Serveur d'exploration sur les relations entre la France et l'Australie

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Fine-Mapping of Common Genetic Variants Associated with Colorectal Tumor Risk Identified Potential Functional Variants

Identifieur interne : 000622 ( Pmc/Corpus ); précédent : 000621; suivant : 000623

Fine-Mapping of Common Genetic Variants Associated with Colorectal Tumor Risk Identified Potential Functional Variants

Auteurs : Mengmeng Du ; Shuo Jiao ; Stephanie A. Bien ; Manish Gala ; Goncalo Abecasis ; Stephane Bezieau ; Hermann Brenner ; Katja Butterbach ; Bette J. Caan ; Christopher S. Carlson ; Graham Casey ; Jenny Chang-Claude ; David V. Conti ; Keith R. Curtis ; David Duggan ; Steven Gallinger ; Robert W. Haile ; Tabitha A. Harrison ; Richard B. Hayes ; Michael Hoffmeister ; John L. Hopper ; Thomas J. Hudson ; Mark A. Jenkins ; Sébastien Küry ; Loic Le Marchand ; Suzanne M. Leal ; Polly A. Newcomb ; Deborah A. Nickerson ; John D. Potter ; Robert E. Schoen ; Fredrick R. Schumacher ; Daniela Seminara ; Martha L. Slattery ; Li Hsu ; Andrew T. Chan ; Emily White ; Sonja I. Berndt ; Ulrike Peters

Source :

RBID : PMC:4933364

Abstract

Genome-wide association studies (GWAS) have identified many common single nucleotide polymorphisms (SNPs) associated with colorectal cancer risk. These SNPs may tag correlated variants with biological importance. Fine-mapping around GWAS loci can facilitate detection of functional candidates and additional independent risk variants. We analyzed 11,900 cases and 14,311 controls in the Genetics and Epidemiology of Colorectal Cancer Consortium and the Colon Cancer Family Registry. To fine-map genomic regions containing all known common risk variants, we imputed high-density genetic data from the 1000 Genomes Project. We tested single-variant associations with colorectal tumor risk for all variants spanning genomic regions 250-kb upstream or downstream of 31 GWAS-identified SNPs (index SNPs). We queried the University of California, Santa Cruz Genome Browser to examine evidence for biological function. Index SNPs did not show the strongest association signals with colorectal tumor risk in their respective genomic regions. Bioinformatics analysis of SNPs showing smaller P-values in each region revealed 21 functional candidates in 12 loci (5q31.1, 8q24, 11q13.4, 11q23, 12p13.32, 12q24.21, 14q22.2, 15q13, 18q21, 19q13.1, 20p12.3, and 20q13.33). We did not observe evidence of additional independent association signals in GWAS-identified regions. Our results support the utility of integrating data from comprehensive fine-mapping with expanding publicly available genomic databases to help clarify GWAS associations and identify functional candidates that warrant more onerous laboratory follow-up. Such efforts may aid the eventual discovery of disease-causing variant(s).


