7 patients data regarding vaccination was unavailable.

    Conclusion: Despite the small number of patients, the absence of adverse events in this case series suggests that immunization with yellow fever vaccine may be safe in IgA deficient patients, excluded other contraindications. More studies are essential to confirm the safety and help the decision-making process regarding the vaccine administration for IgA deficient patients, especially in this yellow fever outbreak scenario.

    (111) Submission ID#601201

    A Case of Immunodeficiency, Centromeric Instability, and Facial Anomalies Syndrome (ICF) with NK Deficiency and Subsequent EBV-driven Diffuse Large B-cell Lymphoma Treated with Bone Marrow Transplant

    Caitlin M. Burk, MD1, Kara E. Coffey, MD2, Emily M. Mace, PhD3, Bret Bostwick, MD4, Ivan K. Chinn, MD5, Zeynep H. Coban-Akdemir, PhD6, Shalini N. Jhangiani, PhD7, James R. Lupski, MD, PhD8, Damara Ortiz, MD9, Jessie L. Barnum, MD10, Steven W. Allen, MD10, Leanna-Marie Robertson, MD2, Jordan S. Orange, MD, PhD11, Hey J. Chong, MD, PhD12

    1Pediatric Resident, UPMC Children's Hospital of Pittsburgh, Pittsburgh, PA

    2Fellow, Allergy and Immunology, University of Pittsburgh School of Medicine, Pittsburgh, PA

    3Assistant Professor, Department of Pediatrics, Columbia University Irving Medical Center, New York, NY

    4Assistant Professor, Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX

    5Assistant Professor, Pediatric Allergy and Immunology, Baylor College of Medicine, Houston, TX

    6Postdoctoral Research Fellow, Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX

    7Project Manager, Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX

    8Professor, Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas, USA

    9Assistant Professor, Pediatric Medical Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA

    10Assistant Professor, Pediatric Hematology and Oncology, University of Pittsburgh School of Medicine, Pittsburgh, PA

    11Professor and Chair, Department of Pediatrics, Columbia University Irving Medical Center, New York, NY

    12Assistant Professor and Chief, Pediatric Allergy and Immunology, University of Pittsburgh School of Medicine, Pittsburgh, PA

    Introduction/Backround: Immunodeficiency, centromeric instability, and facial anomalies syndrome (ICF) is a rare group of autosomal recessive disorders involving the triad of hypogammaglobulinemia, centromeric instability, and facial anomalies. The majority of patients have hypo- or agammaglobulinemia, but T cell defects have also been reported. We present the case of a child with ICF-2 who presented with NK deficiency and ultimately developed an EBV-driven malignancy and was successfully treated with bone marrow transplant.

    Methods: Whole exome sequencing and NK cell function via 51-Cr cytotoxicity assay and phenotyping via flow cytometry were performed at Baylor College of Medicine and Texas Childrens Hospital. Centromeric banding studies were performed at University of Pittsburgh Medical Center.

    Results: The female patient presented at 3 months of age with CMV pneumonitis and persistent CMV viremia requiring treatment followed by prophylaxis with valgancyclovir. She initially had hypogammaglobulinemia and low T, B, and NK cells; she had normal TRECs, lymphocyte mitogen proliferation responses and Zap 70, MHCI and MHCII expression. The hypogammaglobulinemia and T- and B-cell lymphopenia resolved within 9 months after initial presentation as she clinically improved from her CMV infection. She was found to have NK cell deficiency on three separate commercially tested samples. Whole exome sequencing revealed a homozygous variant in ZBTB24 indicative of ICF-2 syndrome that was confirmed with Sanger sequencing (c.1492_1493del, p.Q498Vfs). Repeat NK cell studies confirmed impaired function, and phenotyping showed an increase in CD56-bright and a decrease in CD16-positive cells, suggesting either impaired transition from immature to mature NK cells or impaired survival of mature cells. Her karyotype and centromeric banding studies were normal, as were centromeric instability studies.

    She later developed a memory B-cell defect and presented at 34 months of age with persistent fever, respiratory distress, loss of vaccine titers, hypogammaglobulinemia and low B and T cells. She was found to have EBV viremia and an EBER-positive diffuse large B-cell lymphoma in her right lung. Due to tenuous clinical status, she received rituximab for treatment of EBV prior to definitive lymphoma diagnosis. She was treated with chemotherapy per protocol ANHL1131, group B (pre-phase with COP, courses 1 and 2 with COPADM, and courses 3 and 4 with CYM) and her course was complicated by seizures attributed to methotrexate toxicity. She ultimately underwent reduced intensity conditioning with hydroxyurea, alemtuzumab, fludarabine, mephalan, and thiotepa followed by a CD-34 selected, HLA-matched, unrelated donor peripheral blood stem cell transplant. Her early post-transplant course was complicated by adeno- , EBV, and CMV viremia, all successfully treated with antivirals and a donor lymphocyte infusion. She is now greater than 8 months post-transplant, off immunosuppression with 100% donor engraftment, no evidence of organ toxicity or GVHD, and with excellent immune reconstitution.

    Conclusions: This is the first reported case of impaired NK cell function and phenotype and EBV-driven malignancy in a patient with ICF-2. This case expands the phenotype of ICF-2 and suggests that early bone marrow transplant should be considered in these children. It also demonstrates a novel requirement for ZBTB24 in human NK cell maturation and function.

    (112) Submission ID#601233

    Variant Mutation in PLCG2 Associated with Common Variable Immundeficiency Without Cold Urticaria

    Veronica Solivan-Vargas, MD1, Wilfredo Cosme-Blanco, MD/Phd2, Cristina Ramos-Romey, MD3, Sylvette Nazario-Jimenez, MD4

    1Resident, Hospital Episcopal San Lucas

    2Allergy & Immunology Physician, Veteran Affairs Caribbean Healthcare System

    3Assistant Director of Allergy Immunology Program, University of Puerto Rico

    4Director of Allergy Immunology Program, University of Puerto Rico

    Rationale: Common variable immunodeficiency (CVID) is a disorder that affects the production of immunoglobulins and is associated with development of autoimmunity. Multiple mutations have been described that are associated with CVID, but PLCG2 mutations have only been described in patients with phospholipase C gamma 2 (PLC2) associated antibody deficiency and immune dysregulation (PLAID) and autoinflammatory PLC2 associated antibody deficiency and immune dysregulation (APLAID). We present a case of a 44 y/o male CVID patient with recurrent upper respiratory tract infections, steroid-dependent autoimmune thrombocytopenia, low B cell count, hepatosplenomegaly, and restrictive lung disease. He was found with a variant of unknown significance at the PLCG2 gene. In contrast to PLAID our patient does not exhibit cold urticaria.

    Method: Case presentation of a CVID patient followed in our clinics. Patients chart and previous laboratories were reviewed. Sequence analysis and deletion/duplication CVID panel testing was performed using Invitae©

    Discussion: Genetic testing has revolutionized the diagnosis of immune deficiencies, but variants of unknown significance are being increasingly reported. In this case, a variant of uncertain significance was identified which replaces threonine for alanine at codon 829 of the PLCG2 protein. This codon is located at the SH3 domain, which is part of a region that provides auto-inhibitory enzymatic functions. PLAID mutations have been identified in SH2 domain, but it has been known that both SH2 and SH3 domains facilitate PLCG association with other proteins. Studies with deletion of PLCG2 gene have shown functional abnormalities in B cells, natural killer cells and mast cells. To our knowledge, there has not been any previous report of a CVID patient with a variant mutation at the SH3 domain of the PLCG2 gene without being diagnosed as PLAID or APLAID. Our patient has immunodeficiency, recurrent upper respiratory tract infections, steroid-dependent recurrent autoimmune thrombocytopenia, rheumatoid arthritis, hepatosplenomegaly, early-osteoporosis and restrictive lung disease. He does not have cold urticaria as seen in PLAID, but exhibits autoimmunity not observed in APLAID.

    Conclusion: Conclusion: PLCG2 is an important protein in the pathway of B cell development. A novel mutation in the SH3 domain of the PLCG2 gene may be associated with the CVID phenotype of low B cells and autoimmunity. This could lead to a gain-of-function mutation as seen in PLAID but without early-onset cold urticaria. Functional studies are required to confirm the significance of this mutation.

    (113) Submission ID#601257

    JAK-dependent and Independent Cytokines Drive the Pathogenicity of HLH : Targets for Combination Therapy

    Josée-Anne Joly1, Sara Bourbonnais, MSc2, Alexis Vallée3, Chloé Berthe, MSc2, Hélène Decaluwe, MD, PhD, FRCPC4

    1Master student, CHU Sainte-Justine Research Center, Université de Montréal

    2Research Assistant, CHU Sainte-Justine Research Center, Université de Montréal

    3PhD student, CHU Sainte-Justine Research Center, Université de Montréal

    4Pediatric Immunologist and Clinician Scientist, CHU Sainte-Justine and Université de Montréal

    Primary (or Familial) Hemophagocytic lymphohistiocytosis (HLH) is a rare, life-threatening hyper-inflammatory disease affecting mainly young children. It is caused by mutations in genes involved in the granule-dependent cytotoxic pathway, and is characterized by extreme inflammation and massive tissue infiltration by activated T cells and macrophages. To this day, hematopoietic stem cell transplantation is the only available curative treatment with a transplant-related mortality of 30%. Thus, the development of new, more efficient anti-inflammatory treatments would be a significant advancement in the treatment of HLH. Here, we hypothesize that combination therapies targeting both JAK-dependent and independent cytokines will be more effective than either one alone to reduce the life-threatening symptoms induced by this pathology.

    Using a perforin-deficient mouse model of HLH, we first compared the effect of targeting individual cytokines with blocking antibodies on the progression of the disease. We show that blocking IFNg and IL-18, but not IL-6, significantly reduces the severity of HLH. Targeting the JAK-STAT signalling pathway with ruxolitinib, a specific inhibitor of JAK1 and JAK2, downstream of IFNg and IL-6, but not IL-18, is similarly beneficial. More importantly, combination therapies using ruxolitinib and blocking antibodies to either IFNg or IL-18 show synergistic effects, further mitigating the progression of the disease. These results suggest that JAK-dependent and independent cytokines drive the pathogenicity of HLH in perforin-deficient mice. It further supports that ruxolitinib, although effective in reducing the symptoms of HLH, should be used in combination with anti-IFNg and/or anti-IL-18 antibodies to prevent HLH progression. This is particular relevant since the former were recently approved for the treatment of HLH while the latter (IL-18 binding proteins) are in clinical trials for IL-18-dependent macrophage activation syndromes.

    This project was supported by funds from the Fondation de Cancérologie Charles Bruneau and the Canadian Institutes of Health Research.

    (114) Submission ID#601260

    A Case of Disseminated Pneumocystis Jiroveci in a Non-Human Immunodeficiency Virus Infected Patient

    Aminaa E. Siddiqi, MD1, Joshua Sacha, MD2, Rebecca Saenz, MD, Phd1, Anne Liu, MD3, Christian Kunder, MD4, Gulbu Uzel, MD5, Beth Martin, MD6, David B. Lewis, MD7, Yael Gernez-Goldhammer, MD, PhD8

    1Fellow Physician, Division of Allergy and Immunology, Department of Pediatrics, Stanford School of Medicine

    2Clinician, David Grant USAF Medical Center in Travis AFB, California

    1Fellow Physician, Division of Allergy and Immunology, Department of Pediatrics, Stanford School of Medicine

    3Clinical Assistant Professor, Division of Allergy and Immunology, Department of Pediatrics, Stanford School of Medicine

    4Clinical Assistant Professor, Department of Pathology, Stanford School of Medicine

    5Staff Clinician, Laboratory of Clinical Immunology and Microbiology, National institute of Allergy and Infectious Diseases, NIH, Bethesda, MD, USA

    6Clinical Assistant Professor, Division of Hematology, Department of Hematology, Stanford School of Medicine

    7Professor, Department of Pediatrics, Allergy, Immunology and Rheumatology at the Lucile Salter Packard Childrens Hospital, Stanford School of Medicine

    8Clinical Assistant Professor, Division of Allergy and Immunology, Department of Pediatrics, Stanford University School of Medicine

    Disseminated Pneumocystis jiroveci (PJP) infection is a well described entity in patients with acquired immunodeficiency syndrome (AIDS). Despite the increased risk of opportunistic lung infection in patients with severe T cell dysfunction (e.g. CD40L deficiency) and/or severe CD4 T cell lymphopenia, we are not aware of any reports of disseminated Pneumocystis jiroveci infection in non- human immunodeficiency virus (HIV) patients with primary immunodeficiency (PID).

    We report the first case, to our knowledge, of disseminated PJP in a patient with CVID like/CTLA4 haploinsufficiency. He had been diagnosed with common variable immunodeficiency (CVID) in 2009, approximately eight years prior to being referred to us, and was on intravenous immunoglobulin (IVIG). He was also diagnosed with multilineage Evans syndrome in 2015. His medical history was also significant for potential granulomatous lymphocytic interstitial lung disease (GLILD) (lung biopsy in the remote past with interstitial disease), significant splenomegaly (29.4 cm), severe portal hypertension, nodular liver disease (likely nodular regenerative hyperplasia) complicated by anasarca, history of chronic diarrhea (potential enteropathy), lymphadenopathy s/p biopsy with nodular lymphoid hyperplasia, and a history of multiple pneumonias. In 2017, he had developed disseminated PJP with lung, liver, and bone involvement. The T2 vertebra PJP invasion was confirmed with a bone biopsy; Gomori methenamine silver staining and PCR were performed and concluded PJP. He was treated with Trimethoprim sulfamethoxazole (TMP-SMX) and steroids, then was continued on TMP-SMX prophylaxis. Due to his liver damage and his chronic neutropenia, TMP-SMX was replaced by atovaquone as a secondary prophylaxis for PJP.

    His laboratory studies were significant for an absolute neutrophil count of 1.54 K/uL, absolute lymphocyte count of 0.61 K/uL, hemoglobin of 12.7 g/dL, platelets of 78 K/uL, total bilirubin of 2.3 mg/dL, and LDH of 243 U/L. His HIV PCR was negative. His infectious workup showed negative PCR for Epstein Barr virus, Cytomegalovirus, Herpes virus and Hepatitis B and C. His stool PCR was negative for norovirus. Flow cytometry demonstrated almost absent B cells and decreased T cells count; 586 CD3+ T cells, 12 CD20+ cells, 12 CD19+ cells, 181 CD3+CD8+ cells, 392 CD3+CD4+ cells, 15 CD56+CD16+ cells, 19 CD4+CD45RA, and 380 CD4+CD45RO cells. On IVIG, his IgA, IgG and IgM levels were respectively <8 mg/dL; 995 mg/dL, and IgM 48.4 mg/dL. T-cell receptor beta chain repertoire analysis showed an oligoclonal distribution. Severe combined immunodeficiency panel through Ambry genetic testing was negative as was genetic testing for CD40L deficiency. Given his complex clinical history, whole exome sequencing was obtained and detected an autosomal dominant heterozygous missense mutation (C.436G>A) implicated in CTLA-4 haploinsufficiency and previously reported by Schwab et al.

    Our patient is currently undergoing therapy with abatacept (CTLA-4 fusion protein), which has been reported to improve GLILD, splenomegaly and enteropathy in patients with CTLA-4 haploinsufficiency. He is improving on this regimen. He has met with the Stem Cell transplant team, but at this point of time, due to his abnormal lung function, his liver damage and his significant splenomegaly, he is not a good candidate.

    (115) Submission ID#601261

    Deficiency of the Non-classical Inhibitor of NF-kappaB, IkappaBNS, Causes a Novel Primary Immunodeficiency Due to Dysregulated NF-kappaB Signaling

    Charlotte Slade, MBBS, FRACP, FRCPA1, Maryam Rashidi, MD, PhD2, Tom Scerri, PhD2, Melanie Bahlo, PhD3, Steven Holt, MBBS, PhD, FRCP, FRACP4, Samantha Chan, MD5, James Vince, PhD6, Philip Hodgkin, PhD7, Jo A Douglass, MBBS, MD, FRACP8, Vanessa L. Bryant, BSc (Hons) PhD9

    1Researcher/Clinical Immunologist, Walter & Eliza Hall Institute/Royal Melbourne Hospital

    2Postdoctoral scientist, WEHI

    3Head, Population Health and Immunity, WEHI

    4Head, Department of Nephrology, Royal Melbourne Hospital

    5Clinical Immunologist, Royal Melbourne Hospital

    6Laboratory Head, WEHI

    7Division Head, Immunology, WEHI

    8Head, Clinical Immunology & Allergy, Royal Melbourne Hospital

    9Head, Immunogenetics Research Unit, Walter & Eliza Hall Institute

    Defects in the NF-B signaling pathway are implicated in the pathogenesis of several primary immune deficiencies in humans. The clinical features of these conditions vary significantly, reflecting the complexity of the pathway, and its broad role in innate and adaptive immune responses, and the development and differentiation of lymphoid organs.

    Here we report the first case of a human PID caused by a homozygous mutation in NFKBID in a 30 year-old male. He was the second child of consanguineous parents, and was diagnosed with possible CVID at the age of 16, after recurrent episodes of pneumococcal pneumonia. However the clinical features have evolved over time; he developed severe EBV infection at age 18, causing hepatitis and pancreatitis. At the age of 20, he presented with an ANCA-negative systemic vasculitis, manifesting as pulmonary haemorrhage, and acute necrotizing pauci-immune glomerulonephritis. Pulsed methylprednisolone and cyclophosphamide induced an initial remission, however, relapse a year later led to end-stage renal failure. He is now dialysis-dependent, and due to the underlying PID, and chronic CMV viraemia, is not a candidate for renal transplantation.

    Genomic DNA was subjected to whole-exome sequencing. Variants were filtered using a model of autosomal-recessive inheritance and functional analysis of primary cells was performed. We identified a novel, homozygous, single-base deletion resulting in a frame-shift, and premature stop in NFKBID. NFKBID encodes IBNS, a non-classical inhibitor of NF-B signaling.

    At diagnosis the patient had reduced levels of IgG2, IgA and IgM, elevated IgE, with absent humoral immune responses to pneumococcal polysaccharide vaccine, and an intact response to tetanus. Lymphocyte numbers were initially within normal reference ranges, albeit with an increased proportion of CD4+:CD8+ T cells. However, over time there has been a significant reduction in B cells and CD8+ T cells. CD4+ T cells demonstrated a skewing towards a central memory phenotype (CD45RO+/CCR7+), and CD4 T cell proliferative responses to PHA were comparable to a healthy control. Functional analysis of primary cells from the proband revealed a complete absence of BNS protein expression, dysregulated NF-B signaling, and elevated pro-inflammatory cytokine production. The patient is currently receiving a trial of targeted therapy to modulate the aberrant immune responses.

    This novel PID highlights the importance of regulation of NF-B signalling, in orchestrating an appropriate immune response, maintenance of self-tolerance, and protection against viral pathogens.

    (116) Submission ID#601278

    A Demonstration of the Diagnostic and Clinical Utility of Genomic Sequencing in Primary Immunodeficiency Diseases in Australia

    Charlotte Slade, MBBS, FRACP, FRCPA1, Fiona Moghaddas, PhD2, Sebastian Lunke, BSc, PhD, RCPA3, Zornitza Stark, MA BMBCh DM MBioeth FRACP4, Ingrid Winship, MB ChB, MD, FRACP, FACD, FAICD5, Kirsty West6, Alison Trainer7, Samar Ojaimi, MBBS (Hons) PhD, FRACP FRCPA8, Matthew Hunter, MBChB, FRACP9, Yael Prawer10, Katherine Nicholls, MBBS BMedSc FRACP FRCPA11, Mittal Patel, MBBS FRACP11, Pricilla Auyeung, MBBS PhD FRACP FRCPA11, Kymble Spriggs, MBBS, MPH, GDipClinEd, DTMH, MRCP(UK) FRACP11, Jeremy McComish, MBBS FRACP FRCPA11, Gary Unglik, MBBS(Hons) FRACP FRCPA11, Joseph De Luca11, Samantha Chan, MD11, Giulia Valente12, Anna Jarmolowicz13, Laine Hosking14, Ben van Dort15, Theresa Cole, BM MRCPCH (UK) PhD FRACP14, Joanne Smart, BSc MBBS PhD FRACP16, Sharon Choo, MBBS FRACP FRCPA14, Elly Lynch17, Clara Gaff, BSc(Hons) PhD FHGSA (genetic counselling)18, Seth Masters, BSc (Hons) PhD19, Jo A Douglass, MBBS, MD, FRACP20, Vanessa L. Bryant, BSc (Hons) PhD21

    1Researcher/Clinical Immunologist, Walter & Eliza Hall Institute/Royal Melbourne Hospital

    2Researcher/Clinical Immunologist, Royal Melbourne Hospital

    3Head of the Translational Genomics Unit, Victorian Clinical Genetics Service

    4Clinical Geneticist, Victorian Clinical Genetics Service

    5Head, Clinical Genetics, Royal Melbourne Hospital

    6Associate Genetic Counsellor, Royal Melbourne Hospital

    7Clinical Geneticist, Royal Melbourne Hospital

    8Immunopathologist, Monash Health

    9Head, Monash Genetics Clinic, Monash Health

    10Associate Genetic Counsellor, Monash Health

    11Clinical Immunologist, Royal Melbourne Hospital

    12 Associate Genetic Counsellor, Austin Health

    13Associate Genetic Counsellor, Royal Children's Hosptial

    14Clinical Immunologist, Royal Children's Hosptial

    15Immunology Nurse, Royal Children's Hosptial

    16Head, Clinical Immunology, Royal Children's Hosptial

    17Clinical Project Manager, Melbourne Genomics Health Alliance

    18Executive Director, Melbourne Genomics Health Alliance

    19Head, Masters Lab, Walter & Eliza Hall Institute

    20Head, Clinical Immunology & Allergy, Royal Melbourne Hospital

    21Head, Immunogenetics Research Unit, Walter & Eliza Hall Institute

    Primary Immunodeficiency diseases (PID) are a heterogeneous group of conditions with variable clinical features that are frequently associated with significant diagnostic delay. Accurate diagnosis has significant therapeutic benefit and may lead to personalized therapies. We established the Immunology Flagship of Melbourne Genomics Health Alliance in Australia to determine the clinical utility of genomic sequencing for diagnosis and management of individuals with suspected and confirmed cases of PID.

    198 adults and children with suspected or confirmed PID (n=153), autoinflammatory disease (n=33) and hereditary angioedema (HAE, n=11) were recruited to the Melbourne Genomics Immunology Flagship. Whole-exome sequencing (WES) was performed, with targeted gene analysis. Variant curation and reporting was performed according to the American Council of Medical Genetics guidelines. Overall, WES was diagnostic in 15% (30/198), confirming a preexisting diagnosis in 7% (14/198), and offering a new or more specific diagnosis in 8% (16/198). Variants of uncertain significance were identified in a further 28 patients (14%) in genes known to be associated with their clinical diagnosis, that warrant further functional validation. In the HAE group, diagnosis was confirmed in only 5 patients (45%), suggesting that WES may not be the appropriate technique for genetic diagnosis in this condition. A higher diagnostic rate was observed for autoinflammatory disorders (20%; 8/40) compared to PID (12%; 18/146). Of those who received a diagnosis, immediate changes to patient management and treatment occurred for 17/29 patients (59%), including HSCT for 3 and specific targeted therapy for 11 (38%) individuals.

    We have demonstrated the utility of WES for accurate diagnosis of complex immune diseases, with the potential to change diagnoses, guide therapeutic intervention and provide opportunities for genetic counseling. Further longitudinal analysis will determine clinical outcomes and health economic implications of genomic sequencing for diagnosis and management of immunological conditions in Australia.

    (117) Submission ID#601297

    Ataxia Telangiectasia and Common Variable Immunodeficiency with B-cell Lymphoma in Adolescent

    William Rafael. Marquez, Sr., MD1, Lorenzo Benitez, MD2, Lina Jaramillo, MD3

    1Misericordia Children Hospital, Bogota. Colombia, Pediatric immunologist

    2Fellow pediatric neumology, Bosque University, pediatric neumology

    3professor National University, Misericordia Children Hospital, Bogota. Colombia, pediatric pathologist

    At birth he had neonatal asphyxia and cerebral palsy. At 4 years old he had presented involuntary movements, left paresis, bilateral horizontal nystagmus. At 8 years of age, he had a right nasal obstruction. It was resected by otorhinolist and informed by biopsy: inflammatory polyp and chronic sinusitis. He has had 3 pneumonias, sinusitis and diarrhea.

    At the age of 13 years, the ataxia telangiectasia was confirmed by sequencing with PCR (62 exons, 91711 bp) of the ATM gene: transition G> A, nucleotide position 2250, codon 750, affecting splicing. Alpha fetoprotein 572-606.90 U/ml. Brain MRI, say Cerebellar Atrophy.

    He had IgG 685 mg / dl - 734 mg / dl, IgA 0.00 mg / dl, <1 mg / dl, IgM 268 mg / dl - 315 mg / dl, IgE 0.10 - <1 IU / ml. Subclasses of IgG: IgG3: 0.05 G / dl, IgG4: 0.04 gr/dl, low. IgG anti hepatitis B 6,22. No seroconversion. HIV negative TCD3 + lymphocytes: 32,40%, = 553 cells / mm3, LTCD4 +: 23,78% = 413,21 CEL / mm3, LTCD8 +: 7,69% = 133,5 cells / mm3, CD4 / CD8: 3.09. For all of the above, common variable immunodeficiency was diagnosed. He receives human immunoglobulin.

    At 16, I arrived at this hospital due to fever, respiratory distress and lymphadenopathy in the neck. CT showed ganglionic conglomerate on right side neck.

    Lymph node biopsy: strong tumors with CD20 and BCL2, weak and moderate diffuse PAX-5; Negativity with CD68, CD3 and CD10, and a cell proliferation index with Ki67 of 50%, diagnosis: Diffuse large B cell lymphoma. Treated with rituximab and chemotherapy. Lymphoma completely remitted.

    Conclusion: the association Ataxia Telangiectasia and lymphoma is frequent. By contrast, CVID and Ataxia Telangiectasia are extraordinarily rare.

    (118) Submission ID#601301

    Loss of Donor Chimerism 20 Years After Bone Marrow Transplant for Chronic Granulomatous Disease

    Christa S. Zerbe, MD, MS1, Jennifer Treat, PA-C, MSHS2, Samantha Kreuzburg, BA, RN3, Steven Holland, MD4, Harry L. Malech, MD5

    1Senior Research Physician, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH

    2Physician Assistant, Medical Science & Computing

    3Research Nurse Specialist, National Institutes of Health/National Institute of Allergy and Infectious Diseases

    4Director, Division of Intramural Research, NIAID

    5Chief, Genetic Immunotherapy Section, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    Introduction: Chronic granulomatous disease (CGD) is a primary immunodeficiency wherein affected patients are susceptible recurrent infections caused by specific bacteria and fungi as a result of defective NADPH activity. Additionally, inflammatory complications involving the bowel and lungs can cause significant morbidity. Currently the only proven permanent cure to CGD remains hematopoietic stem cell transplant.

    Case: A 25-year-old patient was diagnosed in infancy with X-linked CGD. At age 5yrs he received a nonmyeloablative peripheral blood stem cell transplant from his 10/10 non-carrier sister as previously reported (NEJM 344:881, 2001). Conditioning was cyclophosphamide (60 mg/kg) on D-6 and D-7; daily fludarabine (25mg/m2) on D-5 through D-1; Antithymocyte globulin at 40mg/kg on D-5 through D-2. Post-transplant immunosuppression consisted of cyclosporine on D-4 through D+100. He received 7.8x106 CD34+ peripheral blood stem cells which were T-cell depleted with 1x105 add back of CD3+ cells on Day 0. After 10 days of neutropenia (ANC <500) there were signs of engraftment. Per protocol, he received donor peripheral-blood lymphocytes containing 2.0x106 CD3+ cells/kg on D+ 30 after transplantation. Since donor T cells constituted less than 60 percent of his circulating CD3+ T cells and he had no graft versus-host disease, he received 1.0¬107 CD3+ cells/kg on D+60. After the discontinuation of cyclosporine, he received a total of three donor-lymphocyte infusions (1.0¬107 CD3+ cells/kg) at 90-day intervals achieving 100% T cell and myeloid engraftment at 26 months post-transplant with no acute nor chronic GvHD. At last follow-up 6 years post-transplant (2004) he had 100% and 98% lymphoid and myeloid peripheral chimerisms, respectively. The patient and family declined further periodic followup. Then, in October 2018 he presented with malaise, cough and fevers. He eventually was found to have a large consolidation and a BAL grew Burkholderia cepacia. His DHR showed 12% activity and peripheral blood myeloid and lymphoid chimerisms were 12% and 60%, respectively.

    Discussion: This late graft failure following peripheral blood transplant occurred following a conditioning regimen which is not the current standard for transplant in CGD. In the case series in which this patients transplant is reported (NEJM 2001), another patients myeloid chimerism fell to 15% by 3 years post-transplant, remaining stable at that level of chimerism without any serious infections over regular periodic follow up to the present time. Current regimens typically include busulfan to enhance engraftment and prevent graft failure. This case reinforces the need for prolonged monitoring of primary immune deficiency patients after transplantation.

    (119) Submission ID#601303

    Atypical Presentation of Complete DiGeorge Syndrome Without Correlating Genetic Defect: Rescued by State Newborn Screening

    Aba Al-Kaabi, MD, FAAP1, Lovya George MD, FAAP2, Erin M. Calhoun, B.S., MD Candidate, Class of 20203, Selina Gierer, DO4

    1Fellow - Allergy and Clinical Immunology, Univeristy of Kansas Medical Center

    2Assistant Professor – Neonatology, Univeristy of Kansas Medical Center

    3medical student, Univeristy of Kansas Medical Center

    4Assistant Professor - Allergy and Clinical Immunology, Univeristy of Kansas Medical Center

    Introduction: With the introduction of severe combined immunodeficiency (SCID) newborn screen (NBS) in the state of Kansas in 2017, a case of complete DiGeorge Syndrome (DGS) was discovered in an infant born to a diabetic mother with atypical features. This is the first DGS case diagnosed after adding the SCID NBS, which emphasizes the need to establish SCID NBS in all 50 states.

    Case presentation: The female infant was born via spontaneous vaginal delivery at 39 1/7 weeks to a 31 year old G1 now P1 mother. Maternal history was significant for chronic hypertension, obesity, insulin dependent type 2 diabetes, anxiety, depression, and scoliosis. The infant was noted to have a left sided abdominal wall defect and hernia, imaging identifying left renal agenesis, and was initially suspicious for VATER syndrome. Fortunately, the infant's SCID NBS revealed low T cell receptor excision circles (TRECS). Her initial white blood cell count was 14.1 with an absolute lymphocyte count of 2.679 K/UL. EBV PCR, CMV PCR, and HIV studies were negative. Chest imaging discovered absent thymus, abnormal vertebrae with only 10 ribs on the right and 12 ribs on the left, and abnormally formed thoracic vertebrae (T7). Echocardiogram detected an atrial septal defect measuring 0.32 cm, possible PFO versus secundum ASD. Endocrinology was consulted for management of labile calcium and phosphorus levels. FISH was negative for 22q11.2 deletion. Microarray revealed a variant of unknown significance arr[GRCh37]2p11.2(86285942_86506132)x3. Sequence analysis of combined and severe immune deficiency genes showed a variant of uncertain significance c.544C>A (p.Leu182Met).

    Management and Outcome: Additional evaluation included: CD3 67UL (1700-3600UL), CD4 51UL (1700-2800UL), CD8 19UL (800-1200UL), CD45RA 14 cells/uL (1100-5200cells/uL), normal CD 19, and CD 16/56, normal immunoglobulin G level, and normal dihydrorhodamine assay. Skeletal survey, CT abdomen and chest, and HLA typing were performed in preparation for thymic transplant.

    Discussion: Patients with complete DGS, a form of SCID found in less than 1 percent of patients with 22qDS, have absent thymus and a T cell count <3 standard deviations below normal for age (typically <50 naïve CD3+ T cells/mm3). In a large series of patients with complete DGS, only 52 percent had an identifiable 22q11.2 deletion [1].

    Infants of a diabetic mother have various genetic and syndromic associations including diabetic embryopathy. [2] Despite the importance of immunological aspects in pregnancy, few studies have reported on the cellular immune modifications of diabetic embryopathy. Diabetes during pregnancy may affect the development of the thymus and thus maturation of the immune system in the offspring. [3]

    The recent addition of a TREC assay to newborn screening can identify such a subset of infants with atypical presentations. SCID NBS uses an assay for TRECs, a biomarker of T cell development. [4-6] This initial presentation now places the immunologist in the role of "first responder" with regard to diagnosis and management of these patients, who may present with atypical features. Newer genetic and molecular techniques now allow for earlier identification of immune defects in such disorders with life-long clinical concerns. [7]

    References:

    1. Markert et al. Review of 54 patients with complete DiGeorge anomaly enrolled in protocols for thymus transplantation: outcome of 44 consecutive transplants. Blood. 2007;109(10):4539. Epub 2007 Feb 6.

    2. Stiehm et al. Steim’s Immune Deficiencies. Academic Press. 2014. Print.

    3. Warncke K et al. Thymus Growth and Fetal Immune Responses in Diabetic Pregnancies. Horm Metab Res. 2017 Nov;49(11):892-898. doi: 10.1055/s-0043-120671. Epub 2017 Nov 14.

    4. Kwan et al. Newborn screening for severe combined immunodeficiency in 11 screening programs in the United States. JAMA. 2014 Aug 20;312(7):729-38. doi: 10.1001/jama.2014.9132. Erratum in: JAMA. 2014 Nov 26;312(20):2169.

    5. Kwan et al. History and current status of newborn screening for severe combined immunodeficiency. Semin Perinatol. 2015 Apr;39(3):194-205. doi: 10.1053/j.semperi.2015.03.004. Epub 2015 Apr 30. Review.

    6. http://primaryimmune.org/idf-advocacy-center/idf-scid-newborn-screening-campaign.

    7. Kuo et al. Immune and Genetic Features of the Chromosome 22q11.2 Deletion (DiGeorge Syndrome). Curr Allergy Asthma Rep. 2018 Oct 30;18(12):75. doi: 10.1007/s11882-018-0823-5.

    (120) Submission ID#601306

    Clinical and Laboratory Features of Thymoma and Immunodeficiency (Good's Syndrome): A Report from the USIDNET Registry and the Mount Sinai Hospital Cohort

    Hsi-en Ho, MD1, Ramsay Fuleihan, MD2, A Wesley. Burks, MD3, Shradha Agarwal, MD4, Charlotte Cunningham-Rundles, MD, PhD5

    1Fellow - Allergy and Clinical Immunology, Icahn School of Medicine at Mount Sinai

    2Professor of Pediatrics, Division of Allergy and Immunology, Northwestern University Feinberg School of Medicine, Chicago, NY

    3Professor of Pediatrics, Division of Allergy and Immunology, UNC School of Medicine

    4Associate Professor, Division of Clinical Immunology, Icahn School of Medicine at Mount Sinai

    5Professor in Medicine, Division of Clinical Immunology, Icahn School of Medicine, Mount Sinai, NY, NY, USA

    Introduction/Background: Goods syndrome is a rare cause of combined B- and T-cell immunodeficiency occurring in association with a thymoma. Affected patents are susceptible to bacterial, fungal, viral, and opportunistic infections. An association with autoimmunity has also been reported. Current knowledge of Goods syndrome is primarily limited to case reports and small series.

