Autoimmune polyendocrine syndrome type 1: an Italian survey on 158 patients

Background Autoimmune Polyglandular Syndrome type 1 (APS-1) is a rare recessive inherited disease, caused by AutoImmune Regulator (AIRE) gene mutations and characterized by three major manifestations: chronic mucocutaneous candidiasis (CMC), chronic hypoparathyroidism (CH) and Addison’s disease (AD). Methods Autoimmune conditions and associated autoantibodies (Abs) were analyzed in 158 Italian patients (103 females and 55 males; F/M 1.9/1) at the onset and during a follow-up of 23.7 ± 15.1 years. AIRE mutations were determined. Results The prevalence of APS-1 was 2.6 cases/million (range 0.5–17 in different regions). At the onset 93% of patients presented with one or more components of the classical triad and 7% with other components. At the end of follow-up, 86.1% had CH, 77.2% AD, 74.7% CMC, 49.5% premature menopause, 29.7% autoimmune intestinal dysfunction, 27.8% autoimmune thyroid diseases, 25.9% autoimmune gastritis/pernicious anemia, 25.3% ectodermal dystrophy, 24% alopecia, 21.5% autoimmune hepatitis, 17% vitiligo, 13.3% cholelithiasis, 5.7% connective diseases, 4.4% asplenia, 2.5% celiac disease and 13.9% cancer. Overall, 991 diseases (6.3 diseases/patient) were found. Interferon-ω Abs (IFNωAbs) were positive in 91.1% of patients. Overall mortality was 14.6%. The AIRE mutation R139X was found in 21.3% of tested alleles, R257X in 11.8%, W78R in 11.4%, C322fsX372 in 8.8%, T16M in 6.2%, R203X in 4%, and A21V in 2.9%. Less frequent mutations were present in 12.9%, very rare in 9.6% while no mutations in 11% of the cases. Conclusions In Italy, APS-1 is a rare disorder presenting with the three major manifestations and associated with different AIRE gene mutations. IFNωAbs are markers of APS-1 and other organ-specific autoantibodies are markers of clinical, subclinical or potential autoimmune conditions. Supplementary Information The online version contains supplementary material available at 10.1007/s40618-021-01585-6.


Introduction
Autoimmune Polyendocrine Syndrome type 1 (APS-1) (OMIM 240300), also termed Autoimmune Poly-Endocrine-Candidiasis-Ectodermal-Dystrophy (APECED) [1] or multiple autoimmune syndrome type 1 (MAS-1) [2,3] is a rare disease with a mean prevalence of 10 cases per million inhabitants [4]. The prevalence varies and is higher in patients with consanguinity or in some particular populations [5]. The highest reported prevalence is in the Iranian Jewish population (1 case per 9000 individuals) [6], followed by Finland (1 case per 25,000 individuals) [1], Norway (1 case per 90,000 individuals) [7] Poland (1 case per 129,000 individuals) [8], and Ireland (1 case per 130,000 individuals) [9] with the lowest prevalence in France (1 case per 500,000 individuals) [10] and in Japan (1 case per 10 million individuals) [5,11]. APS-1 is defined by the presence of at least two of the three major diseases: chronic mucocutaneous candidiasis (CMC), chronic hypoparathyroidism (CH) and Addison's disease (AD) [12]. In addition, patients present with other autoimmune and non-autoimmune diseases and ectodermic dystrophy [1,4,[12][13][14]. In European cohorts, one or more major manifestations presented at the onset in 80-90% of patients while a small proportion (5-20%) presented with other diseases [5,13,15,16]. In contrast, in an American cohort, 40-80% of patients had non-major diseases at manifestation onset and the development of classical manifestations was delayed [17]. These observations have led to establishing new criteria for diagnosing APS-1 which recommend testing for interferon autoantibodies in patients presenting one of the following clinical manifestations: CMC, AD (occurring below 20 years of age), CH, premature ovarian failure (occurring below 30 years of age), enamel hypoplasia, periodic fever with rash, non-infectious keratitis or autoimmune hepatitis. If testing for interferon autoantibodies is not available it is recommended to proceed directly with AIRE gene analysis [4]. APS-1 is caused by autosomal recessive mutations of AIRE gene on chromosome 21 (21q22. 3). AIRE gene plays an essential role in central tolerance. Mutations in the AIRE gene prevent the elimination of self-reactive T cells at central level and induce a Treg defect at peripheral levels [4,18,19]. This leads to the development of multiple autoimmune diseases at a young age [4,20,21]. The most frequent mutation is R257X on exon 6 detectable in 87% of Finnish patients although it was also found in other populations [4,5,15,22,23]. Mutations C322fsX372 or del13 in exon 8 are typical of Anglo-Saxon populations but can also occur in patients of different descent [4,5,[22][23][24]. To date, more than 126 AIRE mutations have been identified [4,5,20]. Although APS-1 is very rare, 20 cohorts were described from 1998 to 2018 [6, 8-11, 15, 17, 20, 24-35] and a total of 568 patients from different populations have been assessed to date (Table 1 and Supplementary Figure 1). Several reports of Italian patients with APS-1 were published from 1974 to 2016 [22,26,[36][37][38][39][40][41][42][43][44][45][46][47][48][49][50][51][52][53]. However, a complete analysis on all the patients with APS-1 in Italy has not been carried out so far. The aim of this study was to collect data on all the APS-1 patients living in Italy.
In collaboration with the Italian Society of Endocrinology (SIE), the Italian Society of Pediatric Endocrinology and  n/a n/a n/a n/a n/a n/a n/a n/a n/a n/a n/a n/a 14 5 n/a n/a n/a n/a n/a n/a n/a n/a n/a n/a n/a n/a n/a n/a n/a n/a

