Abstract 383: Survey of spliceosome gene mutations and associated splicing defects across 33 cancer types
Authors: Seiler Michael , Peng Shouyong , Agrawal Anant , Tsai Jennifer , Palacino James , Buonamici Silvia , Yu Lihua 
Published: July 2017

Abstract
Hotspot mutations in the core spliceosome genes SF3B1, U2AF1, SRSF2 have been reported at high frequency in hematological malignancies and with lower occurrence in solid tumors. The TCGA Exome-seq and RNAseq data from 33 tumor types, the majority solid tumors, provides a unique opportunity to define the landscape of spliceosome mutations and associated splicing alterations in human malignancies.
To this end, we compiled a list of 404 genes known to be involved in splicing. We performed 2 different mutational analyses; for the first analysis each tumor type was evaluated independently using MutSig2CV and 68 significantly mutated genes (q ≤ 0.1) were identified. The second analysis was performed by systematically looking for genes across all tumor types with loss of function (LoF) or hotspot in-frame mutations, which identified an additional 12 genes. Among the 80 genes, EEF1A1, HNRNPCL1, PCBP1, PHF5A and ZC3H4 carry hotspot mutations in addition to the previously reported spliceosome genes SF1, SF3B1, SRSF2 and U2AF1. Interestingly, we observed that known leukemia mutation/deletion near P95 in SRSF2 were also present in uveal melanoma. Furthermore, the 2 hotspot mutations in U2AF1 identified in leukemia and lung adenocarcinoma (LUAD) were detected in an additional 9 tumor types. We identified new SF3B1 hotspot mutations p.L833F, p.E862K, p.E902K/G, and p.R957Q located in the heat domains (HD) 9-12 in AML, bladder (BLCA), and endometrial cancers. Unexpectedly, alternative exon usage was the most common splicing event observed in p.E902K mutant BLCA samples. This observation differs from the reported alternative 3’ splice site usage induced by SF3B1 hotspot mutations located in the HDs 4-8.
The majority of mutated spliceosome genes (52/80) contained LoF mutations across multiple tumor types. In particular, RBM10 showed the highest mutation frequency in LUAD (6.5%) and BLCA (3.8%), and FUBP1 in low grade glioma (8.0%). Differential splicing analysis comparing RBM10 LoF mutant and wild type LUAD identified exon inclusion and intron skipping as major splicing alterations, consistent with data showing RBM10 knock-down induces alternative exon usage within specific genes. Additionally, we identified cassette exon usage as the major splicing alteration in FUBP1 mutant versus wild type glioma samples. FUBP1 has been reported to bind and repress inclusion of AT rich exons, and confirming this finding we observed higher AT content in exons included in FUBP1 LoF mutant samples when compared to unaffected following exons. These findings suggest that LoF mutations in spliceosome genes impact splicing regulation and may play a critical role in cancer.
In conclusion, the landscape of hotspot and LoF mutations in multiple spliceosome genes and associated splicing alterations highlight the increasing importance of the splicing machinery in tumorigenesis in solid tumors beyond hematological malignancies.
Citation Format: Michael Seiler, Shouyong Peng, Anant Agrawal, Jennifer Tsai, James Palacino, Silvia Buonamici, Lihua Yu. Survey of spliceosome gene mutations and associated splicing defects across 33 cancer types [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 383. doi:10.1158/1538-7445.AM2017-383