    Objectives: To examine the spectrum of clinical and laboratory features of a major cohort of Goods syndrome patients in the US.

    Methods: We conducted a retrospective analysis of patients with Goods syndrome in the USIDNET Registry and the Mount Sinai Hospital (MSH) cohort.

    Results: We identified 20 patients with thymoma and hypogammaglobulinemia (USIDNET, n=11; MSH, n=9; median age: 60 years; female: 45%), representing data from 151 patient years. The median age at diagnosis of thymoma and hypogammaglobulinemia were 52 years (range 31-85), and 50.5 years (range 28-86), respectively. Two patients were deceased (at age 65 and 70 years, cause unspecified). All patients had low IgG (median 313mg/dL, range 47-699). IgA and IgM were reduced in 90% and 45% of patients, respectively. Low CD19+ B cells (median 0.5/mm^3, range 0-28) were reported in all available records. The absence of CD19+ B cells was observed up to 21 years post-thymectomy. A wide range of additional laboratory abnormalities were identified: low CD4+ T cells (n=5), low CD8+ T cells (n=2), low CD4/CD8 ratio (n=6), low NK cells (n=6), and absent peripheral eosinophils (n=8). The most common sites of infections were lower respiratory (70%), upper respiratory (55%), and gastrointestinal (35%). In addition, sepsis (15%), meningoencephalitis (5%), osteomyelitis (5%), and urinary tract infection (5%) were also observed. Identifiable infectious agents included: bacteria (35%), virus (35%), fungus (25%), parasites (10%), and protozoa (5%), with opportunistic infections recorded in 25% of patients. Opportunistic infections were significantly associated with absolute CD4 lymphopenia (p=0.048, Fishers exact test). Enterovirus was identified as a previously unreported cause of meningoencephalitis in this population. Autoimmune manifestations were reported in 45% of patients, with a higher prevalence of inflammatory colitis (20%) than previously reported. Hashimoto thyroiditis, fibromyositis, and bronchiolitis obliterans organizing pneumonia (n=1 each) were identified as previously unreported autoimmune/inflammatory conditions in this population. A case of alopecia areata was also observed. Additionally, bronchiectasis was recorded in 20% of patients. All patients were initiated on immunoglobulin replacement, with antibiotics prophylaxis in 20%, and immunosuppressive medications employed in 10% of patients post diagnosis of immunodeficiency.

    Conclusion: Goods syndrome is a combined immunodeficiency, with a wide range of autoimmunity in a subset of patients. We expanded upon the spectrum of associated infectious and inflammatory complications through a major US cohort. Persistent immune dysregulation was observed up to 2 decades post-thymectomy.

    (121) Submission ID#601308

    Immune Dysregulation: Diagnosis of Behcets Disease in an Affected Chronic Granulomatous Disease Carrier

    Aba Al-Kaabi, MD, FAAP1, Jessica Hobson, MD2, John Martinez, MD3

    1Fellow - Allergy and Clinical Immunology, University of Kansas Medical Center

    2Assistant Professor - Allergy and Clinical Immunology, University of Kansas Medical Center

    3Assistant Professor - Allergy and Clinical Immunology, University of Kansas Medical Center

    Introduction: Primary immunodeficiencies (PIDs) constitute a large group of rare disorders that affect the immune systems function. Some PID patients develop autoimmunity in addition to having increased susceptibility to infections due to their impaired immunity [917]. (1)

    Case presentation/ Management: A 43 year old Caucasian female with history of bipolar disorder, Factor V Leiden deficiency, anti thrombin 3 deficiency, pulmonary embolism, endometriosis, and seasonal allergies was evaluated for Chronic Granulomatous Disease (CGD) in 2007. The main symptoms were inflammatory breast lesions necessitating 4 surgeries on the right breast, and back, facial, genital, ocular, mouth, and scalp sores. Biopsy with cultures of the wounds was positive for Corynebacterium, coagulase-negative staphylococcus, enterococcus, bacteroides species, and Provatella. Neutrophil oxidative burst was ordered by the infectious disease specialist and showed normal and abnormal neutrophil populations, a finding consistent with CGD carrier. Patient was started on Interferon gamma-1b after failing multiple courses of antibiotics. Her symptoms were well controlled on Interferon gamma-1b 100mcg/0.5ml SQ every other day, Trimethoprim 100mg tab (2tabs in am and 1 tab in pm), cefixime 400mg once daily, and topical mupirocin as needed except for her recurrent genital ulcers. CGD can be rarely associated with oral ulcers however there is a limited literature describing associated genital ulcers.

    According to the International Study Group diagnostic criteria published in 1990 (2), the patient was diagnosed by a rheumatologist as having Behcets disease (BD). There are no pathognomonic laboratory tests in BD; as a result, the diagnosis is made clinically. Patient failed a trial of colchicine and was later started on Cyclosporine, which resulted in decrease of her mouth and genital ulcers.

    Discussion: BD is a rare disease mostly seen along the Silk Road. The prevalence has been reported as 0.12 (USA) to 370 (in a single village, northern Turkey) for 100 000 inhabitants. (3) CGD is a primary immunodeficiency caused by defects in any of the five subunits of the NADPH oxidase complex responsible for the respiratory burst in phagocytic leukocytes. Patients with CGD are at increased risk of life-threatening infections with catalase-positive bacteria and fungi, and inflammatory complications such as CGD colitis. (4)

    Reports of CGD female carriers with discoid lupus erythematosus, photosensitivity rashes, and other autoimmune phenomena have been published [48,49] (4). To the best of our knowledge, this is the first case to report BD in an affected CGD carrier.

    The treatment of inflammatory disease in patients with CGD poses a difficult balance between therapeutic immunosuppression and the increased risk of severe infection. (5). High dose intravenous immunoglobulin, and targeted therapies such as CTLA4-Ig for T cell mediated pathologies, Rituximab for B-cell mediated pathologies, and anti-TNF for IBD, may be preferable over the broad immunosuppressive activity of glucocorticoids. In addition, emerging evidence suggests that hematopoietic stem cell transplantation has indication for cases that have been difficult to control using immunosuppression. (1) Given all that, our case emphasizes the need to maintain suspicion for autoimmune disorders / immune dysregulation in patients with PID.

    References:

    1) Aziz et al. Cellular and molecular mechanisms of immune dysregulation and autoimmunity. Cell Immunol. 2016 Dec;310:14-26. doi: 10.1016/j.cellimm.2016.08.012. Epub 2016 Aug 27.

    2) Criteria for diagnosis of Behçet's disease. International Study Group for Behçet's Disease. Lancet. 1990;335(8697):1078.

    3) Zouboulis CC (2003) Epidemiology of adamatiades-Behcet's disease. In: Zierhut M, Ohno S (eds) Immunology of Behcet's Disease, pp 1–16. Zeitlinger BV, Lisse, The Netherlands.

    4) Arnold, D and Heimall, J. A Review of Chronic Granulomatous Disease. Adv Ther. 2017 Dec;34(12):2543-2557. doi: 10.1007/s12325-017-0636-2. Epub 2017 Nov 22.

    5) Thomsen IP et al. A Comprehensive Approach to the Management of Children and Adults with Chronic Granulomatous Disease. J Allergy Clin Immunol Pract. 2016 Nov - Dec;4(6):1082-1088. doi: 10.1016/j.jaip.2016.03.021. Epub 2016 May 10.

    (122) Submission ID#601310

    Encephalopathy in an Adolescent with CD40-ligand Deficiency

    Raquel Rozner, MD1, Elizabeth Feuille, MD2, James Bussel, MD3, Luigi D. Notarangelo, MD, PhD4, Charlotte Cunningham-Rundles, MD, PhD5

    1Resident Physician, New York Presbyterian Hospital-Weill Cornell Medical Center

    2Attending Physician, New York Presbyterian Hospital- Weill Cornell Medical Center

    3Professor in Pediatrics, Department of Hematology and Oncology, Weill Cornell Medicine, NY, USA

    4Chief, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    5Professor in Medicine, Division of Clinical Immunology, Icahn School of Medicine, Mount Sinai, NY, NY, USA

    Introduction: CD40-ligand deficiency is an X-linked combined immunodeficiency, characterized by susceptibility to infection, often with associated neutropenia, malignancy, and autoimmunity. Central nervous system (CNS) manifestations are less commonly reported than respiratory or gastrointestinal complications, but are most often attributed to infection. Herein we describe a challenging case of gradual onset episodic memory loss, confusion, and unilateral hemiplegia in a young male with CD40-ligand deficiency.

    Case Presentation: The patient is a 13-year-old male with CD40-ligand deficiency on immunoglobulin replacement therapy presenting with recurrent, episodic altered mental status (AMS) and gradual neurocognitive decline. Initial neurologic symptoms began at age 11 years, and included fever, nausea, and eyelid fluttering. Initial comprehensive infectious workup at this time, including blood and urine cultures, Lyme antibody, serum PCR for HSV, CMV, EBV, respiratory viral PCR including atypical viruses, CSF studies including culture, Lyme EIA, PCRs for enterovirus, VZV, EBV, CMV, HSV1/2 were unrevealing. Electroencephalogram (EEG) and MRI displayed generalized slowing and global atrophy, respectively. Definitive diagnosis was not made. The patient continued to decline with worsening developmental delay and memory loss. One year later, at age 12 years, he had a recurrent episode of AMS with repeat negative infectious workup including blood and urine cultures, respiratory virus PCR including atypical viruses, CSF culture including acid fast bacillus and fungi, cryptococcal antigen, viral encephalitis panel by PCR, and serum PCR for EBV and HHV-6. EEG at this time showed left hemispheric epileptogenic potential, consistent with seizure activity.

    His presentation, at age 13 years, was notable for right-sided hemiplegia with facial numbness, dysarthria, nausea, and fever. He was found to have anello virus on PCR of CSF, abnormal left temporal region on EEG, and global atrophy with stable, diffuse generalized volume loss on MRI. He was diagnosed with occult anello virus-induced encephalitis with hemiplegic migraine and discharged on valproate.

    Discussion: Here we present the first reported case of Anello virus detected by PCR in a CD40-ligand deficient male with neurocognitive manifestations, attributed primarily to hemiplegic migraine. Given the anello virus prevalence and relatively avirulent character, it is presumed to be unlikely culprit for encephalitis; however, the significance of this finding is as yet unknown. This case highlights diagnostic challenges in immunodeficiency: infection may go undetected by standard diagnostic techniques; however, the significance of infections identified with advanced techniques may not yet be understood.

    (123) Submission ID#601323

    Etanercept Use in Refractory Chronic Henoch-Schönlein Purpura

    Sana Habib, MD1, Elif Dokmeci, MD2

    1Pediatric Resident, UNM

    2Assoc Prof, University of New Mexico

    Background: Henoch-Shönlein purpura (HSP) is an IgA-mediated small vessel vasculitis that presents with a tetrad of abdominal pain, arthritis, glomerulonephritis, and purpura. HSP is typically a self-limiting disease of childhood following a viral illness. There is no universal treatment for patients with chronic or recurrent HSP. We report a chronic refractory case of HSP that was successfully treated with a tumor necrosis factor inhibitor (TNFi), Etanercept. Etanercept functions as recombinant protein that consists of a TNF-alpha receptor ligand-binding region that links to the Fc portion of human IgG. It is currently approved for use in 5 diseases: juvenile rheumatoid arthritis, rheumatoid arthritis, ankylosing spondylitis, plaque psoriasis, psoriatic arthritis. TNFi are categorized into two broad categories, recombinant receptors (etanercept) and neutralizing antibodies (ex. infliximab and adalimumab). There have been prior case reports of HSP associated with TNFi agents during the treatment of other autoimmune conditions in the adult population. To our knowledge, there have been 3 prior etanercept related HSP reports, one report associated with adalimumab, and one with infliximab. However, there has been no prior report of etanercept use successfully treating chronic refractory HSP.

    Case Presentation: A 16-year-old Native American male with 3 year history of chronic HSP, HLA-B27 positive, and enthesitis related arthritis who was initially treated with steroids, sulfasalazine and methotrexate for symptoms of joint pain and purpura. His IgA level was 545 mg/dL prior to therapy. Despite treatment for one month of steroids, eight months of sulfasalazine exclusively and eight months of methotrexate and sulfasalazine, he continued to have persistent purpura on bilateral extremities without improvement. He was subsequently initiated on Etanercept 50mg weekly and methotrexate was discontinued. Approximately one month later, his rash significantly improved. His rash and joint pain recurs when he misses a dose of Etanercept. Punch biopsies were taken 3 months after initiation of etanercept. The biopsies of a lesion from his left arm showed early leukocytoclastic vasculitis and from his left leg showed weak granular deposition of IgA, IgM and C3 within vessel walls. There is controversy whether this is a true IgA vasculitis. However, we believe that his clinical presentation and the deposition of IgA and C3 within blood vessel walls seen on biopsy correlates with chronic Henoch-Shönlein purpura.

    Conclusion: There is no standard treatment of chronic HSP, but there are reports of benefit with NSAID and corticosteroids. Per our literature review, there are no prior reports of Etanercept use in the treatment of chronic HSP. TNF inhibitor, Etanercept should be considered as a treatment for chronic refractory HSP in the pediatric population as it has showed rapid resolution of purpura in this case report. Further studies of Etanercept in the treatment of chronic HSP should be conducted given the controversial literature of anti-TNF ab induced HSP during the treatment of other autoimmune diseases.

    (124) Submission ID#601334

    Recurrent Sinusitis in Heterozygous Hemochromatosis; Is It a Risk Factor?

    Osman Dokmeci, MD1

    1Assist Prof, University of New Mexico

    BACKGROUND: Hereditary hemochromatosis (HH) is one of the most common inherited disorders in people of northern European descent. HH patients are at risk for a number of infections including invasive fungal sinus infections. Although clinical manifestations of iron overload appear to be quite uncommon in patients who are heterozygous carriers of HFA mutation, we present cases that appear to suggest an increased risk non allergic rhino-sinusitis.

    CASE REPORT: We present a 66 year old gentleman with perennial colored rhinorrhea, with facial pressure and tenderness, constant post nasal drip, dry cough and bilateral congestion that had been going on for the past several years. He also had a frequent urge to clear his throat and had frequent episodes of sore throat despite having no history of GERD or LPR.

    He reported to have multiple sinus infections every year that would progress to pneumonia and eventually require long courses of oral antibiotics. All started in his 40s intensified in the recent past. He had 3 other siblings; one died in his 40s due to liver complications of HH and had a carrier sister and brother with a hx of sino nasal problems exactly similar to the patients. His exam was remarkable for bilateral narrowed nasal passages and moderate edema of the mucosa. His rhinolaryngoscopy showed significant edema and purulent drainage, most notably from bilateral middle meati.

    His skin test was negative. His CBC showed a WBC count of 6.7/ml with 2% eosinophils and his immunoglobulin panel showed an IgA of 236 mg/dl, IgG of 1190 mg/dl and IgE of 31 mg/dl. Patient was placed on Alkalol sinus rinses and Azelastine nasal spray, which he reported to work pretty well. He left for Costa Rica and is expected to return back with his siblings to A&I clinic in the coming months.

    DISCUSSION: HH is one of the most common inherited disorders in people of northern European descent with an incidence of 1:200 and carrier rate of 1:10.. Most affected HH patients are homozygous for the mutation designated C282Y at the HFE gene located at the 6th chromosome. Unlike hereditary hemochromatosis, clinical manifestations of iron overload appear to be quite uncommon in patients who are heterozygous carriers. HH patients are at risk for a number of infections with bacteria whose virulence is increased in the presence of excess tissue iron. HH is also a risk factor for acute fulminant FRS . Here the mechanism is postulated to be due to quantitative or qualitative neutrophil defects as this condition is mostly seen in patients with DM, aplastic anemia, and can happen in patients undergoing antineoplastic chemotherapy. No known increased susceptibility for infections through either mechanism is postulated for patients with the heterozygous carrier state. Here we present 3 HH carrier patients who present with recurrent rhinosinusitis with no allergen sensitizations and normal IgE levels. Since most fungal immunity is at the tissue level and is cytokine driven, it can be speculated that increased tissue levels of iron might interfere with mechanisms of innate immunity.

    (125) Submission ID#601340

    Compound Heterozygous DOCK8 Mutations in a Patient with pre-B Cell Acute Lymphoblastic Leukemia and EBV-associated Diffuse Large B-cell Lymphoma

    David K. Buchbinder, MD, MSHS1, Ivan Kirov, MD2, Jeffrey Danielson, MS3, Nirali N. Shah, MD4, Alexandra F. Freeman, MD5, Helen C. Su, MD, PhD6

    1Assistant Clinical Professor, Department of Hematology, Children's Hospital of Orange County, Orange, CA, Department of Pediatrics, University of California at Irvine, Orange, CA

    2Clinical Professor, Department of Oncology, Children's Hospital of Orange County, Orange, CA, Department of Pediatrics, University of California at Irvine, Orange, CA

    3Research Staff Member, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD

    4Associate Research Physician, Pediatric Oncology Branch, NCI, NIH, Bethesda, MD

    5Director, Primary Immune Deficiency Clinic, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    6Chief, Human Immunological Diseases Section, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD

    Background: Dedicator of cytokinesis 8 (DOCK8) mutations are associated with a combined immunodeficiency disorder marked by atopic features, infectious susceptibility with a striking preponderance of cutaneous viral disease, and a risk for the development of malignancy including lymphoma. Almost all cases can be diagnosed by documentation of the loss of DOCK8 protein expression.

    Methods: We describe a 22-year-old male with a diagnosis of pre-B cell acute lymphoblastic leukemia (ALL) followed by Epstein-Barr Virus (EBV) associated diffuse large B cell lymphoma (DLBCL). Compound heterozygous mutations in DOCK8 were documented following the completion of whole exome sequencing (WES). The pathogenicity of the variants was assessed. Flow cytometric quantification of intracellular DOCK8 protein was completed. DOCK8 protein function was assessed by evaluating the morphology of patient lymphocytes when migrating in a 3D collagen matrix.

    Results: A concern for a primary immunodeficiency was raised due to a history of recurrent otitis media which began at 12 months of age. By 4 years of age, numerous warts were noted on his fingers; however, they were transient for a duration of only 2 years. No atopic features were appreciated. At 15 years of age, a diagnosis of pre-B cell ALL was made. During ALL therapy, infectious complications were severe including an intestinal perforation, osteomyelitis, and sepsis. At 22 years of age, still in an ongoing remission from his ALL, an incidental finding of a lung nodule led to a diagnosis of EBV-associated DLBCL. During therapy, however, infectious complications were again severe including a soft tissue infection and sepsis. WES was performed and compound heterozygous mutations in DOCK8 (c.1128_1132del and c.4474-1G>C) were documented. Flow cytometric quantification of intracellular DOCK8 protein was normal when compared to a normal control. Nevertheless, additional functional assessment of DOCK8 protein was completed. When migrating through a 3D collagen matrix, 45% of the patient lymphocytes studied demonstrated abnormal elongation (stretch ratio > 8 defined by length/width) compared with 10% of lymphocytes from a normal control. He is being evaluated for hematopoietic stem cell transplant.

    Conclusion: Autosomal recessive mutations in DOCK8 are a rare cause of a combined immunodeficiency marked by atopic features, infectious susceptibility with a striking preponderance of cutaneous viral disease, and a risk for the development of malignancy including lymphoma. Here, pre-B cell ALL followed by the development of a subsequent malignant neoplasm (EBV-associated DLBCL) led to the discovery of DOCK8 deficiency. Hence, as our case underscores, for rare instances of high clinical suspicion despite normal DOCK8 protein expression, additional functional testing is crucial to make a definitive diagnosis and plan treatment. Understanding the spectrum of DOCK8 mutants and their phenotypes will improve our understanding of DOCK8 deficiency.

    (126) Submission ID#601345

    Recalcitrant Abdominal Abscesses in a Patient with Hyperimmunoglobulin E Syndrome

    Anh Nguyen, MD/MPH1, Victoria Dimitriades, MD2

    1Allergy and Immunology Fellow, Division of Rheumatology, Allergy and Clinical Immunology, Department of Internal Medicine, University of California Davis Health

    2Associate Clinical Professor of Pediatrics, Division of Pediatric Allergy, Immunology & Rheumatology, University of California Davis Health

    Background: Autosomal dominant hyperimmunoglobulin E syndrome (AD-HIES) is a rare primary immunodeficiency caused by heterozygous loss-of-function mutations in the signal transducer and activator of transcription 3 (STAT3) gene. AD-HIES classically characterized by recurrent cold Staphylococcal abscesses, pneumonia, eczema, and an elevation of IgE level. Other additional clinical manifestations of HIES have been recognized including skeletal dysplasia (scoliosis, pathologic fractures, delayed dental deciduation), pneumatoceles, coronary-artery aneurysms, brain lesions, and Chiari malformations.

    Objective: To describe a unique case of abdominal abscesses in a patient with AD-HIES.

    Method: A 22-year-old female with known AD-HIES (C.1144 C>T (p.Arg382Trp)) and a complicated history of early Pneumococcal pneumonia and meningococcemia resulting in bilateral amputation below the knees along with loss of several digits, presented for evaluation of skin infection. She had a history of recurrent Staphylococcal skin abscesses and presented with inability to use her prostheses due to pain from inflammation around her amputation sites. She underwent imaging and was found to have bilateral extremity abscesses with an associated osteomyelitis of her L tibia (which was found to be MRSA after incision and drainage). While receiving intravenous antibiotics for her osteomyelitis, she developed intractable abdominal pain. Imaging showed a thick-walled, multi-septated, paranephric abscess as well as several smaller abscesses scattered throughout her abdomen. She underwent multiple drain placements and drainage of retroperitoneal fluid collections via interventional radiology (IR). Purulent fluid from the abdominal abscess drainage grew MRSA. The patient continued to have re-accumulation of abscesses despite multiple drainages. Repeat imaging noted increased paranephric abscesses which were not communicating with drains. Given lack of response to several IR-placed abdominal drains and to 6 weeks of intravenous antibiotics, she had an open surgical washout with minimal improvement. Hospital course was further complicated by development of a left lower lung lobe consolidation and sub-segmental pulmonary embolism necessitating treatment with heparin. Finally, after several weeks of escalating antimicrobial therapy and with additional drain placements, the retroperitoneal abscesses started to recede. Repeat abdominal imaging several months later while asymptomatic revealed slow but continuing resolution of the abscesses.

    Conclusion: The present case raises awareness of an unusual location for infection in a patient with AD-HIES. Although the majority of complications of AD-HIES are sinopulmonary and skin infections, recalcitrant intra-abdominal abscesses should be considered in the differential of infections in HIES.

    (128) Submission ID#601352

    Epidemiology on Primary Immunodeficiencies in Korea: A Systematic Review of Reported Literature and Analysis of Bigdata from National Health Insurance System

    Sohee Son, PhD1, Ji-Man Kang, MD, PhD2, Younsoo Hahn, MD, PhD3, Kang Mo Ahn, MD, PhD1, Yae-Jean Kim, MD, PhD1

    1Department of Pediatrics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea

    2Department of Pediatrics, Severance Children's Hospital, Yonsei University College of Medicine, Seoul, Korea

    3Department of Pediatrics, Chungbuk National University Hospital, Cheongju, Korea

    Introduction/Background: The recent epidemiologic studies have revealed that primary immunodeficiencies (PIDs) are more common than previously thought. However, there are very few data on epidemiology of PIDs in Korea.

    Objectives: We attempted to estimate the PID epidemiology and disease burden in Korea and provide the background information for PID registry for future.

    Methods: To review the previously reported scientific studies, PubMed, KoreanMed, Google Scholar were searched. Any studies on PIDs reported in scientific journal (Korean or International) from January 2001 to November 2018 were searched. Both Korean and English reports were searched. Diagnosis for PID was categorized from group I to group XI according to 2017 IUIS Phenotypic Classification. Study period was divided into two periods: period 1 from 2001 to 2005 and period 2 from 2006 to 2018, because there was a multicenter study to estimate PID epidemiology from 2001 to 2005. In addition, the number of PID patients and the cost for care were estimated among patients who requested reimbursement to Health Insurance Review and Assessment Service (HIRA) Korea for one year in 2017.

    Results: A total of 334 PID patients were identified in 75 reports. One hundred and ninety-nine patients (20 reports) and 135 patients (55 reports) were found in period 1 and period 2, respectively. The PIDs were reported in 11 patients for immunodeficiencies affecting cellular and humoral immunity, 23 patients for combined immunodeficiency with associated or syndromic features, 143 patients for predominantly antibody deficiencies, 33 patients for diseases of immune dysregulation, 113 patients for congenital defects of phagocyte, 1 patient for defects in intrinsic and innate immunity, 4 patients for auto-inflammatory disorders, 6 patients for complement deficiencies, and none for phenocopies of PID.

    From HIRA reimbursement data, the number of PID patients were 42 for combined immunodeficiency, 486 for predominantly antibody deficiency, 47 for common variable immunodeficiency, 135 for functional defect of neutrophils, 238 for immunodeficiency associated with other major defects, 272 for other immunodeficiencies. A total of 1,220 PID patients were treated for 14,316 days and $3,351,678 was reimbursed in 2017.

    Conclusions: We performed a systematic review on published studies for PID in medical journals and national open data system of HIRA to estimate the PID disease burden for the first time in Korea. To obtain more information on true PID epidemiology and disease burden in Korea, a national multicenter study for PID registry is required in the future.

    (129) Submission ID#601355

    Lentiviral Gene Therapy Corrects Platelet Phenotype and Function in Wiskott-Aldrich Patients

    Lucia Sereni, PhD1, Maria Carmina Castiello, PhD2, Dario Di Silvestre, PhD3, Patrizia Della Valle4, Chiara Brombin, PhD5, Francesca Ferrua, MD6, Maria Pia Cicalese, MD, PhD6, Loris Pozzi, MSc4, Maddalena Migliavacca, MD, PhD6, Maria Ester Bernardo, MD6, Claudia Pignata, MD7, Roula Farah, MD8, Lucia Dora Notarangelo, MD9, Nufar Marcus, MD10, Lorella Cattaneo, MD11, Marco Spinelli, MD12, Stefania Giannelli, PhD13, Marita Bosticardo, PhD14, Koen van Rossem, MD, PhD15, Armando D'Angelo, MD16, Alessandro Aiuti, MD, PhD17, Pierluigi Mauri, PhD18

    1Junior Postdoctoral Fellow, San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), Division of Regenerative Medicine, Stem Cells and Gene Therapy, San Raffaele Scientific Institute, Milan, Italy

    2Senior Postdoctoral Fellow, San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), Division of Regenerative Medicine, Stem Cells and Gene Therapy, San Raffaele Scientific Institute, Milan, Italy

    3Researcher, Proteomic and Metabolomic Laboratory, Institute of Biomedical Technologies, National Research Council (ITB-CNR), Segrate (MI), Italy

    4Biologist, Coagulation Service & Thrombosis Research Unit, San Raffaele Scientific Institute, Milan, Italy

    5Researcher, University Centre for Statistics in the Biomedical Sciences (CUSSB), Vita-Salute San Raffaele University, Milano, Italy 5Vita-Salute San Raffaele University, Milan, Italy

    6Clinician, San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), Division of Regenerative Medicine, Stem Cells and Gene Therapy, San Raffaele Scientific Institute, Milan, Italy

    7Clinician, Pediatric Section, Dep of Translational Medical Sciences, University of Naples Federico II, Naples, Italy

    8Clinician, Department of Pediatrics, Division of HematologyOncology. Saint George Hospital University Medical Centre, Beirut, Lebanon

    9Clinician, Pediatric Onco-Haematology and BMT Unit, Children's Hospital, ASST Spedali Civili of Brescia, Brescia, Italy

    10Clinician, Department of Pediatrics B, Schneider Children's Medical Center of Israel, Petach Tikva, Israel

    11Clinician, A.O. SS. Antonio e Biagio C. Arrigo. Alessandria, Italy

    12Clinician, Pediatric Clinic, MBBM Foundation, Maria Letizia Verga Center, Monza, Italy

    13Biologist, San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), Division of Regenerative Medicine, Stem Cells and Gene Therapy, San Raffaele Scientific Institute, Milan, Italy

    14Staff Scientist, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    15Clinical Research Director, Rare Diseases Unit, GlaxoSmithKline, Brentford, UK

    16Clinician, Head of Unit, Coagulation Service & Thrombosis Research Unit, San Raffaele Scientific Institute, Milan, Italy

    17Clinician, Head of Unit, San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), Division of Regenerative Medicine, Stem Cells and Gene Therapy, San Raffaele Scientific Institute, Milan, Italy.

    18Staff Scientist, Head of Unit, Proteomic and Metabolomic Laboratory, Institute of Biomedical Technologies, National Research Council (ITB-CNR), Segrate (MI), Italy

    Anna Villa, MD

    Clinician, Head of Unit, San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), Division of Regenerative Medicine, Stem Cells and Gene Therapy, San Raffaele Scientific Institute, Milan, Italy. 8 Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, Milan, Italy

    Micro-thrombocytopenia is one of the most serious challenges for Wiskott-Aldrich Syndrome (WAS) and X-linked Thrombocytopenia (XLT) patients. Thrombocytopenia leads to severe, potentially life-threatening, bleeding episodes, which require frequent transfusions and account for 23% of deaths in patients experiencing WAS mutations.

    The gold standard treatment for WAS patients is hematopoietic stem cell transplantation (HSCT) from an HLA-identical donor but more recently a number of gene therapy (GT) trials in Europe and USA showed promising results. In particular, it has been shown that WAS patients receiving lentiviral mediated GT, consisting of autologous CD34+ cells transduced with lentiviral vector encoding the human WAS gene under the control of the endogenous promoter, in combination with a reduced intensity conditioning regimen, have a significant increase in platelet (PLT) counts. Even though PLT counts do not reach normal levels, treated patients decreased the severity and frequency of bleedings.

    Here, in a cohort of 4 XLT and 16 WAS patients, fifteen treated with GT, the PLT phenotype and function were analyzed by electron microscopy, flow cytometry and proteomic profile. The aim of the project is to assess the presence of PLT defects in WAS untreated patients and the impact of GT treatment on the correction of PLT behavior.

    We demonstrate that PLTs of untreated WAS patients have reduced size and abnormal ultrastructure along with hyperactivated phenotype at steady state, showing increased expression of CD62P, activated IIb3 integrin and CD40L; conversely, activation response to agonist and aggregation capacity are both decreased. Analyzing PLT samples isolated from treated patients, we found that GT restores PLT size and ultrastructure very early after treatment and reduces the hyperactivated phenotype proportionally to WAS protein (WASp) expression and follow-up length. PLTs isolated from GT treated patients showed a normal activation response to agonists and restored aggregation capacity in 5 out of 7 analysed patients.

    By proteomics, various protein pathways were found downregulated in untreated PLT samples, mainly involving cytoskeletal-rearrangement proteins, integrins, signal transduction molecules, vesicles-transport proteins; additionally, decreased metabolic capacity were observed. These results are in line with the functional defects observed in PLTs in terms of activation and aggregation. Conversely, the expression of protein-pathways found downregulated in untreated patients is comparable to healthy controls in GT-treated PLT samples, reflecting the amelioration of PLT phenotype and function.

    Overall, our study highlights the coexistence of multiple defects in the activation and aggregation responses occurring in WAS patient PLTs in absence of WASp. GT was able to normalize the PLT proteomic profile followed by consequent restoration of PLT ultrastructure and phenotype, suggesting GT is responsible for the observed reduction of bleeding episodes in treated patients.

    (130) Submission ID#601392

    PIK3CD, a Rare Autosomal Dominant Disorder of the Immune System: A Reason for the Use of Next Generation Sequencing

    Jacob L. Barish, MD1, Lyda Cuervo-Pardo, MD2, Mario Rodenas, MD, FAAAAI3

    1Internal Medicine Resident, University of Florida, Department of Internal Medicine

    2Assistant Professor, University of Florida, Division of Rheumatology & Clinical Immunology, Department of Medicine

    3Assistant Professor, University of Florida, Division of Rheumatology & Clinical Immunology, Department of Medicine

    Introduction: PIK3CD is an autosomal dominant genetic disorder of the immune system that results in persistent activation of PI3K. Signaling through PI3K is essential for immune cell regulation of metabolism, migration, proliferation and differentiation, leading patient to present with lymphadenopathy, immunodeficiency and senescent T cells. The mutated protein causes T cells to over activate and mature too quickly leading to their death, this over activation also blocks the maturation of B cells.

    Case presentation: A 51-year-old female with a childhood history of failure to thrive, asthma, chronic rhinitis and Common variable Immunodeficiency on intravenous immunoglobulin replacement, was seen in immunology clinic to establish care. She reported frequent episodes of pneumonia and bronchitis in her childhood. Her family history was significant for family members with leukopenia, but no diagnosed immunodeficiency. Patient had 1 son who did not report symptoms concerning for immunodeficiency. Physical exam was within normal limits with no lymphadenopathy.

    Laboratory examinations exhibited normal IgA (185 mg/dL), IgG (800 mg/dL), and IgM (100 mg/dL). While flow cytometry showed normal absolute CD3 687 (570-2400 cells/uL), CD4 (540 cells/uL), NK Cells (151 cells/uL), CD19 (179 cells/uL), CD45RA (160 cells/uL), CD45RO (311 cells/uL), CD2 (757 cells/uL), and HLA-DR (173 cells/uL), nonswitched memory cells (9 cell/uL) and class-switched memory cells: (15 cells/uL). (4-62 cells/uL). Vaccine response was not pursued as patient had been on IVIG. Genetic testing was pursued, and revealed a mutation in PIK3CD gene, specifically a mutation in the c.2320G>A; p.Val774Met variant (rs370932461). This mutation though seen in databases, is not currently reported in medical literature as associated with this condition. Based on these, CT chest was ordered to screen for bronchiectasis, adenopathy and lymphoma. CT showed no cardiopulmonary disease or adenopathy, but did show an incidental adrenal mass which is now being worked up. While the pattern of inheritance of this mutation is autosomal dominant, her son is asymptomatic and testing of her son has not been pursued, though it was advised for her cousins given history of leukopenia. Patient has continued on IgG replacement therapy.

    Conclusion: Recent publication by the Clinical Immunology Society suggests consideration for next generation sequencing when it can affect future family planning or it has treatment and prognostic implications. This case highlights all aspects of the importance of genetic testing as part of the diagnosis of CVID, since it can affect progeny, it offers the possibility of treatment with immune modulating agents and has implications on screening, since patients are at increased risk for malignancies.