Patients
One hundred and sixty-seven patients with APS-1 were enrolled into the study. Data on age, gender, manifestations, serum biochemistry and serum autoantibody profiles at onset and during a follow-up were collected. The diagnosis of the various diseases was performed according to the criteria previously published [3,54] and summarized in the Table 2. Furthermore, AIRE gene mutations were analyzed. APS-1 was diagnosed using the classical clinical criteria defined in 1980 [12] or according to the new criteria [4]. The ethnic origin and for Italian patients the geographical region of provenance of their families were also recorded.

AIRE gene mutation
All 14 exons of the AIRE gene and their flanking exon-intron boundaries (GenBank accession no. Aj009610) were analyzed in 136 patients and in 170 non affected family members using previously described methods [22]. Data were analyzed for the total Italian cohort and also for groups of patients according to their regional origin.

Autoantibodies
Patients' sera were screened for interferon-ω autoantibodies (IFNωAbs) and for the range of organ and non-organspecific autoantibodies relevant for various autoimmune diseases using methods described previously [26,41,[55][56][57][58][59][60][61][62]. Autoantibody positivity confirmed the diagnosis in patients who presented with clinical autoimmune diseases. Autoantibody positive patients who did not display clinical signs/ symptoms and/or had no biochemical markers of a subclinical disease were periodically followed up to assess the progression to the disease and to calculate the annual incidence of the disease.

Statistical analysis
Comparison of female/male ratios, correlations between autoantibodies and the specific diseases, genotype and phenotype, geographic provenance and phenotype were analyzed by Chi-square test. Statistical significance was determined at p < 0.05. Survival analysis was performed using Kaplan-Meier analysis. Differences between subgroups were tested with Log-Rank and Cox regression.  [63] and is summarized in Table 3. Furthermore, "hot spot areas" for APS-1 were identified in three Italian regions, for example, 12/28 of cases in Sardinia were found in the "Ogliastra" area, 11/24 of Veneto region cases in "Bassano del Grappa" and 12/15 of Apulia cases in "Salento". The patients were followed up from diagnosis of APS-1 until the date of death or to the end of 2019. The mean follow-up period was 23.7 ± 15.1 years (range 1-67 years). The mean age of the patients at the entry and at the end of the follow-up period was 8.3 ± 11.8 years (range 0.5-76) and 32 ± 18 years (range 3-81 years), respectively.

Hypergonadotropic hypogonadism (HH)
Females HH, also known as premature ovarian failure (POF) in females, was found in one 30-year-old patient at the onset of APS-  (Figs. 1, 2). At the onset of DM-1, all affected patients were positive for glutamic acid decarboxylase autoantibodies (GADAbs) and/or islet cell autoantibodies (ICA). Furthermore, 18/64 patients (28.1%) without DM-1 were found positive for ICA and/or GAD-Abs and during follow-up, the annual incidence of clinical DM-1 was 1.1%.