Url:
DOI: 10.1371/journal.pone.0157521
PubMed: 27379672
PubMed Central: 4933364

Links to Exploration step

PMC:4933364

Le document en format XML

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<name sortKey="Jenkins, Mark A" sort="Jenkins, Mark A" uniqKey="Jenkins M" first="Mark A." last="Jenkins">Mark A. Jenkins</name>
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<name sortKey="Kury, Sebastien" sort="Kury, Sebastien" uniqKey="Kury S" first="Sébastien" last="Küry">Sébastien Küry</name>
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<name sortKey="Le Marchand, Loic" sort="Le Marchand, Loic" uniqKey="Le Marchand L" first="Loic" last="Le Marchand">Loic Le Marchand</name>
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<name sortKey="Leal, Suzanne M" sort="Leal, Suzanne M" uniqKey="Leal S" first="Suzanne M." last="Leal">Suzanne M. Leal</name>
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<nlm:aff id="aff019">
<addr-line>Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX, United States of America</addr-line>
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<name sortKey="Newcomb, Polly A" sort="Newcomb, Polly A" uniqKey="Newcomb P" first="Polly A." last="Newcomb">Polly A. Newcomb</name>
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<name sortKey="Schoen, Robert E" sort="Schoen, Robert E" uniqKey="Schoen R" first="Robert E." last="Schoen">Robert E. Schoen</name>
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<name sortKey="Schumacher, Fredrick R" sort="Schumacher, Fredrick R" uniqKey="Schumacher F" first="Fredrick R." last="Schumacher">Fredrick R. Schumacher</name>
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<name sortKey="Seminara, Daniela" sort="Seminara, Daniela" uniqKey="Seminara D" first="Daniela" last="Seminara">Daniela Seminara</name>
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<addr-line>Division of Cancer Control and Population Sciences, National Cancer Institute, Bethesda, MD, United States of America</addr-line>
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<name sortKey="Slattery, Martha L" sort="Slattery, Martha L" uniqKey="Slattery M" first="Martha L." last="Slattery">Martha L. Slattery</name>
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<addr-line>Department of Internal Medicine, University of Utah Health Sciences Center, Salt Lake City, UT, United States of America</addr-line>
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</affiliation>
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<name sortKey="Hsu, Li" sort="Hsu, Li" uniqKey="Hsu L" first="Li" last="Hsu">Li Hsu</name>
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<addr-line>Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, United States of America</addr-line>
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<name sortKey="Chan, Andrew T" sort="Chan, Andrew T" uniqKey="Chan A" first="Andrew T." last="Chan">Andrew T. Chan</name>
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<addr-line>Channing Division of Network Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, MA, United States of America</addr-line>
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<name sortKey="White, Emily" sort="White, Emily" uniqKey="White E" first="Emily" last="White">Emily White</name>
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<addr-line>Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, United States of America</addr-line>
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</affiliation>
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<nlm:aff id="aff003">
<addr-line>School of Public Health, University of Washington, Seattle, WA, United States of America</addr-line>
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<name sortKey="Berndt, Sonja I" sort="Berndt, Sonja I" uniqKey="Berndt S" first="Sonja I." last="Berndt">Sonja I. Berndt</name>
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<addr-line>Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda, MD, United States of America</addr-line>
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<name sortKey="Peters, Ulrike" sort="Peters, Ulrike" uniqKey="Peters U" first="Ulrike" last="Peters">Ulrike Peters</name>
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<addr-line>Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, United States of America</addr-line>
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<addr-line>School of Public Health, University of Washington, Seattle, WA, United States of America</addr-line>