    (131) Submission ID#601396

    Rapid Identification of Patients with RAG Mutations Using Valpha 7.2 Antibody

    Kerry Dobbs, BSc1, Julie E. Niemela, MS, MLS2, Kenneth Olivier, MD, MPH3, Alexandra F. Freeman, MD4, Jenna Bergerson, MD/MPH5, Gregory M. Constantine, MD6, Sergio D. Rosenzweig, MD/PhD7, Luigi D. Notarangelo, MD, PhD8

    1Biologist, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    2Sequencing Laboratory, Team Leader, Immunology Service, Department of Laboratory Medicine, NIH Clinical Center, Bethesda, MD, USA

    3Chief, Pulmonary Branch, NHLBI, NIH, Bethesda, MD, USA

    4Director, Primary Immune Deficiency Clinic, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    5Staff Clinician, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    6Clinical Fellow, Allergy and Immunology, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    7Chief, Immunology Service, Department of Laboratory Medicine, NIH Clinical Center, Bethesda, MD, USA

    8Chief, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    Background: Abnormal V(D) J recombination activity in patients with mutations in the Recombination-Activating Genes 1 and 2 (RAG1/2) results in markedly reduced usage of distal V and J genes at the T cell receptor alpha (TRA) locus. Mucosa-associated invariant T (MAIT) cells express a semi-invariant T cell receptor containing the distal TRAV1-2 gene. MAIT cells can be identified by flow cytometry using a mAb directed against Valpha 7.2, which recognizes the product of the TRAV1-2 gene. By performing high throughput sequencing (HTS) of TRA rearrangements and flow cytometry, we have confirmed lack of T cells using distal Valpha genes in patients with known RAG mutations. We now report that flow cytometry with mAb against Valpha 7.2 successfully identified RAG deficiency in two patients with an atypical presentation.

    Methods: TRA rearrangements were analyzed by HTS using gDNA from sorted T cell subsets from RAG-mutated patients and healthy donors. Distal Valpha usage was measured in whole blood by flow cytometric analysis with an anti-Valpha 7.2 antibody. RAG mutations were detected by Sanger sequencing. Patients were enrolled in NIAID protocol 18-I-0041.

    Results: HTS of TRA rearrangements revealed lack of distal TRAV and TRAJ gene usage in patients with RAG1/2 mutations. The presence of circulating MAIT cells in controls and patients with known RAG1/2 mutations and various clinical phenotypes was analyzed by flow cytometry using mAb against Valpha 7.2. We found a virtual lack of Valpha 7.2 expression in RAG mutated patients (<0.5%) compared to controls (2-8%). We used the Valpha 7.2 assay to test two patients with unknown immunodeficiency manifesting as skin granulomas and autoimmune cytopenia, and found nearly absent expression (0.14% and 0.08%). Targeted sequencing of RAG1/2 revealed that both patients were compound heterozygous for RAG1 mutations: p.R112H/p.C328Y and p.R410W/p.R507Q, respectively.

    Conclusions: Patients with mutations in RAG1/2 demonstrate a skewing of their TCRalpha repertoire. The reduction in recombinase activity in these patients does not allow for rearrangements of the most distal Valpha segments. Rapid identification of patients lacking Valpha 7.2+ T cells by flow cytometry may prompt Sanger sequencing and identification of RAG1/2 mutations in a matter of days. This assay represents a simple but powerful tool to reduce the cost and time associated with other analysis methods.

    Acknowledgements: Supported by DIR/NIAID/NIH.

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    (132) Submission ID#601398

    Human STAT5 Deficiency Results in an Increase of Follicular T Cells Leading to Expanded Germinal Center B Cells and Autoimmunity

    Maria Soledad. Caldirola, MSc, PhD1, María Isabel. Gaillard, MSc2, Norberto Walter Zwirner, PhD3, Liliana Bezrodnik, MD4

    1Biochemist, Grupo de Inmunología Hospital de Niños "R.Gutierrez"- IMIPP-CONICET, Buenos Aires-Argentina

    2Biochemist, Grupo de Inmunología Hospital de Niños

    3Researcher, Instituto de Biología y Medicina Experimental (IBYME-CONICET), Laboratorio de Fisiopatología de la Inmunidad Innata, Buenos Aires, Argentina, Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires,

    4Director, Centro de Inmunología Clínica Dra.Bezrodnik y equipo

    Introduction: The fate of effector T cells is strongly dependent on the expression of Bcl-6 or Blimp-1, which are inhibited reciprocally through a complex signaling pathway. Several studies have shown that Bcl-6 is a key transcription factor for differentiation towards the follicular helper T cells (Tfh) lineage able to collaborate with B lymphocytes (BL). On the contrary, the transcription factor Blimp-1 is highly expressed in T lymphocytes Th1, Th2 and Treg, thus regulating the differentiation towards Tfh. Materials and methods: whole fresh blood and peripheral mononuclear cells from a patient with homozygous mutation in STAT5b were analysed by flow cytometry. Analysis of cTfh (CD4+CD45RA-CXCR5+), cTfh1 (CXCR3+), cTfh17 (CCR6+), cTfh2 (CXCR3-CCR6-), naïve BL (LB IgM+IgD+CD27-), memory (MBL) (LB IgM+ IgD- CD27+), switched (MBL-Sw) (IgD-IgM-) and plasmablast (PBC) (CD27+CD38++) cells was performed. Immunoglobulins were measured in serum. Results: the patient with STAT5b deficiency showed increased values of cTfh (38%) (Healthy donors p10-p90: 7,9-17,8 %) that presented an activated phenotype (ICOS+ and PD-1+) with a skewed to a Th17 profile (CCR6+), consistent with her hipergammaglobulinemia and the marked and sustained increase in the switched MBL and PBC subpopulations in peripheral blood over the years. Discusion: This immunological phenotype described in the patient with STAT5b deficiency could explain in part the pathophysiology of the autoimmune disorders. This patient (as well as the other two patients with mutations in STAT5b previously described by our group), have had chronic hypergammaglobulinemia, autoantibodies and consequently autoimmune processes (psoriasis, hypothyroidism, eczema, alopecia and celiac disease, among others). We believe that the link between this clinical symptomatology and the molecular defect relies in the fact that the absence of STAT5b promotes a greater expression of Bcl-6, which generates a bias towards the production of cTfh cells, that give rise to a greater activation of LB, generation of LBM and plasma cells (dysregulation in the CG), events that manifest as hypergammaglobulinemia and autoimmunity. In summary, we provide promising evidence of the mechanisms that lead to autoimmunity in this type of patients that could also be a consequence of the defect in the regulation of GC, highlighting the crucial role of STAT5b in the humoral immune response and maintenance of the tolerance of the immune system.

    (133) Submission ID#601403

    The First Year: Experience from Mayo Clinic Laboratories After Clinical Implementation of Nine Primary Immunodeficiency Next Generation Sequencing Tests

    Michelle L. Kluge, M.S., LCGC1, Susan A. Lagerstedt, BS2, Laura J. Train, BS3, Linda Hasadsri, MD, Ph.D.4, Ann M. Moyer, MD, Ph.D.4

    1Genetic Counselor, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN

    2Development Technologist Coordinator, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN

    3Development Technologist II, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN

    4Laboratory Director, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN

    Background/Introduction: The term primary immunodeficiencies (PID) encompasses a phenotypically and genetically diverse group of conditions. Genetic testing for these conditions can guide treatment, reduce morbidity and mortality, allow for genetic counseling, and identification of additional at-risk family members. However, this testing can be complicated by a number of factors, including pseudogenes, high homology, methodology limitations, and the heterogeneous nature of PIDs.

    Methods: Mayo Clinic Laboratories launched their first set of nine PID next generation sequencing (NGS) tests approximately one year ago. These tests include one single gene assay for GATA2 deficiency and eight targeted next generation sequencing panels for: atypical hemolytic uremic syndrome (aHUS), autoinflammatory disorders, B-cell disorders, monogenic irritable bowel disease (IBD), phagocytic defects, severe combined immunodeficiencies (SCID), and severe or cyclic neutropenia. Herein we summarize our first year of experience with these NGS tests, with a focus on the eight targeted panel tests.

    Results: From March 2018 through November 2018 we performed testing for 341 cases. Our highest volume of tests was for the aHUS panel (127/341 cases, 41%). A variant was reported in 76/341 cases (22.29%). These variants included variants of uncertain significance, likely pathogenic variants and pathogenic variants. The indication with the highest percentage of cases where a variant was reported was SCID (9/13 cases, 69.23%). The number of cases that were considered solved, where the genotype likely explains the patients phenotype, varied widely by indication. Twenty cases were found to have a pathogenic or likely pathogenic variant or variants; however 2/20 cases were heterozygotes for an autosomal recessive condition and were not considered solved cases. The panel with the highest percentage of solved cases is our SCID panel (4/13 cases, 30.77%). Conversely, we have yet to solve an autoinflammatory, irritable bowel disease, or telomere defects case; however ~20% of cases in each of those three panels have had a variant of uncertain significance reported. We hypothesize that one of the reasons for the low detection rate for these three panels is inappropriate test orders. We are also actively looking for ways to update all 8 panels to increase detection rates and clinical utility, for example expanding the gene list of our IBD panel, including large deletion/duplication detection, and including NCF1, a difficult gene to capture by NGS, on the phagocytic panel. Finally, we present the molecular findings from a number of interesting cases that were solved using our targeted NGS panels.

    Conclusions: The launch of our PID NGS tests in March of 2018 has allowed us to aid patients by confirming diagnoses and providing molecular diagnoses that will enable more accurate genetic counseling and risk assessment. We have also uncovered areas for improvement, both on the clinical side: provider education is important to enable better identification of patients who can benefit from molecular genetic testing for PIDs, and on the laboratory side: introduction of more expanded panels and additional methodologies.

    (134) Submission ID#601405

    Treatment-resistant Autoimmune Cytopenias as a Sign of Primary Immunodeficiency Disorders (PIDs)

    Sara Ciullini Mannurita, MSc1, Enrico Attardi, MD2, Ebe Schiavo, PhD1, Maria Luisa Coniglio, MSc3, Maddalena Bagni, MD2, Marina Vignoli, PhD1, Fabio Tucci, MD4, Marinella Veltroni, MD4, Claudio Favre, MD5, Eleonora Gambineri, MD6

    1Research fellow, University of Florence, Dep. Neurosciences, Psychology, Drug Research and Child Health (NEUROFARBA), Florence, Italy

    2Haematology trainee, AOU Careggi, Haematology Unit, University of Florence, Florence, Italy

    3Biologist, Anna Meyer Children's Hospital, Department of Haematology-Oncology, Florence, Italy

    4Medical doctor, Anna Meyer Children's Hospital, Department of Haematology-Oncology, Florence, Italy

    5Head of Department, Anna Meyer Children's Hospital, Department of Haematology-Oncology, Florence, Italy

    6Associate Professor, Anna Meyer Children's Hospital, Department of Haematology-Oncology - Bone Marrow Transplantation BMT Unit, University of Florence, Dep. Neurosciences, Psychology, Drug Research and Child Health (NEUROFARBA), Florence, Italy

    The progressive decrease of red blood cells, platelets or neutrophils via a self-directed immune process is jointly termed as autoimmune cytopenias. While autoimmune cytopenias, including autoimmune hemolytic anemia (AIHA), immune thrombocytopenic purpura (ITP), and autoimmune neutropenia (AN), are a common presentation of autoimmunity in the general population, they are particularly frequent and can appear as the first sign in patients with primary immunodeficiencies (PIDs). Possible causes of cytopenia in PIDs comprise mainly immune dysregulation, bone marrow failure (BMF) and myelodysplasia.

    Our goal is to investigate possible immune mediated mechanisms underlying chronic cytopenia in children in order to achieve an early diagnosis and consequently offer timely and appropriate therapy.

    We selected 24 patients affected by chronic cytopenia, evaluated with immunophenotyping by flow-cytometry; data were subjected to multivariate analysis by Principal Component Analysis (PCA). Next Generation Sequencing (NGS) analysis of genes frequently implicated in PIDs was performed.

    Among the patients, 5 were affected by bone marrow failure, of which 2 were diagnosed with Fanconi Anemia and severe congenital neutropenia; 12 were affected by immune-mediated cytopenia and 7 by idiopathic cytopenia. The immunephenotyping showed a typical pattern of CD8 T cell subpopulations expression in patients compared with healthy donors with an increase of naïve T cells and a reduction of central memory (CM) and effector memory (EM) T cells levels. We observed a decrease in total B cells, B switched and B memory cells and an increase in CD21low cells. PCA showed an overlap between groups, however it revealed a peculiar trend of some single patient, suggesting the pathway involved in immune defect.

    Preliminary results from NGS studies revealed genetic variations in genes previously associated with PIDs in 10 out of 11 patients investigated. In particular we identify one patient with a mutation in FAS, one with a mutation in AIRE and one with a mutation in IKAROS. Concerning the remaining patients further studies are ongoing to validate the pathogenicity of the genetic variations.

    PCA is a very effective tool to analyze several parameters at the same time, highlighting patients whose phenotype shows the main peculiarities. The presence of specific lymphocyte subpopulation patterns can be important indicators of immune-mediated cytopenias and helpful signs of specific PIDs that should promptly be investigated with genetic analysis.

    (135) Submission ID#601434

    What We Are Missing with PID Exomes, Including Poorly Covered Exons

    Sarah E. Henrickson, MD, PhD1, Manish Butte, MD, PhD2

    1Attending Physician, The Children's Hospital of Philadelphia, Divsion of Allergy Immunology

    2Division of Allergy/Immunology Chair, Division of Immunology, Allergy, and Rheumatology, Dept. of Pediatrics and Jeffrey Modell Diagnos-tic and Research Center, University of California, Los Angeles

    The rapid of discovery of novel, monogenic primary immunodeficiencies has been made possible by the broad availability of clinical whole exome sequencing (WES). However, clinical WES has major shortcomings that should be understood by practicing immunologists. Focusing on the 2017 IUIS list of ~330 monogenic primary immunodeficiency genes, we show here limitations in coverage that could significantly impact clinical interpretation. On the Agilent Whole Exome capture kit, the most common WES platform, there are a number of genes with exons that are poorly covered. Specifically, there are at least 94 genes with less than 100% exonic coverage, 26 with less than 99% coverage and 5 with less than 90% coverage (e.g. IKBKB, NCF1, TACI, UNC93B1 and TBX1). Beyond this challenging technical issue, there are more subtle issues as well. These include the presence of pseudogenes in at least 17 of our genes (e.g. AK2, C1QBP, CD46, CFTR, CR2, MSN, NCF1, NCSTN, IKBKG, NHP2, PMS2, PTEN, RNASEH2C, RPS, SBDS and WAS), which can make accurate sequencing very challenging. Finally, there are many known causative intronic (e.g. BTK, CTLA-4, WASP) and copy number variant mutations (e.g. RAG1 and XIAP) as well as large deletions (e.g. DOCK8) that we cannot expect to be optimally covered using WES. This list of genes requires consideration even with a negative exome and may require additional approaches including Whole Genome Sequencing, Sanger sequencing, CNV arrays and/or long-read NGS sequencing. WES is a powerful genomic diagnostic tool, but to avoid missing key diagnostic insights using these alternative approaches may be critical when certain genes are in the differential diagnosis. Going forward, as PID phenotypes continue to broaden, these issues remain fundamentally important even if these genes are not obviously implicated in a given clinical phenotype.

    (136) Submission ID#601465

    Recurrent Viral Encephalitis in a Child with Several Variants of Uncertain Significance in Primary Immunodeficiency Genes

    Sara Sussman, MD1, Zoya Treyster, MD2, Artemio M. Jongco, III, MD, PhD, MPH3

    1Fellow, Department of Pediatrics, Zucker School of Medicine at Hofstra Northwell School of Medicine

    2Fellow, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    3Assistant Professor of Medicine and Pediatrics, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell; Center for Health Innovations and Outcomes Research, Feinstein Institute for Medical Research, Manhasset, NY

    More physicians are utilizing targeted genetic panels to reach a definitive diagnosis for their patients with immunodeficiency. However, this increase in testing also has led to the discovery of many more variants of uncertain significance (VUS) in the genes tested. These findings can often leave the patient and the physician with more questions than answers. We present a patient with recurrent infections found to have multiple variants of uncertain significance in several genes associated with primary immunodeficiency.

    A 13-year-old female who was diagnosed with Crohns disease at age 9 after intestinal perforation and jejunal resection experienced two discrete episodes of Epstein Barr Virus (EBV) meningoencephalitis and septic shock. The first episode was diagnosed when patient had fever and altered mental status and occurred prior to her Crohns disease diagnosis and the second episode was complicated with altered mental status, disseminated intravascular coagulation (DIC) and hypotension requiring PICU admission. Aside from these two major infections, the family denied any other infections requiring antibiotics in the last 5 years and reported a remote history of repeated streptococcal pharyngitis that have not recurred. Immunology was consulted at the time of the second episode of meningoencephalitis and work up was mainly unremarkable with normal immunoglobulins, adequate vaccine response to Hib, tetanus, diphtheria, rubella, measles and pneumococcus (18 out of 22 protective titers). She had normal T cell numbers with slightly decreased natural killer numbers for age. Neutrophil studies showed normal Dihydrorhodamine (DHR) analysis, glucose-6-phosphate dehydrogenase levels and Myeloperoxidase (MPO) stain. Commercial testing of her Toll like Receptors (1-8) showed normal function. Invitae Primary Immunodeficiency Panel demonstrated a heterozygous variant in NOD2 (c2.104C>T; p.Arg702Trp) as well as heterozygous variants of uncertain significance in IL7R (c.662G>T; p.Ser221Ile) and TLR3 (c.889C>G; p.Leu297Val). The patients NOD2 variant is known to be associated with an increased risk for Crohns disease.

    Even with our patients presentation with recurrent severe viral infections and IBD, it is not immediately clear how these genetic results explain the pathology. Innate immune defects probably contribute to her presentation and it is currently unclear if and how the combination of multiple genetic variants has left her immunologically vulnerable. We use this case to demonstrate that even when genetic testing does not elucidate a clear-cut diagnosis of primary immunodeficiency, it can still provide helpful insight into a patients underlying immune phenotype.

    (137) Submission ID#601470

    Quercetin Halts Abnormal IL-1beta and IL-18 Production: A Natural Calm for XIAP Deficiency?

    Erika Owsley, B.S.1, Vijaya Chaturvedi, B.S.1, Michael Jordan, MD2, Catherine Terrell, B.S.1, Parinda Mehta, MD3, Rebecca A. Marsh, MD4

    1Research Associate, Cincinnati Children's Hospital

    2Professor of Pediatrics, Divisions of Immunobiology, and Bone Marrow Transplantation and Immune Deficiency Cincinnati Children's Hospital

    3Professor, University of Cincinnati

    4Associate Professor, Division of Bone Marrow Transplantation and Immune Deficiency, Cincinnati Childrens Hospital Medical Center

    Introduction: XIAP Deficiency is a rare primary immune deficiency characterized by hemophagocytic lymphohistiocytosis, recurrent fever and inflammatory syndromes, inflammatory bowel disease, hypogammaglobulinemia, recurrent infections, and other manifestations. Loss of XIAP results in abnormal TNF receptor signaling and NLRP3 inflammasome actvity which leads to dysregulated production of IL-1beta and IL-18. We hypothesized that suppressing the NLRP3 inflammasome with either targeted deletion or pharmacologic inhibition would suppress abnormal production and secretion of inflammatory IL-1beta and IL-18.

    Methods: Bone marrow derived macrophages (BMDMs) from control, XIAP-deficient, and XIAP and NLRP3 double knock-out mice were derived with 1 week of culture in L929-cell conditioned media. BMDMs were stimulated with a variety of TLR agonists or TNF-alpha, with or without a variety of inhibitors including the NLRP3 inhibitor MCC950, the cathepsin B inhibitor CA-074, and quercetin, which is a natural flavonoid (antioxidant) found in many fruits and vegetables, and available as a nutritional supplement. IL-1beta, IL-18, and TNF-alpha were measured in supernatants by ELISA, and cell death was evaluated by flow cytometry using PI exclusion.

    Results: As expected, BMDMs from XIAP deficient mice had markedly increased TLR-agonist- or TNF-alpha-induced IL-1beta production compared to normal BMDMs. Genetic deletion of NLRP3 and the pre-treatment of cells with the NLRP3 inhibitor MCC950 greatly reduced abnormal IL-1beta production; residual production of IL-1beta could be inhibited by caspase-8 inhibition. Pre-treatment of cells with the cathepsin B inhibitor CA-074 also decreased cytokine production but was toxic at higher concentrations. Quercetin reliably abrogated IL-1beta, and also IL-18. Quercetin was found to inhibit priming of the NLRP3 inflammasome (decreased upregulation of pro-IL1beta and NLRP3) and also decreased TNF-alpha secretion following TLR agonist stimulation.

    Conclusion: Quercetin suppresses the NLRP3 inflammasome and may be a promising therapeutic option for patients with XIAP deficiency. It prevents IL-1beta and IL-18 secretion. It is a particularly appealing option given that it is a naturally occurring antioxidant, has a great safety profile, and is readily available as a nutritional supplement. Human studies are needed.

    (139) Submission ID#601493

    Single Cell RNAseq Analysis Reveals Profound Abnormalities in the Distribution and Diversity of Thymic Epithelial Cells in Wild-type and Rag1 Mutant Mice

    Francesca Pala, PhD1, Andrew Martins, PhD2, John Tsang, PhD3, Luigi D. Notarangelo, MD, PhD4, Marita Bosticardo, PhD5

    1Post-Doctoral Fellow, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    2Research Fellow, LABORATORY OF IMMUNE SYSTEM BIOLOGY, NIAID, NIH

    3Investigator, LABORATORY OF IMMUNE SYSTEM BIOLOGY, NIAID, NIH

    4Chief, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    5 Staff Scientist, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    Background: The thymus contains a heterogeneous population of stromal cells which orchestrate T cell development and selection. Recently, single cell RNA sequencing (scRNAseq) analysis in mice has disclosed an unexpected complexity of thymic stromal cells, and medullary thymic epithelial cells (mTECs) in particular. However, the developmental origin, hierarchy, and function of these subpopulations remain ill-defined. Moreover, although cortical TECs (cTECs) are thought to represent a more homogeneous population, their characterization has been largely restricted to the adult thymus. We have previously shown that impaired lymphostromal cross-talk in the thymus of patients with combined immunodeficiency (and of corresponding mouse models) is associated with abnormalities of thymic architecture and TEC maturation. Here, we sought to compare TEC distribution and gene expression in wild-type (WT) and in mice carrying Rag1 hypomorphic mutations observed in patients with combined immune deficiency and immune dysregulation.

    Methods: Multi-color flow cytometry and scRNAseq were used to analyze composition and distribution of cTEC and mTEC subpopulations in WT and Rag1 mutant mice at various weeks of age (NIAID animal protocol: LCIM-6E).

    Results: We observed that Rag1 mutant mice have an excess of cTECs, and that their mTEC compartment is predominantly represented by cells with high levels of MHC class II (MHC-II) expression, recapitulating the phenotype of neonatal WT thymi. While MHC-IIhi mTECs are thought to represent a minor fraction of mTECs in adult WT mice and include mature AIRE+ cells, a relative abundance of MHC-IIhi mTECs is observed also at neonatal age, where they are thought to represent immature mTECs. To define more precisely TEC maturation, we performed scRNAseq on sorted CD45- EPCAM+ cells, and identified 8 and 10 distinct clusters of TECs in WT and Rag1 mutant mice, respectively. A large proportion of cells in Rag1 mutant mice could be ascribed to the cTEC compartment, confirming our previous flow cytometry and histopathology results. Furthermore, scRNAseq analysis also disclosed a different distribution of mTEC subsets in WT and Rag1 mutant mice. To address the hypothesis that this difference in cTEC and mTEC abundance and subset distribution may reflect different maturation stages in TEC development in WT and Rag1 mutant mice, we will perform lineage tracing and transplantation experiments, and we will also extend TEC scRNAseq analysis to WT and mutant mice of embryonic and neonatal age. In parallel, to evaluate the contribution of thymocyte maturation in shaping the stromal populations, scRNAseq will be performed on thymocytes.

    Conclusions: We have further refined the complexity of TECs, and shown that impaired development of T cells in combined immune deficiency (as exemplified by Rag1 mutant mice) has profound effects on the composition and maturation of TECs and may thus contribute to abnormalities of immune tolerance that are often associated with these conditions.

    (140) Submission ID#601506

    Multigenicity of the Deficit of the Immune System: Novel Frontiers of Primary Immunodeficiencies (PIDs)

    Ebe Schiavo, PhD1, Sara Ciullini Mannurita, MSc1, Maria Luisa Coniglio, MSc2, Enrico Attardi, MD3, Marina Vignoli, PhD4, Laura Luti, MD5, Annalisa Tondo, MD6, Ilaria Fotzi, MD6, Elena Chiocca, MD6, Fabio Tucci, MD6, Marinella Veltroni, MD6, Claudio Favre, MD7, Eleonora Gambineri, MD8

    1Research fellow, University of Florence, Dep. Neurosciences, Psychology, Drug Research and Child Health (NEUROFARBA), Florence, Italy.

    2Biologist, Anna Meyer Children's Hospital, Department of Haematology-Oncology, Florence, Italy.

    3Haematology trainee, AOU Careggi, Haematology Unit, University of Florence, Florence, Italy.

    4Research fellow, University of Florence, Dep. Neurosciences, Psychology, Drug Research and Child Health (NEUROFARBA), Florence, Italy.

    5Medical doctor, Pediatric Hematology Oncology, Bone Marrow Transplant, S. Chiara Hospital, Pisa, Italy.

    6Medical doctor, Anna Meyer Children's Hospital, Department of Haematology-Oncology, Florence, Italy.

    7Head of Department, Anna Meyer Children's Hospital, Department of Haematology-Oncology, Florence, Italy.

    8Associate Professor, Anna Meyer Children's Hospital, Department of Haematology-Oncology - Bone Marrow Transplantation BMT Unit, University of Florence, Dep. Neurosciences, Psychology, Drug Research and Child Health (NEUROFARBA), Florence, Italy.

    The advent of next-generation sequencing (NGS), with the development of whole-exome sequencing (WES) in particular, has allowed the identification of unknown genetic lesions for many diseases and the implementation of specific therapeutic strategies. Primary immunodeficiencies (PIDs) are a group of rare diseases which have benefited from NGS, with the discovery and molecular characterization of previously genetically undefined diseases and the identification of novel molecules involved in the regulation of the immune system.

    PIDs are often associated with autoimmune disease due to the dysregulation of the immune system as a whole. The clinical phenotypes are heterogeneous and often overlapping. While a monogenic cause of disease has been identified in a most subsets of patients, the recent application of whole-genome sequencing has found that a polygenic cause is likely.

    Our aim is to investigate the genetic background of patients with immunedysregulations and autoimmunity and to evaluate the possible pathogenicity of the identified gene variants through extensive functional studies.

    We select 19 patients with sign of immunedysregulation and autoimmunity, extended immunophenotyping and next-generation sequencing (NGS) analysis of 50 genes frequently implicated in PIDs was performed.

    In six of them we identify a single gene as responsible of the clinical feature. In particular, we identify two patients with gain of function mutation in STAT3, one patient with a mutation in CTLA4, one patient with an activating PIK3CD mutation, one with a RAG1 mutation and one with a FAS mutation. In most of them variants in multiple genes have been detected. Interestingly, we find that some genes are recurrently mutated in more then one patient such as WAS, DOCK8, CASP10, CASP8, NFATC2 and FCGR3A. Further studies are ongoing to validate the effect of the variations identified.

    Our results strongly suggest that the old hypothesis, based on a single gene mutation as a cause of illness, should be revised in favor of the concept that "is the sum that causes the effect" and that a different point of view on PIDs now seems inevitable.

    (141) Submission ID#601507

    Dose and Clinical Outcomes in Patients with CVID and Bronchiectasis Receiving Immunoglobulin Replacement Therapy in the Home

    Allyson Checkley, PhD1, Loretta Kristofek, BSN, RN2, William Bolgar, PharmD3, Luqman Seidu, MD4

    1Research and Registry Program Advisor, Coram CVS Specialty Infusion Services

    2Clinical Service Liaison, Coram CVS Specialty Infusion Services

    3VP Clinical Operations, Coram CVS Specialty Infusion Services

    4Physician, Omni Allergy, Immunology, and Asthma

    Introduction/Background: Immunoglobulin replacement therapy (IGRT) may be optimized to reduce the severity and incidence of infections and potentially delay or abrogate the development of pulmonary complications of primary immune deficiencies. Pulmonary complications including bronchiectasis are common in common variable immune deficiency (CVID) and contribute significantly to morbidity and mortality in these patients. It remains unclear whether continued obstructive bronchial changes are a result of repeated respiratory infections, associated inflammation and immune dysregulation, or simply lung-damage that is irreversible by the time therapy is initiated. It has also been suggested that under-treatment in addition to the diagnostic delay may contribute to the development of bronchiectasis in patients with PID. Lower serum IgG levels with any given dose of immunoglobulin replacement therapy have been demonstrated in patients with bronchiectasis compared to those PID patients without this complication. In addition, earlier studies have shown that greater doses of Ig (600 mg/kg/ month) may reduce the frequency and duration of infections and help prevent or slow progression of chronic lung disease.

    Objective: To evaluate the prevalence of bronchiectasis in a cohort of patients with a diagnosis of CVID and identify associated Ig dosing patterns and clinical outcomes.

    Methods: Data were analyzed from the IDEaL (Immunoglobulin, Diagnosis, Evaluation, and key Learnings) Patient Registry. This is a prospective, longitudinal registry study of patients receiving Ig replacement therapy in the home or ambulatory infusion suite with one national home infusion provider. Nursing and pharmacy standard of care forms were collected, and dose, infection rate, and prevalence of bronchiectasis were evaluated in patients with a diagnosis of CVID (ICD-10 codes: D83.9, D83.1)

    Results: There were 310 patients in the Registry with CVID, 14 (4.5%) of which bronchiectasis was also observed. Seventy-nine percent (n=246) of the study population was female, and 50% (n=7) of the cases of bronchiectasis were observed in females. The mean age of the patients with concurrent bronchiectasis was 65±15.8 at start of care compared to 57±15.8 in those without this observed bronchial obstruction. Most bronchiectasis patients (n=11) received IGRT subcutaneously every week with a mean dose of 123.8±22.8 mg/kg/wk. The mean dose of Ig in the 3 remaining patients receiving Ig intravenously was 506.8±82.0 mg/kg/month. The average annual rate of infection in IVIG and SCIG patients with bronchiectasis was 1.6±1.0 and 2.2±1.3, respectively, however many were serious bacterial infections. At time of analysis, 7 of the bronchiectasis patients remained active in the registry and 7 had withdrawn. Reasons for withdrawal included stopping IGRT due to the following: patient decision (n=3), physician decision (n=1) insurance change (n=1), and patient expired (n=2).

    Conclusions: There were 14 documented cases of bronchiectasis in our cohort of CVID registry patients, and dosing patterns aligned with standard doses despite the presence of bronchial obstruction. Further studies are necessary to assess evolution of lung damage with respect to Ig dosing in patients with CVID.

    (142) Submission ID#601511

    NGS Reveals Repertoire Restriction of Treg Cells in APDS1 Patients

    Ottavia M Delmonte, MD, PhD1, Riccardo Castagnoli, MD2, Stephen Daley, DVM3, Kerry Dobbs, BSc4, Marita Bosticardo, PhD5, Su Hua, PhD6, Gulbu Uzel, MD7, Luigi D. Notarangelo, MD, PhD8

    1Staff Clinician, 1 Laboratory of Clinical Immunology and Microbiology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD.

    2Research fellow, 1 Laboratory of Clinical Immunology and Microbiology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD.

    3Senior Research Fellow, Infection and Immunity Program, Monash Biomedicine Discovery Institute and Department of Biochemistry and Molecular Biology, Monash University, Melbourne, VIC, Australia

    4Biologist, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    5Staff Scientist, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    6Staff Scientist, Immunopathogenesis Unit, LCIM, NIAID, NIH

    7Staff Clinician, Laboratory of Clinical Immunology and Microbiology, National institute of Allergy and Infectious Diseases, NIH, Bethesda, MD, USA

    8Chief, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    Background: Activated phosphoinositide 3-kinase syndrome type 1 (APDS1) is a combined immunodeficiency resulting from gain-of-function (GOF) mutations in PIK3CD, the gene encoding the catalytic subunit of phosphoinositide 3-kinase (PI3K). This form of PID is characterized by recurrent respiratory tract infections, susceptibility to herpes virus infections, impaired antibody responses, lymphoproliferation and autoimmunity. Previous studies showed that patients with APDS1 have B cell defects that contribute to the clinical phenotype. Furthermore, these patients display T cell abnormalities, including increased numbers of memory T cells and T follicular helper cells (Tfh), reduction of naïve T cells and impaired T regulatory cell (Treg) function. Whether these T cell abnormalities are also associated with perturbations of T cell repertoire in unknown.

    Objective: We aimed to investigate the effects of increased PI3K signaling on the T-cell repertoire of patients with APDS.

    Methods: High throughput sequencing was used to study composition and diversity of T-cell receptor (TRA) and T-cell receptor (TRB) repertoire in sorted Treg, Tfh, conventional CD4+ (Tconv), and CD8+ T cells from 4 patients with PIK3CD GOF mutations and healthy controls.

    Results: Treg cells of patients with APDS1 show restriction of TRA and TRB repertoire diversity, and increased clonality. No repertoire restriction was detected in Tfh, Tconv, and CD8+ T cells from the same patients. However, the TRB repertoire of Treg and CD8+ cells was enriched for the presence of hydrophobic amino acids in position 6 and 7 of the CDR3, a biomarker of self-reactivity.

    Conclusion: These data demonstrate that the T-cell repertoire of patients with APDS1 is characterized by a molecular signature that may contribute to the increased rate of autoimmunity associated with this condition. Furthermore, our result support the notion that the PI3K pathway is a key regulator of Treg cell development and homeostasis in humans.

    figureal

    (143) Submission ID#601524

    Combined Immunodeficiency in a Patient with a Heterozygous TNFRSF13B (TACI) Variant

    Amanda V. Grippen Goddard, DO1, Mohini Pathria, MD1, Flavia Hoyte, MD2, Rafeul Alam, MD, PhD3

    1Allergy/Immunology Fellow, National Jewish Health

    2Associate Professor, National Jewish Health

    3Professor, National Jewish Health

    Introduction/Background: Monogenetic defects are responsible for approximately 10% of cases of common variable immunodeficiency disorder (CVID). In most cases of CVID, there is a polygenic mode of inheritance. Variants in TNFRSF13B (TACI) are seen in 7-10% of CVID patients, but also in asymptomatic relatives and in about 1-2% of normal individuals who are heterozygotes. This highlights the incomplete penetrance of TACI mutations and suggests that heterozygous mutations increase the risk, but are neither necessary nor sufficient to cause CVID. TACI mutations are known to mainly result in defects in B-cell switching and differentiation; however, some of these patients are also found to have T-cell defects.

    Objectives: 1) Illustrate the polygenic nature of CVID. 2) Discuss predominantly T-cell immunodeficiency in a patient with a heterozygous TACI variant.