Pituitary insufficiency
None of the 158 patients presented with pituitary insufficiency at the onset of APS-

Autoimmune gastritis (AG)/pernicious anemia (PA)
At the onset of APS-1, AG was observed in one 4-year-old patient. However, 41/158 (25.9%) patients developed AG at a mean age of 23 ± 13 years (range 4-56) by the end of follow-up (Figs. 1, 2). Macrocytic anemia suggesting PA was detected in 21/41 patients with AG and was diagnosed at a mean age of 18.5 years (Fig. 2). Out of 38 patients with AG tested for parietal cell autoantibodies (PCA), 27 (71%) were positive. Intrinsic factor autoantibodies (IFA) were tested in 21 patients with PA and were positive in 13 (62%).

Vitiligo
At the onset of APS-1, 3/158 (2%) patients had vitiligo (isolated in one, associated with other diseases in two) that developed at a mean age of 4 ± 1.7 years. However, by the end of follow-up, vitiligo developed in 27 (17.1%) subjects at a mean age of 17 ± 15 years (range 3-62 years) (Figs. 1,  2). Melanin-producing cell autoantibodies (MPCAbs) were found in 10/18 tested patients with vitiligo and in one without vitiligo who developed vitiligo 10 years later.

Alopecia
Only 3/158 (2%) patients at the onset of APS-1 had alopecia which presented in association with other diseases at 3 years of age in all three subjects. In all, 38/158 (24%) patients developed alopecia by the end of follow-up at a mean age of 13 ± 10 years (range 2-40 years) (Figs. 1, 2).

Autoimmune intestinal dysfunction (AID)
Two APS-  (Figs. 1, 2). TPHAbs were positive in 31/37 (83.8%) and AADC in 36/37 (97.3%) patients with diarrhea or obstinate constipation. Gastric and duodenal biopsies in eight autoantibody-positive patients with diarrhea or obstinate constipation showed the presence of CD3 + / CD8 + intraepithelial lymphocytes with a reduced number of enterochromaffin cells. The mean serum levels of serotonin in these patients were low of 0.16 ± 0.14 μM/L (normal range: 0.28-1.7 μM/L). The mean serotonin levels in other eight TPHAb-positive patients without diarrhea or obstinate constipation were within the lower levels of normal range at 0.58 ± 0.41 μM/L suggesting a subclinical AID.

Autoimmune hepatitis (AIH)
AIH was diagnosed at the onset of APS-1 in 4/158 patients (2.5%) and was isolated in 3 patients while associated with other manifestations in one. The mean age at diagnosis was 2.8 ± 2.2 years. By the end of follow-up, 34 patients (21.5%) developed AIH at a mean age of 13.1 ± 9.2 years (range 1-32 years) (Figs. 1, 2). Liver-kidney microsomal autoantibodies (LKMAbs) were positive in 13/24 patients (66.6%) with AIH, while 3/7 patients (43%) were positive for smooth muscle antibodies. The severity of the disease varied with some subjects having moderate to high levels of liver enzymes and responding to immunosuppression therapy while some (n = 4) progressing to acute fulminant hepatitis causing death.

Spleen atrophy (SA)
None of the patients had SA at the onset of APS-1 while during follow-up, SA was detected in 7/158 patients (4.4%) at a mean age of 25 ± 10 years (range 5-33 years) (Figs. 1, 2). Other very rare diseases Some of the patients in this study presented with other very rare diseases such as exocrine pancreas insufficiency, tubulointerstitial nephritis, hypopotassemia with apparent mineralocorticoid excess or sensitivity, deficit of IgA, metaphyseal dysplasia, vasculitis, autoimmune hemolytic anemia, cutaneous rash with fever, ocular myasthenia, obstructive pulmonary disease positive for potassium channel regulator autoantibodies (KCNR-GAbs), psoriasis, pure red cell aplasia, posterior reversible encephalopathy syndrome, autoimmune demyelinating disease, cystic fibrosis, retinitis pigmentosa, chronic urticaria, lichen ruber planus, polyneuropathy, epilepsy, Hirschsprung's disease, renal agenesia, toxic epidermal necrolysis, HCV and autoimmune myocarditis.

Prevalence of other non-autoimmune diseases at the onset and by the end of follow-up
Overall diseases in patients with APS-1 at the end of follow-up By the end of the mean 23.7 ± 15.1 years (range 1-67 years) of follow-up, 991 diseases were documented in the Italian cohort of APS-1 patients with a mean of 6.3 per patient, ranging widely from 1 to 16 diseases/patient (Fig. 4).

