Gastroenterology

Gastroenterology

Volume 160, Issue 4, March 2021, Pages 1164-1178.e6
Gastroenterology

Original Research
Full Report: Basic and Translational—Alimentary Tract
Identifying Novel Susceptibility Genes for Colorectal Cancer Risk From a Transcriptome-Wide Association Study of 125,478 Subjects

https://doi.org/10.1053/j.gastro.2020.08.062Get rights and content

Background And Aims

Susceptibility genes and the underlying mechanisms for the majority of risk loci identified by genome-wide association studies (GWAS) for colorectal cancer (CRC) risk remain largely unknown. We conducted a transcriptome-wide association study (TWAS) to identify putative susceptibility genes.

Methods

Gene-expression prediction models were built using transcriptome and genetic data from the 284 normal transverse colon tissues of European descendants from the Genotype-Tissue Expression (GTEx), and model performance was evaluated using data from The Cancer Genome Atlas (n = 355). We applied the gene-expression prediction models and GWAS data to evaluate associations of genetically predicted gene-expression with CRC risk in 58,131 CRC cases and 67,347 controls of European ancestry. Dual-luciferase reporter assays and knockdown experiments in CRC cells and tumor xenografts were conducted.

Results

We identified 25 genes associated with CRC risk at a Bonferroni-corrected threshold of P < 9.1 × 10–6, including genes in 4 novel loci, PYGL (14q22.1), RPL28 (19q13.42), CAPN12 (19q13.2), MYH7B (20q11.22), and MAP1L3CA (20q11.22). In 9 known GWAS-identified loci, we uncovered 9 genes that have not been reported previously, whereas 4 genes remained statistically significant after adjusting for the lead risk variant of the locus. Through colocalization analysis in GWAS loci, we additionally identified 12 putative susceptibility genes that were supported by TWAS analysis at P < .01. We showed that risk allele of the lead risk variant rs1741640 affected the promoter activity of CABLES2. Knockdown experiments confirmed that CABLES2 plays a vital role in colorectal carcinogenesis.

Conclusions

Our study reveals new putative susceptibility genes and provides new insight into the biological mechanisms underlying CRC development.

Section snippets

Genome-Wide Association Study Data From Study Cohorts

The study utilized GWAS summary statistics data from 125,478 individuals of European descent (58,131 CRC cases and 67,347 controls) from 6 cohort studies. The cohort studies include Genetics and Epidemiology of Colorectal Cancer Consortium (n = 11,958 case, n = 14,740 controls), CORECT (n = 22,911 case, n = 14,311 controls), CROC (n = 12,007 cases, n = 12,000 controls), CONE (n = 4439 cases, n = 4115 controls), CORSA (n = 1460 cases, n = 774 controls), and UKBio (n = 5356 cases, n = 21,407

Gene Expression Predicted by Flanking Genetic Variants

We used transcriptome and genotype data from the GTEx to build gene-expression prediction models for normal transverse colon tissues from European descendants (n = 284) (see Methods). A total of 16,082 models were built for genes predicted by flanking genetic variants (flanking ±1 Mb region) using the elastic net approach.36,49 Our results showed that the expression levels of 9503 genes could be predicted by local genetic variants at R2 > 0.01 (10% correlation), with a median of 27 variants per

Discussion

Utilizing data from a large-scale GWAS collaboration of GTEx and TCGA, we conducted a TWAS analysis to search for susceptibility genes for CRC risk. We disclosed 25 putative CRC predisposition genes, including 6 novel genes located in the regions far away from any previously reported susceptibility loci for CRC risk, and 9 novel genes as targets for established CRC GWAS loci. We have also identified an additional 48 genes with genetically predicted expressions associated with CRC risk using a

Acknowledgments

The authors thank the CLUE cohort GTEx, TCGA, ENCODE, and Roadmap for providing valuable data resources for the research. The authors thank Drs Lang Wu, Yingchang Lu, and Chenjie Zeng for valuable discussion and Marshal Younger for assistance with editing and manuscript preparation.

CRediT Authorship Contributions

Xingyi Guo, PhD (Conceptualization: Lead; Data curation: Lead; Formal analysis: Lead; Funding acquisition: Lead; Methodology: Lead; Resources: Lead; Supervision: Lead; Writing – original draft: Lead)

Weiqiang Lin, PhD

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    Conflicts of interest The authors disclose no conflicts.