    Results: A 26-year-old, Mexican male, was diagnosed with Tuberculosis at the age of 6, shortly after moving to the United States. He received one year of antimycobacterial therapy. He continued to have frequent, but mild, respiratory infections that resulted in approximately 2 missed school days per month, but did not require any hospitalizations. At the age of 22, he was diagnosed with multi-lobar pneumonia, which progressed to sepsis and hypoxic respiratory failure requiring intubation. One year later, he developed a second episode of pneumonia with sepsis requiring ICU admission. Subsequently, an immunodeficiency evaluation revealed an IgA of 40 mg/dL (70-400), IgG of 667 mg/dL (700-1600), IgM of 31 mg/dL (60-300), protection against 7/14 streptococcal pneumoniae serotypes, and protective Tetanus and Diptheria IgG antibodies (2.4 and 11.5 IU/mL, respectively). Quantitative lymphocytes subsets were significantly abnormal. They showed a CD45 of 0.47 K/mcL (0.82-2.84), CD3 of 300 cells/mcL (550-2202), CD4 of 197 cells/mcL (385-1437), CD8 of 97 cells/mcL (199-846) and CD 19 of 26 cells/mcL (91-409). Immature B cells (CD38lo/CD21lo) were elevated at 13.9% (0.5-8.0). Lymphocyte proliferation in response to PHA, Con A and PWM was 4-10x lower in the patient as compared to a healthy control. He was subsequently started on intravenous immunoglobulin (IVIG) replacement therapy and continued this for 1 year without any infections. Due to an insurance related lapse of IVIG infusion for 8 months, he developed another multi-lobar pneumonia requiring hospitalization. He is currently back on monthly IVIG replacement therapy and doing well. Comprehensive genetic testing for 207 known primary Immunodeficiency genes revealed one likely pathogenic heterozygous variant, c.310T>C (p.Cys104Arg) in TNFRSF13B (TACI), and three additional variants of uncertain significance: c.31G>A (p.Asp11Asn) in RAC2, c.237G>C (p.Gln79His) in TRAF3IP2, and c.4720G>A (p.Ala1574Thr) in VPS13B.

    Conclusions: Features that stand out in our patient are 1). Recurrent bacterial infection in the setting of normal antibody response and mildly depressed IgA, IgM and IgG; and 2). Absence of viral and fungal infections despite markedly reduced CD4 and CD8 T cell number and proliferation. Although TACI mutations can result in B and T cell deficiencies, it is unlikely that a heterozygous TACI variant alone can explain this unusual clinical manifestation in our patient. We speculate that epistasis with other immune gene variant(s) contributed to this complex phenotype.

    (144) Submission ID#601532

    Diagnostic Challenges in Hospitalized Patients with Suspected Inborn Errors of Immunity in a Reference Center in the Southwest of Colombia

    Andres F. Zea-Vera, MD, PhD1, Fabio S. Vargas-Cely2, Vanessa Montoya-Lozano3

    1Assistant Professor, Universidad del Valle. Hospital Universitario del Valle.

    2Medicine Student, Universidad del Valle

    3Nursing Student, Universidad del Valle

    Introduction: The Hospital Universitario del Valle (located in Cali, Colombia) is a reference center for the low income people of the southwest of Colombia with a population of influence close to 4.5 million people. The low number of clinical Immunologist in Colombia represents a huge challenge for the field.

    Results: Since June 2016, sixty one (61) hospitalized patients have been evaluated by the Clinical Immunology service. The most common causes of consultation are recurrent infection syndrome, disseminated tuberculosis, hypergammaglobulinemia, refractory autoimmune disease and early age malignancies. The main diagnoses at admission were complicated pneumonia, meningitis and opportunistic infections in HIV negative patients.

    Patients were classified as Primary Immunodeficiencies (10 patients 16%), suspected Immunodeficiencies or patients in follow up after discharge (27 patients 44%) and secondary Immune disorders due to autoimmunity, malignancy, immunosuppressant therapy or chronic infections (24 patients 60%).

    According to the IUIS-2017 classification, 10 patients with confirm Inborn Errors of Immunity (PID) were diagnosed: II. CID with associated or syndromic features (4), III. Predominantly Antibody deficiencies (2), I. Immunodeficiencies affecting cellular and humoral immunity (1), VII. Auto-inflammatory disorders (2), IX. Phenocopies of PID (1). Two non related cases of Ataxia-Telangiectasia and one case of Schimke syndrome (SMARCAL1 compound heterozygous mutation) were diagnosed in the last year. We observed a wide range of age (we evaluate adult and pediatric population) with a Male:Female ratio close to 1:1

    Conclusions: The Hospital Universitario del Valle - Clinical Immunology inpatient service had increased the opportunity for the subsidiary health care system patients in Colombia. Interestingly our main PID group was combined Immunodeficiency with associated or syndromic features opposite to previous reports. Molecular and functional testing diagnosis is a growing necessity in Colombia. Future characterization of patients with PID is necessary to reduce complications.

    (145) Submission ID#601541

    Diagnostic Yield of a Next-Generation Sequencing Panel for Primary Immunodeficiencies in a Cohort of Pediatric Patients with Immunohematologic Disorders

    Elizabeth A. Varga, MS, LGC1, Kristin Zajo, MA, MS, LGC2, Melissa Rose, DO3, Benjamin Prince, MD, MSCI4

    1Genetic Counselor, Co-Director of Personalized Medicine, Division of Hematology/Oncology/BMT and the Institute for Genomic Medicine, Nationwide Childrens Hospital

    2Genetic Counselor, Division of Hematology/Oncology/BMT, Nationwide Children's Hospital

    3Director, Cytopenia and Bone Marrow Failure Program, Division of Hematology/Oncology/BMT, Nationwide Children's Hospital

    4Assistant Professor of Pediatrics, Division of Allergy-Immunology, Nationwide Children's Hospital,

    Immunohematologic disorders encompass a broad array of clinical conditions in which hematologic manifestations, usually cytopenias, are caused by aberrant immune responses. These often lead to the development of neutropenia, hemolytic anemia, or thrombocytopenia, either separately or in combination. Common underlying mechanisms include immunodeficiency, immune dysregulation, and systemic autoimmunity. Clinical diagnosis of these disorders is complicated by overlapping phenotypes.

    In April 2017, a 207-gene next generation sequencing (NGS) panel inclusive of copy number variation analysis was launched by a commercial laboratory to facilitate clinical diagnosis of primary immunodeficiency (PID), monogenic autoimmunity and autoinflammatory disorders. We assessed the outcomes of genetic testing utilizing this panel on a cohort of pediatric patients with immunohematologic phenotypes evaluated at our tertiary care center during an 18-month period (5/1/17-10/31/18). Eligible subjects were evaluated by at least two of three providers from a multidisciplinary pediatric hematology-immunology team, including a hematology physician, immunology physician and a geneticist or genetic counselor.

    Twenty-three patients met inclusion criteria; 20 (87%) were Caucasian, 12 (52%) were male with an average age of 11.7 years. The two most common phenotypic diagnoses included cytopenias, single- or multi-lineage (leukopenia, neutropenia, anemia, thrombocytopenia) primarily attributed to autoimmune causes or hypogammaglobulinemia. Five (22%) were given a definitive genetic diagnosis as a result of panel testing, though in two of these cases, the causative mutations were listed as variants of uncertain significance (VUS). Diagnoses included common variable immunodeficiency due to a pathogenic variant in NFKB2, STAT3 multiorgan autoimmunity due to gain-of-function mutation, and familial cold autoinflammatory syndrome due to a pathogenic mutation in NLRP12. Biallelic DNMT3B VUS were found in a patient whose phenotype and further laboratory studies (including karyotype) were consistent with immunodeficiency-centromeric instability, facial anomalies syndrome. Further, a STAT3 VUS was identified in a patient with multiorgan autoimmunity and his father with hypothyroidism; studies from an outside research laboratory were consistent with gain-of-function with this variant (private communication). An additional three patients had VUS identified that were suspected to be related to their phenotype, prompting eligibility for research studies. Four (17%) patients had increased risk alleles in NOD2, conferring an increased risk of Crohns disease. Three (13%) patients had pathogenic or likely pathogenic carrier findings warranting genetic counseling. In addition, 47 VUS (an average of 2 per patient) thought to be unrelated to phenotype were identified, necessitating further investigation and counseling.

    The use of an NGS panel in a cohort of pediatric patients with immunohematologic disorders led to a definitive diagnosis in 22% of previously undiagnosed patients and prompted further research investigation in several more. Genetic testing also led to the identification of clinically significant carrier findings, risk alleles and 47 VUS unrelated to phenotype, necessitating genetic counseling. Our experience illustrates the value of genetic testing for diagnosis of immunohematologic disorders, and the importance of multidisciplinary care, including genetic counseling, for the proper evaluation and management of these patients.

    (146) Submission ID#601557

    Reduced-intensity, T Cell-replete, Alternative Donor Allogeneic Hematopoietic Cell Transplantation with Post-transplantation Cyclophosphamide Is Safe and Effective for Primary Immune Deficiencies

    Orly R. Klein, MD1, Dimana Dimitrova, MD2, Ellen Carroll, RN3, Kenneth R. Cooke, MD4, Stephanie N. Hicks, RN5, Christopher G. Kanakry, MD6, Howard M. Lederman, MD, PhD7, Jennifer Sadler, RN5, Elias T. Zambidis, MD, PhD8, Heather J. Symons, MD, MHS9, Jennifer A. Kanakry, MD10

    1Instructor, Oncology and Pediatrics, Johns Hopkins University School of Medicine

    2Medical Officer, National Institutes of Health, National Cancer Institute, Experimental Transplantation and Immunology Branch

    3Transplant Coordinator, National Institutes of Health, National Cancer Institute, Experimental Transplantation and Immunology Branch

    4Professor, Oncology and Pediatrics, Johns Hopkins University School of Medicine

    5Research Nurse, National Institutes of Health, National Cancer Institute, Experimental Transplantation and Immunology Branch

    6Tenure-track Investigator, Lasker Clinical Research Scholar, National Institutes of Health, National Cancer Institute, Experimental Transplantation and Immunology Branch

    7Professor of Pediatrics, Medicine and Pathology, Division of Pediatric Allergy and Immunology at Johns Hopkins University School of Medicine

    8Associate Professor, Oncology and Pediatrics, Johns Hopkins University School of Medicine

    9Assistant Professor, Oncology and Pediatrics, Johns Hopkins University School of Medicine

    10Clinical Head of Transplant, National Institutes of Health, National Cancer Institute, Experimental Transplantation and Immunology Branch

    Background: Allogeneic hematopoietic cell transplantation (alloHCT) is curative for primary immune deficiencies (PID). However, many patients lack a fully-matched unaffected sibling, or may have an unknown underlying genetic defect, rendering it undesirable to use related donors. Many PID patients have significant comorbidities at the time they are referred to alloHCT, precluding the use of myeloablative conditioning. The use of alternative donors with reduced-intensity conditioning (RIC) has historically led to increased rates of graft failure, graft-versus-host disease (GVHD), and transplant-related mortality (TRM). Post-transplantation cyclophosphamide (PTCy) as GVHD prophylaxis immunomodulates the graft through the preferential sparing of regulatory T cells and hematopoietic stem cells from its cytotoxic effects, thus allowing for robust donor engraftment that overcomes the HLA barrier while effectively preventing severe acute and chronic GVHD. We report the outcomes of two institutions using a RIC alloHCT regimen with alternative donors and PTCy in patients with PID.

    Design: We transplanted 35 PID patients (Table 1) using alternative donors and RIC, either serotherapy-free (n=21) or alemtuzumab-based (n=14). All patients received PTCy for GVHD prophylaxis on days +3 and +4, either alone (n=3), or combined with mycophenolate mofetil and either sirolimus (n=21) or tacrolimus (n=11). Donors included haploidentical family members (n=16), matched unrelated (n=15), and mismatched unrelated (n=4). Stem cell source was T cell-replete bone marrow (n=33) or peripheral blood stem cells (n=2).

    Results: The median follow-up is 17 months (range 0.5-8 years). At 17 months, overall survival is 91%, and event-free survival (defined as alive without graft failure) is 83%. The median days of neutrophil and platelet engraftment are 17 (range 14-42) and 28 (range 15-110), respectively. There were 10 patients who developed acute GVHD, grade 1 (n=5) or grade 2 (n=5), and there were no cases of grade 3 or 4 aGVHD. Seven of eight patients treated with systemic corticosteroids responded, and one was corticosteroid-dependent, then responded to second-line therapy. One patient developed skin-only chronic GVHD, which responded to corticosteroids and PUVA light therapy. Five patients developed graft failure, either primary (n=1) or secondary (n=4), and four were successfully re-transplanted and remain engrafted. One patient with secondary graft failure had autologous recovery and has not required a second alloHCT given some durable infection control gained during initial engraftment. There were three deaths prior to day 180 due to infection, and one death at 1.5 years secondary to presumed overdose. In ongoing follow-up of engrafted survivors (n=30), evidence of phenotype reversal has been demonstrated in all patients, with complete or ongoing resolution of some or all of their underlying disease manifestations, including infection, transfusion-dependence, autoimmunity, malignancy, and/or immune dysregulation.

    Discussion: We have observed high rates of engraftment, low rates and severity of acute and chronic GVHD, and low TRM in 35 patients with PID transplanted using alternative donors, RIC, and PTCy-based GVHD prophylaxis. RIC alloHCT with PTCy shows promise for curing PID, and its use minimizes toxicity and widely expands the donor pool, thus allowing us to offer this curative therapy to many more patients with PID.

    Table 1. Patient and donor characteristics.

      Patients
    (n=35)
    Male, n (%) 25 (71%)
    Age at time of HCT in years, median (range) 13 (0.5-54)
    Diagnosis, n  
     Unknown primary immunodeficiency 8
     Chronic granulomatous disease 8
     MAGT1 deficiency 3
     PI3KCD gain of function 3
     RAG1/2 deficiency, hypomorphic 2
     XIAP deficiency 2
     STAT3 deficiency 1
     IFNGR1 deficiency 1
     CTLA4 haploinsufficiency 1
     IL10R1 deficiency 1
     NFKB1 haploinsufficiency 1
     NFKBIA gain of function 1
     Wiskott-Aldrich syndrome 1
     ADA2 deficiency 1
     IPEX syndrome 1
    Allograft type, n (%)  
     T cell-replete bone marrow 33 (94%)
     T cell-replete peripheral blood stem cells 2 (6%)
    Donor source, n (%)  
     HLA-haploidentical 16 (46%)
     HLA-matched unrelated 15 (43%)
     HLA-mismatched unrelated 4 (11%)

    Abbreviations: HCT, hematopoietic cell transplantation; IPEX, immunodysregulation polyendocrinopathy enteropathy X-linked; HLA, human leukocyte antigen

    (147) Submission ID#601564

    Generating a CYBB-KO THP-1 Cell Line Model for Studying Auto-inflammation in Chronic Granulomatous Disease

    Aissa Benyoucef1, Lorie Marchitto2, Fabien Touzot, MD, PhD3

    1Research Assistant, CHU Ste-justine

    2Master Student, CHU Ste-Justine

    3Department of Pediatrics, CHU Ste-Justine, University of Montreal, Montreal, QC, Canada

    Chronic granulomatous disease (CGD) is a primary immune disorder that involves mutations in the nicotinamide adenine dinucleotides (NADPH) oxidase complex (Deffert, Cachat, & Krause, 2014). Two-third of CGD cases are caused by loss-of-function mutations in the CYBB gene that encodes the gp91pox subunit of the NADPH. The increased in patients' life expectancy thanks to progress in diagnosis and management has underlined the burden of inflammatory manifestations occurring independently of infectious agents (Dunogue et al., 2017; Marciano et al., 2018). CGD patients develop inflammatory granulomatous disorders, notably colitis, as a consequence of a dysregulated inflammasome activation. The treatment of inflammatory manifestations remains challenging, as it can be associated with an increased risk of infections. Thus, understanding the pathophysiological mechanism of auto-inflammation in CGD could help improve the therapeutic arsenal for the management of these manifestations. To reveal the precise pathophysiological mechanism of auto-inflammation in CGD, we have developed a cellular model that reproduces the CGD phenotype in phagocytic cell. Through CRISPR-Cas9 gene-editing we generated a THP-1 cell line harboring the previously described mutation c.90_92delCCGinsGGT (p.Tyr30Ter) in the CYBB gene responsible for gp91phox knock-out by early termination of translation. This cell line recapitulates the phenotype of CGD phagocytes: (i) decreased H2O2 production (ii) and enhanced inflammatory responses after PMA stimulation as evidenced by increased IL-1, IL-6 and TNFa secretion levels (Kuijpers & Lutter, 2012). These features were rescued by complementation through lentiviral transduction of a wild type CYBB gene. This new model will help us to investigate the auto-inflammation reported in CGD patients and also to propose new therapeutic targets of inflammatory manifestations in this disorder.

    Deffert, C., Cachat, J., & Krause, K. H. (2014). Phagocyte NADPH oxidase, chronic granulomatous disease and mycobacterial infections. Cell Microbiol, 16(8), 1168-1178. doi:10.1111/cmi.12322

    Dunogue, B., Pilmis, B., Mahlaoui, N., Elie, C., Coignard-Biehler, H., Amazzough, K., . . . Lortholary, O. (2017). Chronic Granulomatous Disease in Patients Reaching Adulthood: A Nationwide Study in France. Clin Infect Dis, 64(6), 767-775. doi:10.1093/cid/ciw837

    Kuijpers, T., & Lutter, R. (2012). Inflammation and repeated infections in CGD: two sides of a coin. Cell Mol Life Sci, 69(1), 7-15. doi:10.1007/s00018-011-0834-z

    Marciano, B. E., Zerbe, C. S., Falcone, E. L., Ding, L., DeRavin, S. S., Daub, J., . . . Holland, S. M. (2018). X-linked carriers of chronic granulomatous disease: Illness, lyonization, and stability. J Allergy Clin Immunol, 141(1), 365-371. doi:10.1016/j.jaci.2017.04.035

    (148) Submission ID#601588

    Genetic Testing Reveals a Homozygous RTEL1 Mutation in a 12 Month Old Female with Pancytopenia, Failure to Thrive and Low Immunoglobulins

    Miriam Samstein, MD, PhD1, Idil Ezhuthachan, MD1, Sherry Farzan, MD2, Artemio M. Jongco, III, MD, PhD, MPH3

    1Fellow, Northwell Health

    2Attending, Northwell Health

    3Assistant Professor of Medicine and Pediatrics, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell; Center for Health Innovations and Outcomes Research, Feinstein Institute for Medical Research, Manhasset, NY

    Introduction/background: Dyskeratosis congenita (DC) is a disease of short telomeres characterized by bone marrow failure, immune dysfunction and somatic abnormalities including abnormal skin pigmentation, nail dystrophy and oral leukoplakia. Several genes have been implicated in the pathogenesis of DC including RTEL1 and ACD. Inheritance patterns include autosomal dominant, autosomal recessive and X-linked. Successive generations of affected individuals can be born with progressively shorter telomeres. While classic presentations include skin findings, bone marrow failure can be the first presenting sign. Hoyeraal-Hreidarsson Syndrome is a severe form of DC with symptoms beginning in early childhood and including microcephaly, cerebellar hypoplasia, intrauterine growth retardation (IUGR), severe immunodeficiency and early-onset progressive bone marrow failure.

    Objectives: Understand the clinical features of DC and Hoyeraal-Hreidarsson syndrome

    Describe a novel mutation leading to a DC like phenotype

    Highlight the utility of genetic testing for severely ill patients with immune disorders

    Methods: This is a case presentation of a patient with immunodeficiency and bone marrow failure found to have a homozygous variant of uncertain significance (VUS) in RTEL1 on genetic testing with the Invitae 207 gene primary immunodeficiency panel.

    Results: The patient is a female born at 38 weeks as the product of a pregnancy complicated by IUGR. Zika testing, head ultrasound and NY state newborn screen were normal. She began to fall off the growth curve and had watery diarrhea at age 4 months despite high calorie formula implementation. At age 9 months she began having low grade fevers and was found to have a hemoglobin of 4 prompting urgent hospital admission for transfusion. Hematology sent testing for bone marrow failure syndromes, with suspicion for Diamond Blackfan anemia. She required PRBC transfusion roughly once every 2-3 weeks. At 11 months of age she became pancytopenic and had an urgent hospital admission. Immunology and GI were first consulted at this time. Due to persistently poor weight gain GI performed a colonoscopy which revealed IBD. Initial immune evaluation revealed hypogammaglobulinemia (IgG = 82 mg/dL, IgA = 7 mg/dL, and IgM = 32 mg/dL), nonprotective titers to HiB, pneumococcus, and diphtheria despite vaccination, and the patient received IVIG. She was also panlymphopenic (CD3 = 1092, CD4 = 837, CD8=253, CD19 = 31, CD16/56=7). Invitae primary immune deficiency panel demonstrated a homozygous variant of uncertain significance in RTEL1 (c.1742T>C p.Leu581Pro) as well as a heterozygous mutation in ACD c.262G>C (p.Ala88Pro), CARD14 (c.1192G>C (p.GLu398Gln), EPG5 (c.4231C>A(p.Leu1411Ile). She was also heterozygous for a pathogenic variant of SLC37A4 (c.1043delCT(p.Leu348Valfs*53).

    Conclusions: In appropriately selected patients genetic testing can shed light on previously unidentified immune deficiencies. Although the clinical features of DC were present in this patient, the rarity of Hoyeraal-Hreidarsson syndrome makes it a difficult to make diagnosis and these patients are typically diagnosed with other bone marrow failure syndromes or idiopathic anemia. Prompt evaluation by an immunologist for babies with failure to thrive and bone marrow failure is warranted.

    (149) Submission ID#601601

    Chronic Rhinosinusitis and Nasal Polyposis in Patients with IRAK-4 Deficiency

    Sara Seghezzo, MD1, Lauren Sanchez, MD2, Dylan K. Chan, MD, PhD3, Kristina W. Rosbe, MD4, Morna J. Dorsey, MD, MMSc5

    1Clinical Fellow, Department of Pediatrics, Division of Allergy, Immunology, and Bone Marrow Transplant, University of California, San Francisco

    2Assistant Clinical Professor, Department of Pediatrics, Division of Allergy, Immunology, and Bone Marrow Transplant, University of California, San Francisco

    3Assistant Professor, Clinical Professor, Department of Pediatric Otolaryngology and Pediatrics, University of California San Francisco, San Francisco, CA

    4Clinical Professor, Department of Pediatric Otolaryngology and Pediatrics, University of California San Francisco, San Francisco, CA

    5Pediatric Immunologist and Allergist, Department of Pediatrics, Division of Allergy, Immunology, and Bone Marrow Transplant, University of California San Francisco, San Francisco, CA

    Introduction: Homozygous or compound heterozygous mutations in IRAK4 underlie IRAK-4 deficiency, a rare autosomal recessive immunodeficiency that results in impaired toll-like receptor (TLR) and Interleukin-1 (IL-1) driven responses. Children with IRAK-4 deficiency are predisposed to recurrent and invasive infections secondary to Streptococcus pneumoniae, Staphylococcus aureus and other pyogenic bacteria with high mortality rates in early childhood. The frequency and severity of infections is thought to decrease with age due to the acquisition of humoral immunity and immunologic memory, however due to the rarity of the disease, the natural history of this condition beyond early childhood is not well described.

    Objectives: We present three unrelated IRAK-4 deficient patients with persistent chronic rhinosinusitis with nasal polyposis that developed in childhood.

    Cases: Patient 1 is a 15 y/o male with compound heterozygous mutations in IRAK4 (p.G75Afs*14/c.717-1G>T) with a history of recurrent S. pneumoniae osteomyelitis (left hip at age 9 and left knee at age 10) and C. septicum sepsis at age 9 following acute bowel perforation. Additionally, he experienced recurrent AOM during infancy and recurrent UTI since age 9. Despite prophylactic antibiotics and IVIG, he has had recurrent polymicrobial (MRSA, S. pneumoniae, H. influenzae, P. aeruginosa, A. fumigatus) rhinosinusitis with nasal polyposis since age 4 refractory to medical management requiring surgical intervention and prolonged courses of IV antibiotics.

    Patient 2 is an 11 y/o female with homozygous deletions (exons 10-12) in IRAK4 with a history of ruptured appendicitis complicated by Pseudomonas abscess and bacteremia at age 2, culture-negative sepsis with septic arthritis and osteomyelitis of the right leg at age 3, and septic shock secondary to MSSA bacteremia complicated by rhabdomyolysis and DIC at age 5. She has a history of chronic rhinosinusitis, and despite IVIG and prophylactic antibiotics, she developed polymicrobial (H. influenzae, B. fragilis) rhinosinusitis with associated nasal polyposis pending surgical management.

    Patient 3 is a 10 y/o female with homozygous mutations in IRAK4 (Q293X/Q293X on exon 8) with a history of S. pneumoniae meningitis at 3 months, M. catarrhalis epiglottitis and neck cellulitis at 4 months, RSV bronchiolitis at 6 months, Enterococcus bacteremia at 8 months, S. pneumoniae sepsis at age 2 and Streptococcus lymphadenitis at age 9. Despite IVIG and prophylactic antibiotics, she developed recurrent polymicrobial (H. influenzae, B. fragilis, MSSA, V. cholera, P. aeruginosa, A. fumigatus) rhinosinusitis refractory to medical management requiring surgical intervention and IV antibiotics.

    Conclusions: In our centers experience, IRAK-4 deficient patients continue to suffer from infectious complications, most prominently recurrent polymicrobial sinus infections beyond early childhood. The consistent presence of sinonasal polyps in these children is unusual, as it is not typically found in uncomplicated pediatric chronic rhinosinusitis. These infections have occurred despite antimicrobial prophylaxis and IVIG, highlighting the role of IRAK-4 in sinopulmonary epithelium. Additionally, the infectious organisms identified in our patient cohort are not commonly associated with IRAK-4 deficiency. Further study of chronic rhinosinusitis and nasal polyposis in a larger cohort of IRAK-4 deficient patients and other innate immunodeficiencies may help identify pathways for targeted treatment of these patients.

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    (150) Submission ID#601621

    Osteomyelitis in Chronic Granulomatous Disease: Experience from a Tertiary Care Centre in North-West India

    Johnson Nameirakpam1, Pandiarajan Vignesh, MD;DM (Pediatric Clinical Immunology and Rheumatology)2, Amit Rawat, MD (Pathology) PDCC (Laboratory Immunology) PDCC (Nephropathology) MAMS3, Deepti Suri, MD4, Anju Gupta5, Surjit Singh, MD; DCH (Lon.); FRCP (Lon.); FRCPCH (Lon.); FAMS6

    1DM Fellow Pediatric Clinical Immunology and Rheumatology, Allergy and Immunology Unit, Advanced Pediatrics Centre, Postgraduate Institute of Medical Education and Research

    2Assisstant professor, department of pediatrics, Postgraduate Institute of Medical Education & Research

    3Professor of Pediatric Allergy and Immunology, Paediatric Allergy Immunology Unit, Department of Paediatrics, Advanced Paediatric Centre, Postgraduate Institute of Medical Education & Research

    4Consultant, Allergy immunology unit, Department of Pediatrics, Advanced Pediatrics Centre, Postgraduate Institute of Medical Education and Research, Chandigarh, India, Dept. of Pediatrics, Allergy- Immunology Unit, Postgraduate Institute of Medical Education and Research

    5Professor, Allergy and Immunology unit, Advanced pediatrics Centre, , Postgraduate Institute of Medical Education and Research, Chandigarh, India

    6Head, Department of Pediatrics and Chief, Allergy Immunology Unit, Advanced Pediatrics Centre,Principal Investigator, Indian Council of Medical Research (ICMR) Centre for Advanced Research in Primary Immunodeficiency DiseasesVice-President, Indian Rh, Postgraduate Institute of Medical Education & Research

    Introduction: Chronic granulomatous disease (CGD) is an inherited phagocytic defect associated with inability to clear catalase positive organisms. Infections in patients with CGD are severe and recalcitrant. Commonest infections are pulmonary followed by soft tissue infections and suppurative lymphadenitis. Osteomyelitis is an uncommon infection in patients with CGD. It poses several diagnostic and therapeutic challenge. We herein report our experience of osteomyelitis in CGD over the last 10 years.

    Material and methods: Review of records was carried out to describe the profile of osteomyelitis in cohort of patients with CGD at Pediatric Immunodeficiency Clinic, Advanced Pediatrics Centre, Postgraduate Institute of Medical Education and Research, Chandigarh, India. The diagnosis of CGD was based on Nitroblue tetrazolium dye reduction test (NBT) and Dihydrorhodamine reduction (DHR) assay.

    Results: Of the 63 patients with CGD, 8 (12.7%) had osteomyelitis (6 males and 2 females; age range 1- 10 years). Most patients had their first episode of serious infection in early childhood (mean age: 1.5 years). Stimulation index (SI) of DHR assay ranged from 1 to 4.58. Mutational analysis was done in 5/8 patients (3 X-linked; 2 autosomal recessive). Site of involvement was variable ribs- 4; vertebrae- 2; radius- 1; skull- 2; tibia- 1. Aspergillus fumigatus was the most common isolate (62%; 5/8); others had Aspergillus flavus, Aspergillus terreus and Serratia marcescens each. All 4 patients with rib osteomyelitis had concurrent pneumonia, and fungus was isolated in all of them (Aspergillus fumigatus- 2, Aspergillus flavus- 1, Zygomyces spp.- 1). Antifungals (intravenous amphotericin B) were given for a duration of 4-6 weeks and were followed by oral voriconazole in therapeutic doses for 3 to 6 months in majority of them. Debridement and resection of ribs was required in one patient, while other patients were managed conservatively. Out of 8 patients, 2 (25%) succumbed to pneumonia and respiratory failure.

    Conclusion: Osteomyelitis in the context of CGD is usually caused by Aspergillus spp. Involvement of ribs and vertebra usually occurs with the contiguous spread of infection from the lungs. Therapy often requires prolonged duration of anti-microbials, and may require surgical debridement in addition to it.

    (151) Submission ID#601626

    A 29-year-old Woman with History of Hypogammaglobulinemia and Acute Liver Failure

    Beth K. Thielen, MD, PhD1, Timothy Moss, MD, PhD2

    1Fellow, University of Minnesota, Departments of Internal Medicine and Pediatrics, Divisions of Infectious Diseases and International Medicine and Pediatric Infectious Diseases and Immunology

    2Assistant Professor, University of Minnesota, Department of Pediatrics, Division of Genetics and Metabolism

    A 29-year-old woman with a 7-month history of nausea, vomiting, and abdominal pain was admitted to an outside hospital with new onset of jaundice and anasarca. Liver biopsy was thought most consistent with alcoholic steatohepatitis, and she was discharged with counseling on alcohol cessation and medical management of liver disease. She presented to our facility for a second opinion. Over the following days, she developed further rise in direct hyperbilirubinemia up to 19.2 mg/dL, new coagulopathy with an INR 2.06 and hypoalbuminemia to 1.7 mg/dL in the absence of ongoing alcohol consumption. Liver sonography revealed course echotexture and patent vessels. PCRs directed against multiple hepatotropic viruses were negative and copper studies were normal. Due to a history of moderate alcohol consumption, she was started on high-dose corticosteroids due to a presumptive diagnosis of alcoholic hepatitis.

    Additional history raised concern for a possible primary immunodeficiency, including idiopathic thrombocytopenic purpura at 11 years of age, multiple episodes of sinusitis treated with antibiotics and sinus surgery, one episode of suspected bacterial pneumonia, and one hospitalization for influenza A during which she developed neutropenia. In her 20s, she developed refractory genital warts, prompting infectious diseases evaluation. Initial immune evaluation had revealed low immunoglobulins (IgA <7 mg/dL, IgG 198 mg/dL, IgM 13 mg/dL) with very low responses to tetanus and diphtheria, despite a recent booster dose, and B and T cell lymphopenia (CD19+ 89 cells/μL, CD3+ 567 cells/μL, CD4+ 345v, CD 8+ 244 cells/μL, CD16/56+ 236 cells/μL); antigen and mitogen proliferation were not assessed. Intravenous immunoglobulin replacement was initiated but discontinued by the patient due to infusion-related adverse effects, and she was lost to follow up until she presented with liver failure. Both parents were deceased from cardiovascular disease in their 40s and she had no siblings. She had limited knowledge of family history but no known immune diseases.

    Due to suspicion for genetic etiology of immune disorder and liver disease, we performed next-generation sequencing of a panel of over 200 genes implicated in primary immune deficiencies. Patient was heterozygous for a nucleotide substation (c.1752+1G>A) within a splice site at the exon 16/intron 16 boundary of the NFKB1 gene. During the hospitalization, immunoglobulin replacement and trimethoprim-sulfamethoxazole prophylaxis were initiated. An attempt was made to refer the patient for additional immunological evaluation and transplantation evaluation but unfortunately, she developed worsening liver failure and multiple complications, including extended-spectrum beta-lactamase (ESBL)-producing E. coli bacteremia, hypotension requiring vasopressors and extensive bowel ischemia, and died in the hospital.

    In summary, this case highlights both the risk of diagnostic delay in adult patients presenting with a primary immune deficiency and potential for genetic testing to clarify the diagnosis. While the particular genetic change has not been described, other splice site and predicted loss-of-function mutations have been reported as pathogenic in this gene, which have been implicated in autosomal dominant common variable immunodeficiency. This case further expands on the genetic causes and spectrum of disease associated with changes in the NFKB1 gene.

    (152) Submission ID#601723

    Acquired Immunodeficiency - More Than Meets the BMI (Body Mass Index)

    Laura E. Maurer, MD, MPH1, Victor P. Bilan, MD2

    1Resident Physician, Yale New Haven Hospital

    2Chief Resident for Quality and Safety, West Haven Veterans Affairs Hospital

    Introduction: Malnutrition and micronutrient deficiency are underrecognized causes of acquired immunodeficiency in adults, and may occur even in patients with high body mass index (BMI).

    Methods: A 46-year-old woman with a medical history significant for one remote urinary tract infection presented to the emergency department after sudden onset of severe right flank pain. The pain was accompanied by urinary frequency and not relieved by ibuprofen; she denied fevers or chills. She was diagnosed with pyelonephritis and discharged on ciprofloxacin, which was later changed to trimethoprim-sulfamethoxazole after her culture grew resistant E. coli. Her pain continued despite treatment, prompting her to return to the hospital three days later.

    Upon presentation, she was afebrile with blood pressure of 128/88 mmHg and heart rate of 86 bpm. Her body mass index was 32.4 kg/m^2. Her physical exam was otherwise notable for right costovertebral angle tenderness. Laboratory studies revealed a leukocyte count of 14,300/ul with 83% neutrophils; alkaline phosphatase of 146 units/L and albumin of 2.7 g/dL, but otherwise normal liver function tests; normal lactic acid; and urinalysis with 3,000 WBC/hpf, 40 RBC/hpf, moderate bacteria, and the presence of WBC clumps. CT scan of the abdomen and pelvis demonstrated an obstructing 13 mm right renal stone with hydronephrosis and a right renal abscess contiguous with a right-sided hepatic abscess measuring 7.8 x 6.0 x 7.5 cm. She was treated with ceftriaxone and metronidazole, and underwent imaging-guided drainage of the abscesses. Abscess cultures again grew resistant E. coli. She was discharged from the hospital with drains in place and a plan to continue trimethoprim-sulfamethoxazole until definitive management of her nephrolithiasis with ureteroscopy and lithotripsy.

    Discussion: There remained the question of how an ostensibly immunocompetent patient had developed such severe intraabdominal infection with little systemic inflammatory response (e.g. no fever and only mild leukocytosis). A HIV antibody screen was negative. On further interview, she described a 200lb intentional weight loss over the preceding 2 years, accomplished by dietary restriction to less than 600 calories per day. Nutritional assays revealed prealbumin, vitamin C, and vitamin B6 levels below the threshold of detection. She had low-normal B12 and B1. Out of concern for an acquired immunodeficiency resulting from malnutrition with micronutrient deficiency, balanced nutrition was discussed with the patient who agreed to liberalize her diet.