Prevalence of IFNωAbs in patients with APS-1
IFNωAbs were positive in 103/113 (91.2%) patients tested. However, the 10 IFNωAb-negative patients were tested after a mean period of 25 years from the onset of the disease.

Discussion
Our study describes 158 well-characterized Italian patients with APS-1 who represent the largest one nation cohort of patients with APS-1 published to date. Previously large cohorts of 91 Finnish [15] or 112 Russian patients have been reported [33]. In all 568, APS-1 patients from 20 different cohorts have been described previously (Table 1 and supplementary Figure 1) and with addition of 158 patients from our study the total number of well-characterized patient cohorts with APS-1 has reached 726. In addition to the cohort studies, there are a number of case reports on the individual or familial presentations of APS-1 in different countries. Therefore, the overall number of APS-1 patients worldwide could be estimated at approximately 1000 individuals. In our study, patients were recruited from specialist endocrine centers across Italy allowing for the first time an estimate of the prevalence of APS-1 in Italy at 2.6 cases/million, i.e., approximately 1 case per 384,615 individuals. Although the number of patients enrolled was relatively high, the prevalence of APS-1 in Italy is lower than in Iranian Jews, Finland, Norway, Poland and Ireland [1, 5-11, 15, 20] while higher than in France and Japan. Our study showed large variations in the prevalence of APS-1 in different regions of Italy. For example, the highest prevalence was noted in Sardinia with 17 cases/million (1 case per 58,823 inhabitants), followed by Veneto with 4.7, Sicily with 3.9 and Apulia with 3.7 cases/million, respectively. In contrast, lower prevalence (0.6-0.5 cases/million) was observed in the other regions (Table 2). Furthermore, three "hot spot areas" for APS-1 were identified in Italy (Fig. 5), and in those areas AIRE gene screening in the healthy population would be helpful in detecting asymptomatic carriers who could then be offered appropriate genetic counseling. In this study, more than 93% of patients presented with one of the classical triad diseases at onset while 7% manifested with other diseases. A similar clinical presentation has been described in European populations [5,15]. However, in the cohorts of American patients [17,35], non-major component diseases were noted at APS-1 onset in 40-80% of the cases.
In the majority of our Italian patients who presented with the classical triad at onset, the first most frequently manifesting disease was CMC followed by CH and then AD. However, by the end of a long follow-up, the most frequent disease was CH diagnosed in 86% of the patients at a mean age of 11 years, followed by AD in 77% of the patients at a mean age of 16.3 years and CMC in 76% of the patients which presented at a younger mean age of 9.1 years. The highest prevalence of CMC, CH and AD was in patients from the South and Apulia and the lowest from the central regions (Fig. 3). These differences may be related to the differences in genetic background of populations in these regions. Subsequent to initial presentation of the three major manifestations of APS-1, many other autoimmune and nonautoimmune diseases developed throughout patients' lives. APS-1 is a most complex and puzzling autoimmune syndrome due to the great variety and severity of autoimmune and non-autoimmune comorbidities. In our cohort, patients manifested in total 991 diseases with an average of 6.3 conditions per patient. However, 4 exceptional patients in our study developed up to 16 diseases per patient representing the highest co-morbidity in APS-1 reported to date [19].
IFNωAbs were found to be the best serological marker of APS-1 regardless of the comorbidities and the duration of the syndrome as previously described [4,20,[33][34][35]48]. This confirms that IFNωAbs are a diagnostic marker for APS-1 and can be particularly helpful in early diagnosis in patients presenting with a single manifestation of APS-1 [4].
Patients in our study developed cancers at a relatively young age of which approximately half were SCC of the mouth, tongue, esophagus, probably related to the CMC or gastric adenocarcinoma probably related to AG and this is consistent with previous reports [65]. In the case of other cancers, LGLL is a monoclonal lymphoproliferative disease characterized by persistent and indolent lymphocytosis described in only 0.2 and 0.7 cases/million in the USA and Netherlands, respectively [64], and associated with autoimmune diseases (rheumatoid arthritis, Sjogren's syndrome, autoimmune cytopenia, hemolytic anemia, or thrombocytopenia) [66,67]. In our study, LGLL was found in 3/158 (1.9%) of APS-1 patients and this represent the highest frequency of this disorder in an autoimmune syndrome. Only two case reports of LGLL in APS-1 were published so far [68,69]. Finally, one exceptional case developed two adenocarcinomas of the colon not previously reported in this syndrome.
