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Epigenomic Promoter Alterations Amplify Gene Isoform and Immunogenic Diversity in Gastric Adenocarcinoma

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Epigenomic Promoter Alterations Amplify Gene Isoform and Immunogenic Diversity in Gastric Adenocarcinoma Published OnlineFirst March 20, 2017; DOI: 10.1158/2159-8290.CD-16-1022 RESEARCH ARTICLE Epigenomic Promoter Alterations Amplify Gene Isoform and Immunogenic Diversity in Gastric Adenocarcinoma Aditi Qamra1,2, Manjie Xing3,4, Nisha Padmanabhan3, Jeffrey Jun Ting Kwok5, Shenli Zhang3, Chang Xu3, Yan Shan Leong6, Ai Ping Lee Lim1, Qianqao Tang7, Wen Fong Ooi1, Joyce Suling Lin1, Tannistha Nandi1, Xiaosai Yao1, Xuewen Ong3, Minghui Lee3, Su Ting Tay3, Angie Tan Lay Keng3, Erna Gondo Santoso7, Cedric Chuan Young Ng7, Alvin Ng3,4, Apinya Jusakul3, Duane Smoot8, Hassan Ashktorab9, Sun Young Rha10, Khay Guan Yeoh11,12, Wei Peng Yong13, Pierce K.H. Chow14,15, Weng Hoong Chan16, Hock Soo Ong16, Khee Chee Soo15, Kyoung-Mee Kim17, Wai Keong Wong16, Steven G. Rozen3,18, Bin Tean Teh3,6,7,18, Dennis Kappei6, Jeeyun Lee19, John Connolly5,20, and Patrick Tan1,3,6,18,21 ABSTRACT Promoter elements play important roles in isoform and cell type–specific expression. We surveyed the epigenomic promoter landscape of gastric adenocarcinoma, analyz- ing 110 chromatin profiles (H3K4me3, H3K4me1, H3K27ac) of primary gastric cancers, gastric cancer lines, and nonmalignant gastric tissues. We identified nearly 2,000 promoter alterations (somatic promot- ers), many deregulated in various epithelial malignancies and mapping frequently to alternative promoters within the same gene, generating potential pro-oncogenic isoforms (RASA3). Somatic promoter– associated N-terminal peptides displaying relative depletion in tumors exhibited high-affinity MHC bind- ing predictions and elicited potent T-cell responses in vitro, suggesting a mechanism for reducing tumor antigenicity. In multiple patient cohorts, gastric cancers with high somatic promoter usage also displayed reduced T-cell cytolytic marker expression. Somatic promoters are enriched in PRC2 occupancy, display sensitivity to EZH2 therapeutic inhibition, and are associated with novel cancer-associated transcripts. By generating tumor-specific isoforms and decreasing tumor antigenicity, epigenomic promoter alterations may thus drive intrinsic tumorigenesis and also allow nascent cancers to evade host immunity. SIGNIFICANCE: We apply epigenomic profiling to demarcate the promoter landscape of gastriccancer . Many tumor-specific promoters activate different promoters in the same gene, some generating pro-oncogenic isoforms. Tumor-specific promoters also reduce tumor antigenicity by causing relative depletion of immunogenic peptides, contributing to cancer immunoediting and allowing tumors to evade host immune attack. Cancer Discov; 7(6); 1–22. ©2017 AACR. 1Cancer Therapeutics and Stratified Oncology, Genome Institute of Singapore. 16Department of Upper Gastrointestinal & Bariatric Surgery, Singapore, Singapore. 2Department of Physiology, Yong Loo Lin School of Singapore General Hospital, Singapore. 17Department of Pathology & Trans- Medicine, National University of Singapore, Singapore. 3Cancer and Stem Cell lational Genomics, Samsung Medical Center, Sungkyunkwan University Biology Program, Duke-NUS Medical School, Singapore. 4NUS Graduate School of Medicine, Seoul, Korea. 18SingHealth/Duke-NUS Institute of Pre- School for Integrative Sciences and Engineering, National University of cision Medicine, National Heart Centre Singapore, Singapore. 19Department Singapore, Singapore. 5Institute of Molecular and Cell Biology, Agency for of Medicine, Division of Hematology-Oncology, Samsung Medical Center, Science, Technology and Research, Singapore. 6Cancer Science Institute Sungkyunkwan University School of Medicine, Seoul, Korea. 20Institute of of Singapore, National University of Singapore, Singapore. 7Laboratory Biomedical Studies, Baylor University, Waco, Texas. 21Cellular and Molecular of Cancer Epigenome, Department of Medical Sciences, National Cancer Research, National Cancer Centre, Singapore. 8 Centre, Singapore. Department of Internal Medicine, Meharry Medical Col- Note: Supplementary data for this article are available at Cancer Discovery 9 lege, Nashville, Tennessee. Department of Medicine, Howard University, Online (http://cancerdiscovery.aacrjournals.org/). Washington, DC. 10Department of Internal Medicine, Yonsei University College of Medicine, Seoul, South Korea. 11Department of Medicine, Yong A. Qamra and M. Xing contributed equally to this article. Loo Lin School of Medicine, National University of Singapore and National Corresponding Author: Patrick Tan, Duke-NUS Medical School, 8 College University Health System, Singapore. 12Department of Gastroenterology Road, Singapore 169857, Singapore. Phone: 65-6516-1783; Fax: 65-6221- & Hepatology, National University Hospital, Singapore. 