    Conclusion: This case illustrates that marked acquired immunosuppression can result from malnourished and undernourished states. This phenomenon is well recognized in young children and in resource-limited settings, but may occur even in the presence of normal or high body mass index. It further demonstrates the importance of routinely taking a thorough dietary history and encouraging healthy eating practices as part of usual clinical anticipatory guidance.

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    (153) Submission ID#601826

    Centralized Sequencing Initiative at NIAID: Year 1

    Magdalena A. Walkiewicz, Ph.D.1, Morgan Similuk, ScM2, Celine Hong, Ph.D.3, Leila Jamal, MSc., Ph.D.4, Haley Hullfish, B.S.5, Jia Yan, MSc., Ph.D.6, Patty Littel, B.S.7, Sandhya Xirasagar, Ph.D.8, Adriana Almeida de Jesus, MD9, Elise Ferre, PA-C, MPH10, Raphaela Goldbach-Mansky, MD, Ph.D.11, Michail Lionakis, MD, Sc.D12, Steven M. Holland, MD13

    1ABMG certified Clinical Molecular Geneticist, National Institute of Allergy and Infectious Diseases (NIAID)

    2Genetic Counselor, Molecular Development of the Immune System Section, Laboratory of Immune System Biology, NIAID, National Institutes of Health, Bethesda, MD, USA

    3N/A, NHGRI

    4Genetic Counselor, NIAID

    5Research Fellow, National Institute of Allergy and Infectious Disease (NIAID)

    6Clinical Protocol Coordinator and Genetic Counselor, NIAID

    7RN, NIAID

    8Health Scientist, NIAID

    9Staff Clinician, NIAID

    10Physician Assistant, Fungal Pathogenesis Section, Laboratory of Clinical Immunology & Microbiology (LCIM) , National Institute of Allergy & Infectious Diseases (NIAID), NIH

    11Clinical Investigator, NIAID

    12Chief, Fungal Pathogenesis Section, National Institute of Allergy and Infectious Diseases (NIAID) National Institutes of Health (NIH)

    13Director, Division of Intramural Research, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health

    Background: The past decade has brought dozens of new Mendelian disorders of immunity. Yet, the genetic contribution(s) to diverse disorders of the immune system remain largely unelucidated. The majority of research participants referred to the National Institute of Allergy and Infectious Diseases (NIAID) for what may be a Mendelian disorder evade molecular diagnosis. Making progress in this area requires a coordinated, systematic, and transparent approach to clinical genomics research which leverages the unique environment at the National Institutes of Health Clinical Center (NIH CC).

    Methods/Design: This study is designed to systematically apply exome sequencing and related technologies with clinical grade interpretation and reporting to NIAID research participants at the NIH CC under a single protocol in order to facilitate research and clinical genetics care across NIAID. We are recruiting approximately 1000 participants per year from approximately 35 intramural clinical investigators. We generate genomic data, collect standardized phenotyping and report clinical interpretation in the medical record, all while providing linked genetic counseling.

    Results: To date, we consented 1287 participants, we sent out 1058 samples for exome sequencing and 183 samples underwent copy number variant analysis. We have completed analysis for 359 families (502 individuals) and finalized and resulted 177 cases. Here we present a case series illustrating some of our findings. Case 1: A 10-year-old female was referred to NIAID for neonatal onset multisystem inflammatory disease (NOMID). Developmental delay and mild intellectual disability were appreciated on clinical evaluation. Exome sequencing detected a mosaic novel likely pathogenic variant in NLRP3. Chromosomal Microarray Analysis (CMA) showed a ~5 Mb interstitial deletion of chromosome 12 previously associated with developmental delay and intellectual disability. Case 2: A 10-year-old Ukrainian male was referred to NIAID for the clinical diagnosis of autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED). Exome sequencing and CMA did not detect pathogenic variants in AIRE, but did find a de novo variant in FAM111B. Defects in FAM111B are associated with poikiloderma with tendon contractures, myopathy, and pulmonary fibrosis (POIKTMP). The clinical features of the patient were consistent with POIKMP. Case 3: A 63-year-old man had a history of brain, liver and kidney nocardiosis, disseminated MAC infection, prostate cancer and lymphoma. Family history was significant for prostate cancer. Exome sequencing showed a heterozygous pathogenic variant in BRCA2, associated with susceptibility to breast-ovarian, male breast, pancreatic and prostate cancer.

    Conclusion: This case series illustrates that multiple diagnoses, unexpected diagnoses, secondary genomic findings, and data sharing helped identify variants in candidate genes. Process standardization supports data integrity and efficiency while accommodating the need for investigator flexibility and providing tailored patient care.

    (154) Submission ID#601828

    T-cell Receptor Repertoire Clonality in Peripheral Blood and Affected Tissue in Activated PI3 Kinase Delta Syndrome (APDS)

    Sara Barmettler, MD1, James M. Heather, PhD2, Jocelyn R. Farmer, MD/PhD3, Gabriel Wong, MD/PhD4, Mark Cobbold, MD/PhD5

    1Attending Physician, Massachusetts General Hospital

    2Research Fellow, Massachusetts General Hospital

    3Instructor, Massachusetts General Hospital

    4Physician, University of Birmingham

    5Associate Professor, Massachusetts General Hospital

    Rationale: Activated PI3 kinase delta syndrome (APDS) is a primary immunodeficiency caused by dominant mutations that increase activity of phosphoinositide-3-kinase (PI3K). The catalytic subunit p110 is mainly expressed in cells of the hematopoietic system, primarily lymphocytes and myeloid cells, and mutations affect both B- and T-cells. We sought to further evaluate the role of the T-cell receptor (TCR) repertoire in immune dysregulation and the pathogenesis of autoimmunity and lymphoproliferation in patients with APDS.

    Methods: We evaluated the TCR repertoire in the peripheral blood in 3 patients with PIK3CD mutations and compared these to the peripheral TCR repertoire in 26 patients with common variable immunodeficiency (CVID) and 50 healthy controls to investigate the role of the TCR in disease. The TCR repertoire in affected tissue of 2 patients with PIK3CD mutations was also evaluated (tissue included lymph nodes for both patients, in addition to gastrointestinal tract and lung tissue in one patient). A fixed number of TCRs were subsampled (35,000 for blood and 5,000 for tissue) and diversity was calculated using the Gini and Shannon indexes.

    Results: Using the Shannon and Gini diversity indexes, the TCR repertoire in patients with PIK3CD mutations had less diversity/increased clonality as compared to healthy controls and those with CVID (Figure 1). For the two APDS patients with biopsy tissue available for analysis, the diversity of the TCRs in tissue was increased as compared to the peripheral blood TCR repertoire (Figure 2).

    Conclusions: PI3K plays an important role in the development and function of both B- and T-cells. Patients with APDS were found to have decreased TCR repertoire diversity in the circulating T-cell compartment compared to healthy controls and other CVID patients. The increased TCR diversity in the affected tissues compared to peripheral blood implicates the PI3K/AKT signaling pathway with T-cell trafficking and tissue immune homeostasis, and suggests this pathway may play a role in the development of inflammatory and lymphoproliferative complications in these patients.

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    (155) Submission ID#601830

    Hyperactivated PI3Kd Promotes Self and Commensal Reactivity at the Expense of Optimal Humoral Immunity

    Silvia Preite, PhD1, Jennifer L. Cannons, PhD2, Andrea J. Radtke, PhD3, Ivan Vujkovic-Cvijin, PhD4, Julio Gomez-Rodriguez, PhD2, Stefano Volpi, MD5, Bonnie Huang, PhD6, Cheng Jun7, Nicholas Collins, PhD8, Kerry Dobbs, BSc9, Julie Reilley10, Quan-Zhen Li, MD, PhD11, Stefania Pittaluga, MD, PhD12, Gulbu Uzel, MD13, Luigi D. Notarangelo, MD, PhD14, Yasmine Belkaid, PhD15, Ronald N. Germain, MD, PhD16, Pamela L. Schwartzberg, MD, PhD17

    1Postdoctoral fellow, National Human Genome Research Institute. Laboratory of Immune System Biology, NIAID, NIH, Bethesda, MD, USA

    2Staff Scientist, National Human Genome Research Institute. Laboratory of Immune System Biology, NIAID, NIH, Bethesda, MD, USA

    3Postdoc, Laboratory of Immune System Biology, NIAID, NIH, Bethesda, MD, USA

    4Postdoc, Laboratory of Parasitic Diseases, NIAID, NIH, Bethesda, MD, USA

    5Researcher, Clinica Pediatrica e Reumatologia, Centro per le Malattie Autoinfiammatorie e Immunodeficienze, Istituto Giannina Gaslini e Università degli Studi di Genova, Genoa, Italy

    6Postdoc, National Human Genome Research Institute. Laboratory of Immune System Biology, NIAID, NIH, Bethesda, MD, USA

    7Technician, National Human Genome Research Institute, NIH, Bethesda, MD, USA

    8Postdoc, Laboratory of Parasitic Diseases, NIAID, NIH

    9Biologist, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    10Biologist, National Human Genome Research Institute. Laboratory of Immune System Biology, NIAID, NIH, Bethesda, MD, USA

    11Associate Professor, Microarray Core Facility and Department of Immunology, University of Texas Southwestern Medical Center, Dallas, TX, USA

    12Senior Research Physician, Laboratory of Pathology, National Cancer Institute, NIH, Bethesda, MD, USA

    13Staff Clinician, Laboratory of Clinical Immunology and Microbiology, National institute of Allergy and Infectious Diseases, NIH, Bethesda, MD, USA

    14Chief, Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    15Senior Investigator, Laboratory of Parasitic Diseases. Microbiome Program, NIAID, NIH, Bethesda, MD, USA

    16Chief, Laboratory of Immune System Biology. Chief, Lymphocyte Biology Section, Laboratory of Immune System Biology, NIAID, NIH, Bethesda, MD, USA.

    17Senior Investigator, National Human Genome Research Institute. Laboratory of Immune System Biology, NIAID, NIH, MD, USA

    Gain-of-function mutations in PI3Kd result in a human primary immunodeficiency, named APDS (Activated PI3K-delta syndrome), characterized by lymphopenia, lymphoproliferation, respiratory infections and inefficient responses to vaccination. However, what promotes these immune disturbances at the cellular and molecular level remains unknown. We have recently published a mouse model that recapitulates major features of this disease and used this model and patient samples to probe how hyperactive PI3Kd fosters aberrant humoral immunity. We found that mutant PI3Kd alters the intrinsic function of T and B cells, leading to ICOS-independent increases in T follicular helper (Tfh) and germinal center (GC) B cells, disorganized GCs, and poor class-switched antigen-specific responses to immunization. These phenotypes were associated with increased phosphorylation of AKT and S6 in T and B cells, and lower threshold of activation, with altered regulation of FOXO1 and BCL2 family members. Moreover, B cells showed enhanced responsiveness and proliferation to both antigens and innate stimuli, accompanied by reduced cell death. Strikingly, aberrant responses were accompanied by increased reactivity to gut bacteria, and a broad increase in autoantibodies that were dependent on commensal microbial stimulation, as demonstrated by striking reduction of self-reactivity upon antibiotic treatment in mutant mice. We now have further examined B cell function in these mice and demonstrate that altered FOXO1 plays a major role in disruption of both B and T cell function. We further provide evidence for altered activation of metabolic pathways in B cells, compared to WT cells, that may contribute to the dysregulated B cell reactivity. Our findings suggest that proper PI3Kd regulation is critical for ensuring optimal host-protective humoral immunity despite tonic stimulation from the commensal microbiome.

    This research was supported in part by the Intramural Research Program of the NIH, NHGRI and NIAID.

    (156) Submission ID#601877

    The B-cell Subset Mileu of Autoimmune Cytopenias in Primary Immunodeficiency

    Travis Sifers1, Charlotte Cunningham-Rundles, MD, PhD2

    1Fellow-in-Training, Mount Sinai School of Medicine, Ichan School of Medicine at Mount Sinai

    2Professor in Medicine, Division of Clinical Immunology, Icahn School of Medicine, Mount Sinai, NY, NY, USA

    Autoimmune cytopenias are seen in a significant proportion of patients with immunodeficiencies affecting antibody production. Previous B-cell maturation studies using fluorescence-activated cell sorting (FACS) have associated various phenotypes of primary immunodeficiency diseases affecting antibody production with differing levels of B-cell differentiation. In this study we analyzed the peripheral B-cell compartment of 84 patients with a hypogammaglobulinemia and >1% B-cells with and without a history of autoimmune cytopenias. B-cells were isolated from peripheral blood using monoclonal anti-CD19 and these cells were gated to identify the proportion of memory B cell (CD19+CD27+), IgM+ memory B (CD27+IgM+), marginal zone B-cells (IgM+IgD+), isotype-switched memory B-cells (CD27+IgM-IgD-) and transitional cells (IgMhiCD38hi).

    PID patients with a history of AIC had decreased proportions of total CD27+ B-cell (11.6% vs 25.6%; p=0.0003) and IgM memory B cells (8.3% vs 18.4%; p = 0.0018). Conversely, the proportion of marginal zone B-cells was increased in this group (82.0% vs 66.5%; p = 0.0043).

    Consistent with previous reporting, the proportion of isotype-switched memory B-cells was significantly lower in the AIC group (0.75% vs 2.8%; p = 0.0003). Statistically significant inter-group difference was not seen within the transitional B-cell subset.

    Our data suggest that maturation arrest of marginal zone (CD27+IgM+IgD+) B-cells may be implicated in the development of autoimmune cytopenias in humoral immunodeficiency.

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    (157) Submission ID#601907

    XLA Presenting as HHV-6 Meningitis and Pseudomonas Meningitis/Bacteremia in a Patient Without History of Recurrent Infections

    Melissa A. Cardenas-Morales, MD1, Camile Ortega, D.O.2, Jose Calderon, MD3, Paul Martinez, MD4, Vivian Hernandez-Trujillo, MD3

    1Allergy and Immunology Fellow, Nicklaus Childrens Hospital

    2Allergy and Immunology Fellow, Nicklaus Children's Hospital

    3Allergy Immunology Attending, Nicklaus Children's Hospital

    4Critical Care Physician, Nicklaus Childrens Hospital

    Introduction: X-linked agammaglobinemia (XLA) is a primary immunodeficiency due to the inheritance of a pathologic variant of the Bruton Tyrosine Kinase (BTK) protein. The condition usually manifests within the first 2 years of life. An estimated 60% of patients present with a severe, life-threatening infection. Invasive Pseudomonas aeruginosa infection is uncommon in immunocompetent hosts. Several cases of XLA patients presenting as pseudomonas sepsis, meningitis, and bacteremia have been reported.

    While severe viral infections may also be an initial presentation of primary immunodeficiency, an immune evaluation is not always obtained in this scenario. Patients with XLA have an increased susceptibility to severe enterovirus infections, manifesting as chronic meningoencephalitis, which can be fatal.

    The following case describes a patient with newly diagnosed XLA presenting as suspected coxsackievirus and confirmed HHV-6 meningitis, Pseudomonas meningitis and bacteremia. This may be the first reported new diagnosis of XLA presenting with both severe bacterial and viral co-infection.

    Case Description: A 2 year old, partially vaccinated, Hispanic male with a history of febrile seizures presented to the emergency room with fever, oliguria, watery diarrhea, lethargy, meningismus, ecthyma gangrenosum and lower abdominal pain. Eight days prior to presentation, he was seen by his pediatrician for facial rash and low grade temperature, and was diagnosed with hand-foot-and mouth disease. He worsened on empiric antibiotics. He had no history of sinopulmonary infections. He did not attend daycare. His vaccines were delayed due to parental choice, and he had not received live vaccines (Rotavirus, MMR or VZV). Full sepsis evaluation was performed. CSF demonstrated pleocytosis, and he was started on empiric antibiotics and transferred to PICU. Due to worsening abdominal pain, CT of the abdomen was performed, which was consistent with ruptured appendicitis and septic emboli at the lung bases. CSF PCR panel was positive for HHV-6 and he was started on Gancyclovir. CSF and blood cultures subsequently grew Pseudomonas aeruginosa. Immune evaluation was performed. Serum immunoglobulins were undetectable. In addition to IV antibiotics, he received 500 mg/kg IVIG and lymphocyte subsets revealed profound B cell lymphopenia (0.23 %, 5 cells/uL). BTK protein analysis revealed hemizygous BTK pathogenic variant confirming the diagnosis of X-Linked agammaglobulinemia. The hospital course was further complicated by brain abscesses and pyoventriculitis. He was treated with 3 additional doses of 500 mg/kg IVIG and IV antibiotics. Repeat MRI of the brain nearly 4 weeks after admission demonstrated significant improvement. There was significant clinical recovery. He was discharged home at baseline neurological status. His IgG level upon discharge home was 605 mg/dL with the plan to increase dose to 600 mg/kg per month with close monitoring.

    Conclusion: Both severe opportunistic bacterial infections and severe viral infections as the initial presentation of XLA have been well reported in the literature. This case describes the first reported severe Pseudomonas aeruginosa and HHV-6 co-infection in a newly diagnosed XLA patient. This case further highlights the necessity for an increased index of suspicion of primary immunodeficiency in a patient who presents with a severe first infection, despite lack of recurrent infections.

    (158) Submission ID#601911

    Missense Variants as a Contributing Cause to DOCK8 Immune Deficiency

    Haley Hullfish, B.S.1, Morgan Similuk, ScM2, Huie Jing, PhD3, Jeffrey Danielson, MS4, Leila Jamal, MSc., Ph.D.5, Celine Hong, Ph.D.6, Jia Yan, MSc., Ph.D.7, Steven M. Holland, MD8, Alexandra F. Freeman, MD9, Magdalena A. Walkiewicz, Ph.D.10, Helen C. Su, MD, PhD11

    1Research Fellow, National Institute of Allergy and Infectious Disease (NIAID)

    2Genetic Counselor, Molecular Development of the Immune System Section, Laboratory of Immune System Biology, NIAID, National Institutes of Health, Bethesda, MD, USA

    3Research Staff Member, NIAID

    4Research Staff Member, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD

    5Genetic Counselor, NIAID

    6N/A, NHGRI

    7Clinical Protocol Coordinator and Genetic Counselor, NIAID

    8Director, Division of Intramural Research, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health

    9Director, Primary Immune Deficiency Clinic, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    10ABMG certified Clinical Molecular Geneticist, National Institute of Allergy and Infectious Diseases (NIAID)

    11Chief, Human Immunological Diseases Section, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD

    We present two patients with DOCK8 deficiency due to compound heterozygous variants including a copy number loss at chromosome band 9p24.3 spanning approximately .107 Mb with partial deletion of the DOCK8 gene and a novel c.2603C>T (p.Ser868Leu) missense variant [chr9:379933 (GRCh37) NM_203447] in DOCK8. Functional data is presented to support the pathogenicity of the missense change, along with a review of the literature on DOCK8 variants.

    The proband is a 14-year-old female with elevated serum IgE, severe atopic dermatitis, mild persistent asthma, food allergies, and seasonal allergic rhinitis. She is currently healthy following haploidentical bone marrow transplant in June 2018. She has a 17-year-old brother with DOCK8 deficiency with the same compound heterozygous variants. The brother had later onset of symptoms and a milder presentation of intermittent asthma and seasonal allergic rhinitis. Each of the parents is heterozygous for one of the two variants.

    We evaluated the pathogenicity of the c.2603C>T missense variant with western blots of DOCK8 protein expression, intracellular flow cytometry, and DOCK8 stretch assays. Flow cytometry showed decreased DOCK8 protein expression and stretch assays revealed T cells that were stretched in collagen gels.

    Notably, DOCK8 is a large gene containing 47 exons spanning 190 kb and it is relatively common to be a carrier of a rare missense change. In fact, gnomAD has approximately 1500 individuals with rare (<0.002 frequency) missense alleles in DOCK8. Therefore, it is important to demonstrate the potential pathogenicity of any given rare missense change, since few pathogenic missense variants in DOCK8 have been reported. Of the 168 published DOCK8 variants listed in the Human Gene Mutation Database (HGMD) only 13 are missense. The majority are gross deletions, 97 of which were reported in HGMD. The remaining reported DOCK8 variants include 19 nonsense, 15 splicing, 13 small deletions (all frameshifting), 3 small insertions (all frameshifting), 2 small indels, and 5 gross insertions/duplications.

    This case demonstrates the relatively infrequent but important contribution of missense changes to pathogenic DOCK8 alleles. Functional validation of missense alleles is critical in the complex evaluation of DOCK8 deficiency.

    (159) Submission ID#601984

    IgD Class Switched B Cells in Patients with Common Variable Immunodeficiency

    Taissa de Matos. Kasahara1, Sudhir Gupta, MD2

    1PhD student, State University of Rio de Janeiro and University of Californis Irvine

    2Professor, University of California at Irvine, Irvine, CA, USA

    Introduction/Background: Common variable immunodeficiency (CVID) is the most frequent form of primary hypogammaglobulinemia with decreased serum IgG and IgA levels and variable levels of IgM in adults. In addition to decreased serum immunoglobulins, 25-30% of CVID patients present autoimmune manifestations. The mechanisms that lead to a breakdown of self-tolerance in CVID are not completely understood. However some differences in B and T cells subsets and autoreactive B and T cells can be detected. Elevated expression of surface IgD and downregulation of IgM receptor are hallmarks of anergic naïve B cells that contain autoreactive receptors in human peripheral blood. Moreover, memory B cells that have class switched to IgD and present an IgD+IgM- phenotype are also highly reactive to self-antigens in healthy individuals. The role of these autoreactive naïve and memory B cells in the immunopathogenesis of CVID has not been evaluated. Here we investigated the frequency of CD27- and CD27+ B cells expressing IgD and IgM in peripheral blood of CVID patients.

    Methods: Peripheral blood mononuclear cells (PBMC) from CVID patients (n=29) and health subjects (n=32) were separated by FicollHypaque and incubated with anti-human CD19-PerCP, CD27-FITC, IgD-BV510 and IgM-APC to identify different subsets of B cells by flow cytometry. CD19+CD27-IgD+IgM- and CD19+CD27-IgD+IgM+ B cells were sorted, loaded with CFSE and cultured with CpG and ant-CD40 for 5 days to evaluate the proliferation.

    Results: Among the compartment of CD27- B cells, CVID patients showed an increased frequency of IgD+IgM+ cells and a lower frequency of IgD-IgM- cells as compared to control group. No differences were observed in the frequency of IgD+IgM- cells in CD27- B cells between CVID patients and controls. In contrast, in the compartment of CD27+ B cells, CVID patients showed an increased frequency of IgD+IgM-, IgD+IgM+ and IgD-IgM+ cells and a lower frequency of IgD-IgM- cells when compared to health subjects. When the patients were divided in two groups based on autoimmune manifestations, the group with autoimmune disease showed an increased frequency of IgD+IgM+ and IgD-IgM+ cells in CD27- B cells when compared to the control groups. Both patient groups showed an increased frequency of IgD+IgM-, IgD+IgM+ and IgD-IgM+ cells and a lower frequency of IgD-IgM- cells when compared to health subjects. Regarding the proliferation, naïve B cells from CVID patients showed a reduced proliferative capacity in response to in vitro stimulation as compared with naïve B cells from health subjects.

    Conclusion: Our results suggest that the increase of CD27+IgD+IgM- B cells can be related to the susceptibility of autoimmunity in CVID patients.

    (160) Submission ID#601996

    A Case Review of IgG4 Related Disease

    Blake A. Thompson, MD1, Lyda Cuervo-Pardo, MD2, Mario Rodenas, MD, FAAAAI2

    1Internal Medicine Resident, University of Florida

    2Assistant Professor, University of Florida, Division of Rheumatology & Clinical Immunology, Department of Medicine

    Introduction: Immunoglobulin G4-related disease (IgG4-RD) is a group of immune-mediated conditions where tissues are affected with dense lymphoplasmacytic infiltrations with a predominance of IgG4-positive plasma cells and storiform fibrosis, usually in the setting of elevated serum concentrations of IgG4. Common presentations include autoimmune pancreatitis, sclerosing cholangitis, retroperitoneal fibrosis, salivary gland disease, and orbital disease, among others. Symptoms of asthma or allergy are present in approximately 40 percent of patients and they typically exhibit a good initial therapeutic response to glucocorticoids.

    Case presentation: A 61-year-old female with a history of gastroparesis, cutaneous lupus erythematosus and suspected autoimmune pancreatitis was referred to Allergy/Immunology clinic for evaluation of elevated IgG4.

    She reported a 15-year history of recurrent abdominal pain attributed to recurrent pancreatitis based on previous mild lipase elevations. Prior endoscopic ultrasound (EUS) of the pancreas revealed edema. There was concern for gallstone pancreatitis but ERCP followed by cholecystectomy, biliary and pancreatic sphincterotomy had no change in her symptoms. In 2016, she was noted to have a positive ANA and high serum IgG4, per patient (values from OSH records could not be obtained). Symptoms improved with a course of steroids, hence suspicion for autoimmune pancreatitis.

    In 2018 she developed a rash on her arms and face. Biopsies of the affected areas revealed cutaneous lupus erythematosus on the arms and a basal cell carcinoma on the face, which was excised. ANA was only 1:80 at that time.

    At the visit, she complained of severe allergic rhinitis, joint pains, as well as a malar rash, which responded to intermittent courses of prednisone by prior providers.

    Laboratories obtained at initial visit were significant for thrombocytopenia (135 thou/cu mm), positive Lupus anticoagulant (56 sec) and elevated IgG4 (95 mg/dL; normal range 4-86 mg/dL). C3, C4, C1q, ANA, anti-double stranded DNA, anti-Smith antibodies, antiphospholipid panel, UPEP and SPEP were all unremarkable. CT chest and abdomen were also normal.

    Given the patient's history of cutaneous lupus erythematosus, Plaquenil was started as a steroid sparing agent. EUS of the pancreas with possible biopsy was ordered in an attempt to obtain a histopathologic diagnosis of IgG4-RD.

    Conclusion: This case exhibits the association between elevated IgG4, pancreatitis of unknown origin, allergic rhinitis, and cutaneous lupus erythematosus, highlighting the value of identifying a pathologic connection between seemingly unrelated disorders in patients with elevated IgG4, as they may be manifestations of IgG4-RD. In order to make the diagnosis, histopathologic findings showcasing lymphoplasmacytic tissue infiltration consisting mainly of IgG4-positive plasma cells and small lymphocytes is essential.

    The majority of patients respond to glucocorticoids, and while the duration of response is variable, most patients flare during or after glucocorticoids are tapered, as noted in this patient. Rituximab has been shown to be effective in some patients and will be considered in this patient if symptoms persist.

    (161) Submission ID#602042

    Newborn Infant with Purine Nucleoside Phosphorylase (PNP) Deficiency and Congenital Cytomegalovirus (CMV) Infection

    Benjamin Rahoy, DO1, Sachit Patel, MD2, Shirley Delair, MD, MPH3, Michael Hershfield, MD4, Hana Niebur, MD5

    1Pediatric Resident, University of Nebraska Medical Center

    2Clinical Director, Pediatric Blood and Marrow Transplantation; Assistant Professor, Division of, Hematology/Oncology, University of Nebraska Medical Center

    3Associate Professor, Division of Pediatric Infectious Diseases, University of Nebraska Medical Center

    4Professor of Medicine, Professor of Biochemistry, Duke University Medical Center

    5Assistant Professor, Division of Pediatric Allergy/Immunology, University of Nebraska Medical Center

    Rationale: PNP deficiency is an autosomal recessive disorder due to defective purine metabolism leading to Severe Combined Immunodeficiency (SCID) and neurological deterioration. Newborn screening utilizing T-Cell Receptor Excision Circle (TREC) assay can detect affected patients before complications arise. Herein, we describe an infant initially identified by newborn screening with PNP deficiency and congenital CMV, a previously unreported presentation.

    Methods: CMV quantitative PCR (qPCR) was performed by Nebraska Medicine, PNP enzyme activity by Duke and genetic sequencing by Invitae.

    Results: A small for gestational age (SGA) male infant was reported to have an abnormal TREC assay on day of life (DOL) 7. He was hospitalized for further evaluation. Initial studies revealed profound lymphopenia, normal lymphocyte proliferation to mitogens and no evidence of maternal engraftment. Additionally on DOL 10, he had CMV viremia and viruria; thus with SGA, failed unilateral hearing screen and head ultrasound with bilateral parenchymal calcifications, congenital CMV was suspected. PNP enzyme activity was abnormal. CMV treatment was initiated with ganciclovir on DOL 10. Foscarnet was added on DOL 13. CMV qPCR levels decreased below the limit of detection by DOL 30. Genetic testing found a pathogenic homozygous mutation in PNP (c.286-18G>A). The infant has a 10/10 HLA-matched, unaffected, CMV positive sibling and will proceed to hematopoietic stem cell transplantation.

    Conclusions: To our knowledge, this is the first reported case of PNP deficiency identified through newborn screening. This novel case of congenital CMV and PNP deficiency highlights the importance of CMV screening and need for treatment strategies for congenital CMV in SCID.

    (162) Submission ID#602066

    A Clinical Genomic Research Ecosystem Maximizes Data and Improves Patient Care

    Morgan Similuk, ScM1, Leila Jamal, MSc., Ph.D.2, Haley Hullfish, B.S.3, Sandhya Xirasagar, PhD4, Magdalena A. Walkiewicz, Ph.D.5, Steven Holland, MD6

    1Genetic Counselor, Molecular Development of the Immune System Section, Laboratory of Immune System Biology, NIAID, National Institutes of Health, Bethesda, MD, USA

    2Genetic Counselor, NIAID

    3Research fellow, National Institute of Allergy and Infectious Disease (NIAID)

    4GRIS program lead, NIAID

    5ABMG certified Clinical Molecular Geneticist, National Institute of Allergy and Infectious Diseases (NIAID)

    6Scientific Director, NIAID, NIH

    Despite a dramatic increase in the use of next generation sequencing over the last decade, the majority of the more than 50 million identified human genomic variants do not have well-established clinical implications. Progress is being made on this complex challenge through multiple approaches, including data sharing. To maximize our understanding of genomic data, platforms that enable effective and responsible data-sharing are essential. This means that genotypic and phenotypic data must be findable, accessible, interoperable, and reusable under conditions that are ethical and transparent.

    To highlight innovations in data-sharing and their potential to advance discovery, we present three data-sharing mechanisms. For each platform, we will present a case highlighting its key functionality and discuss opportunities and challenges that may arise as each platform is scaled up.

    (1.) Genomic Research Integration System (GRIS) is a collaboration-engendering web application that facilitates the identification of genetic variants associated with rare immunological disorders. Users can access integrated and standardized phenotypic and genomic data that is analyzable within the platform. GRIS enables systematic and automated capturing, and links patient data from disconnected systems and paper-based records. Standardized annotations allow for the comparison of data from different clinical studies. The main goal of this tool is discoverability of other affected individuals enrolled in separate protocols within the NIAID intramural research program. This internal database was used to find a second family with a rare variant in a candidate gene.

    (2.) The Genomic Ascertainment Cohort (TGAC) is a resource that aims to improve our understanding of the phenotypic consequence of genetic variation by providing access to aggregate, de-identified genomic data from large NIH intramural and related cohorts. Participants have provided informed consent to be re-contacted for additional phenotyping in the future. The main goal of this tool is to enable further study of the clinical consequence of variants in a large, unbiased cohort of patients ascertained for many indications. This database was used to investigate findings in participants with previously published pathogenic variants in genes associated with primary immune deficiency based on medical record review.

    (3.) ClinGen is dedicated to building an authoritative central resource that defines the clinical relevance of genes and variants for precision medicine and research. Through the sharing of genetic and health data, ClinGen seeks to answer whether a given gene is associated with a disease (clinical validity)?; whether a given variant is causative (pathogenicity)?; and whether the information is actionable (clinical utility)? This resource is meant to convene disease- and gene-specific expert groups to curate the medical literature on Mendelian disease to better define gene-disease and variant-disease relationships using many lines of evidence. This resource was used to clarify clinical validity of disease-gene assertions.

    Together these efforts help create a clinical research ecosystem that maximizes the value of clinical research data and ultimately improves patient care.

    This research was supported by the Intramural Research Program of the NIH, NIAID.

    (163) Submission ID#602340

    Immunodeficiency in Elderly: Data from the USIDNET Registry

    Charmi Patel, MD1, Hannah Wright2, Ramsay Fuleihan, MD3, Charlotte Cunningham-Rundles, MD, PhD4, Daniel Suez, MD5, Artemio M. Jongco, III, MD, PhD, MPH6

    1Attending physician, Donald Barbara Zucker School of Medicine at Hofstra/Northwell

    2Research Data Analyst, USIDNET Consortium, National Institute of Allergy and Infectious Diseases (NIAID), Towson, MD

    3Professor of Pediatrics, Division of Allergy and Immunology, Northwestern University Feinberg School of Medicine, Chicago, NY

    4Professor in Medicine, Division of Clinical Immunology, Icahn School of Medicine, Mount Sinai, NY, NY, USA

    5President, Allergy, Asthma & Immunology Clinic, PA

    6Assistant Professor of Medicine and Pediatrics, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell; Center for Health Innovations and Outcomes Research, Feinstein Institute for Medical Research, Manhasset, NY

    Introduction: According to the Population Reference Bureau, the number of elderly Americans, defined as age 65 and older, is projected to more than double from 46 million to 98 million by 2060, rising from 15% to 24% of the total population. The impact of immunodeficiency in this important segment of the population remains understudied.

    Methods: The USIDNET Registry was queried to obtain demographic, clinical data of elderly patients defined as age 65 and older. Descriptive analyses were performed on the data.

    Results: 373 participants (7.2%) were eligible out of 5176 total registry participants. The median age of the cohort was 70 years and predominantly female (74.7%) and White (78.0%) with a median BMI of 26.6 ± 6.6.The majority (81.8%) of subjects were living. Humoral deficiencies comprised the majority of diagnoses (94.6%), with Common Variable Immune Deficiency being the most frequent (76.9%). Of the remaining non-humoral diagnoses, immune dysregulation (1.3%) and immunodeficiency with myelodysplasia (1.1%) were the most frequent. The majority (79.1%) of subjects reported having received immunoglobulin replacement therapy (IGRT) at some point, with 51.7% reporting via IV route. Of the 1275 infections that occurred in this cohort, sinopulmonary infections were the most commonly reported, specifically sinusitis (18.5%), pneumonia (13.8%), upper respiratory infection (6.7%), and otitis media (5.5%). In this cohort, 107 autoimmune, 49 cardiovascular, and 11 granulomatous complications were reported . The number of patients with malignancy was 89, with some patients diagnosed with multiple malignant disorders. Of the reported malignancies, the majority (69.9%) were solid tumors.

    Conclusions: Compared to the age-matched non-immunodeficiency United States population, this cohort had more females 74.7% (USIDNET) versus 56.0% (US population) and fewer whites 78.0% (USIDNET) vs 86.0% (US population. Humoral immunodeficiencies, specifically CVID, were most common diagnoses, similar to other age groups of immunodeficiency patients. Majority of these patients have received IGRT, with approximately half via IV route. This cohort reported living with a variety of non-infectious complications, including autoimmunity and malignancies. More research which specifically focuses on elderly patients with immunodeficiency is needed.