The mortality in APS-1 patients was significantly higher compared to the general Italian population [64] in agreement with previous reports on European [20,[70][71][72] and non-European cohorts [34,35]. Mortality was highest in the North-East regions compared to the other regions in Italy. In approximately one-third of our patients, death was caused by malignancies while in the other two-third causes were adrenal crises, fulminant hepatitis, or generalized candidiasis.
In addition to IFNωAbs, we have also assessed the value of other serological markers of autoimmune diseases. We confirmed the role of NALP-5Abs as markers of CH; however, this test is not widely available outside research laboratories [61]. ACA and/or 21-OHAbs showed excellent diagnostic sensitivity and were detected in the great majority of APS-1 patients with AD within 2 years of onset. Furthermore, they were also found in approximately half of APS-1 patients without AD and conferred a high risk (86.2%) of progression to AD in APS-1 compared to a low risk (25%) in patients with other types of APS [73]. As AD in APS-1 tends to develop after CMC and/or CH, it would seem appropriate to test for ACA and/or 21-OHAbs in all patients with CMC and/or CH. This strategy would alert physicians to the high probability of progression to clinical AD for patients positive for these autoantibodies and would be helpful in preventing unexpected life-threatening adrenal crisis events [74]. Nearly, half of the females with APS-1 in our cohort developed POF (Fig. 1). This is in agreement with previous cohort studies where POF was among the most frequent of the non-classical components of APS-1 (Table 1 and supplementary Figure 1). In our study, the great majority of the patients with POF were positive for StCA and/or 17αOHAbs and/or SCCAbs which are considered diagnostic for autoimmune oophoritis without the need for ovarian biopsy or further genetic investigations [75][76][77]. Furthermore, approximately half of the young females with APS-1 who had normal menses and were positive for StCA and/or 17αOHAbs and/or SCCAbs developed POF during follow-up with an annual incidence of 5.6%. This confirmed that these autoantibodies are also good predictive markers of POF [77]. In addition, StCA, and/or 17αOHAbs and/or SCCAbs were positive in 60% of the males with APS-1 of whom none had HH at the time of testing and during the follow-up only one patient developed HH. Consequently, in this study, positivity for autoantibodies to steroid-producing cells/steroidogenic enzymes was associated with a very different risk of HH in females and males in agreement with previous studies [77,78].
Almost one-third of APS-1 patients had AITD which presented in most cases as CT and only exceptionally as GD consistent with previous reports [15]. Some patients with normal thyroid function were positive for thyroid autoantibodies at entry into the study and during the follow-up, developed hypothyroidism with an annual incidence of 4.5%. This would suggest that thyroid autoantibodies could be considered as markers of progressing to thyroid dysfunction also in APS-1.
During the follow-up, a third of the patients with APS-1 manifested diarrhea or constipation and the great majority of them were positive for TPHAbs and/or AADCAbs. These Abs correlated with lymphocytic infiltration of the gastric and duodenal mucosa, reduced number of enterochromaffin cells and with the low serum serotonin levels consistent with AID. Our study also confirmed that TPHAbs and/ or AADCAbs are markers of a subclinical AID as reported previously [79,80].
In cohorts described previously to date, 3-33% of patients with APS-1 had DM-1 (Table 1 and supplementary Figure 1). In our cohort, DM-1 was found in 8% of patients who were all positive for diabetes-associated autoantibodies. This represents one of the lowest co-occurrences of DM-1 and APS-1 reported thus far. Although 28% of our APS-1 patients without DM-1 were positive for ICA and/or GAD-Abs the annual risk of developing clinical DM-1 at the end of follow-up was very low (1%) compared to patients with other autoimmune diseases [81] or to healthy children [82]. This difference risk may be related to the absence of specific HLA genotypes predisposing to DM-1 in our APS-1 patients. In addition, GADAbs in patients with APS-1 may recognize different epitopes compared to those recognized by patients with DM-1 without APS-1 [83].
AG was observed in a quarter of our patients and a half of these manifested with PA at a very young age although PA usually manifests in elderly individuals. In addition, onefifth of the APS-1 patients had AIH occurring at a mean age of 13 years. The majority of our patients were responsive to immunosuppression therapy but four patients died from fulminant hepatitis. CD has been reported in 5-14% of APS-1 patients in different cohorts (Table 1 and Supplementary Figure 1) while in our study, only 3% of patients were affected by CD which may be related to the absence of HLA genes predisposing to CD in our population.