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<title xml:lang="en" level="a" type="main">Fine-Mapping of Common Genetic Variants Associated with Colorectal Tumor Risk Identified Potential Functional Variants</title>
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<addr-line>Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, United States of America</addr-line>
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<name sortKey="Jiao, Shuo" sort="Jiao, Shuo" uniqKey="Jiao S" first="Shuo" last="Jiao">Shuo Jiao</name>
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<name sortKey="Bien, Stephanie A" sort="Bien, Stephanie A" uniqKey="Bien S" first="Stephanie A." last="Bien">Stephanie A. Bien</name>
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<addr-line>School of Public Health, University of Washington, Seattle, WA, United States of America</addr-line>
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<name sortKey="Gala, Manish" sort="Gala, Manish" uniqKey="Gala M" first="Manish" last="Gala">Manish Gala</name>
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<addr-line>Division of Gastroenterology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, United States of America</addr-line>
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<name sortKey="Abecasis, Goncalo" sort="Abecasis, Goncalo" uniqKey="Abecasis G" first="Goncalo" last="Abecasis">Goncalo Abecasis</name>
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<addr-line>Department of Biostatistics, University of Michigan School of Public Health, Ann Arbor, MI, United States of America</addr-line>
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<author>
<name sortKey="Bezieau, Stephane" sort="Bezieau, Stephane" uniqKey="Bezieau S" first="Stephane" last="Bezieau">Stephane Bezieau</name>
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<name sortKey="Brenner, Hermann" sort="Brenner, Hermann" uniqKey="Brenner H" first="Hermann" last="Brenner">Hermann Brenner</name>
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</nlm:aff>
</affiliation>
<affiliation>
<nlm:aff id="aff008">
<addr-line>German Cancer Consortium (DKTK), Heidelberg, Germany</addr-line>
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<name sortKey="Butterbach, Katja" sort="Butterbach, Katja" uniqKey="Butterbach K" first="Katja" last="Butterbach">Katja Butterbach</name>
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<nlm:aff id="aff007">
<addr-line>Division of Clinical Epidemiology and Aging Research, German Cancer Research Center (DKFZ), Heidelberg, Germany</addr-line>
</nlm:aff>
</affiliation>
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<name sortKey="Caan, Bette J" sort="Caan, Bette J" uniqKey="Caan B" first="Bette J." last="Caan">Bette J. Caan</name>
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<nlm:aff id="aff009">
<addr-line>Division of Research, Kaiser Permanente Medical Care Program of Northern California, Oakland, CA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
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<name sortKey="Carlson, Christopher S" sort="Carlson, Christopher S" uniqKey="Carlson C" first="Christopher S." last="Carlson">Christopher S. Carlson</name>
<affiliation>
<nlm:aff id="aff002">
<addr-line>Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Casey, Graham" sort="Casey, Graham" uniqKey="Casey G" first="Graham" last="Casey">Graham Casey</name>
<affiliation>
<nlm:aff id="aff010">
<addr-line>Keck School of Medicine, University of Southern California, Los Angeles, CA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Chang Claude, Jenny" sort="Chang Claude, Jenny" uniqKey="Chang Claude J" first="Jenny" last="Chang-Claude">Jenny Chang-Claude</name>
<affiliation>
<nlm:aff id="aff011">
<addr-line>Division of Cancer Epidemiology, German Cancer Research Center (DKFZ), Heidelberg, Germany</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Conti, David V" sort="Conti, David V" uniqKey="Conti D" first="David V." last="Conti">David V. Conti</name>
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<nlm:aff id="aff010">
<addr-line>Keck School of Medicine, University of Southern California, Los Angeles, CA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Curtis, Keith R" sort="Curtis, Keith R" uniqKey="Curtis K" first="Keith R." last="Curtis">Keith R. Curtis</name>
<affiliation>
<nlm:aff id="aff002">
<addr-line>Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Duggan, David" sort="Duggan, David" uniqKey="Duggan D" first="David" last="Duggan">David Duggan</name>
<affiliation>
<nlm:aff id="aff012">