    Funding The data analyses were conducted using the Advanced Computing Center for Research and Education at Vanderbilt University. This research is supported primarily by the grant from US National Institutes of Health (NIH) grant R37 CA227130 to Xingyi Guo and R01 CA188214 to Wei Zheng. Genetics and Epidemiology of Colorectal Cancer Consortium: National Cancer Institute (NCI), NIH, US Department of Health and Human Services (U01 CA164930, U01 CA137088, R01 CA059045, R21 CA191312, R01 CA201407). Genotyping/Sequencing services were provided by the Center for Inherited Disease Research (CIDR) (X01-HG008596 and X01-HG007585). CIDR is fully funded through a federal contract from the NIH to The Johns Hopkins University, contract number HHSN268201200008I. This research was funded in part through the NIH/NCI Cancer Center Support Grant P30 CA015704. PCS is supported by the NIH (R01 CA160356, R01 CA193677, R01 CA204279 and R01 CA143237). ASTERISK: a Hospital Clinical Research Program (PHRC-BRD09/C) from the University Hospital Center of Nantes and supported by the Regional Council of Pays de la Loire, the Groupement des Entreprises Françaises dans la Lutte contre le Cancer, the Association Anne de Bretagne Génétique and the Ligue Régionale Contre le Cancer. The ATBC Study is supported by the Intramural Research Program of the US NCI, NIH, and by US Public Health Service contract HHSN261201500005C from the NCI, Department of Health and Human Services. CLUE II: This research was funded by the American Institute for Cancer Research and the NCI (P30 CA006973 to W.G. Nelson). COLO2&3: NIH (R01 CA60987). ColoCare: This work was supported by the NIH (grant numbers R01 CA189184 (Li/Ulrich), U01 CA206110 (Ulrich/Li/Siegel/Figueireido/Colditz), 2P30CA015704-40 (Gilliland), R01 CA207371 (Ulrich/Li)), the Matthias Lackas-Foundation, the German Consortium for Translational Cancer Research, and the EU TRANSCAN initiative. The Colon Cancer Family Registry (CFR) Illumina GWAS was supported by funding from the NCI, NIH (grant numbers U01 CA122839, R01 CA143247). The Colon CFR/CORECT Affymetrix Axiom GWAS and OncoArray GWAS were supported by funding from NCI, NIH (grant number U19 CA148107 to S Gruber). The Colon CFR participant recruitment and collection of data and biospecimens used in this study were supported by the NCI, NIH (grant number UM1 CA167551) and through cooperative agreements with the following Colon CFR centers: Australasian Colorectal Cancer Family Registry (NCI/NIH grant numbers U01 CA074778 and U01/U24 CA097735), USC Consortium Colorectal Cancer Family Registry (NCI/NIH grant numbers U01/U24 CA074799), Mayo Clinic Cooperative Family Registry for Colon Cancer Studies (NCI/NIH grant number U01/U24 CA074800), Ontario Familial Colorectal Cancer Registry (NCI/NIH grant number U01/U24 CA074783), Seattle Colorectal Cancer Family Registry (NCI/NIH grant number U01/U24 CA074794), and University of Hawaii Colorectal Cancer Family Registry (NCI/NIH grant number U01/U24 CA074806), Additional support for case ascertainment was provided by the Surveillance, Epidemiology and End Results Program of the NCI to Fred Hutchinson Cancer Research Center (Control Nos. N01-CN-67009 and N01-PC-35142, and Contract No. HHSN2612013000121), the Hawai‘i Department of Health (Control Nos. N01-PC-67001 and N01-PC-35137, and Contract No. HHSN26120100037C, and the California Department of Public Health (contracts HHSN261201000035C awarded to the University of Southern California, and the following state cancer registries: AZ, CO, MN, NC, NH, and by the Victoria Cancer Registry and Ontario Cancer Registry. COLON: The COLON study is sponsored by Wereld Kanker Onderzoek Funds, including funds from grant 2014/1179 as part of the World Cancer Research Fund International Regular Grant Programme, by Alpe d’Huzes and the Dutch Cancer Society (UM 2012–5653, UW 2013-5927, UW2015-7946), and by TRANSCAN (JTC2012-MetaboCCC, JTC2013-FOCUS). The NQplus study is sponsored by a ZonMW investment grant (98-10030); by PREVIEW, the project PREVention of diabetes through lifestyle intervention and population studies in Europe and around the World (PREVIEW) project, which received funding from the European Union Seventh Framework Programme (FP7/2007–2013) under grant no. 312057; by funds from TI Food and Nutrition (cardiovascular health theme), a public–private partnership on pre-competitive research in food and nutrition; and by FOODBALL, the Food Biomarker Alliance, a project from JPI Healthy Diet for a Healthy Life. Colorectal Cancer Transdisciplinary (CORECT) Study: The CORECT Study was supported by the NCI, NIH, US Department of Health and Human Services (grant numbers U19 CA148107, R01 CA81488, P30 CA014089, R01 CA197350, P01 CA196569, R01 CA201407, and R01 CA143237) and National Institutes of Environmental Health Sciences, NIH (grant number T32 ES013678). CORSA: This study was funded by FFG BRIDGE (grant 829675 to Andrea Gsur), the “Herzfelder’sche Familienstiftung” (grant to Andrea Gsur) and was supported by COST Action BM1206. Cancer Prevention Study-II (CPS-II): The American Cancer Society funds the creation, maintenance, and updating of the CPS-II cohort. This study was conducted with Institutional Review Board approval. Colorectal Cancer Genetics & Genomics, Spanish study, was supported by Instituto de Salud Carlos III, co-funded by FEDER funds –a way to build Europe– (grants PI14-613 and PI09-1286), Agency for Management of University and Research Grants of the Catalan Government (grant 2017SGR723), and Junta de Castilla y León (grant LE22A10-2). Sample collection of this work was supported by the Xarxa de Bancs de Tumors de Catalunya sponsored by Pla Director d’Oncología de Catalunya (XBTC), Plataforma Biobancos PT13/0010/0013 and ICOBIOBANC, sponsored by the Catalan Institute of Oncology. Czech Republic CCS: This work was supported by the Czech Science Foundation (grants 17-16857S and 18-09709S), the Grant Agency of the Ministry of Health of the Czech Republic (grants 15-27580A and 17-30920A), the Charles University Research Centre program (UNCE/MED/006), the Charles University Research Fund (Progres Q39 and Q28), and MEYS CR, financed from EFRR (CZ.02.1.01/0.0/0.0/16_019/0000787). DACHS: This work was supported by the German Research Council (BR 1704/6-1, BR 1704/6-3, BR 1704/6-4, BR 1704/6-6, CH 117/1-1, HO 5117/2-1, HE 5998/2-1, KL 2354/3-1, RO 2270/8-1 and BR 1704/17-1), the Interdisciplinary Research Program of the National Center for Tumor Diseases, Germany, and the German Federal Ministry of Education and Research (01KH0404, 01ER0814, 01ER0815, 01ER1505A and 01ER1505B). DALS: NIH (R01 CA48998 to M. L. Slattery). EDRN: This work is funded and supported by the NCI, EDRN grant (U01 CA 84968-06). EPIC: The coordination of EPIC is financially supported by the School of Public Health, Imperial College London and the International Agency for Research on Cancer. The national cohorts are supported by Danish Cancer Society (Denmark); Ligue Contre le Cancer, Institut Gustave Roussy, Mutuelle Générale de l’Education Nationale, Institut National de la Santé et de la Recherche Médicale (France); German Cancer Aid, German Cancer Research Center, Federal Ministry of Education and Research, Deutsche Krebshilfe, Deutsches Krebsforschungszentrum and Federal Ministry of Education and Research (Germany); the Hellenic Health Foundation (Greece); Associazione Italiana per la Ricerca sul Cancro-AIRC-Italy and National Research Council (Italy); Dutch Ministry of Public Health, Welfare and Sports, Netherlands Cancer Registry, LK Research Funds, Dutch Prevention Funds, Dutch ZON (Zorg Onderzoek Nederland), World Cancer Research Fund, Statistics Netherlands (The Netherlands); ERC-2009-AdG 232997 and Nordforsk, Nordic Centre of Excellence programme on Food, Nutrition and Health (Norway); Health Research Fund, PI13/00061 to Granada, PI13/01162 to EPIC-Murcia, Regional Governments of Andalucía, Asturias, Basque Country, Murcia and Navarra, ISCIII RETIC (RD06/0020) (Spain); Swedish Cancer Society, Swedish Research Council and County Councils of Skåne and Västerbotten (Sweden); Cancer Research UK (14136 to EPIC-Norfolk; C570/A16491 and C8221/A19170 to EPIC-Oxford), Medical Research Council (1000143 to EPIC-Norfolk, MR/M012190/1 to EPIC-Oxford) (United Kingdom).