13Department of 2402; E-mail: [email protected] Haematology-Oncology, National University Hospital of Singapore, Singa- doi: 10.1158/2159-8290.CD-16-1022 pore. 14Department of General Surgery, Singapore General Hospital, Singa- pore. 15Division of Surgical Oncology, National Cancer Centre Singapore, ©2017 American Association for Cancer Research. OF1 | CANCER DISCOVERY JUNE 2017 www.aacrjournals.org Downloaded from cancerdiscovery.aacrjournals.org on September 30, 2021. © 2017 American Association for Cancer Research. Published OnlineFirst March 20, 2017; DOI: 10.1158/2159-8290.CD-16-1022 INTRODUCTION of biological, functional, and regulatory diversity, as current estimates suggest that 30% to 50% of genes in the human Gastric cancer is the third leading cause of global cancer genome are associated with multiple promoters (15), which mortality with high prevalence in many East Asian countries can be selectively activated as a function of developmental lin- (1). Patients with gastric cancer often present with late-stage eage and cellular state (16). Differential usage of alternative disease (2, 3), and clinical management remains challenging, promoters causes the generation of distinct 5′ untranslated as exemplified by several recent negative phase II and phase regions (5′ UTR) and first exons in transcripts, which in turn III clinical trials (4–7). At the molecular level, studies have can influence mRNA expression levels (17), translational identified characteristic gene mutations (8, 9), copy-number efficiencies (18, 19), and the generation of different protein alterations, gene fusions (10), and transcriptional patterns in isoforms through gain and loss of 5′ coding domains (15, 20). gastric cancer (11, 12). However, few of these have been clini- In cancer, alternative promoters in genes such as ALK (21), cally translated into targeted therapies, with the exception of TP53 (22), LEF1 (23), and CYP19A1 (24) have been reported, HER2-positive gastric cancer and trastuzumab (13). There is producing cancer-specific isoform variants with oncogenic thus a strong need for additional and more comprehensive properties. To date, promoter alterations in cancer have been explorations of gastric cancer, as these may highlight new bio- largely studied on a gene-by-gene basis, and very little is markers for disease detection, predicting patient prognosis or known about the global extent of promoter-level diversity in responses to therapy, as well as new therapeutic modalities. gastric cancer and other solid malignancies. Promoter elements are cis-regulatory elements that func- Promoters in the genome can be experimentally identified tion to link gene transcription initiation to upstream reg- by various methods. Broadly divided into RNA-based or epi- ulatory stimuli, integrating inputs from diverse signaling genomic approaches, the former involves technologies such pathways (14). Promoters represent an important reservoir as RNA sequencing (RNA-seq), CAP analysis gene expression JUNE 2017 CANCER DISCOVERY | OF2 Downloaded from cancerdiscovery.aacrjournals.org on September 30, 2021. © 2017 American Association for Cancer Research. Published OnlineFirst March 20, 2017; DOI: 10.1158/2159-8290.CD-16-1022 RESEARCH ARTICLE Qamra et al. (CAGE), and global run-on sequencing (25–27). For the latter, To enable accurate promoter identification, we inte- active promoters have been shown to exhibit characteristic grated data from multiple histone modifications, selecting chromatin modifications, specifically H3K4me3 positivity, H3K4me3 regions simultaneously codepleted for H3K4me1 H3K27ac positivity, and H3K4me1 depletion (28–31). Com- (“H3K4me3hi/H3K4me1lo regions”; Supplementary Fig. S1; pared with transcriptome sequencing (25), using histone Methods; ref. 42). Comparisons against data from exter- modifications to identify promoters carries certain advan- nal sources, including GENCODE reference transcripts, tages. First, epigenome-guided promoter identification allows ENCODE chromatin state models, and CAGE databases, genomic localization of the promoter element itself, rather validated the vast majority of H3K4me3hi/H3K4me1lo regions than the ensuing transcript product. Second, particularly for as true promoter elements (Supplementary Text; Supplemen- clinical samples, epigenome-guided promoter identification tary Fig. S1). Because primary gastric tissues comprise several is less prone to transcript degradation artifacts caused by 5′ different tissue types, including epithelial cells, immune cells, RNA exonucleases (32). Epigenome-marked promoters may and stroma, we further confirmed that our promoter profiles also highlight transcript classes not easily detectible by other were reflective of bona fide gastric epithelia by comparison means, such as promoters originating via recapping events, against Epigenome Roadmap data for gastric
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