<addr-line>Translational Genomics Research Institute, Phoenix, Arizona, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Gallinger, Steven" sort="Gallinger, Steven" uniqKey="Gallinger S" first="Steven" last="Gallinger">Steven Gallinger</name>
<affiliation>
<nlm:aff id="aff013">
<addr-line>Department of Surgery, Mount Sinai Hospital, Toronto, ON, Canada</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Haile, Robert W" sort="Haile, Robert W" uniqKey="Haile R" first="Robert W." last="Haile">Robert W. Haile</name>
<affiliation>
<nlm:aff id="aff010">
<addr-line>Keck School of Medicine, University of Southern California, Los Angeles, CA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Harrison, Tabitha A" sort="Harrison, Tabitha A" uniqKey="Harrison T" first="Tabitha A." last="Harrison">Tabitha A. Harrison</name>
<affiliation>
<nlm:aff id="aff002">
<addr-line>Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Hayes, Richard B" sort="Hayes, Richard B" uniqKey="Hayes R" first="Richard B." last="Hayes">Richard B. Hayes</name>
<affiliation>
<nlm:aff id="aff014">
<addr-line>Division of Epidemiology, Department of Population Health, New York University School of Medicine, New York, NY, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Hoffmeister, Michael" sort="Hoffmeister, Michael" uniqKey="Hoffmeister M" first="Michael" last="Hoffmeister">Michael Hoffmeister</name>
<affiliation>
<nlm:aff id="aff007">
<addr-line>Division of Clinical Epidemiology and Aging Research, German Cancer Research Center (DKFZ), Heidelberg, Germany</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Hopper, John L" sort="Hopper, John L" uniqKey="Hopper J" first="John L." last="Hopper">John L. Hopper</name>
<affiliation>
<nlm:aff id="aff015">
<addr-line>Melbourne School of Population and Global Health, The University of Melbourne, Melbourne, Victoria, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Hudson, Thomas J" sort="Hudson, Thomas J" uniqKey="Hudson T" first="Thomas J." last="Hudson">Thomas J. Hudson</name>
<affiliation>
<nlm:aff id="aff016">
<addr-line>Ontario Institute for Cancer Research, Toronto, ON, Canada</addr-line>
</nlm:aff>
</affiliation>
<affiliation>
<nlm:aff id="aff017">
<addr-line>Departments of Medical Biophysics and Molecular Genetics, University of Toronto, Toronto, ON, Canada</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Jenkins, Mark A" sort="Jenkins, Mark A" uniqKey="Jenkins M" first="Mark A." last="Jenkins">Mark A. Jenkins</name>
<affiliation>
<nlm:aff id="aff015">
<addr-line>Melbourne School of Population and Global Health, The University of Melbourne, Melbourne, Victoria, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Kury, Sebastien" sort="Kury, Sebastien" uniqKey="Kury S" first="Sébastien" last="Küry">Sébastien Küry</name>
<affiliation>
<nlm:aff id="aff006">
<addr-line>Service de Génétique Médicale, CHU Nantes, Nantes, France</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Le Marchand, Loic" sort="Le Marchand, Loic" uniqKey="Le Marchand L" first="Loic" last="Le Marchand">Loic Le Marchand</name>
<affiliation>
<nlm:aff id="aff018">
<addr-line>Epidemiology Program, University of Hawaii Cancer Center, Honolulu, HI, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Leal, Suzanne M" sort="Leal, Suzanne M" uniqKey="Leal S" first="Suzanne M." last="Leal">Suzanne M. Leal</name>
<affiliation>
<nlm:aff id="aff019">
<addr-line>Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Newcomb, Polly A" sort="Newcomb, Polly A" uniqKey="Newcomb P" first="Polly A." last="Newcomb">Polly A. Newcomb</name>
<affiliation>
<nlm:aff id="aff002">
<addr-line>Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, United States of America</addr-line>
</nlm:aff>
</affiliation>
<affiliation>
<nlm:aff id="aff003">
<addr-line>School of Public Health, University of Washington, Seattle, WA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Nickerson, Deborah A" sort="Nickerson, Deborah A" uniqKey="Nickerson D" first="Deborah A." last="Nickerson">Deborah A. Nickerson</name>
<affiliation>
<nlm:aff id="aff020">
<addr-line>Genome Sciences, University of Washington, Seattle, WA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Potter, John D" sort="Potter, John D" uniqKey="Potter J" first="John D." last="Potter">John D. Potter</name>
<affiliation>
<nlm:aff id="aff002">
<addr-line>Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, United States of America</addr-line>
</nlm:aff>
</affiliation>
<affiliation>
<nlm:aff id="aff003">
<addr-line>School of Public Health, University of Washington, Seattle, WA, United States of America</addr-line>
</nlm:aff>
</affiliation>
<affiliation>
<nlm:aff id="aff021">
