P-182 Examination of Crohn's Disease Variants in a Fraternal Twin Family Using Exome Sequencing and Inflammatory Bowel Disease-Associated Single Nucleotide Polymorphisms. (March 2016)

• Record Type: Journal Article
• Authors: Choi, Christine; Coble, Joel; Schneper, Lisa; Berg, Arthur; Harris, Leonard; Hegarty, John; Deiling, Sue; Broach, James; Koltun, Walter
• Is Part Of: Inflammatory bowel diseases. Volume 22(2016:Mar.)Supplement 1
• Publication Date: 2016-03
• Subjects: Inflammatory bowel diseases -- Periodicals; Colitis, Ulcerative -- Periodicals; Crohn Disease -- Periodicals; Inflammatory Bowel Diseases -- Periodicals; 616.344
• DOI: 10.1097/01.MIB.0000480296.86595.4c
• Languages: English
• ISSNs: 1078-0998

Abstract

Abstract : Background: A unique opportunity for genetic investigation was presented by a 3-generation family with fraternal twins, one of whom had Crohn's disease (CD) with 2 affected children and the other who was healthy with 3 unaffected children, suggesting an autosomal dominant inheritance. This study tested the hypothesis that inflammatory bowel disease (IBD)-associated single nucleotide polymorphism (SNP) genotyping and exome sequencing methods may be combined to identify and/or determine new genetic variants and/or genotypes that correlate with diseased versus non-diseased family members. Methods: A total of 13 family members, spanning 3-generations, which included 3 diseased and 10 non-diseased individuals were recruited into the institution's Colorectal Biobank. The age of onset of CD for the index fraternal twin was 29, while her diseased son and daughter were age 19 and 25, respectively. The analysis first was done using a customized Illumina microarray containing 324 IBD-associated SNPs for 11 family members. In the second analysis, genomic DNA from 6 critical individuals (mother of fraternal twins [generation 1], fraternal twins [generation 2], affected children [generation 3] and their father [generation 2]) was subjected to whole exome sequencing (WES; >40× coverage) using the Nextera Rapid WES expanded library capture kit and run on an Illumina HiSeq2500. Sequence data was aligned to b37 using bwa 0.7.3a and subsequent realignment and joint variant calling was performed using GATK (v.2.7.4) best practice guidelines. Further filtering criteria was initially based on disease segregation in the pedigree and genotype quality scores of >15, depth >10 and allele balance >0.35 in the index patient and <0.35 in non-diseased individuals. Initially, 17 variants identified by segregating with disease and having a predicted damaging effect (e.g., nonsynonymous, CADD score), expression pattern and potential biological relevance, were targeted for validation by Sanger sequencing and/or Taqman assay. Variants were analyzed in all 13 recruited family members to confirm the presence and determine the segregation pattern of any specific single nucleotide variant (SNV). Results: None of the 324 IBD-associated SNPs defined by the custom microarray or the 17 variants identified by exome sequencing correlated according to diseased versus non-diseased family members in an autosomal dominant fashion. Conclusions: Our current analysis did not suggest an autosomal dominant pattern of inheritance. Additional haplotype analysis (specific combinations of risk alleles) of multiple IBD-SNPs may reveal disease-specific genotypes. Further exome sequencing of the unaffected father and siblings of the index patient may be instrumental in identifying variants contributing to disease. Factors adversely influencing this analysis include possible environmental factors and potential delayed presentation of disease in the presumed unaffected family members. Further analysis of this family is ongoing with the aim to discover either novel variants and/or specific combinations of risk alleles that correlate with disease presence.