O-003 Understanding the Relevance of Whole Exome Sequencing Identified Variants in Patients with Very Early-Onset-IBD.

BACKGROUND: Very early onset inflammatory bowel disease (VEO-IBD) is frequently considered a different disease process than older onset IBD. The severe phenotype and young age of onset suggest a more pronounced genetic susceptibility and dysregulated immune response. We hypothesized that rare or novel variants involving pathways in barrier defense, autoimmunity as well as both B and T cell development and activation, were enriched in patients with VEO-IBD. In turn, these variants result in altered gene expression, impaired immunological responses, and aberrant host-microbe interactions. Our primary aim is to identify dysregulation in the gene expression profiles of VEO-IBD patients, to determine the down-stream impact of candidate causal variants on the affected immune pathways and to delineate their role in disease pathogenesis.

METHODS: IBD patients 5 years and younger, and parents were recruited. Exome capture was performed by Agilent SureSelect V4, and sequencing was accomplished using the Illumina HiSeq platform. Following functional annotation, single base substitutions likely to alter protein function, such as missense and loss of function mutations, were kept for subsequent analysis. RNA isolated from whole blood collected in Tempus tubes was hybridized to Illumina HumanHT-12v4 arrays to evaluate the downstream effects of the variants on the mRNA product through changes in gene expression.

RESULTS: Three hundred eleven samples were analyzed, including 71 trios. Patients' age ranged from 3 weeks to 4 years. 4 trios were analyzed for gene expression. WES identified compound heterozygous mutations in FAT1, and a heterozygous variant in BIRC6 in a male diagnosed with severe VEO-IBD at age 2, unresponsive to medical and surgical therapy. FAT1 is an antagonist of caspase-8, and BIRC6 is a member of the Inhibitors of Apoptosis protein (IAP) family. Gene expression demonstrated upregulation of genes involved in leukocyte apoptosis, including CASP7 and CASP8. This increased apoptotic activity is similar to the mechanism seen in XIAP deficiency, a known primary immunodeficiency that can result in VEO-IBD. In addition, there was an upregulation of NAIP, and subsequent down-stream increased anti-microbial activity. NAIP is a component of the NLRC4 inflammasome.

CONCLUSIONS: The association of candidate risk variants with gene expression provides a powerful method to mine WES identified variants. This case illustrates the ability to detect downstream effects of suspected causal variants. Currently there is a paucity of data utilizing this strategy in patients with VEO-IBD. This systems biology-based approach may provide further understanding of the interacting pathways responsible for a subset of disease in VEO-IBD.