<addr-line>Centre for Public Health Research, Massey University, Wellington, New Zealand</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Schoen, Robert E" sort="Schoen, Robert E" uniqKey="Schoen R" first="Robert E." last="Schoen">Robert E. Schoen</name>
<affiliation>
<nlm:aff id="aff022">
<addr-line>Department of Medicine and Epidemiology, University of Pittsburgh Medical Center, Pittsburgh, PA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Schumacher, Fredrick R" sort="Schumacher, Fredrick R" uniqKey="Schumacher F" first="Fredrick R." last="Schumacher">Fredrick R. Schumacher</name>
<affiliation>
<nlm:aff id="aff010">
<addr-line>Keck School of Medicine, University of Southern California, Los Angeles, CA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Seminara, Daniela" sort="Seminara, Daniela" uniqKey="Seminara D" first="Daniela" last="Seminara">Daniela Seminara</name>
<affiliation>
<nlm:aff id="aff023">
<addr-line>Division of Cancer Control and Population Sciences, National Cancer Institute, Bethesda, MD, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Slattery, Martha L" sort="Slattery, Martha L" uniqKey="Slattery M" first="Martha L." last="Slattery">Martha L. Slattery</name>
<affiliation>
<nlm:aff id="aff024">
<addr-line>Department of Internal Medicine, University of Utah Health Sciences Center, Salt Lake City, UT, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Hsu, Li" sort="Hsu, Li" uniqKey="Hsu L" first="Li" last="Hsu">Li Hsu</name>
<affiliation>
<nlm:aff id="aff002">
<addr-line>Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Chan, Andrew T" sort="Chan, Andrew T" uniqKey="Chan A" first="Andrew T." last="Chan">Andrew T. Chan</name>
<affiliation>
<nlm:aff id="aff004">
<addr-line>Division of Gastroenterology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, United States of America</addr-line>
</nlm:aff>
</affiliation>
<affiliation>
<nlm:aff id="aff025">
<addr-line>Channing Division of Network Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, MA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="White, Emily" sort="White, Emily" uniqKey="White E" first="Emily" last="White">Emily White</name>
<affiliation>
<nlm:aff id="aff002">
<addr-line>Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, United States of America</addr-line>
</nlm:aff>
</affiliation>
<affiliation>
<nlm:aff id="aff003">
<addr-line>School of Public Health, University of Washington, Seattle, WA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Berndt, Sonja I" sort="Berndt, Sonja I" uniqKey="Berndt S" first="Sonja I." last="Berndt">Sonja I. Berndt</name>
<affiliation>
<nlm:aff id="aff026">
<addr-line>Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda, MD, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Peters, Ulrike" sort="Peters, Ulrike" uniqKey="Peters U" first="Ulrike" last="Peters">Ulrike Peters</name>
<affiliation>
<nlm:aff id="aff002">
<addr-line>Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, United States of America</addr-line>
</nlm:aff>
</affiliation>
<affiliation>
<nlm:aff id="aff003">
<addr-line>School of Public Health, University of Washington, Seattle, WA, United States of America</addr-line>
</nlm:aff>
</affiliation>
</author>
</analytic>
<series>
<title level="j">PLoS ONE</title>
<idno type="eISSN">1932-6203</idno>
<imprint>
<date when="2016">2016</date>
</imprint>
</series>
</biblStruct>
</sourceDesc>
</fileDesc>
<profileDesc>
<textClass></textClass>
</profileDesc>
</teiHeader>
<front>
<div type="abstract" xml:lang="en">
<p>Genome-wide association studies (GWAS) have identified many common single nucleotide polymorphisms (SNPs) associated with colorectal cancer risk. These SNPs may tag correlated variants with biological importance. Fine-mapping around GWAS loci can facilitate detection of functional candidates and additional independent risk variants. We analyzed 11,900 cases and 14,311 controls in the Genetics and Epidemiology of Colorectal Cancer Consortium and the Colon Cancer Family Registry. To fine-map genomic regions containing all known common risk variants, we imputed high-density genetic data from the 1000 Genomes Project. We tested single-variant associations with colorectal tumor risk for all variants spanning genomic regions 250-kb upstream or downstream of 31 GWAS-identified SNPs (index SNPs). We queried the University of California, Santa Cruz Genome Browser to examine evidence for biological function. Index SNPs did not show the strongest association signals with colorectal tumor risk in their respective genomic regions. Bioinformatics analysis of SNPs showing smaller
