Abstract 1929: Pan-Cancer analysis of the effects of splice-altering variants on mRNA splicing and stability
Authors: Jayasinghe Reyka G. , Huang Kuan-lin , Ning Jie , Wyczalkowski Matthew , Lu Charles , Xie Mingchao , Wendl Michael , McLellan Michael , Ye Kai , Ding Li 
Published: August 2015

Abstract
Sequences surrounding splice junctions function to recruit splicing elements to promote and suppress the inclusion of an adjacent exon. Mutations disrupting motifs around splice regions and the creation of de novo splice sites have not been functionally characterized genome-wide. Furthermore, many variants that alter the mRNA isoform have been shown to introduce splicing defects, such as premature termination codons (PTCs), which are degraded via nonsense-mediated decay (NMD). Using cancer samples across multiple cancer types, we have the unique opportunity to study the effects of somatic and germline events introduced throughout the genome and with the large dataset provided by The Cancer Genome Atlas (TCGA), we have the power to identify significant events that are influencing mRNA splicing and stability.
In this study we aim to characterize splice altering variants (SAVs) and nonsense mutations that directly effect exon inclusion and mRNA degradation, respectively, utilizing the TCGA dataset consisting of both DNA-Seq (whole genome and/or exome sequencing) and RNA-seq from approximately 8,000 tumors representing 24 major cancer types. We have identified 23,615 somatic splice site mutations and 71,009 somatic nonsense mutations in 24 cancer types. For each variant, we have collected the following expression data derived from the RNA-seq: exon, splice junction, isoform, and gene; defining the expression signature of each variant. Expression signatures of known SAVs will facilitate the identification of novel and mis-classified missense and silent mutations affecting splicing. Exploring the distribution of mutations and their correlation with the aforementioned expression data will allow us to establish commonalities and differences that may be indicative of cancer type, molecular subtype, or clinical characteristics. Furthermore our position dependent analysis of nonsense mutations and their correlation with degradation will enhance our ability to determine significant sites influencing mRNA isoform presence. We hypothesize that some intronic/exonic variants outside the canonical splice site will influence exon inclusion and alter the ratio of mRNA isoforms present.
Our analysis has identified a number of SAVs, including variants that were mis-classified as missense mutations such as c.190 in BRCA1. The germline coding mutation was found to strengthen a cryptic splice site and discovered to have a higher variant allele fraction in the tumor relative to the normal tissue in two ovarian samples. In conclusion our findings and the continued development of our project will contribute to improving current annotation methods and broaden our understanding of variants that affect splicing and their biological contribution to mRNA isoforms selection in the cell.
Citation Format: Reyka G. Jayasinghe, Kuan-lin Huang, Jie Ning, Matthew Wyczalkowski, Charles Lu, Mingchao Xie, Michael Wendl, Michael McLellan, Kai Ye, Li Ding. Pan-Cancer analysis of the effects of splice-altering variants on mRNA splicing and stability. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1929. doi:10.1158/1538-7445.AM2015-1929