    Disclaimer: Where authors are identified as personnel of the International Agency for Research on Cancer/World Health Organization, the authors alone are responsible for the views expressed in this article and they do not necessarily represent the decisions, policy or views of the International Agency for Research on Cancer/World Health Organization. EPICOLON: This work was supported by grants from Fondo de Investigación Sanitaria/FEDER (PI08/0024, PI08/1276, PS09/02368, P111/00219, PI11/00681, PI14/00173, PI14/00230, PI17/00509, 17/00878, Acción Transversal de Cáncer), Xunta de Galicia (PGIDIT07PXIB9101209PR), Ministerio de Economia y Competitividad (SAF07-64873, SAF 2010-19273, SAF2014-54453R), Fundación Científica de la Asociación Española contra el Cáncer (GCB13131592CAST), Beca Grupo de Trabajo “Oncología” AEG (Asociación Española de Gastroenterología), Fundación Privada Olga Torres, FP7 CHIBCHA Consortium, Agència de Gestió d’Ajuts Universitaris i de Recerca (Generalitat de Catalunya, 2014SGR135, 2014SGR255, 2017SGR21, 2017SGR653), Catalan Tumour Bank Network (Pla Director d’Oncologia, Generalitat de Catalunya), PERIS (SLT002/16/00398, Generalitat de Catalunya), CERCA Programme (Generalitat de Catalunya) and COST Action BM1206. CIBERehd is funded by the Instituto de Salud Carlos III. ESTHER/VERDI. This work was supported by grants from the Baden-Württemberg Ministry of Science, Research and Arts and the German Cancer Aid. Harvard cohorts (HPFS, NHS, PHS): HPFS is supported by the NIH (P01 CA055075, UM1 CA167552, U01 CA167552, R01 CA137178, R01 CA151993, R35 CA197735, K07 CA190673, and P50 CA127003), NHS by the NIH (R01 CA137178, P01 CA087969, UM1 CA186107, R01 CA151993, R35 CA197735, K07 CA190673, and P50 CA127003) and PHS by the NIH (R01 CA042182). Hawaii Adenoma Study: NCI grants R01 CA72520. Hwasun Cancer Epidemiology Study–Colon and Rectum Cancer: grants from Chonnam National University Hwasun Hospital (HCRI15011-1), and the NIH (R01 CA188214). Kentucky: This work was supported by the following grant support: Clinical Investigator Award from Damon Runyon Cancer Research Foundation (CI-8); NCI R01CA136726. The Leeds Colorectal Cancer Study was funded by the Food Standards Agency and Cancer Research UK Programme Award (C588/A19167). Melbourne Collaborative Cohort Study (MCCS) cohort recruitment was funded by VicHealth and Cancer Council Victoria. The MCCS was further supported by Australian National Health and Medical Research Council grants 209057, 396414, and 1074383, and by infrastructure provided by Cancer Council Victoria. Cases and their vital statuses were ascertained through the Victorian Cancer Registry and the Australian Institute of Health and Welfare, including the National Death Index and the Australian Cancer Database. MEC: NIH (R37 CA54281, P01 CA033619, R01 CA063464, and U01 CA164973). MECC: This work was supported by the NIH, US Department of Health and Human Services (R01 CA81488 to SBG, R01 CA197350 to SBG, U19 CA148107 to SBG, N01 CN043302 assigned to SBG, 5P01 CA196569 to WG, P30 CA014089 to SBG, R01 CA144040 to SDM, and P50 CA150964 to SDM), as well as funding from the Ravitz Foundation, the Irving Weinstein Foundation, the Anton B. Burg Foundation, the Jane and Kris Popovich Chair in Cancer Research, and a generous gift from Daniel and Maryann Fong. Memorial Sloan Kettering Cancer Center: The work at Memorial Sloan Kettering Cancer Center in New York was supported by the Robert and Kate Niehaus Center for Inherited Cancer Genomics and the Romeo Milio Foundation. It is also supported by the NCI-designated Comprehensive Cancer Center (grant number P30 CA008748). NCCCS I & II: We acknowledge funding support for this project from the NIH, R01 CA66635 and