    (164) Submission ID#602465

    A Rare Case of Helicobacter Bilis Chronic Complicated Osteomyelitis with Pyomyositis and Cellulitis in a Patient with XLA Agammaglobulinemia : Discussion of Challenges in Diagnosis and Management

    Candace Rypien, MD1, Nicola A. Wright, MD2, Luis Murguia-Favela, MD3, Andrea Fong, MD4, Dan Gregson, MD5

    1Pediatric Infectious Disease, Alberta Children's Hospital

    2Associate Professor, Department of Pediatrics, Alberta Children's Hospital, Calgary, Alberta, Canada

    3Clinical Assistant Professor, University of Calgary

    4Physician, University of Saskatchewan

    5Clinical Microbiologist and Infectious Disease Physician, University of Calgary

    X-linked agammaglobulinemia (XLA) is a primary immunodeficiency caused by mutations in the Bruton tyrosine kinase gene which leads to B cell maturation failure and defective antibody production. This puts patients at risk of recurrent sinopulmonary infections, gastrointestinal infections, and recurrent skin infections including infections caused by Helicobacter sp.

    Helicobacter sp are gram negative bacilli commonly found in the gastrointestinal tract of various animals. Helicobacter sp. have been linked with gastritis most notably Helicobacter pylori causing gastric ulcers in humans. Helicobacter sp. has been found in rare cases to cause disseminated infections including pyodermic gangrenosum and cellulitis notably in patients with agammaglobulinemia.

    Infections caused by Helicobacter bilis are challenging to diagnosis due to difficulties with culturing the pathogen as well as poor guidelines for antimicrobial management.

    Case Report:

    The patient was diagnosed with X-linked agammaglobulinemia at the age of 16 months with a history of recurrent sinusitis and was started on IVIG q3weeks. Despite regular IVIG, he developed bronchiectasis. At 11 years of age in 2013, he developed a chronic rash around his left knee resembling erythema nodosum. By 2014, he had developed a left knee effusion associated with left sided calf pain.

    His knee pain was found to improve during courses of ciprofloxacin to treat recurrent lung infections. Given case report data of H. pylori causing erythema nodosum in patients with agammaglobulinemia, he was treated empirically for an H. pylori infections with no improvement. In 2015 he was found to have progressive cellulitis with pyomyositis of the left leg. A skin biopsy of a calf nodule was found to be culture negative but 16S PCR was positive for H. bilis. He was started on treatment with ertapenem and levofloxacin with subsequent resolution of his rash.

    His left ankle pain progressed and by late 2015 and was found to have possible osteomyelitis of the left ankle on MRI. In 2016 he was found to be bacteremic with H bilis. Due to progressive symptoms with significant impact on function and rising inflammatory markers despite 12 months of antimicrobial treatment, doxycycline and flagyl were added leading to clinical improvement and normalization of his inflammatory markers. He was continued on oral doxycycline and flagyl for 12 months for a chronic osteomyelitis.

    Discussion:

    H. bilis is a slow growing pathogen which is challenging to culture in the laboratory often requiring special agar plates and prolonged incubation. In patients with agammaglobulinemia and associated chronic skin infections or erythema nodosuma, H bilis should be suspected as a possible pathogen. Due to challenges with culturing, 16S PCR or amplification of the 16S ribosomal subunit should be considered to try to identify the pathogen.

    There are poorly delineated clinical antimicrobial breakpoints to help guide therapy with minimal evidence. Case reports suggest prolonged therapy with aminoglycosides and penicillin. Other studies have successfully treated patients with a carbapenem, azithromycin and levofloxacin. In the absence of sensitivity data, prolonged treatment (12months) should be considered with a combination of antimicrobials. Patients should be followed closely as recurrent infections are not uncommon.

    (165) Submission ID#604074

    Clinical Phenotyping of a DOCK8 Deficiency Cohort

    Alexandra F. Freeman, MD1, Nirali N. Shah, MD2, Amanda Urban, DNP, CRNP3, Dennis Hickstein, MD4, Helen C. Su, MD, PhD5

    1Director, Primary Immune Deficiency Clinic, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    2Associate Research Physician, Pediatric Oncology Branch, NCI, NIH, Bethesda, MD

    3Nurse Practitioner, Clinical Monitoring Research Program Directorate, Frederick National Laboratory for Cancer Research sponsored by the National Cancer Institute

    4Senior Investigator, NCI, NIH

    5Chief, Human Immunological Diseases Section, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD

    Introduction: DOCK8 deficiency is a combined immunodeficiency characterized by eczema, recurrent sinopulmonary infections, viral skin infections, malignancy and early mortality. In recent years, liver disease and vasculopathy have been increasingly recognized as a complication of DOCK8 deficiency. We clinically characterized our cohort of DOCK8 deficient patients, with a specific focus on these newly identified areas of disease involvement.

    Methods: Chart reviews were performed on patients seen at NIH with genetic and clinical diagnosis of DOCK8 deficiency. Patients were all enrolled on IRB approved NIAID protocols.

    Results: We identified 52 patients from 40 families with DOCK8 deficiency in our NIH cohort, ranging in age from 6-44 years. Of the 40 families, 17 had homozygous mutations. Of the 52 patients, food allergy was diagnosed in 31 (60%), eczema in 49 (94%), and asthma in 30 (58%). Chronic or recurrent viral skin infections were seen in 49/52 (94%). Chronic EBV viremia by PCR positivity was seen in 18/46 patients (39%); only 2 patients were known to be EBV immune without viremia. CMV viremia was infrequent. Sinopulmonary infections were common, with bronchiectasis occurring in 23 /50 (46%) with available imaging. Liver disease was diagnosed in 14 (27%), with 7 having biliary tract abnormalities on imaging and stool positive for Cryptosporidia; most patients with Cryptosporidia were without diarrhea. The incidence of Cryptosporidia is likely under-represented due to more recent availability of sensitive assays for Cryptosporidia detection. Other liver abnormalities included fatty liver, metastatic disease from malignancy and medication related hepatitis. Vasculopathy, predominantly of the aorta and cerebral arteries, was diagnosed in 7, with patients in the last 5 years being prospectively imaged. Autoimmunity was rare (5%) including autoimmune cytopenias and hypothyroidism. 36 of 50 with follow-up are alive (70%) with age range 6-44 years. Of the 36 living patients, 28 (78%) have had a HSCT. Causes of deaths include malignancy (6), infection (1), and HSCT complications (7). Long-term follow-up of patients with HSCT (up to 6 years) has revealed resolution of the infection susceptibility and eczema, no new cancers, and stabilization of vasculopathy.

    Conclusions: In addition to the well described manifestations of DOCK8 deficiency including eczema, allergy, recurrent sinopulmonary infections, skin viral infections and malignancy, our cohort revealed a relatively high incidence of liver disease, frequently associated with stool positivity for Cryptosporidia, as well as vasculopathy. Both of these clinical manifestations should be considered during preparation for HSCT as they may affect management through transplant. Autoimmunity has likely been over-estimated in prior descriptions of DOCK8 deficiency. Long-term follow-up after HSCT is needed to determine the prognosis from the vasculopathy, liver disease, and malignancy risk.

    (166) Submission ID#604115

    Transcriptome Analysis Reveals an Important Role for EXTL3 in Human Hematopoietic Cell Differentiation

    Yasuhiro Yamazaki1, Stefano Volpi2, Luigi D. Notarangelo1

    1Laboratory of Clinical Immunology and Microbiology, IDGS, DIR, NIAID, NIH, Bethesda, MD, USA

    2U.O. Clinica Pediatrica e Reumatologia, IRCCS Istituto Giannina Gaslini, Genova, Italy

    Introduction/Background: EXTL3 (Exostosin Like Glycosyltransferase 3) is an exostosin family member which initiates heparan sulfate (HS) chain biosynthesis and elongation. We have reported homozygous EXTL3 hypomorphic mutation (R339W) as a cause of immuno-osseous-dysplasia syndrome. Fourteen patients who have EXTL3 homozygous mutation were reported so far. Eight of them manifested T cell lymphopenia, and 5 presented with severe combined immunodeficiency (SCID) or Omenn syndrome. Using patient-derived induced pluripotent stem cells (iPSCs) as a model, we have previously reported that EXTL3 mutations affect differentiation to thymic epithelial progenitor cells as well as expansion of hematopoietic progenitor cells. Consistent with the latter, previous studies have suggested that mutations in other genes involved in HS biosynthesis affect hematopoietic stem cell (HSC) differentiation. However, the exact mechanisms by which EXTL3 mutations affect hematopoiesis are not known.

    Objectives: We tried to clarify gene expression difference in HSCs derived from wild-type, EXTL3 hypomorphic and EXTL3 knock-out (KO) human iPSCs.

    Methods: The control BJ iPSC line was engineered by CRISPR/Cas9 gene targeting. EXTL3 KO iPSCs were obtained which carried compound heterozygous EXTL3 mutations (c.1003_1004insT; c.1005_1006insGATATTT). HSC differentiation was induced using the STEMdiff hematopoietic kit (STEMCELL technologies). Bulk RNA from each iPS cells and each differentiated CD34+CD43+CD45+ was analyzed by RNA sequencing.

    Results: As compared to control iPSCs, patient-derived cells showed slightly lower capacity to generate CD34+CD43+CD45+ cells. On the other hand, EXTL3 KO cells showed no differentiation into CD34+CD43+CD45+ cells. Gene Set Enrichment Analysis showed enriched expression of genes involved in hematopoietic progenitor cell differentiation, regulation of hemopoiesis, and positive regulation of hemopoiesis in both control and patient-derived CD34+CD43+CD45+ cells compared to parental iPSCs. Moreover, these gene sets were more abundantly enriched in control than in patient-derived CD34+CD43+CD45+ cells. The gene set of Response to type I interferon was significantly enriched in control versus patient-derived CD34+CD43+CD45+ cells.

    Conclusions: These results confirm that EXTL3 plays an important role for HSC homeostasis in human cells. Because type 1 interferons play a role in HSC proliferation, the decreased type I interferon signature may account for the reduced number of HSCs that we have previously reported upon in vitro differentiation of EXTL3-mutated versus control-derived iPSCs.

    This study was supported by the Division of Intramural Research, NIAID, NIH, under protocol 16-I-N139.

    (167) Submission ID#604171

    A Case of Autoinflammatory Syndrome with Osteoporosis and Specific Antibody Deficiency

    Irina Dawson, MD1, Mark Ballow, MD2, Lori Broderick, MD, PhD3, Jolan Walter, MD, PhD4

    1Allergy and Immunology Fellow, Division of Allergy and Immunology, Department of Pediatrics, University of South Florida, St Petersburg, FL

    2Associate Professor, Division of Allergy and Immunology, Department of Pediatrics, University of South Florida, St Petersburg, FL

    3Assistant Professor, Division of Allergy, Immunology and Rheumatology, Department of Pediatrics, University of California, San Diego, CA

    4Associate Professor, Robert A. Good Endowed Chair and Division Chief, Division of Pediatric Allergy & Immunology, Department of Pediatrics, University of South Florida, Johns Hopkins All Children's Hospital, St. Petersburg, FL.

    Autoinflammatory syndromes are inherited disorders with an exaggerated inflammatory response with no specific trigger. The clinical phenotypes of variants of autoinflammatory syndromes may overlap.

    We report a case of a 13 year old male with prior diagnosis of specific antibody deficiency, Periodic Fever, Aphthous Stomatitis, Pharyngitis, Cervical Adenitis (PFAPA) syndrome, arthralgia and moderate atopic dermatitis.

    He was diagnosed at 3 years of age with specific antibody deficiency based on persistently low pneumococcal titers against repeat immunizations. Due to recurrent infections, he was placed on immunoglobulin replacement therapy (IgRT) at 8 years of age. IgRT was discontinued at 13 years of age due to full resolution in infections and patient demonstrated robust response to immunizations. Patient had lifelong history of recurrent fevers (every 5 weeks) associated with pharyngitis and aphthous ulcers consistent with diagnosis of PFAPA. As he became older these episodes became less frequent. Last episode of fever was over a year ago. The father had similar symptoms of recurrent fevers and oral ulcers as a child but currently remains asymptomatic. Paternal grandfather died of kidney disease.

    Patient has been generally in good health until recent year with intermittent abdominal pain, arthralgia and several long bone fractures with no history of prior trauma. A bone density scan revealed osteopenia and osteoporosis with a Z score of -2.2 of lumbar spine, -4.0 of left femoral neck, -3.1 of left hip.

    Given history of familial autoinflammatory disease, and antibody deficiency genetic testing was obtained which identified a pathogenic heterozygous variant of TACI and MEFV c.2082G>A (p.Met694lle). TACI mutation has been linked to antibody deficiency syndromes. Genetic study for family members is pending.

    The MEFV gene is associated with autosomal recessive familial Mediterranean fever (FMF) and has been reported in autosomal dominant FMF as well. FMF is characterized by recurrent episodes of fever associated with serositis, arthralgia, and arthritis. Patients with FMF have elevation in acute phase reactants during attacks with most returning to normal levels during the episode-free periods. Multiple studies have shown that patient with FMF have lower bone mineral density and Z-scores than the general population. Inflammation in FMF is thought to be mediated by several different cytokines (IL-1, IL-2, IL-6, IL-7, IL-8, IL-11, IL-15 and TNF-). These same cytokines play a role in osteoclast activity and bone resorption. It has been suggested chronic inflammation during acute attacks and subclinical inflammation during the disease-free period lead to bone loss and osteoporosis. Regular use of colchicine, the main treatment for FMF, may slow down osteoporosis.

    Beside careful monitoring of clinical and laboratory phenotype, genetic evaluation is an important step in distinguishing between overlapping entities and can prevent complication and promote targeted intervention.

    (168) Submission ID#604373

    When Mosquito Bite Allergy Is Treated with Bone Marrow Transplantation (BMT)

    Joseph A. Church, MD1, Ronald M. Ferdman, MD2, Neena Kapoor, MD3

    1Professor, Pediatrics, Children's Hospital Los Angeles and Keck School of Medicine of U.S.C.

    2Associate Professor, Pediatrics, Children's Hospital Los Angeles and Keck School of Medicine of U.S.C.

    3Professor, Pediatrics, Children's Hospital Los Angeles and Keck Schood of Medicine of U.S.C.

    A 5 year old previously healthy boy was referred for periodic fever/PFAPA and mosquito bite hypersensitivity. Eight weeks earlier he developed fever to 104F, mouth sores and exudative tonsillitis; a rapid strep screen was negative. One week later he developed moderate cervical lymphadenopathy and had a positive EBV Early Antigen antibody.. One month later he had several severe local reactions to mosquito bites. Each manifested 6-8 cm of erythema and induration with a 1+ cm bullae which left an ulcer after rupture and healed with a hypopigmented scar. The bites were accompanied by fever to 104F for 4 days. One febrile episode was treated with low dose prednisolone for presumed PFAPA, and the fever resolved within hours. His past history was positive for nasal allergy and mild asthma. His parents are not related: mom is of European-Indonesian and dad European-African (Creole ancestry. Testing prior to this visit showed normal IgG, IgA and IgM, elevated IgE (12,000 U/L) and normal CBC. Lymphocyte subsets revealed CD3+ 23% (1538/mcL), CD4+ 17% (1109/mcL), CD8+ 6% (363/mcL), CD19+ 9% (587/mcL), NK cells 67% (4435/mcL). On examination he appeared well with height at 86th%ile and weight at 58th%ile. There was no lymphadenopathy, hepatosplenomegaly or inflammed skin lesions; there was a 1cm round scar on the right plantar surface at the site of a prior mosquito bite. Laboratory studies confirmed NK lymphocytosis 64% (5459/mcL) and elevated IgE (29,600 U/L). Lymphoproliferation to mitogens, CD3/CD28, CMV and HSV were normal, but absent to tetanus and candida antigens. EBV antibodies reflected past infection (VCA-IgG+, VCA-IgM-, EBNA+); quantitative EBV PCR was >5,000,000 copies/mL whole blood. NK cytotoxicity and CD107a expression were decreased. Bone marrow NK analysis suggested conality. The patient was diagnosed with "hypersensitivity to mosquito bites with EBV-associated T-/NK lymphoproliferation." This disorder represents a subset of chronic active EBV (CAEBV) that is rarely seen outside of East Asia. The lack of organomegaly or lymphadenopathy with hyper-IgE and NK lymphocytosis and decreased NK function support the likelihood that NK cells are the target of EBV infection in this patient. This diagnosis may be a precursor to hemophagocytosis, liver necrosis or lymphoma/leukemia, and the only curative treatment is bone marrow transplantation. The patient's sister is a 10/10 HLA match. She is seropositive for past EBV infection, and she has no history of extreme reactions to mosquito bites. Genetic mutations that cause familial hemophagocytic lymphohistiocytosis have not been reported in CAEBV, and to the best of our knowledge familial cases of this disorder have not been identified. The response to BMT in this patient is pending.

    (169) Submission ID#604432

    Epidemiology of Anti-epileptic Drug Induced Hypogammaglobulinemia in a Tertiary Care Network

    ErinMarie Kimbrough, MD1, Keith Sacco, MD1, Ismael Carrillo-Martin, MD2, Natalia Chamorro-Pareja, MD3, Daniela Haehn, MD4, Alexei Gonzalez Estrada, MD5

    1Resident, Department of Medicine, Mayo Clinic Jacksonville

    2Research Trainee, Department of Medicine,Mayo Clinic Jacksonville

    3Research Trainee, Division of Pulmonary, Allergy, and Sleep Medicine, Mayo Clinic Jacksonville

    4Research Trainee, Department of Anesthesia and Perioperative Medicine, Mayo Clinic Jacksonville

    5Division of Pulmonary, Allergy, and Sleep Medicine, Mayo Clinic Jacksonville

    Introduction/Background: A number of case reports have described symptomatic hypogammaglobulinemia following administration of anti-epileptic drugs (AEDs), specifically lamotrigine, carbamazepine, and levetiracetam. The mechanism by which symptomatic hypogammaglobulinemia develops is unclear. We evaluated the prevalence and the clinical significance of hypogammaglobulinemia associated with use of these AEDs.

    Objectives: Our aim was to characterize the prevalence of AED-induced hypogammaglobulinemia, identify specific AEDs associated with hypogammaglobulinemia, and characterize the timeline to development of hypogammaglobulinemia after initiation of therapy.

    Methods: A retrospective, multicenter, electronic medical record review spanning 18 years identified patients with hypogammaglobulinemia who were on AED therapy (lamotrigine, carbamazepine, or levetiracetam). Patients were excluded if they had a pre-existing primary immunodeficiency (PID), malignancy, protein-losing enteropathy, or significant proteinuria. Patients on chronic immunosuppressive therapy, those without laboratory criteria for hypogammaglobulinemia, or those on one of the AEDs for less than one month were also excluded.

    Results: Of the 316 cases reviewed, 5 patients met our inclusion criteria. The median age was 35; 80% were adults, 80% were female, and 80% were white. Lamotrigine was implicated in 3/5 of the cases, carbamazepine in 2/5, and levetiracetam in 1/5. Tetanus and pneumococcal titers were available for 4/5 patients. Of those patients, 3/4 had protective titers to both per report with responses to >70% of the serotypes. Only one patient reported severe, recurrent infections while the remaining four had little to no symptoms. Interestingly, the patient with severe infections did have protective titers. Of the five laboratory proven hypogammaglobulinemia patients, one died of an infection, two have continued on the medication due to refractory seizures responsive only to these medications, and two are currently being tapered off of their AED.

    Conclusion: While it appears that AED-induced hypogammaglobulinemia is quite rare, it should be considered in a patient without other secondary causes of hypogammaglobulinemia on AED therapy. Many antiepileptics downregulate NFkB signaling suggestive that patients who develop symptomatic hypogammaglobulinemia may have hypomorphic mutations in the NFkB signaling pathway.

    (170) Submission ID#604503

    Autoimmune Lymphoproliferative Syndrome with Histopathologic Features of Castleman Disease

    Rachelle Lo, MD1, Dita Gratzinger, MD, PhD2, Elizabeth Keiser, MD, MPH3, Kay Chang, MD4, Yael Gernez-Goldhammer, MD, PhD5

    1Fellow, Allergy and Immunology, Division of Allergy and Immunology, Department of Pediatrics, Stanford University School of Medicine

    2Associate Professor, Department of Pathology, Stanford University School of Medicine

    3Fellow,General Surgical Pathology, Department of Pathology, Stanford University School of Medicine

    4Professor, Professor of Otolaryngology and Pediatrics, Stanford University, Department of Otolaryngology, Lucile Packard Children's Hospital at Stanford, Division of Pediatric Otolaryngology

    5Clinical Assistant Professor, Division of Allergy and Immunology, Department of Pediatrics, Stanford University School of Medicine

    Autoimmune Lymphoproliferative Syndrome (ALPS) results from defective apoptosis of lymphocytes mediated through the Fas/Fas ligand (FasL) pathway. The hallmark lab finding is an expansion of T cells that express the alpha/beta T cell receptor, but lack both CD4 and CD8 (double negative T cells) in the setting of normal or elevated lymphocyte counts. Patients present with chronic, nonmalignant, noninfectious lymphadenopathy or splenomegaly. For definitive diagnosis, patients need to have (1) a pathogenic mutation in FAS, FAS ligand or caspase 10 or (2) a defective FAS-induced lymphocyte apoptosis. We describe a probable case of ALPS with heterozygous mutation in FAS c.287A>G(p.His96Arg), a variant that has not been previously reported (his lymphocyte apoptosis assay is pending). Unique to this case is the patients Castleman disease-like features on pathology.

    A 15 year-old male referred from hematology clinic presented with an 8 year history of chronic lymphadenopathy, splenomegaly, anemia, and no underlying diagnosis. Malignancy had previously been excluded by bone marrow aspirate and biopsy 8 years prior. However, he had a right sided lymph node that had increased in size for the past 4 months. He was otherwise asymptomatic. A lymph node biopsy 7 years prior was reportedly normal. His exam demonstrated significant bilateral lymphadenopathy, greater on right, with an approximately 8 x 6 cm mobile right neck mass. He had splenomegaly palpated 7 cm down and across to midline. He was therefore admitted for excisional lymph node biopsy to evaluate for possible malignancy and labs were sent to evaluate for ALPS.

    Labs were supportive of ALPS. He had elevated T cell receptor alpha beta double negative T cells (TCR a/b DNTCs) in blood (10.5%). B12 level was elevated (>1000 pg/mL). Plasma soluble FASL level was elevated (5517 pg/mL). Interleukin-10 (IL-10) and IL-18 levels were elevated (88 and 909 pg/mL respectively). He had multilineage cytopenias: anemia with Hgb of 9.5 g/dL and neutropenia (absolute neutrophil count of 1380 K/uL). He had hypergammaglobulinemia with an IgG level of 2010 mg/dL. Broad infectious work-up was negative, including HIV, QuantiFERON, Cocci, Bartonella, Toxoplasma, Coxiella burnetii, EBV PCR and, CMV IgM.

    Lymph node biopsy showed no evidence of malignancy. Immunostains and flow cytometry showed the presence of expanded TCR a/b DNTCs in the lymph node, consistent with ALPS. Interestingly, lymph node histology showed morphologic features typical of plasma cell variant Castleman disease. Numerous Castleman-like follicles showed typical regressive changes with onion-skinning morphology. Paracortical hyperplasia with sheets of plasma cells was noted. There was negative staining for HHV8 (a well-known cause of plasma cell variant Castleman disease).

    The diagnosis of idiopathic multicentric HHV8-negative Castleman disease was excluded by definition in the setting of ALPS, per evidence-based consensus criteria published in 2017. In addition, our patient did not show any symptoms typically associated with it, such as fever, night sweats, weight loss, weakness or fatigue. Should his FAS-induced lymphocyte apoptosis be defective (in 2 separate assays), this would confirm his ALPS-FAS diagnosis and we would start the patient on sirolimus.

    (171) Submission ID#604721

    SLC46A1 Deficiency Presenting as Hypogammaglobulinemia and Wide Clinical Manifestations

    Agostina Llarens, MD1, Carolina Dorfman, MD1, Daniela Di Giovanni, MD2, Andrea Gomez Raccio, MD2, Gisela Seminario, MD2, María Isabel. Gaillard, MSc3, Patricia Carabajal, MD4

    1Physician – Immunology trainer, Children's Hospital Ricardo Gutierrez

    2Physician – Immunologist, Children's Hospital Ricardo Gutierrez

    3Biochemist, Grupo de Inmunología Hospital de Niños

    4Head of Immunology Unit, Children' s Hospital Ricardo Gutierrez

    Introduction: SLC46A1 gene encodes the proto-couple folate transporter (PCFT), which supports intestinal folate uptake, and participates in folate transport into the central nervous system. SLC46A1 mutations cause PCFT defects, resulting in low folate levels in serum and cerebrospinal fluid. Hereditary folate malabsorption (HFM) is a rare, autosomal recessive disorder with PCFT deficiency resulting in cerebral folate deficiency. Most of the patients present megaloblastic anaemia, moderate pancytopenia in the first few months of life, failure to thrive, diarrhoea and/or later onset neurological symptoms including seizures and developmental delay.

    Immunodeficiency in HFM can manifest itself with hypogammaglobulinemia with normal T-cell function. B-cell precursor compartment seems to be particularly vulnerable to folate deficiency in some HFM patients. This immunodeficiency can be restored with specific treatment with folic acid.

    Aim: To describe a female patient with a homozygous pathological variation in the SLC46A1 gene.

    Results: A 17 months old girl, born of non-consanguineous parents. She started at 3 months old with diarrhoea due to rotavirus, low weight and bicytopenia with normal bone marrow aspiration. She presented low levels of folic acid 1.5ng/ml (NV 3.1-20.5 ng/ml) at first thought due to secondary to malnutrition. Treatment with folic acid supplementation was administrated, improving platelets counts. At 5 months old she presented steatorrhea with severe perianal panniculitis which required surgical treatment. No germs were rescued after a skin biopsy. Moreover, she suffered from a respiratory infection due to Picornavirus with two episodes of pneumothorax which required intensive care. At that moment IVIG treatment was administered due to hypogammaglobulinemia and clinical severity.

    Chronic diarrhoea worsened with bloody depositions. Three rectal ulcers were found in the gut biopsy. Bowel inflammatory disease was suspected and mesalazine administration was started with weight improvement.

    Furthermore, at 10 months old she presented 3 status epilepticus, with pathological EEG and normal MRI; one of them related to a CMV infection, successfully treated.

    In the immunological evaluation IgG and IgA were low with normal IgM and IgD. The protein-antibody response was not evaluated. She presented normal lymphocyte and T cells extended populations, T cells proliferation assay, DHR, Treg cells, complement, CD107a expression, alpha-fetoprotein, without autoantibodies

    A molecular panel testing was done by NGS and a homozygous variant in SLC46A1 gene was found, causing impaired intestinal folate absorption.

    Conclusion: HFM should be considered in the diagnosis of patients with cytopenias and hypogammaglobulinemia in order to provide specific treatment.

    HFM has wide clinical manifestations, not only with megaloblastic anaemia and neurological impairment but also with gastrointestinal and skin manifestations. With folate treatment, clinical and immunological defects can be normalized.

    (172) Submission ID#604754

    A Case Report of Focal Epithelial Hyperplasia (Hecks Disease) with Elevated Tumor Necrosis Factor Alpha

    Zoya Treyster, MD1, Sara Sussman, MD2, David Rosenthal, DO, PhD3

    1Fellow, Division of Allergy & Immunology, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    2Fellow, Department of Pediatrics, Zucker School of Medicine at Hofstra Northwell School of Medicine

    3Assistant Professor of Medicine and Pediatrics, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell

    Introduction: Multifocal epithelial hyperplasia (MEH), or Hecks Disease, is a rare, benign infection of the mucosa caused by human papilloma virus (HPV). Clinically, MEH manifests as numerous painless, soft, sessile papules or plaques, and typically occurs in the labial, lingual, and buccal mucosa. MEH lesions are usually associated with HPV types 13 and 32, and seen more commonly in patients of Caribbean or Central/South American descent. Prior studies in adults have shown that Tumor Necrosis Factor Alpha (TNF) promotes HPV, and may influence duration of HPV infection.

    Case: We present a five-year-old full term male of Haitian descent referred for assessment of multiple flesh colored, papular lesions on the buccal and labial mucosa that had persisted and quantitatively increased over one year, although some lesions regressed. He had no pain or difficulty eating. Medical history significant for one seizure; negative for infection. No family history of infection, immunodeficiency, consanguinity, or miscarriage. Head and neck examination failed to reveal cervical lymphadenopathy, masses, or hypertrophy in the salivary glands. Intraoral examination revealed multiple papular nodules, mostly flat although some were corrugated. The greatest concentration was noted on the lower left labial surface extending to the mucosal vermillion interface, not involving the vermillion or commissure region. Lesions extended into the mandibular vestibule and the left buccal mucosa. No other lesions were noted on extremities, genitalia, or any other visualized mucosal surface. Based on history and exam, he was diagnosed with MEH. White blood cell count, neutrophils, lymphocytes, CD4 and CD8 T cell, B cell, NK cell enumeration, and immunoglobulin panel were normal for age. Tetanus and Streptococcus pneumoniae titers were protective. Cytomegalovirus IgG and IgM were negative. Epstein-Barr Virus IgG was positive, IgM and Early Antigen Ab negative. Serology was significant for elevated TNF (84 pg/mL; reference range <22pg/mL) while interferon gamma and interleukins 1, 2, 4, 5, 6, 8, 10, 12, 13, and 17 were normal, as was IL-2 receptor CD25. One month after the initial visit, lesions were stable and unchanged. Nine-valent HPV vaccination was considered, but not administered.

    Conclusions: MEH is a rare but benign disease caused by HPV. Awareness of the disease and its course is important to prevent unnecessary expanded immunodeficiency work-up and possible procedures to eliminate lesions. Although mucosal immunity can be site specific, especially with HPV, our understanding of T-cell cytokine and chemokine responses to HPV in cervical and laryngeal lesions may be instructive. The mechanism which allows HPV persistence in MEH is not characterized, but it likely is due to increased viral persistence and an inability for the host immune response to successfully induce viral latency and successful containment. Elevated TNF levels, with normal levels of IL-2, IL-6, IL-8, IL-10, may correlate with decreased clearance of HPV and prolonged duration of MEH. It remains unclear if viral persistence is the cause of, or the sequela of, increased TNF. Longitudinal monitoring of cytokine (TNF, IL-2, IL-6, IL-8, IL-10) and chemokine (CCL17, CCL18, CCL19, CCL20, CCL21, and CCL22) serum concentrations may be useful biomarkers for disease resolution.

    (173) Submission ID#604905

    Patient Education with a Self-Efficacy Focus for Adult Autosomal Dominant Hyper IgE Syndrome Patients

    Amanda Urban, DNP, CRNP1, Dirk A. Darnell, MA, RN2, Ladan Foruraghi, CRNP3, Alexandra F. Freeman, MD4

    1Nurse Practitioner, Clinical Monitoring Research Program Directorate, Frederick National Laboratory for Cancer Research sponsored by the National Cancer Institute

    2Nurse Case Manager, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    3Nurse Practitioner, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    4Director, Primary Immune Deficiency Clinic, Laboratory of Clinical Immunology and Microbiology, NIAID, NIH, Bethesda, MD, USA

    Introduction: Autosomal dominant Hyper IgE (Jobs) Syndrome is a rare primary immunodeficiency characterized by eczema and sinopulmonary infections as well as musculoskeletal and vascular complications. As in all chronic illnesses, patient education is an ongoing need. In the rare disease population, patient education is especially important as patients must be able to explain their unique healthcare concerns in a variety of medical settings. We focused on AD-HIES, due to our relatively large cohort of patients, the frequent lack of classic signs of illness often impairing diagnosis of severe infection, and the diverse non-immunologic clinical features of this disease.

    Objectives: We aimed to increase understanding of the clinical manifestations of AD-HIES to promote earlier recognition of symptoms and to increase self-efficacy for symptom management in the adult HIES population.

    Methods: Adult patients were asked to participate in a patient education project. Demographic information was collected from participants. They also completed a 12-item multiple choice test about symptom recognition in AD-HIES and PROMIS Self-Efficacy for Managing Symptoms, an 8-item validated survey. Then, patient education handouts that focused on pulmonary symptoms, eczema, bone health, and cardiovascular complications were reviewed with the participant. Six weeks later, participants were asked to repeat the 12-item test and the self-efficacy Survey. The demographic information, test, and self-efficacy were collected anonymously.

    Results: 33 participants provided demographic information, completed the test and the Self-Efficacy Survey. Of the 33 participants, 15 were male and 17 were female. Participants ranged in age from 18 to 66 years. 22/33 (67%) reported looking for information about AD-HIES using search engines and most patients (91%) report that they have been given information about AD-HIES from a doctor. 19/33 (58%) participants identified pulmonary symptoms as the symptom that concerns them most and 10/33 (30%) participants identified more than one symptom of concern. 25 participants returned the second test and second survey. The mean test score increased from 9.08 to 10.28 with 23/25 participants achieving a score of 9/12 or higher. The self-efficacy scores were unchanged with a mean score of 50.08 before reviewing the patient education handouts and 50.13 after.

    Conclusions: Participant feedback to this project was generally positive. AD-HIES patients are seeking information and an educational intervention can improve their understanding of disease. Self-Efficacy results were mixed and unchanged overall, but suggest that AD-HIES patients manage symptoms as well as other patients with chronic illnesses. Patient education should continue at each encounter. This project can be expanded to include more topics, pediatric patients, and other rare disease populations. Funded by the NCI Contract No. HHSN261200800001E

    (174) Submission ID#605066

    T and NK Cell Dysfunction Arising from BCL11B Deficiency

    Samuel Chiang, PhD1, Sharat Chandra, MD, MRCPCH2, Vijaya Chaturvedi, B.S.3, Erika Owsley, B.S.3, Jack Bleesing, MD, PhD4, Brian Dawson, PhD5, Rebecca A. Marsh, MD6, Miao Sun, PhD7

    1Research Associate, Cincinnati Childrens Hospital Medical Center

    2Assistant Professor, UC Department of Pediatrics, Division of Bone Marrow Transplantation and Immune Deficiency, Cincinnati Childrens

    3Research Associate, Cincinnati Children's Hospital

    4Professor, Division of Bone Marrow Transplantation and Immune Deficiency, Cincinnati Childrens

    5Director, Clinical Laboratories, Division of Human Genetics

    6Associate Professor, Division of Bone Marrow Transplantation and Immune Deficiency, Cincinnati Childrens Hospital Medical Center

    7Assistant Director, Clinical Laboratories, Division of Human Genetics

    Introduction: BCL11B plays an important role in the development and maintenance of the immune system and the central nervous system. Expression of BCL11B represses NK and myeloid factors while inducing T cell lineage genes in thymocytes at the DN2 stage. Conditional loss of Bcl11b expression in murine thymocytes leads to T cell deficiency while complete knockout of Bcl11b was fatal within a few days of birth. Recently, specific heterozygous BCL11B mutations have been reported in 11 individuals with global development delay. However, only 2 of these cases, both carrying heterozygous missense variants, had low TREC values with 4 other cases having frequent infections. Little is known regarding the impact of BCL11B on human NK and T cell function.

