Author Manuscript Published OnlineFirst on February 24, 2021; DOI: 10.1158/1055-9965.EPI-20-1457 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Rare variants in the DNA repair pathway and the risk of colorectal cancer Marco Matejcic1, Hiba A. Shaban1, Melanie W. Quintana2, Fredrick R. Schumacher3,4, Christopher K. Edlund5, Leah Naghi6, Rish K. Pai7, Robert W. Haile8, A. Joan Levine8, Daniel D. Buchanan9,10,11, Mark A. Jenkins12, Jane C. Figueiredo13, Gad Rennert14, Stephen B. Gruber15, Li Li16, Graham Casey17, David V. Conti18†, Stephanie L. Schmit1,19† † These authors contributed equally to this work. Affiliations 1 Department of Cancer Epidemiology, Moffitt Cancer Center, Tampa, FL 33612, USA 2 Berry Consultants, Austin, TX, 78746, USA 3 Department of Population and Quantitative Health Sciences, Case Western Reserve University, Cleveland, OH 44106, USA 4 Seidman Cancer Center, University Hospitals, Cleveland, OH 44106, USA 5 Department of Preventive Medicine, USC Norris Comprehensive Cancer Center, Keck School of Medicine, University of Southern California, Los Angeles, CA, USA 6 Department of Medicine, Montefiore Medical Center, Albert Einstein College of Medicine, NY 10467, USA 7 Department of Laboratory Medicine and Pathology, Mayo Clinic Arizona, Scottsdale, AZ 85259, USA 8 Department of Medicine, Research Center for Health Equity, Cedars-Sinai Samuel Oschin Comprehensive Cancer Center, Los Angeles, CA 90048, USA 9 Colorectal Oncogenomics Group, Department of Clinical Pathology, The University of Melbourne, Parkville, Victoria 3010, Australia 10 Victorian Comprehensive Cancer Centre, University of Melbourne, Centre for Cancer Research, Parkville, Victoria 3010, Australia 1 Downloaded from cebp.aacrjournals.org on September 29, 2021. © 2021 American Association for Cancer Research. Author Manuscript Published OnlineFirst on February 24, 2021; DOI: 10.1158/1055-9965.EPI-20-1457 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. 11 Genomic Medicine and Family Cancer Clinic, Royal Melbourne Hospital, Parkville, Victoria 3010, Australia 12 Centre for Epidemiology and Biostatistics, Melbourne School of Population and Global Health, The University of Melbourne, Melbourne, Victoria, Australia 13 Samuel Oschin Comprehensive Cancer Institute, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA 14 Clalit National Cancer Control Center, Carmel Medical Center and Technion Faculty of Medicine, Haifa 34362, Israel 15 Center for Precision Medicine, City of Hope, Duarte, CA 91010, USA 16 Department of Family Medicine, University of Virginia, Charlottesville, VA 22904, USA 17 Center for Public Health Genomics, University of Virginia, Charlottesville, VA 22908, USA 18 Department of Preventive Medicine, Division of Biostatistics, University of Southern California, Los Angeles, CA 90033, USA 19 Department of Gastrointestinal Oncology, Moffitt Cancer Center, Tampa, FL 33612, USA Running title Rare exome-wide variation and colorectal cancer risk Keywords Colorectal cancer, rare variants, DNA repair pathway, exome-wide association analysis, genetic epidemiology 2 Downloaded from cebp.aacrjournals.org on September 29, 2021. © 2021 American Association for Cancer Research. Author Manuscript Published OnlineFirst on February 24, 2021; DOI: 10.1158/1055-9965.EPI-20-1457 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Financial support The Colon Cancer Family Registry (CCFR) was supported by the National Cancer Institute (NCI), of the National Institutes of Health (NIH) under award number U01 CA167551. Additional support for case ascertainment was provided from the Surveillance, Epidemiology, and End Results (SEER) Program of the NCI and the following U.S. state cancer registries: AZ, CO, MN, NC, NH; and by the Victoria Cancer Registry (Australia) and Ontario Cancer Registry (Canada). The content of this manuscript does not necessarily reflect the views or policies of the National Cancer Institute or any of the collaborating centers in the Colon CFR, nor does mention of trade names, commercial products, or organizations imply endorsement by the US Government, any cancer registry, or the Colon CFR. The Kentucky Case-Control Study (KY) was supported by: Damon Runyon Cancer Research Foundation Clinical Investigator Award, CI-8 (Li Li); the Case Center for Transdisciplinary Research on Energetics and Cancer, U54 CA-116867-01 (Li Li); National Cancer Institute K22 Award, K22 CA120545-01 (Li Li); State of Ohio Biomedical Research and Technology Transfer Commission; and the National Cancer Institute Award R25 CA094186-06. The Molecular Epidemiology of Colorectal Cancer Study (MECC) was supported by the National Cancer Institute at the National Institutes of Health: R01 CA197350, R01 CA81488, P30 CA014089, U19 CA148107 (Stephen B. Gruber), P01 CA196569 and a generous gift from Daniel and Maryann Fong. This work was also supported by the National Human Genome Research Institute at the National Institutes of Health [T32 HG000040] and the National Institute of Environmental Health Sciences at the National Institutes of Health [T32 ES013678]. Corresponding author Stephanie Schmit, PhD, MPH Genomic Medicine Institute 3 Downloaded from cebp.aacrjournals.org on September 29, 2021. © 2021 American Association for Cancer Research. Author Manuscript Published OnlineFirst on February 24, 2021; DOI: 10.1158/1055-9965.EPI-20-1457 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Cleveland Clinic 9500 Euclid Avenue / NE50 Cleveland, OH 44195 Phone: +1 216 444 3173 Email: [email protected] Conflict of interest disclosure The authors declare no potential conflicts of interest. Word count: 4,261 Total no. tables: 3 Total no. figures: 2 4 Downloaded from cebp.aacrjournals.org on September 29, 2021. © 2021 American Association for Cancer Research. Author Manuscript Published OnlineFirst on February 24, 2021; DOI: 10.1158/1055-9965.EPI-20-1457 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Abstract Background. Inherited susceptibility is an important contributor to colorectal cancer (CRC) risk and rare variants in key genes or pathways could account in part for the missing proportion of CRC heritability. Methods. We conducted an exome-wide association study including 2,327 cases and 2,966 controls of European ancestry from three large epidemiologic studies. Single variant associations were tested using logistic regression models adjusting for appropriate study-specific covariates. In addition, we examined the aggregate effects of rare coding variation at the gene and pathway levels using Bayesian model uncertainty techniques. Results. In an exome-wide gene-level analysis, we identified ST6GALNAC2 as the top associated gene based on the Bayesian Risk Index (BRI) method (summary Bayes Factor[BF]BRI= 2604.23). A rare coding variant in this gene, rs139401613, was the top associated variant (P=1.01x10-6) in an exome-wide single variant analysis. Pathway-level association analyses based on the integrative Bayesian Risk Index (iBRI) method found extreme evidence of association with the DNA repair pathway (BFiBRI=17852.4), specifically with the non-homologous end joining (NHEJ, BFiBRI=437.95) and nucleotide excision repair (NER, BFiBRI=36.96) subpathways. The iBRI method also identified RPA2, PRKDC, ERCC5, ERCC8 as the top associated DNA repair genes (summary BFiBRI≥10), with rs28988897, rs8178232, rs141369732, rs201642761 being the most likely associated variants in these genes, respectively. Conclusion. We identified novel variants and genes associated with CRC risk and provided additional evidence for a role of DNA repair in CRC tumorigenesis. Impact. This study provides new insights into the genetic predisposition to CRC which has potential for translation into improved risk prediction. 5 Downloaded from cebp.aacrjournals.org on September 29, 2021. © 2021 American Association for Cancer Research. Author Manuscript Published OnlineFirst on February 24, 2021; DOI: 10.1158/1055-9965.EPI-20-1457 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Introduction Inherited genetic factors play an important role in the etiology of colorectal cancer (CRC) (1). Genome-wide association studies (GWAS) in CRC have identified approximately 140 common risk loci predominantly in European and East Asian populations (2-4). However, risk variants, both high- and low-penetrance, collectively account for only a small proportion of the estimated familial risk (3). Part of the unexplained proportion of CRC heritability may be attributable to rare variants in genes/pathways with established or suspected roles in colorectal carcinogenesis, such as DNA repair, TGF-β signaling, vitamin D, and folate metabolism (5-8). Rare variants by definition occur with a <1% frequency in the population and therefore large sample sizes are required to detect rare risk variants with modest effect sizes. Yet, next-generation sequencing approaches have enabled successful identification of population-specific rare variants and assessed their contributions to CRC risk. A recent exome sequencing study of early-onset CRC cases identified rare, highly penetrant pathogenic variants in 16% of cases, including novel variants in POT1, POLE2, and MRE11 (9). Whole-genome sequencing of sporadic CRC cases identified