P30 DK034987. NFCCR: This work was supported by an Interdisciplinary Health Research Team award from the Canadian Institutes of Health Research (CRT 43821); the NIH, US Department of Health and Human Serivces (U01 CA74783); and NCI of Canada grants (18223 and 18226). Funding was provided to Michael O. Woods by the Canadian Cancer Society Research Institute. Northern Sweden Health and Disease Study (NSHDS) investigators thank the Biobank Research Unit at Umeå University, the Västerbotten Intervention Programme, the Northern Sweden MONICA study and Region Västerbotten for providing data and samples and acknowledge the contribution from Biobank Sweden, supported by the Swedish Research Council (VR 2017-00650). This research was also supported by funding to BVG from the Swedish Cancer Society (CAN 2017/581); the Swedish Research Council (VR 2017-01737); Region Västerbotten (VLL-841671, VLL-833291); the Lion’s Cancer Research Foundation (several grants), the Faculty of Medicine and Insamlingsstiftelsen at Umeå University; and the Margareta Dannborg Memorial Fund. Ontario Registry for Studies of Familial Colorectal Cancer (OFCCR): NIH, through funding allocated to the OFCCR (U01 CA074783); see Colon Cancer Family Registry section above. Additional funding toward genetic analyses of OFCCR includes the Ontario Research Fund, the Canadian Institutes of Health Research, and the Ontario Institute for Cancer Research, through generous support from the Ontario Ministry of Research and Innovation. Ohio State University Wexner Medical Center: Ohio Colorectal Cancer Prevention Initiative funding was provided by Pelotonia and HNPCC funding was provided by the NCI (CA16058 and CA67941). PLCO: Intramural Research Program of the Division of Cancer Epidemiology and Genetics and supported by contracts from the Division of Cancer Prevention, NCI, NIH, Department of Health and Human Services. Funding was provided by NIH, Genes, Environment and Health Initiative (GEI) Z01 CP 010200, NIH U01 HG004446, and NIH GEI U01 HG 004438. PMH: NIH (R01 CA076366 to P.A. Newcomb). SEARCH: The University of Cambridge has received salary support in respect of PDPP from the NHS in the East of England through the Clinical Academic Reserve. Cancer Research UK (C490/A16561); the UK National Institute for Health Research Biomedical Research Centres at the University of Cambridge. The Selenium and Vitamin E Cancer Prevention Trial (SELECT) was supported by the NCI of the NIH under award numbers UM1CA182883 and U10CA37429. SMS: This work was supported by the NCI (grant P01 CA074184 to J.D.P. and P.A.N., grants R01 CA097325, R03 CA153323, and K05 CA152715 to P.A.N., and the National Center for Advancing Translational Sciences at the NIH (grant KL2 TR000421 to A.N.B.-H.) The Swedish Low-risk Colorectal Cancer Study: The study was supported by grants from the Swedish research council; K2015-55X-22674-01-4, K2008-55X-20157-03-3, K2006-72X-20157-01-2 and the Stockholm County Council (ALF project). Swedish Mammography Cohort and Cohort of Swedish Men: This work is supported by grants from the Swedish Cancer Foundation, the Swedish Research Council for the Swedish Infrastructure for Medical Population-based Life-course Environmental Research (SIMPLER) and Karolinska Institute´s Distinguished Professor Award to Alicja Wolk. VITAL: NIH (K05 CA154337). Women’s Health Initiative (WHI): The WHI program is funded by the National Heart, Lung, and Blood Institute, NIH, US Department of Health and Human Services through contracts HHSN268201100046C, HHSN268201100001C, HHSN268201100002C, HHSN268201100003C, HHSN268201100004C, and HHSN271201100004C.

    ∗

    Authors share co-first authorship.

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