    Methods: We identified a novel heterozygous truncating mutation in BCL11B in an infant who was first detected by TREC newborn screening. She subsequently developed severe autoimmune hemolytic anemia at the age of 3 months. We used standard immunoblotting and flow cytometry methods to assess protein expression and the impact of this BCL11B mutant on T cell and NK cell development and function.

    Results: The patient has a novel single base-pair deletion in the BCL11B gene, which is predicted to produce a truncated protein with the loss of 3 of 6 zinc finger domains in BCL11B. Immunoblotting of T cell blast lysates revealed a reduced BCL11B expression in the patient consistent with the heterozygous defect in BCL11B but also generated a novel band with a smaller molecular weight that we postulate represents the truncated protein product. While mitogen responses to ConA and PHA were normal, both CD4+ and CD8+ T cell counts were decreased, especially CD4+ naïve and CD4+CD31+ naïve T cells, suggesting reduced thymic output. The function of TH1 cells was skewed with reduced IL-2 production but increased IFN levels after PMA and ionomycin stimulation. Moreover, T regulatory cell counts were below normal range. NK cell counts were normal but these were mostly CD56bright NK cells. Of the few CD56dim NK cells that presented, approximately half did not express CD16, the Fc receptor for ADCC. Perforin was only present in CD16 expressing NK cells. As such, anti-CD16 stimulation understandably led to low but not defective NK cell degranulation. Function after stimulation with K562 cells was normal when controlled for NK cell counts.

    Conclusion: We report a novel BCL11B truncating mutation with a leaky SCID phenotype that manifested with T-cell lymphopenia and autoimmunity. Lowered thymic-derived naïve T and regulatory T cells, skewed TH1 cytokine response, and incomplete NK cell development suggests that BCL11B is important for the development and differentiation of multiple lymphocyte lineages.

    (175) Submission ID#605266

    Refractory Giardia Infection in a Patient with Common Variable Immune Deficiency

    Ekta Kakkar, MD1, Laila Woc-Colburn, MD2, Joud Hajjar, MD, MS3

    1Allergy and Clinical Immunology Fellow, National Jewish Health and The University of Colorado

    2Associate Professor in Infectious Diseases, Baylor College of Medicine

    3Assistant Professor, Baylor College of Medicine, 1Texas Childrens Hospital Center for Human Immunobiology and Division of Immunology, Allergy and Rheumatology

    Introduction: Chronic diarrhea is one of the most common gastrointestinal complaints in patients with common variable immune deficiency (CVID) and can lead to life-threatening complications such as malabsorption and malnutrition. Chronic diarrhea in CVID could be caused by infections, an inflammatory bowel disease-like picture, as well as malignancy. Giardia lamblia is one of the most common parasites causing diarrhea in CVID (up to 40%), and can be refractory in these patients, leading to villous atrophy, weight loss, and failure to thrive.

    Case Report: A 41-year-old female with a history of CVID presents with chronic diarrhea and significant weight loss. Her CVID was diagnosed by hypogammaglobulinemia (low levels of IgG, IgM, and IgA), inadequate responses to protein and polysaccharide-based vaccines, decreased memory B cells (CD19+CD27+ 0.5%), and recurrent sinopulmonary infections. She was started on immune globulin replacement therapy and had significant improvement in her rate of infections. Four years before her presentation to our center, she developed chronic, severe diarrhea. Work up revealed Giardia lamblia infection on endoscopy and colonoscopy. Biopsy showed intraepithelial lymphocytes, villous blunting, and atrophic gastritis with rare plasma cells concerning for non-infectious enteropathy related to her CVID, in addition to the high burden of Giardia organisms. She was initially treated with metronidazole for several weeks. However, her diarrhea did not improve, and she developed significant peripheral neuropathy leading to lower extremity weakness and limited mobility. Her diarrhea persisted and was associated with approximately a 20-pound weight loss. Repeat endoscopy and colonoscopy two years later showed persistent high burden Giardiasis of the small intestine, as well as reactive lymphocytic infiltrates and atrophic gastritis. She was treated with nitazoxanide but continued to have diarrhea, and her stool continued to show trophozoites. Given the significant inflammation and the lack of response to multiple antiparasitic agents, she was referred to our center for further evaluation. She was started on oral budesonide (9 mg daily) and oral immune globulin (20 grams weekly for 12 weeks). With this regimen, she had significant improvement in her diarrhea with a 10-pound weight gain. Repeat colonoscopy showed considerable improvement in inflammation and resolution of her Giardia infection, though her stool antigen continues to be positive.

    Conclusions: Persistent diarrhea in our patient is most likely due to a combination of CVID enteropathy and Giardiasis. A prolonged course of metronidazole and later nitazoxanide did not control her diarrhea and led to significant side effects. Switching to an immunomodulatory approach significantly decreased the inflammation in her bowel and may even have helped to reduce the burden of Giardia in the gut. Targeting both underlying bowel inflammation as well as active infection in CVID patients with chronic diarrhea might be needed to control symptoms.

    (176) Submission ID#605352

    Sphingosine-1-phosphate Lyase Deficiency Identified by Newborn T Cell Receptor Excision Circle Screening for Severe Combined Immunodeficiency

    Cullen M. Dutmer, MD, Austin A. Larson, MD, Elena WY. Hsieh, MD

    1Assistant Professor, Children's Hospital Colorado - University of Colorado School of Medicine

    Introduction: Sphingosine-1-phosphate (S1P) is a lipid chemoattractant that is critical for lymphocyte egress from lymphoid organs. Following a S1P concentration gradient maintained by S1P lyase ubiquitously expressed in tissues, lymphocytes within lymphoid organs are drawn to efferent lymph and blood unless their S1P receptor is internalized or downregulated. Owing to diminished degradation of not only S1P, but also other sphingoid bases, deleterious mutations in SGPL1 (encoding S1P lyase) perturb sphingolipid catabolism in numerous tissues. Correspondingly, human S1P lyase deficiency results in multiorgan dysfunction including kidney, skin, endocrine gland, and neurologic impairment alongside expected lymphopenia. Although severe T cell lymphopenia (<300 cells/microliter) rivaling that of severe combined immunodeficiency (SCID) can be seen in patients with S1P lyase deficiency, no such patients have been identified by newborn screening of T cell receptor excision circle (TREC) counts, which are a surrogate measure of effective T cell production. Herein, we describe an infant boy with an undetectable TREC count at birth who was found to have two novel, biallelic SGPL1 mutations resulting in S1P lyase deficiency.

    Case Description: A 1-day-old boy with a preceding history of fetal hydrops is born at a gestational age of 36 weeks and presents with renal failure, anasarca, and respiratory failure. TREC analysis of a dried blood spot obtained at 24 hours of life reveals zero copies/microliter. Subsequent peripheral blood studies show profound lymphopenia, with diminished CD3+ T (129/microliter; 96 CD4+, 27 CD8+), CD19+ B (130/microliter), and CD16/56+ natural killer (124/microliter) cell counts. Recent thymic emigrants are reduced (11.3% of CD4+ T cells are CD45RA+CD31+), as is the ratio of naïve-to-memory CD4+ T cells (63% CD45RA+, 37% CD45RO+). Expedited whole genome sequencing identifies two novel variants in SGPL1 a paternally inherited splice site variant (c.1566+2T>C) predicted to impact a canonical splice donor site, and a maternally inherited missense change (c.854G>A; p.Cys285Tyr) located in a well-established functional domain of S1P. In addition to nephrotic syndrome and lymphopenia, the patient displays evidence of adrenal insufficiency and has increased plasma levels of sphingoid bases and ceramides. Before further analyses could be pursued, the infant dies at 40 days of age due to ongoing complications of renal failure and eventual cardiorespiratory failure.

    Summary: We report the first case of S1P lyase deficiency identified by newborn TREC screening for SCID. As SGPL1 is not included in most commercially-available, SCID-tailored gene panels, S1P lyase deficiency would be missed by conventional genetic testing. Therefore, analysis for variants in SGPL1 should be considered in neonates with low-to-undetectable TREC counts, nephrotic syndrome, and other suggestive sequelae.

    (177) Submission ID#605673

    Case of WHIM Syndrome with Unique CXCR4 Variant

    Hassan A. Ahmad, MD1, G. Wendell Richmond, MD2

    1Allergy/Immunology Fellow, Rush University Medical Center

    2Allergist/Immunologist, Rush University Medical Center

    Introduction/Background: WHIM Syndrome (warts, hypogammaglobulinemia, recurrent infections, and myelokathexis) is a rare autosomal dominant primary immunodeficiency. It is caused by a defect in the gene encoding the chemokine receptor CXCR4. This receptor, along with the associated ligand CXCL12, regulates leukocyte migration. We present the case of a 40-year-old female, who presented after she self-identified the signature signs of WHIM syndrome in herself and multiple family members.

    Objectives: We present the case of a 40-year-old female who presented with a history of recurrent warts, leukopenia of unknown cause, and recurrent infections as a child. As a child, she experienced multiple ear and sinus infections, along recurrent warts on her upper and lower extremities that have persisted to this day. Furthermore, during a routine examination when she was 14-years-old, she had a complete blood count drawn significant for leukopenia. No further workup was undertaken at that time. When continued leukopenia was noted at the age of 30, referral to a hematologist and a bone marrow biopsy was completed. Bone marrow was significant for myelokathexis with borderline hypercellular marrow for patient age (80% cellularity), and normal cell line quantity. A trial of neupoegen was undertaken, without significant improvement. Her family history is significant for father and brother with both leukopenia and recurrent warts.

    Results: Genetic analysis showed a heterozygous pathogenic variant in the CXCR4 gene, C.1012_1015dup (p.Ser339Phe fs*6). Recent complete blood count was significant for a total WBC count of 1.0 K/uL, with a differential consisting of 30% neutrophils and 57% lymphocytes. Lymphocyte subsets were significant for quantitatively low CD3+, CD8+ and CD19+ subsets, with normal numbers of CD4+ and NK cells. Immunoglobulin levels revealed an IgG of 835 mg/dL, IgA of 145 mg/dL, and IgM of 54 mg/dL; IgG anti-diphtheria and tetanus titers were protective, however, none of the 23 S. pneumoniae serotype titers were > 1.3 ug/mL. Mitogen (PHA, ConA and PWM) and antigen (Candida and tetanus) stimulation of lymphocytes were normal for all stimuli.

    Conclusions: We present the case of a 40-year-old female with a history of recurrent infections, warts, and myelokathexis. On genetic analysis, she is noted to have a pathogenic mutation of the CXCR4 gene. The substitution of a phenylalanine for a serine decreases one of the seven serine phosphorylation sites in the carboxy tail of the molecule that occurs upon binding to its ligand, CXCL12 (SDF1). Additionally, the variation generates a premature stop condon terminating the remainder of the carboxy terminal amino acids including Ser346-7, known to have a role in carboxy terminial beta-arrestin binding. Failure to generate adequate beta-arrestin binding sites leads to prolonged CXCR4 CXCL12 interaction resulting in myelokathexis.

    (178) Submission ID#605697

    Low Dose Azithromycin Prophylaxis Reduces Respiratory Exacerbations in Patients Affected by Primary Antibody Deficiencies : A Multicenter, Double-blind, Placebo-controlled Randomized Clinical Trial

    Cinzia Milito, MD, PhD1, Federica Pulvirenti, MD, PhD2, Francesco Cinetto, MD, PhD3, Maria Carrabba, MD, PhD4, Giovanna Fabio, MD, PhD4, Andrea Matucci, MD5, Giuseppe Spadaro, MD6, Baldassarre Martire, MD7, Alessandro Plebani, MD, PhD8, Carlo Agostini, MD, Ph9, Stefano Tabolli, MD, PhD10, Isabella Quinti, MD, PhD10

    1Department of Molecular Medicine Sapienza University of Rome

    2Department of Molecular Medicine Sapienza University of Rome, Rome, Italy

    3Clinical Immunology- Padova Univ. Hospital, Dpt. of Medicine-DIMED, Padua, Italy, Padua, Italy

    4Fond. IRCCS Ca' Granda Ospedale Maggiore Policlinico, Dpt of Internal Medicine, Milan, Italy, Milan, Italy

    5Immunoallergology Unit- Policlinico di Careggi- Dpt. of Biomedicine, Firenze, Italy

    6Allergy and Clinical Immunology- Univ. of Naples Federico II-, Dpt. of Translational Medical Sciences, Naples, Italy

    7Pediatric Hospital, Bari, Italy

    8Pediatrics Clinic and Institute for Molecular Medicine A. Nocivelli- Univ. of Brescia, Dpt. of Clinical and Experimental Sciences-, Brescia, Italy, Brescia, Italy

    9Clinical Immunology- Padova Univ. Hospital, Dpt. of Medicine-DIMED, Padua, Italy, Padua, Italy

    10Department of Molecular Medicine Sapienza University of Rome, Rome, Italy

    Background: Lacking protective antibodies, patients with Primary Antibody Deficiencies (PAD) suffer from frequent respiratory infections leading to chronic pulmonary damage. Macrolides prophylaxis has been proven effective to successfully manage chronic lung diseases as cystic fibrosis, bronchiectasis, COPD. We conducted a trial to evaluate the efficacy and safety of orally low-dose azithromycin prophylaxis when added to the usual care in PAD patients.

    Methods: A 3-year, phase II, prospective, multicenter, randomized, double-blind, placebo-controlled trial on PAD patients (age 18-74 years) with chronic infection-related pulmonary disease. Patients received azithromycin 250 mg or placebo once daily three-times a week for 24 months. The primary outcome was the decrease of annual episodes of respiratory exacerbations. Secondary endpoints included: time to the first exacerbation, number of hospitalizations, additional doses of antibiotics, Health Related Quality of Life measures, and safety.

    Results: Forty-four patients received azithromycin and 45 patients received placebo. The mean number of exacerbations was 3·6 per patient-year (95%CI 2·5-4·7) in the azithromycin arm, and 5·2 (95%CI 4·1-6·4) in the placebo arm (p=0·02). In the azithromycin group the HR for having an acute exacerbation was 0·5 (95%CI 0,3-0·9, p=0,03) and the HR for hospitalization was 0.5 (95%CI 0,2-1·1) (p=0·04). The rate of additional antibiotic treatment per patient-year was 2·3 (95%CI 2·1-3·4) in the intervention and 3·6 (95%CI 2·9-4·3) in placebo groups (p=0·004). Improvement in HRQofL was observed in intervention group. Azithromycins safety prole was comparable with placebo.

    Conclusion: In PAD with respiratory exacerbation, azithromycin prophylaxis led to reduction of exacerbation episodes, of additional courses of antibiotics, and of risk of hospitalization. Given the deleterious effects of respiratory diseases adding azithromycin to PAD treatment should be considered as a valuable option.

    (179) Submission ID#605776

    Severe Necrotic Reaction to 23-valent Polysaccharide Pneumococcal Vaccine in a Patient with STAT3 Deficiency

    Mervin C. Piñones, MD1, Cecilia Vizcaya, MD2, Guillermo Matamala3, Rodrigo Hoyos, MD4, Arturo Borzutzky, MD4

    1Pediatric Immunology And Rheumatology Fellow, Pontificia Universidad Católica de Chile

    2Pediatric Infectious Disease Specialist, Pontificia Universidad Catolica de Chile

    3Biotechnology, Biotechnologist at Immunology Laboratory, Pontificia Universidad Catolica de Chile

    4Pediatric Immunologist, Pontificia Universidad Catolica de Chile

    Background: The autosomal-dominant hyper-IgE syndrome (HIES), is a primary immunodeficiency caused by mutations in signal transducer and activator of transcription 3 (STAT3) that leads to defective Th17 immunity. Adverse reactions following 23-valent pneumococcal polysaccharide vaccine (PPSV23) have been reported in 75% of STAT3-HIES patients, including severe local reactions that appear to be specific to this vaccine.

    Case report: We present the case of a six-year-old girl, second child of non-consanguineous healthy parents, that developed an extensive inflammatory skin reaction at the vaccination site following a single dose of PPSV23. The vaccine was prescribed due to history of recurrent respiratory tract infections and an incomplete vaccine calendar with no previously administered pneumococcal vaccines. The reaction began after 2 hours with local erythema and edema at vaccination site, expanding in 48 hours to a phlyctenular lesion with no well-defined borders. Within the first 3 weeks, it progressively evolved to a deep necrotic lesion that required surgical debridement. The subsequent skin defect required surgical repair with a split-thickness skin graft from her right thigh as the donor site. The complete wound healing process took about 5 months, leaving a large scar (figure).

    The patient had a longstanding history of recurrent infections with multiple hospitalizations including severe neonatal pneumonia that required respiratory support, a colon perforation with secondary peritonitis and septic shock that required a hemicolectomy at 8 months of age, recurrent oral candidiasis, recurrent pneumonias of different lobes, recurrent acute otitis media, a cervical phlegmon, three episodes of dental abscess and multiple kidney abscesses due to Gram-negative bacteria treated with intravenous antibiotics and surgical drainage. Family history is notable for an older sibling that died due to sudden infant death syndrome. The patients mother has large and wide nose suggestive of STAT3-HIES phenotype, but no history of infections. Immunological work up showed mild eosinophilia (850 cells/mm3), elevated IgE (1850 mg/dl), normal IgG, IgA, IgM and lymphocyte subsets (CD3, CD4, CD8, CD16, CD56). Peripheral Th17 cells were markedly decreased (0.6% vs. 3.7% of normal control). Specific pneumococcal antibodies evaluated 1 month after PSV23 revealed 5/10 serotypes in protective levels. High resolution thorax CT showed multilobar bronchiectasis. Echocardiogram and total spine x-rays were normal. STAT3-HIES was suspected with a National Institutes of Health score of 40. A novel heterozygous missense variant in STAT3 affecting the SRC homology 2 (SH2) domain (p.Lys591Glu) was found by next-generation panel sequencing. A variant in the same position (p.Lys591Met) has been previously reported in a HIES patient (ClinVar). Currently, she is on monthly IVIG and prophylactic antibiotics (cotrimoxazole, azithromycin and fluconazole).

    Conclusions: The case presented raises awareness on the risk of severe local adverse reactions to PPSV23 in STAT3-HIES patients. The etiology of such reactions is unclear and warrants further study. The benefits and risks of immunizing STAT3-HIES patients with PPSV23 should be weighed carefully by medical providers.

    Acknowledgments. Genetic sequencing was kindly provided by Drs. Raif Geha and Janet Chou at the Division of Immunology, Allergy, Rheumatology and Dermatology, Boston Children's Hospital, Harvard Medical School.

    figureas

    (180) Submission ID#605833

    DOCK8 Immunodeficiency in a Malay Family from Malaysia: a Family Study

    Intan Juliana Abdul Hamid, MD, MMed, PhD1, Nik Khairulddin Nik Yusoff, MBBS, MRCP, FRCPCH, DTM&H2, Mariana Daud, MD, Mmed3, Siti Mardhiana Binti. Mohamad, MD, PhD4, Ilie Fadzilah Hashim, BSc, MSc, PhD5, Zarina Thasneem Zainudeen, BSc, MSc, Dphil6, Bina Menon, MBChB, MRCP, DTM&H7, Elena Sigmund, BSc, MSc, PhD8, Bodo Grimbacher, MD9, Amir Hamzah Abdul Latiff, MBBS, MMed, MRCP, FACAAI, FAAAA10, Lokman Mohd Noh, MBBS, DCH,MRCP,FRCPE,Cert.11

    1Paediatric Immunologist, Primary Immunodeficiency Diseases Group, Regenerative Medicine Cluster, Institut Perubatan & Pergigian Termaju, Universiti Sains Malaysia

    2Consultant Paediatric Infectious Disease, Department of Paediatrics, Hospital Raja Perempuan Zainab II, Kota Bharu, Kelantan, Malaysia

    3Fellowship in Paediatric Respiratory and Sleep Medicine, Consultant Paediatric Respiratory,

    Department of Paediatrics, Hospital Raja Perempuan Zainab II, Kota Bharu, Kelantan, Malaysia

    4Clinical Scientist, Primary Immunodeficiency Diseases Group, Regenerative Medicine Cluster, Institut Perubatan & Pergigian Termaju, Universiti Sains Malaysia

    5Scientist, Primary Immunodeficiency Diseases Group, Regenerative Medicine Cluster, Institut Perubatan & Pergigian Termaju, Universiti Sains Malaysia

    6Scientist, Primary Immunodeficiency Diseases Group, Regenerative Medicine Cluster, Institut Perubatan & Pergigian Termaju, Universiti Sains Malaysia

    7Consultant Paediatrician, Institut Pediatrik, Hospital Kuala Lumpur, Malaysia

    8Scientist, Centre of Chronic Immunodeficiency, Universitatsklinikum Freiburg, Germany

    9Scientific Director and Consultant, Centre of Chronic Immunodeficiency, Universitatsklinikum Freiburg, Germany

    10Clinical Immunologist and Allergy, Allergy and Immunology Centre, Pantai Hospital, Kuala Lumpur, Malaysia

    11Fellowship Immunology, Paediatric Immunologist Consultant, Hospital Kuala Lumpur, Jalan Pahang, 50586 Kuala Lumpur, Wilayah Persekutuan

    Abstract (Max 500 words)

    Introduction: DOCK8 deficiency is a rare primary immunodeficiency characterized by susceptibility to viral infections, atopic eczema, defective T-cell activation and TH17 differentiation, impaired eosinophil homeostasis and dysregulation of IgE. To date, there are no reported cases from Malaysia.

    Objective: We aimed to describe the clinical, immunological profile and mutational analysis of three siblings of consanguineous parents, presented with Hyper-IgE and lymphopenia between the years 1998 and 2012, which were solved by mutational analysis of the second and third siblings.

    Methods: Clinical data and investigation results were collated from the medical record. Scoring of the symptoms and physical examination findings using NIH score was performed. T, B, NK lymphocyte subsets and serum IgG, IgA, IgM, total IgE quantification, lymphocyte proliferation test and pneumococcal specific antibody response were performed. Mutational analyses were performed in Freiburg, Germany.

    Result: Three siblings presented at different time points over a 20-year span with raised IgE levels, recurrent infections, eczema, hyper-eosinophilia and bronchiectasis. The NIH scores for hyper-IgE syndrome (HIES) ranged from 39 54. We also documented two serious infections in the siblings, which were disseminated Cryptococcus neoformans and Salmonella sp. Immunological results showed T-cell lymphopenia, defective T-cell proliferation, decreased IgM, raised IgE, hyper-eosinophilia and defective pneumococcal antibody responses present but not in all 3 siblings. We identified a large deletion in DOCK8 starting from exon 30-48 in 2 of the siblings from mutational analysis performed. We will proceed with next generation sequencing and DOCK8 protein assay in Malaysia to further characterize the defect.

    Conclusion: Our on-going study is the first description of DOCK8 in a family from Malaysia. The diagnosis of DOCK8 should be suspected in cases with raised IgE levels, recurrent infections and lymphopenia, despite no warts infection in the history. This study emphasized the importance of international research collaboration and networking in solving complicated cases.

    The following grants are acknowledged: 1. RUI 1.1001/CIPPT/812036 (USM) 2. BMBF 01 EO003 (Freiburg)

    The authors would like to thank the Director General of Health of Malaysia for permission to publish this scientific presentation.

    Key words

    Primary immunodeficiency, DOCK8 deficiency, Idiopathic CD4 lymphopenia, Malaysia, South East Asia

    (181) Submission ID#605966

    A Family with Hypogammaglobulinemia, ACTH Deficiency, Ectodermal Dysplasia and a Novel NFKB2 Mutation

    Linda Geberzahn, MD1, Marc Bienias, MD1, Angela Rösen-Wolff, MD, PhD2, Nicole Toepfner, MD3, Angela Huebner, MD4, Eva-Maria Jacobsen, PhD5, Mingyan Fang, PhD6, MinAe Lee-Kirsch, MD, PhD7, Joachim Roesler, MD8, Catharina Schuetz, MD, MSc9

    1Resident, Department of Pediatrics, Medizinische Fakultät Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany

    2Senior Scientist, Department of Pediatrics, Medizinische Fakultät Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany

    3Resident, Department of Pediatrics, Medizinische Fakultät Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany

    4Consultant, Department of Pediatrics, Medizinische Fakultät Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany

    5Biologist, Pediatrics, Ulm University Medical Center, Ulm, Germany

    6Scientist, 3BGI Genomics, Shenzhen, China

    7Senior Scientist, Department of Pediatrics, Medizinische Fakultät Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany

    8Senior Physician, Department of Pediatrics, Medizinische Fakultät Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany

    9Consultant, Department of Pediatrics, Medizinische Fakultät Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany

    The index patient presented at the age of 4 years with increased susceptibility to lower airway and gastrointestinal infections (hospital admissions 5x/year until puberty). She suffered from mumps and varicella disease despite immunization, as well as from recurrent local, partially destructive HSV infections. She was diagnosed with common variable immunodeficiency (CVID) at age 13 and started on immunoglobulin replacement therapy. Following a hypoglycemic seizure at age 20, the patient was diagnosed with isolated ACTH insufficiency with secondary adrenal insufficiency requiring hormone substitution. During and following her first pregnancy at age 25, she suffered from recurrent bronchopneumonias including Pneumocystis jirovecii infection, resulting in bronchiectases documented on chest CT at age 30. Currently, chronic lung disease is severely limiting her quality of life (Table 1). Her daughter was noticed to be hypogammaglobulinemic soon after birth and failed to develop antibody responses to inactivated vaccines. She was started on immunoglobulin replacement therapy. She has not suffered from severe lower airway infections, but developed alopecia totalis at age 10 and nail dystrophy.

    Whole exome sequencing revealed a heterozygous c.2553_2554insACCCGAG (p.Lys855ProfsTer33, NM_001077494) mutation in exon 22 of NFKB2 in both mother and daughter. This monoallelic loss-of function frameshift mutation was not found in gnomad, GVS Washington or ClinVar databases. As previously published, a monoallelic mutation in this c-terminal domain leads to impaired phosphorylation and subsequent reduced nuclear translocation of the NFKB2/p52 active form. Pediatricians and internal specialists need to be aware of the combination of hypogammaglobulinemia, ACTH deficiency, immune dysregulation and ectodermal dysplasia which is unusual for CVID - possibly indicating NFKB2 deficiency. This clinical syndrome may overlap with symptoms and signs found in both APECED/AIRE (AR) and EDA-ID/NFKBIA (AD) deficiencies. Besides IG and hormone replacement therapy, curative treatment with hematopoietic stem cell transplantation is a therapeutic option for patients with NFKB2 deficiency, although the experience is limited.

    Table 1. Clinical and immunological features of index patient and daughter

      viral infections bacterial infections end organ damage immune dysregulation immunology
    mother mumps and varicella disease (despite immunization), recurrent HSV1 infections (face, index finger) tracheobronchitis
    (H. influenza)
    P. jirovecii pneumonia
    bronchiectases, COPD, loss of distal phalanx (Dig.II) following HSV infection, ulceration of vocal chords, hypopituitarism, vasculitis of lower legs IgG 3.44 g/l, IgA < 0.26 g/l, IgM 0.45 g/l, no protective tetanus and diphtheria antibodies following regular immunization, absence of autoantibodies, initally low switched memory B- cells, total loss of peripheral B-cells
    daughter recurrent upper and lower respiratory infections, recurrent HSV1 infections periorally   mild restrictive lung disease, alopecia totalis, nail dystrophy IgG 2.33 g/l, IgA 0.23 g/l, IgM 0.34 g/l,
    B-cells 1900/μl, no protective tetanus, measles or HiB antibodies (but positive to rubella)

    (182) Submission ID#605974

    Infection Rates and Tolerability in Patients with Primary Immunodeficiency Diseases Treated with Three Different Immunoglobulin Administration Modalities

    Richard L. Wasserman, MD, PhD1, Sudhir Gupta, MD2, Mark R. Stein, MD3, Christopher J. Rabbat, PhD4, Werner Engl, PhD5, Heinz Leibl, PhD6, Leman Yel, MD7

    1Allergist/immunologist, Allergy Partners of North Texas Research, Dallas, TX, USA

    2Professor, University of California at Irvine, Irvine, CA, USA

    3Physician, Allergy & immunology, Allergy Section, Good Samaritan Medical Center, West Palm Beach, FL, USA

    4Director of Medical Affairs, Shire, Chicago, IL, USA (Affiliation at the time of the study)

    5Assoc Dir Biostatistics, Pharmacometrics & Pre-Clinical Biostatistics, Shire, Vienna, Austria

    6Sr Medical Director, Global Development Leader, IG, Clinical Research Immunology

    Shire, Vienna, Austria

    7Sr Medical Director, Global Development Leader, IG, Clinical Research Immunology

    Shire, Cambridge, MA, USA

    Introduction: The modes of immunoglobulin (IG) administration for primary immunodeficiency diseases (PIDD) differ in pharmacokinetics, infusion parameters, and tolerability. During 3 consecutive clinical studies, a cohort of 30 patients with PIDD experienced all 3 modes of administration with the same IG 10% product in sequence from intravenous (IV) to subcutaneous (SC), then to hyaluronidase-facilitated SC (IGHy), providing a unique opportunity to assess each administration modality within the same patient cohort treated and observed at the same sites. Here we report the rates of infections stratified by IgG trough levels, and the rates of adverse events (AEs) with the 3 modes of IG administration (IVIG, SCIG, IGHy) within this patient cohort.

    Design and Methods: This analysis included patients with PIDD aged 4 years who participated in 3 clinical studies: in Study 1 (NCT00546871) patients received IVIG 10% every 34 weeks followed by weekly SCIG 10%; in Study 2 (NCT00814320), patients were treated with IGHy every 34 weeks; in Study 3 (NCT01175313; extension of Study 2), patients continued with the same IGHy dose. To assess a potential association between the administration route at comparable IgG trough levels and the infection rate, IgG trough levels were categorized as 500 <700mg/dL, 700<900mg/dL, 900<1100mg/dL, 1100<1300 mg/dL, 1300 <1500 mg/dL and 1500 mg/dL. Periods where patients had trough levels within these strata were assessed, and the infection frequency was calculated. The time periods for this analysis were 3 months for IVIG and 12 months each for IGHy and SCIG 10% (2.25 years) treatments. In order to account for differences in the frequency of administration, rates of systemic and local AEs were assessed as AEs/patient-year for each mode of therapy.

    Results: For IgG trough levels of <1500 mg/dL, the associated annual infection rates were lower or similar for IGHy than SCIG (2.3 vs 3.5 [1300<1500 mg/dL]; 2.6 vs 3.8 [1100<1300 mg/dL]; 3.6 vs 6.2 [900<1100 mg/dL]; 1.4 vs 5.0 [700<900 mg/dL]; 2.0 vs 2.0 [500<700 mg/dL). For IgG trough levels 1500 mg/dL, the annual infection rate (95% CI) appeared to be lower for SCIG versus IGHy treatment (2.6 [1.83.7] vs 4.2 [1.210.3]) with shorter periods of observation and wide 95% confidence intervals. The rates of causally related systemic AEs/patient-year were lowest during IGHy (0.88) versus SCIG (1.93) and IVIG (5.60) treatment; the majority of causally related systemic AEs were mild. The rate of headaches/patient-year, the most common systemic AE, was lowest during IGHy treatment (0.21) versus SCIG (0.45) and IVIG (1.95). The rates of causally related local AEs/patient-year for IGHy, SCIG, and IVIG were 1.57, 0.92, and 0.13, respectively.

    Conclusion: Evaluation of the patient cohort in 3 consecutive studies over 2.25 years resulted in lower rates of causally related systemic AEs per patient-year during the IGHy treatment period compared with SCIG. Lower or similar infection rates were found during the IGHy treatment period compared with SCIG treatment at the same IgG trough levels (<1500 mg/dL). This observation warrants further investigation.

    (183) Submission ID#606027

    Syntaxina11 Mutation with Normal CD107a Surface Expression

    Carolina Dorfman, MD1, Agostina Llarens, MD1, Daniela Di Giovanni, MD2, Gisela Seminario, MD3, Andrea Gomez Raccio, MD3, Guadalupe Rodruiguez Broggi, MSc4, Patricia Carabajal, MD5

    1Physician – Immunology trainer, Children's Hospital Ricardo Gutierrez

    2Physician – Immunologist, Hospital de Niños Ricardo Gutierrez

    3Physician – Immunologist, Children's Hospital Ricardo Gutierrez

    4Biochemist, Hospital de Niños Ricardo Gutierrez

    5Head of Immunology Unit, Children' s Hospital Ricardo Gutierrez

    Introduction: Familial hemophagocytic lymphohistiocytosis (FHL) is a severe immune dysregulatory syndrome, inherited in an autosomal recessive way, caused by an impaired T and natural killer (NK) cell cytotoxicity. This results in an uncontrolled T cell and macrophage activation with hypercytokinemia.

    Syntaxin 11 (STX11) is one of the known genes of FHL. It is expressed in monocytes, NK cells and cytotoxic T cells and is involved in vesicle priming and membrane fusion.

    The clinical manifestations are fever, hepatosplenomegaly and cytopenias. Neurological features usually present later and are associated with poor prognosis. The disease is often triggered by infections, most commonly viral.

    Cytopenias, hyperferritinemia, hypertriglyceridemia and hypofibrinogenemia are usually present, along with CD25 soluble in blood and cerebrospinal fluid. Bone marrow may demonstrate hemophagocytosis. Specific functional tests can be done to make an approach to the primary defect. CD107a is present on the membrane of secretory granules within T and NK cells. Absence of CD107a expression on the cell surface after activation, suggests a defect in secretory granule migration, docking, priming or fusion.

    The treatment involves the control of infections and immune dysregulation with chemotherapeutic regimens followed by definitive treatment with hematopoietic stem cell transplant (HSCT).

    Aim: To describe a female patient with a pathogenic variation in STX11 with normal CD107a expression.

    Results: She was a 2 years old female, the 5th daughter of non-consanguineous parents, without relevant personal or family records. She was admitted due to a prolonged febrile syndrome, lymphoproliferation, pancytopenia and hepatitis, with HHV6 rescued in bone marrow and blood. Gancyclovir treatment started with good response. She was admitted one month later with similar clinical symptoms with relapsed HHV6 infection. Furthermore, hemophagocytosis was found in the bone marrow and evaluation of NK cell cytotoxicity demonstrated slightly reduced cytotoxic activity. Functional studies for primary FHL were performed: perforin expression and CD107a surface expression were normal. She fulfilled criteria of FHL, and treatment with gancyclovir and steroids was administered. Despite this treatment, she persisted with activated macrophagic parameters, and started with HLH2014 treatment protocol. She improved the clinical symptoms and laboratory parameters, but persisted with HHV6 low viremia. Three months later, when immunosupression was decreased, she was readmitted with similar clinical manifestations and added neurological symptoms (facial paralysis, abnormal movements and sleep tendency). Cerebral spinal fluid was pathological with HHV6 positive rescue. Immunosupresive treatment was adjusted, but HHV6 copies in blood increased markedly. Foscarnet treatment was administered and immunosupression was suspended for 2 days in order to control viral infection. Unfortunately the patient died 6 days later.

    Although specific functional tests were normal, sequencing of STX11 gene by NGS revealed a homozygous variation in c.581_584delTGCC, which is a previously reported mutation responsible for FHL.

    Conclusion: Despite the fact that CD107a was normal, the strong clinical and laboratory results must keep the FHL diagnosis in mind and intensive treatment should be early administered; in order to give the patient the opportunity to achieve the curative treatment.