Our study shows that patients with APS-1 exhibit a wide range of specific autoimmune disorders some of which are rare. Autoantibody tests for these rare conditions are often unavailable in routine diagnostic laboratories and it would be appropriate that these should be provided by specialized, designated reference laboratories in different countries to assist in a complete diagnostic work-up of APS-1.
In this study, 13% of families had at least one other family member with APS-1 and this is one of the highest prevalence of the same autoimmune syndrome in non-twin relatives. It is consistent with the high genetic risk for APS-1 associated with AIRE gene mutations. Consanguinity of parents was found in 5% of families in our study in contrast to earlier reports on different populations [6,31].
Our cohort of patients displayed different AIRE gene mutations some of which were clustered in different geographical regions. This differs from the Finnish, Iranian Jewish or other populations where one specific mutation was predominantly found in the great majority of patients [6,15]. In Italy, the most prevalent mutation was R139X detected in 21.5% of the alleles overall; however, it was predominant in Sardinia being present in 95% of patients. The next most prevalent mutation was R257X found in 11.8% of alleles overall while being the most frequent in Veneto and North-East areas. R257X mutation is the most common among the Finnish APS-1 population [15]. In our cohort, W78R was found in 11.5% of the alleles overall and it was carried by most of the patients from Apulia where it was first detected [42] and in the south of Italy. C322fsX372 was found in 8.1% of APS-1 patients typically in the Veneto and North-East populations. This is a common mutation in patients with an Anglo-Saxon background [21]. In our study, we have identified three novel mutations: S107C and Q108fs on exon 3 in patients from Sicily and c1314-1326del13insGT in a patient from Campania. In our study, 8 patients had only one allelic mutation as reported in other populations [33]. Seven of these had one non-dominant mutation while one patient from central Italy had a dominant mutation G228W described previously [41,84]. The only other dominant mutation associated with APS-1 reported to date was p.C311Y (p932G>A) found in a patient from North Africa [85]. The relatively wide variations in AIRE gene mutations among Italian APS-1 patients may reflect the heterogeneity of the founder genes derived from different migrant populations passing through or settling and living in Italy over time. Furthermore, this study found that some of the mutations were typical for specific geographical regions of Italy. Therefore, recording a detailed family history is essential for patients being tested for AIRE gene mutations. In our study, 10 APS-1 patients had no detectable mutations in the AIRE gene in agreement with other populations [15,84] and this suggests that not yet identified genes could be involved in the development of APS-1. The AIRE gene is strongly implicated in the regulation of organ-specific antigen expression in medullary thymic epithelial cells and in the T cell tolerance [4,19]. However, as suggested by [86], although AIRE is an important transcriptional controller, other controllers should work independently in promiscuous gene expression (PGE). In the light of the foregoing, on a speculative basis, it may be suggested that defects of AIRE partners or of other PGE controllers could be responsible for the development of clinical APECED manifestations in patients that do not harbor AIRE gene mutations. In future studies, on exome sequencing may be useful in extending our understanding of the genetic causes of APS-1. In previous studies, a correlation between genotype and phenotype has been reported in some patients with APS-1 [6,20]. For example, in the Norwegian population some gene deletions were associated with the onset of APS-1 later in life and a milder phenotype while some genotypes which lead to formation of truncated proteins appeared to be associated with CMC and AD [20]. In contrast, some patients carrying the same mutation (even siblings) presented different clinical manifestations and experienced different courses of the disease [18,87]. Among our Italian patients, there were some direct genotype-phenotype associations for R139X, R257X, W78R, T16M and R203X. However, some siblings in our study harboring the same AIRE mutation presented remarkably different phenotypes. This suggests that in addition to key AIRE gene mutation other genetic, environmental or stochastic factors may impact on the phenotypic expression of APS-1 [20].
Overall, our study is a detailed analysis of all available information on the largest cohort to date of patients with APS-1 from a single national population. As such, this study should enhance the current understanding of the complexity of APS-1 and help in the diagnosis and management of this rare disease ultimately benefiting the affected patients and their families.