Very Early Onset Inflammatory Bowel Disease Associated with Aberrant Trafficking of IL-10R1 and Cure by T Cell Replete Haploidentical Bone Marrow Transplantation
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Loss-of-function mutations in IL10 and IL10R cause very early onset inflammatory bowel disease (VEO-IBD). Here, we investigated the molecular pathomechanism of a novel intronic IL10RA mutation and describe a new therapeutic approach of T cell replete haploidentical hematopoietic stem cell transplantation (HSCT).
Clinical data were collected by chart review. Genotypes of IL10 and IL10R genes were determined by Sanger sequencing. Expression and function of mutated IL-10R1 were assessed by quantitative PCR, Western blot analysis, enzyme-linked immunosorbent assays, confocal microscopy, and flow cytometry.
We identified a novel homozygous point mutation in intron 3 of the IL10RA (c.368-10C > G) in three related children with VEO-IBD. Bioinformatical analysis predicted an additional 3′ splice site created by the mutation. Quantitative PCR analysis showed normal mRNA expression of mutated IL10RA. Sequencing of the patient’s cDNA revealed an insertion of the last nine nucleotides of intron 3 as a result of aberrant splicing. Structure-based modeling suggested misfolding of mutated IL-10R1. Western blot analysis demonstrated a different N-linked glycosylation pattern of mutated protein. Immunofluorescence and FACS analysis revealed impaired expression of mutated IL-10R1 at the plasma membrane. In the absence of HLA-identical donors, T cell replete haploidentical HSCT was successfully performed in two patients.
Our findings expand the spectrum of IL10R mutations in VEO-IBD and emphasize the need for genetic diagnosis of mutations in conserved non-coding sequences of candidate genes. Transplantation of haploidentical stem cells represents a curative therapy in IL-10R-deficient patients, but may be complicated by non-engraftment.
KeywordsInterleukin-10 very early onset inflammatory bowel disease children hematopoietic stem cell transplantation
enzyme-linked immunosorbent assay
Interleukin-10 receptor 1
peripheral blood mononuclear cells
tumor necrosis factor-α
very early onset inflammatory bowel disease
We thank the patients, the interdisciplinary medical team, and clinicians involved in the care of these patients. The study is supported by grants from DFG (Gottfried-Wilhelm-Leibniz Program), BioSysNet, SFB1054, the Helmsley Charitable Trust, the Care-for-Rare Foundation (fellowship to D.K. and D.M.), and the DFG grant BO 2512/6-1 to J.B..
D.M. is a student of the PhD program Molecular Medicine at Hannover Biomedical Research School, Germany.
Conflict of Interest
The authors declare no competing financial interests.
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