    (184) Submission ID#606063

    Specific Functional Gammopathy Underlying Infectious Susceptibility in a Patient with Autoimmune-Polyendocrinopathy-Candidiasis-Ectodermal Dystrophy (APECED)

    Gregory M. Constantine, MD1, Elise Ferre, PA-C, MPH2, Michail Lionakis, MD, Sc.D.3

    1Clinical Fellow, Fungal Pathogenesis Section, Laboratory of Clinical Immunology & Microbiology (LCIM) , National Institute of Allergy & Infectious Diseases (NIAID), NIH

    2Physician Assistant, Fungal Pathogenesis Section, Laboratory of Clinical Immunology & Microbiology (LCIM) , National Institute of Allergy & Infectious Diseases (NIAID), NIH

    3Senior Investigator, Fungal Pathogenesis Section, Laboratory of Clinical Immunology & Microbiology (LCIM) , National Institute of Allergy & Infectious Diseases (NIAID), NIH

    Introduction/Background: Autoimmune-Polyendocrinopathy-Candidiasis-Ectodermal Dystrophy (APECED) is a rare inherited primary immunodeficiency disorder resulting from biallelic mutations in the AIRE gene. Although classically characterized by chronic mucocutaneous candidiasis (CMC), hypoparathyroidism and primary adrenal insufficiency (Addisons Disease), recent evidence has demonstrated broader disease manifestations including: urticarial-like eruption, autoimmune hepatitis, intestinal dysfunction, gastritis, Sjogrens-like syndrome and pneumonitis. Clinical observations have also identified a subset of individuals who suffer from recurrent sinopulmonary infections.

    Objectives: To report and characterize the clinical course of a patient with APECED and specific antibody deficiency.

    Methods: Retrospective chart review was performed. The patient was enrolled in NIAID IRB-approved protocol 11-I-0187.

    Results: The patient is a 13 year-old-girl with APECED caused by homozygous AIRE c.967_979del13, who manifested CMC, hypoparathyroidism, adrenal insufficiency, Sjogrens-like syndrome, autoimmune hepatitis, intestinal dysfunction and autoimmune pneumonitis. She suffered from recurrent sinusitis and severe pneumonias requiring hospitalization and administration of intravenous antibiotics several times per year. At age 9, she presented to our institution with fever and cough, a computed tomography (CT) of the chest revealed bilateral pulmonary infiltrates and bronchiectasis. Bronchoscopy showed mucopurulent secretions in the bilateral lower lobes with culture of the bronchoalveolar lavage fluid growing Streptococcus pneumoniae.

    Further evaluation for an underlying disorder such as primary ciliary dyskinesia and cystic fibrosis including exome sequencing and sweat chloride testing was unrevealing. Quantitative immunoglobulins were normal. Despite prior vaccination, specific antibody testing showed negative rubeola IgG and protective levels (> 1.3 mcg/mL) to only 3 of 23 pneumococcal serotypes. Lymphocyte enumeration showed normal B cell subsets. As approximately 15% of APECED patients may experience asplenia, splenic ultrasound was performed confirming the presence of a 7 cm spleen and peripheral blood smear did not reveal Howell-Jolly bodies. Serotyping of the S. pneumoniae isolate confirmed serotype 33F, which is part of the 23-valent vaccine. Follow up vaccine challenge with the 23 valent pneumococcal polysaccharide vaccine showed an inadequate response. Hence, she was started on monthly immunoglobulin replacement and over the following 4 years she has experienced a single methicillin sensitive Staphylococcus aureus pneumonia. She has missed very few school days and other parameters including linear growth have improved, she is now along the fifth percentile for height and along the tenth percentile for weight. Although she continues to experience intermittent cough she remains active participating in sports without limitation.

    Conclusions: We report the evaluation, treatment and outcome of a patient with APECED complicated by autoimmune pneumonitis and specific antibody deficiency. As infectious susceptibility of APECED classically pertains to the signature infectious disease, CMC, patients with invasive or recurrent infections should be evaluated for underlying immune deficiency. Investigation should include assessment for asplenia, quantitative immunoglobulins and specific antibodies with response to antigens. In patients with predominate respiratory symptoms, autoimmune pneumonitis should be evaluated given the near 40% prevalence of pneumonitis observed in American APECED patients.

    Acknowledgements: Supported by DIR/NIAID/NIH

    (185) Submission ID#606159

    Two Siblings with a Delayed/Late-Onset Presentation of Combined Immunodeficiency Due to Adenosine Deaminase Deficiency

    Tamara C. Pozos, MD, PhD1, Manar Abdalgani, MBBS2, Michael Hershfield, MD3

    1Medical Director, Clinical Immunology, Children's Hospital Minnesota

    2Clinical Immunologist, Children's Hospital Minnesota

    3Professor of Medicine, Professor of Biochemistry, Duke University Medical Center

    BACKGROUND: Complete deficiency of adenosine deaminase (ADA) is an autosomally inherited condition that causes one type of SCID secondary to the accumulation of metabolites toxic to lymphocytes. Less severe genetic changes resulting in detectable ADA result in a milder, clinically heterogeneous combined immunodeficiency.

    CASE PRESENTATIONS: Patient 1: A 16 year old Somali-American girl presented with progressive pulmonary disease. Past history included recurrent severe pneumonias, eczema and recurrent Staphylococcal infections. Genetic testing for HyperIgE syndrome was negative. She developed autoimmune hypothyroidism at age 7 years and warts covering both hands at age 10 years. On presentation, she had a normal-to-elevated IgG with some therapeutic vaccine titers, normal IgM and IgA. IgE was elevated at 7381 IU/ml. B cells were essentially absent: absolute CD19+ 1cell/ul. NK cells were also low, absolute 39cells/ul. Absolute number of total T cells was normal with decreased CD4+ Tcells. Lymphocyte proliferation responses to mitogens were significantly decreased, and responses to tetanus antigen were absent.

    ADA activity was deficient (0.4 nmol/h/mg) with only modestly increased red cell dAXP% (ratio of dAXP/RBC-AXP 6.3%), consistent with delayed/late onset ADA deficiency. After 17 weeks of PEG-ADA injections, dAXP% was zero. B cell counts improved though to normal levels. IgE increased after therapy began, peak 13,000IU/ml, and then slowly decreased. Her cough and dyspnea improved, though pulmonary function tests still show severe airflow obstruction.

    Genetic testing revealed a homozygous missense mutation in ADA exon 6, V177M (c.529G>A), a previously identified change shown in vitro to produce 0.1-0.4% of wild type ADA activity (Arredondo-Vega, F. et al, Am J. Hum Genet 1998; 63:1049-59). Patient 1s sister is homozygous for the same mutations.

    Patient 2: Though considered by family to be a healthy 11 year old, her past history included a left axillary abscess at age 2 years, eczema, recalcitrant tinea capitus, and 3 warts. She had no pulmonary disease. She had short stature. Labs revealed normal IgG, IgM and IgA. All vaccine titers were strongly therapeutic. IgE was elevated at 1210IU/ml and initially increased with PEG-ADA injections. Like her sister, B cells at presentation were nearly absent with absolute CD19+ 12 cells/ul. T and NK cells were normal. Also like her sister and their mother who is a carrier, Patient 2 had autoimmune hypothyroidism. After 7 weekly PEG-ADA injections, she had rapid decrease in percentage dAXP/total RBC AXP, from 6.3% to 1.0% and absolute B cell number increased to 92.

    CONCLUSIONS: We report two siblings with identical genetic changes in ADA and identical presenting ratios of dAXP/total RBC-AXP. Both had dysgammaglobulinemia and B lymphopenia, atopy and autoimmunity on presentation. However, the 16 year old has had significantly more medical complications and has shown a slower improvement with replacement therapy compared to her younger sister.

    (186) Submission ID#606161

    Isolated Central Nervous System Disease in Familial Hemophagocytic Lymphohistiocytosis a Multicenter Case Series

    Annaliesse Blincoe, MBChB FRACP1, Maximilian Heeg, MD2, Patrick Campbell, MD, PhD3, Amer Khojah, MD4, Marisa Klein-Gitelman, MD5, Julie-An Talano, MD6, Claire Booth, MBBs PhD7, Despina Moshous, MD, PhD8, Fabien Touzot, MD, PhD9, Arjan Lankester, MD, PhD10, Jacques Rivière, MD11, Maria Caterina Putti, MD12, Sarah Maier, MD13, Kai Lehmberg, MD14, Itziar Astigarraga, MD15, Steven M. Holland, MD16, Rebecca A. Marsh, MD17, Stephan Ehl, MD18, Elie Haddad, MD, PhD19

    1Department of Pediatrics, CHU Sainte-Justine, University of Montreal, Montreal, QC, Canada

    2Institute for Immunodeficiency, Center for Chronic Immunodeficiency. Center for Pediatrics and Adolescent Medicine, University of Freiburg, Freiburg, Germany

    3Paediatric Haematologist-Oncologist, St. Jude Children's Research Hospital

    4Department of Rheumatology, Ann and Robert H Lurie Children's Hopital and Children's Hospital of Chicago, Chicago, IL

    5Division Head, Rheumatology, Professor of Pediatrics, Northwestern University Feinberg School of Medicine Chicago, IL

    6Associate Professor, Pediatric Haematology and Oncology, Children's Hospital of Wisconsin-Milwaukee Campus, Milwaukee, WI

    7Honorary Consultant in Paediatric Immunology, Great Ormond Street Hospital for Children, London, UK

    8Paediatric Immunology, Haematology and Rheumatology Unit, Hôpital Necker Enfants Malades, Paris, France

    9Department of Pediatrics, CHU Ste-Justine, University of Montreal, Montreal, QC, Canada

    10Professor of Pediatrics and Stem Cell Transplantation, University of Leiden Medical Centre, Leiden, Netherlands

    11Pediatric Infectious Diseases and Immunology, Vall d'Hebron University Hospital, Barcelona, Spain

    12Pediatric Haematologist-Oncologist, Department of Pediatrics, University of Padua Medical School

    13Pediatric Haematology and Oncology, University Medical Center Hamburg Eppendorf, Germany

    14Paediatric Haematology and Oncology, University Medical Center Hamburg Eppendorf, Germany

    15Pediatric Oncology, Hospital Universitario Cruces , Barakaldo, Spain

    16Director, Division of Intramural Research, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health

    17Associate Professor, Division of Bone Marrow Transplantation and Immune Deficiency, Cincinnati Childrens Hospital Medical Center

    18Institute for Immunodeficiency, Center for Chronic Immunodeficiency. Center for Pediatrics and Adolescent Medicine, University of Freiburg, Freiburg, Germany

    19Department of Pediatrics. Department of Microbiology, Infectiology and Immunology, CHU SainteJustine, University of Montreal, Montreal, QC, Canada

    Familial hemophagocytic lymphohistiocytosis (FHLH) is a rare, inherited syndrome of immune dysregulation, characterised by genetic predisposition to a systemic hyper-inflammatory disease. Whilst central nervous system (CNS) involvement is well described in FHLH, there have been very few case reports of isolated CNS-HLH in the absence of any systemic features. This clinical presentation may be under-recognised, leading to delays in diagnosis and appropriate treatment. We present 9 new cases together with 11 cases identified in the literature, to describe the clinical presentation, treatment and outcomes.

    Patients and Methods: Patients were identified and retrospective data collected through collaborative efforts at North American and European clinical immunology, haematology/oncology and bone marrow transplant centres and through review of previously reported cases in the literature. Inclusion criteria; (1) confirmed diagnosis of FHLH with pathological FHLH gene mutation; (2) any neurological symptoms at time of diagnosis; (3) absence of systemic HLH according to HLH-2004 criteria, in particular absence of bilineage cytopenia and splenomegaly; (4) features of systemic HLH fulfilling HLH-2004 diagnostic criteria no earlier than 3 months after initial neurological presentation.

    Results: Nine new patients and 11 patients from the literature were identified. The mean age at time of onset of neurological symptoms was 6.2 years. The most common neurological manifestations were ataxia/gait disturbance (65%) and seizures (50%) with a mean delay from onset of neurological symptoms to confirmed FHLH of 15.8 months. Diffuse multifocal white matter changes and cerebellar involvement were the most common CNS MRI findings at 14/17 (82.4%) and 8/17 (47%) respectively. The mean CSF cell count was 15.9 x 106/L and mean CSF protein 106 mg/dL. Epstein-Barr virus was identified as a trigger in 2 patients, and varicella zoster virus in 1 patient. Seven patients eventually developed systemic HLH manifestations at a mean time of 17.4 months (range 4-28 months) after initial neurological symptoms. Mutations in PRFI were detected in 15 patients (75%), RAB27a mutation in 3 (15%) and UNC13D in 2 (10%). Six patients carried the R225W mutation in perforin.

    Nineteen patients received systemic HLH-directed chemo/immunosuppressive therapy. Two patients received intrathecal methotrexate and hydrocortisone. Of the 13 patients treated with HSCT (7 new, 6 literature), 7 patients (53.8%) had improved neurological outcome, 3 patients (23.1%) had stable neurological findings, and one patient had CNS relapse at time of follow-up. There were no cases of neurological improvement in patients who did not receive HSCT. The overall mortality was 35% (2 new and 5 literature patients).

    Conclusion: Our study describes the variable clinical presentation and findings in patients with isolated CNS-HLH. We propose that genetic cytotoxicity defects should be considered in any patient with unexplained and progressive neurological symptoms, even in the absence of ANY signs of systemic inflammation. Neurological improvement was observed in patients who received HSCT, emphasising the need for timely institution of treatment in order to improve outcomes. Our study also identified a predominance of PRF1 mutations; however, larger studies would be required to further evaluate the significance of this finding.

    (187) Submission ID#606194

    Lupus-like Syndrome in a Patient with NOD2-associated Autoinflammatory Disease

    Lyda Cuervo-Pardo, MD1, Mario Rodenas, MD, FAAAAI1, Mingjia Li2, Shuhong Han, Ph.D.3, Haoyang Zhuang, Ph.D.3, Westley Reeves, MD4

    1Assistant Professor, University of Florida, Division of Rheumatology & Clinical Immunology, Department of Medicine

    2Undergraduate Student, Division of Rheumatology, Allergy, & Immunology, University of Florida

    3Assistant Professor, Division of Rheumatology, Allergy, & Immunology, University of Florida

    4Professor and Chief, University of Florida, Division of Rheumatology & Clinical Immunology, Department of Medicine

    Introduction: Autoinflammatory diseases are genetically heterogeneous disorders of innate immunity characterized by recurrent fever, rash, and/or serositis, which generally are considered distinct from autoimmune diseases. We report a case of a patient with lupus-like disease and a mutation of nucleotide-binding oligomerization domain-containing protein 2 (NOD2 R702W, Yao syndrome) suggestive of an overlap between autoinflammatory and autoimmunity processes.

    Case Presentation: A 72-year-old man was evaluated for recurrent pleural effusions, morning stiffness, erythematous rashes, and fever up to 103°C. History was notable for Hashimotos thyroiditis and multiple admissions for presumed pneumonia with recurrent bilateral lung infiltrates and pleural effusions. Transbronchial biopsy showed nonspecific pneumonitis and organizing pneumonia. Antinuclear and anti-dsDNA antibodies were positive. He received prednisone for presumed lupus pneumonitis leading to improvement. Prednisone was tapered and hydroxychloroquine was started, but his fevers, pleuritic pain and pleural effusion reoccurred. Genetic testing revealed a NOD2 sequent variant (R702W) associated with autoinflammatory disease. Hydroxychloroquine was stopped and colchicine was added to his regimen, allowing prednisone to be tapered without recurrence of symptoms. Further immunological testing revealed increased signaling through the type I interferon receptor (interferon signature).

    Conclusion: Although this patient had several clinical (serositis, arthralgia) and immunological (antinuclear and anti-dsDNA antibodies, interferon signature) manifestations of lupus, his clinical presentation also was consistent with Yao syndrome. In retrospect, he had been having recurrent inflammatory symptoms for many years. Recent studies in both mice and humans suggest that inflammasome activation and IL-1 production are involved in the pathogenesis of lupus. This case provides further support for the idea that lupus and Hashimotos thyroiditis, prototypical autoimmune diseases, may have overlapping autoinflammatory features.

    (188) Submission ID#606243

    Immunological and Genetic Outcomes of Infants with Positive Newborn Screening for Severe Combined Immunodeficiency (SCID)

    Vasudha Mantravadi, MD1, Jeffrey J. Bednarski, MD, PhD2, Michelle A. Ritter, RN3, Megan A. Cooper, MD, PhD4, Maleewan Kitcharoensakkul, MD5

    1Resident Physician, Department of Pediatrics, Washington University School of Medicine, St. Louis, Missouri, 63110

    2Assistant Professor of Pediatrics, Hematology and Oncology, The Division of Hematology and Oncology, Department of Pediatrics, Washington University School of Medicine, St. Louis, Missouri, 63110

    3Research Nurse Coordinator, The Division of Pediatric Rheumatology, Department of Pediatrics, Washington University School of Medicine, St. Louis, Missouri, 63110

    4Associate Professor, Rheumatology, The Division of Pediatric Rheumatology, Department of Pediatrics, Washington University School of Medicine, St. Louis, Missouri, 63110

    5Assistant Professor of Pediatrics, Rheumatology and Allergy, Immunology and Pulmonary Medicine, The Division of Allergy, Immunology & Pulmonary Medicine, Department of Pediatrics, Washington University School of Medicine, St. Louis, Missouri, 63110

    Background: The implementation of severe combined immunodeficiency (SCID) newborn screening by TREC assay has played a pivotal role in identifying these patients early in life. The screen has also led to the identification of infants with other immunologic abnormalities, of which the clinical implications have been unclear and there are limited data on their outcomes.

    Objective: To review immunologic and genetic outcomes of infants referred to an immunology service of a tertiary care center with abnormal newborn SCID screens.

    Methods: We retrospectively reviewed charts of infants with positive SCID screen from July 2014 to November 2018. We excluded patients who had positive screen at <36 weeks corrected gestational age. We classified outcomes into 3 groups including SCID, non-SCID T-cell lymphopenia (NSCID-TCL) and normal T-cell count. Idiopathic T-cell lymphopenia was defined as NSCID-TCL (CD3+ < 2,500 cells/mcl) with negative chromosome microarray and negative whole exome sequencing/or genetic panel (either GeneDx® SCID panel or Invitae® Primary Immunodeficiency panel).

    Results: Of 119 infants, 78% were male, 56% were Caucasian, and 37% were African-American. Fifty-four % and 46% of infants were identified by Illinois and Missouri screens, respectively. The mean age at initial evaluation was 22 days (4-122 days). 69% of infants had a normal T-cell count (N=80) or normal repeat newborn screen (N=2), 25% had NSCID-TCL, including mild (CD3+ 1,500-2,500 cells/mcl, N=20) and moderate (CD3+ 300-1,500 cells/mcl, N=10) TCL, and 6% had SCID (N=5), leaky SCID (N=1) or complete DiGeorge (N=1). Genetic etiologies of NSCID-TCL included 22q11 deletion (N=4), trisomy 21 (N=1), and mutations of TBX1 (N=2), FOXN1 (N=1), and CD3E (N=1). Three of these infants had novel variants at the time of diagnosis. Secondary causes of TCL were identified in 1 infant (thoracic infantile fibrosarcoma). One infant had idiopathic TCL. Eighteen infants with NSCID-TCL were followed clinically without complete genetic testing performed. For SCID, mutations were found in JAK3 (N=2), ADA (N=1), IL2RG (N=1), and RAG1 (N=1). The patient with leaky SCID had negative whole exome sequencing. All patients with SCID and leaky SCID underwent hematopoietic stem cell transplantation at a median age of 5 weeks (3 weeks - 4 months), with successful engraftment in all but 1 patient. Of 19 idiopathic and NSCID-TCL cases followed clinically, 12 had at least one follow-up visit at median age 5 months (2.6 months 2.2 years) and the majority had improved or stable lymphocyte count without serious infections requiring intravenous antibiotics, though 1 had a hospitalization for RSV infection. The MYSM1 patient died after cord blood transplant from unclear etiology. Our study had limitations. Half of infants with NSCID-TCL did not have a complete genetic workup, and only a fifth of patients with NSCID-TCL were inpatients, potentially explaining the relatively low number of infants with secondary lymphopenia.

    Conclusions: In our cohort, one-fourth of infants with abnormal SCID screen had NSCID-TCL. Although the majority of NSCID-TCL did well, approximately one-third of them had underlying genetic abnormalities associated with their T-cell lymphopenia.

    (189) Submission ID#606320

    Nucleus-retained WASP Is Deleterious to T-cell Development

    Carole Le Coz, PhD in immunology1, Tanner Robertson2, Caroline Khanna3, Andrew Sy, MD4, David K. Buchbinder, MD, MSHS5, Janis Burkhar, PhD6, Neil Romberg, MD7

    1Postdoc, Children's Hospital of Philadelphia

    2PhD student, University of Pennsylvania

    3Research Assistant, Children's Hospital of Philadelphia

    4Fellow, Children's Hospital of Orange County, Orange, CA, Department of Pediatrics, University of California at Irvine, Orange, CA

    5Assistant Clinical Professor, Department of Hematology, Children's Hospital of Orange County, Orange, CA, Department of Pediatrics, University of California at Irvine, Orange, CA

    6Professor, University of Pennsylvania Perelman School of Medicine

    7Assistant Professor, Children's hospital of Philadelphia

    Cytosolic Wiskott-Aldrich Syndrome protein (WASp) regulates actin cytoskeleton reorganization but also enters the nucleus to affect gene transcription. Mice and humans without WASp develop normal numbers of naïve T cells but are susceptible to infections by viruses and encapsulated bacteria. Although opportunistic mycobacterial infections are reported in severe combined immune deficiencies (SCID) and more specific Th1-associated monogenic diseases, they have not been previously reported in WASp-deficient patients.

    Here we report two unrelated kindreds presenting with opportunistic mycobacterial infections (M. bovis and M. gordonea) that carry nearly identical mutations in the gene encoding WASp (WAS). In one kindred affected males presented as infants with T-B+NK+ SCID, thrombocytopenia and eczema. In the other kindred affected males presented with abundant but dysfunctional T cells. Thrombocytopenia and eczema were present in both groups. In each case mutant WAS encoded truncated WASp lacking its cofilin and the acidic domains (WASp CA). In addition to an inability to normally polymerize actin, primary patient cells retained WASp CA entirely within their nuclei. There it appeared conformationally open and constitutively active even in the absence of stimulation. These data suggest conformationally open WASP retained in the nucleus of T cells alters transcriptional programs leading to new and severe infectious phenotypes.

    (190) Submission ID#606328

    CVID Plasma Promotes Early Commitment to the Follicular Lineage

    Caroline Khanna1, Carole Le Coz2, Neil Romberg, MD3

    1Research Assistant, Children's Hospital of Philadelphia

    2PhD in Immunology, Postdoc, Children's Hospital of Philadelphia

    3Assistant Professor, Children's hospital of Philadelphia

    Background: Many CVID patients with undetectable serum IgA concentrations are endotoxemic and possess enlarged pools of circulating T follicular helper (Tfh) cells.

    Objective: To determine if endotoxemia and other plasma-soluble factors promote follicular T cell differentiation.

    Methods: We cultured healthy donor (HD) naïve CD4+ T cells for 5 days in CVID patient plasma, CVID plasma treated with polymyxin B to neutralize endotoxin, or fetal bovine serum spiked with LPS. After 5 days we measured the frequency of cells expressing the Tfh markers CXCR5 and PD1. We also measured concentrations of cytokines known to promote Tfh differentiation, comparing levels in CVID patient plasma samples that induced expression of Tfh markers with samples that did not induce them.

    Results: We found greater frequencies of HD naïve CD4+ T cells expressed Tfh markers when cultured in plasma from IgA deficient CVID patients, than IgA sufficient patients or healthy donors. These differences disappeared upon addition of polymyxin B to patient plasma samples and could be recapitulated by replacing plasma with LPS spiked FBS.

    In addition to LPS, patient plasma samples that best promoted Tfh marker expression contained higher concentrations of Activin A but not the traditional Tfh-differentiating cytokines IL-12 and IL-6.

    Conclusion: Endotoxin, which circulates in IgA deficient CVID patients, promotes naïve T-cell commitment to the follicular lineage directly and through enhanced release of Activin A.

    (191) Submission ID#606365

    Infusion Parameters of Patients with Primary Immunodeficiency by Previous Immunoglobulin Routes of Administration Among Enrollees in a Patient Program Initiated on Ig20Gly

    Lisa Meckley, PhD1, Yanyu Wu, PhD2, Spiros Tzivelekis, MSc3, Andre Gladiator, PhD4,

    1Director, GHEORE, Shire

    2Lead, Health Economics and Outcomes Analytics, Shire

    3ORE Lead, ORE Immunology & Opthalmolgy, Shire

    4Global Medical Lead Immunology - Global Medical Affairs, Shire

    Rationale: HCUVP is a patient product-introduction program that provides free-of-charge Cuvitru® (immune globulin subcutaneous [human], 20% solution; Ig20Gly) for the first 4 infusions to eligible patients who have primary immunodeficiency disease (PID). Using data from the ongoing HCUVP, this analysis described the clinical characteristics and infusion parameters of patients initiated on Ig20Gly based on their previous immunoglobulin (IG) routes of administration.

    Methods: Patients 2 years of age who had a primary ICD-10 diagnosis code of PID and had no current or prior use of Ig20Gly at the time of program enrollment were eligible for HCUVP. This analysis included data from patients enrolled in HCUVP who received the first Ig20Gly infusion between January 1, 2017 and September 1, 2017. Infusions after October 31, 2017 were excluded. Descriptive statistics were calculated for demographic, clinical, and prescribed infusion characteristics for patients previously treated with intravenous IG (IVIG) and subcutaneous IG (SCIG), respectively.

    Results: A total of 420 patients who had previously been treated with IVIG or SCIG completed all 4 infusions and were eligible for this analysis. Prior to enrolling in HCUVP, 268 patients had received IVIG, of whom 46 patients (17%) were < 18 years old, and 152 patients had received SCIG, of whom 17 patients (11%) were < 18 years old. A greater percentage of patients who switched from SCIG infused biweekly (54 of 152, 36%) compared with those who switched from IVIG (64 of 268, 24%). The mean dose administered during the final infusion was higher in patients who previously received SCIG compared with those who previously received IVIG (15.3 g vs 14.5 g, respectively), despite the SCIG group having a lower mean weight (SCIG: 74.1 kg [SD 23.2]; IVIG: 77.2 kg [27.2]) and fewer patients < 18 years. (Both weight and age can influence dosage). The mean number of infusion sites per infusion (SCIG: 2.1; IVIG: 2.0), mean infusion rate per site (SCIG: 41.5 mL/hr/site; IVIG: 42.9 mL/hr/site), and mean infusion volume per site (SCIG: 37.6 mL/site; IVIG 37.1 mL/site) were comparable between both groups. By the third of a total of 4 infusions, 91% and 94% of patients reached their maximum infusion rate when switching from IVIG or SCIG to Ig20Gly, respectively.

    Conclusion: Among patients enrolled in an Ig20Gly product-introduction program, those who had previously received SCIG were more likely to infuse biweekly and receive a slightly higher mean dose during the final infusion than patients who had previously received IVIG. However, the number of infusion sites, infusion rates per site, infusion volume per site, and percentage of patients reaching their maximum infusion rate were similar regardless of patients previous routes of IG administration.

    Funding: This research was sponsored by Shire.

    (192) Submission ID#606375

    A Literature Review on Shared Decision-Making (SDM) to Inform the Development of an SDM Tool in Primary Immunodeficiency Diseases

    Lisa Meckley, PhD1, Ihor Sehinovych, PharmD2, Spiros Tzivelekis, MSc3

    1Director, GHEORE, Shire

    2Medical Lead – HyQvia, Shire

    3ORE Lead, ORE Immunology & Opthalmolgy, Shire

    Background: Shared decision-making (SDM) is an interactive process that allows patients and their physicians to choose treatments that align with patients preferences. For patients with primary immunodeficiency diseases (PIDs) who require immunoglobulin replacement therapy (IGRT), SDM may help individualize IGRT to clinical needs and lifestyles.

    Objective: To summarize results of a targeted literature review on SDM models and their impact on clinical outcomes and to introduce a novel SDM tool for PID

    Methods: Focused searches for articles in English were conducted in EMBASE and MEDLINE (date range: January 1, 1999 to August 15, 2018). The search targeted the key elements of SDM (defined as the meaningful exchange of information between patient and physician and identification of issues most important to patients) and the impact of SDM on clinical outcomes. Relevant literature was examined for the current state of SDM in PID and used to inform the development of a novel SDM tool.

    Results: The search identified 4,730 records with SDM in the title or abstract. A broad range of therapeutic areas (primarily chronic diseases) was represented, and publication frequency had increased with time. Focused searches identified 159 articles that discussed key elements of SDM. Common elements of SDM included recognizing the decision; two-way sharing of knowledge between physician and patient; expression of patient values and preferences; weighing the options; and making and implementing the decision. The impacts of SDM on clinical outcomes were discussed in 59 studies; 15 were reviewed in detail in acute (n = 7), chronic (n = 5), and general/other (n = 3) conditions. Two studies suggested that SDM may improve clinical outcomes in chronic diseases. In 4 studies, patients who participated in SDM used fewer diagnostic tests and medications and underwent fewer intensive tests and treatments for acute illness. Positive effects of SDM were reported in mental health settings, for patients with chronic illness or making longer-term decisions, and in cases in which SDM interventions occurred over multiple sessions. No studies were found that evaluated SDM in the treatment of PIDD. In a survey of US immunologists (n = 15), participants acknowledged the value of SDM; however, in another survey, patient preferences for IGRT were not the same as what physicians perceived of their patients preferences, underscoring a need for SDM in PID.

    Conclusions: SDM has been widely studied and increasingly implemented in health care decisions globally; however, its effects on key patient outcomes are not well understood, and there are currently no known SDM applications in PID. The key findings from this review support the applicability of SDM in PID and highlight the need for a novel tool to help patients recognize their own priorities and needs, and to ensure these guide important clinical decisions such as IGRT selection. Based on these findings, an SDM tool and accompanying discussion guide for clinicians is being developed to facilitate SDM in PID.

    Funding: This research was sponsored by Shire.

    (193) Submission ID#606400

    Treatment of Adenosine Deaminase Severe Combined Immunodeficiency with Pegylated Recombinant Adenosine Deaminase. a Clinical Trial of Patients Transitioned from Pegademase to Elapegademase-lvlr

    Morna J. Dorsey, MD, MMSc1, Tracy Fausnight, MD2, Heather Lehman, MD, FAAAAI3, Neena Kapoor, MD4, Arye Rubinstein, MD5, Giuseppe Testa, B.S.6, Joseph M. Wiley, MD7,

    1Pediatric Immunologist and Allergist, Department of Pediatrics, Division of Allergy, Immunology, and Bone Marrow Transplant, University of California San Francisco, San Francisco, CA

    2Associate Professor, Pediatric and Adult Allergy/Immunology, Penn State College of Medicine

    3Associate Professor of Pediatrics, Chief Division of Allergy/Immunology and Rheumatology, Children's Hospital of Buffalo

    4Professor, Pediatrics, Children's Hospital Los Angeles and Keck Schood of Medicine of U.S.C.

    5Professor, Pediatric Allergy/Immunology, Albert Einstein College of Medicine

    6VP Product Development, Leadiant Biosciences, Inc.

    7VP Medical Affairs, Drug Safety and Pharmacovigilance, Leadiant Biosciences, Inc.

    Introduction: Accumulation of intracellular adenosine and deoxyadenosine nucleotides (dAXP) due to adenosine deaminase deficiency results in profound lymphopenia and severe combined immunodeficiency. Left untreated this form of SCID is uniformly fatal. While allogeneic hematopoietic cell transplant (HCT) and autologous gene corrected stem cell therapy (GT) are potential cures for ADA-SCID , initiating enzyme replacement therapy (ERT) immediately upon diagnosis regardless of definitive treatment is standard of care. HCT and GT are not therapeutic options for all ADA-SCID patients and ERT offers immediate therapeutic intervention for these patients leading to partial immune reconstitution, and durable survival in most patients treated. Adagen (pegademase), approved by the FDA in 1990 in the USA, is a pegylated bovine ADA (nADA) with the enzyme harvested from bovine intestines. This unsustainable production process led to the development of a recombinant enzyme source based on the bovine protein sequence and an improved pegylated linker by using succinimidyl carbamate (RevcoviTM- (elapegademase-lvlr).

    Methods: A Phase II/III clinical trial was performed at 5 US sites under institutional IRB approval. Eligible ADA-SCID subjects were stable on Adagen and without complicating underlying conditions. Demographics, medical history, lymphocyte counts, immunoglobulin levels, trough plasma ADA activity and RBC dAXP measurements were collected. Patients were treated with Adagen as a single, weekly IM dose adjusted to achieve a trough plasma ADA activity of > 15 mmol/hr/L and RBC dAXP < 0.02 mmol/L (protocol target levels). Once patients had achieved this level (3-9 weeks), a seven-day PK on Adagen was done and the patients were transitioned to Revcovi based on the formula for enzyme equivalent activity of 1mg Revcovi = 150 Units Adagen. After 5 weeks on Revcovi, trough ADA and dAXP were assessed and a seven-day pharmacokinetic study was conducted at week 9. Patients were assessed periodically for clinical and laboratory values and evaluation of the study endpoints was done at week 21. Subjects subsequently continued on Revcovi and were assessed periodically.

    Results: Six patients, ages 16-37 entered the trial with initial Adagen dosing at 7.7-42.9U/kg/wk (see Table 1). Adagen dosing was adjusted to target endpoints of ADA trough activity (>15mmol/hr/L) and RBC dAXP (<0.02 mml/l). Patients transitioned to weekly Revcovi using the aforementioned conversion formula at doses of 0.17-0.285 mg/kg/wk. The drug was well tolerated with the most frequent adverse events (AEs) being cough and vomiting. There were no drug attributable serious AEs. Patients achieved trough RBC dAXP < 0.02 mml/L (primary endpoint) in 69/71 measurements through week 21 and trough plasma ADA > 15mmol/hr/L (secondary endpoint) in 68/74 measurements. Total lymphocyte counts increased from a mean of 0.79 x109/L at start of Revcovi to 0.92 x 109/L at week 21.

    Conclusion: Revcovi appears safe in ADA-SCID patients, provides adequate detoxification (dAXP<0.02mmol/L) and sustained trough plasma ADA activity at ~ 2X trough activity provided by Adagen with improved lymphocyte counts. Revcovi received FDA approval for treatment of ADA-SCID on 10/5/18.

    Table 1 Study STP-2279-002: Patient Data (Six Patients)
    Patient ID Gender/Age/Race b Start of Adagen Lead In Phase a   Adagen PK Week Revcovi Tm Treatment Weeks 1 Through 21 d
    Weekly Dose (U/kg)b Trough ADA Activity (mmol/hr/L) Weekly Dose (U/kg)b Trough ADA Activity (mmol/hr/L) Erythrocyte
    dAXP (mmol/L)
    Total Lymphocytes (x10 9 /L) Weekly Dose (mg/kg) b No. of Trough ADA Activity Values >15 mmol/hr/L No. of Erythrocyte
    dAXP Values < 0.02 mmol/L