<italic>P</italic>
-values in each region revealed 21 functional candidates in 12 loci (5q31.1, 8q24, 11q13.4, 11q23, 12p13.32, 12q24.21, 14q22.2, 15q13, 18q21, 19q13.1, 20p12.3, and 20q13.33). We did not observe evidence of additional independent association signals in GWAS-identified regions. Our results support the utility of integrating data from comprehensive fine-mapping with expanding publicly available genomic databases to help clarify GWAS associations and identify functional candidates that warrant more onerous laboratory follow-up. Such efforts may aid the eventual discovery of disease-causing variant(s).</p>
</div>
</front>
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<article-title>Fine-Mapping of Common Genetic Variants Associated with Colorectal Tumor Risk Identified Potential Functional Variants</article-title>
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</title-group>
<contrib-group>
<contrib contrib-type="author" equal-contrib="yes">
<name>
<surname>Du</surname>
<given-names>Mengmeng</given-names>
</name>
<xref ref-type="aff" rid="aff001">
<sup>1</sup>
</xref>
<xref ref-type="aff" rid="aff002">
<sup>2</sup>
</xref>
<xref ref-type="corresp" rid="cor001">*</xref>
</contrib>
<contrib contrib-type="author" equal-contrib="yes">
<name>
<surname>Jiao</surname>
<given-names>Shuo</given-names>
</name>
<xref ref-type="aff" rid="aff002">
<sup>2</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<contrib-id contrib-id-type="orcid" authenticated="false">http://orcid.org/0000-0002-8713-4583</contrib-id>
<name>
<surname>Bien</surname>
<given-names>Stephanie A.</given-names>
</name>
<xref ref-type="aff" rid="aff002">
<sup>2</sup>
</xref>
<xref ref-type="aff" rid="aff003">
<sup>3</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Gala</surname>
<given-names>Manish</given-names>
</name>
<xref ref-type="aff" rid="aff004">
<sup>4</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Abecasis</surname>
<given-names>Goncalo</given-names>
</name>
<xref ref-type="aff" rid="aff005">
<sup>5</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Bezieau</surname>
<given-names>Stephane</given-names>
</name>
<xref ref-type="aff" rid="aff006">
<sup>6</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Brenner</surname>
<given-names>Hermann</given-names>
</name>
<xref ref-type="aff" rid="aff007">
<sup>7</sup>
</xref>
<xref ref-type="aff" rid="aff008">
<sup>8</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Butterbach</surname>
<given-names>Katja</given-names>
</name>
<xref ref-type="aff" rid="aff007">
<sup>7</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Caan</surname>
<given-names>Bette J.</given-names>
</name>
<xref ref-type="aff" rid="aff009">
<sup>9</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Carlson</surname>
<given-names>Christopher S.</given-names>
</name>
<xref ref-type="aff" rid="aff002">
<sup>2</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Casey</surname>
<given-names>Graham</given-names>
</name>
<xref ref-type="aff" rid="aff010">
<sup>10</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Chang-Claude</surname>
<given-names>Jenny</given-names>
</name>
<xref ref-type="aff" rid="aff011">
<sup>11</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Conti</surname>
<given-names>David V.</given-names>
</name>
<xref ref-type="aff" rid="aff010">
<sup>10</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Curtis</surname>
<given-names>Keith R.</given-names>
</name>
<xref ref-type="aff" rid="aff002">
<sup>2</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Duggan</surname>
<given-names>David</given-names>
</name>
<xref ref-type="aff" rid="aff012">
<sup>12</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Gallinger</surname>
<given-names>Steven</given-names>
</name>
<xref ref-type="aff" rid="aff013">
<sup>13</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Haile</surname>
<given-names>Robert W.</given-names>
</name>
<xref ref-type="aff" rid="aff010">
<sup>10</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Harrison</surname>
<given-names>Tabitha A.</given-names>
</name>
<xref ref-type="aff" rid="aff002">
<sup>2</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Hayes</surname>
<given-names>Richard B.</given-names>
</name>
<xref ref-type="aff" rid="aff014">
<sup>14</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Hoffmeister</surname>
<given-names>Michael</given-names>
</name>
<xref ref-type="aff" rid="aff007">
<sup>7</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Hopper</surname>
<given-names>John L.</given-names>
</name>
<xref ref-type="aff" rid="aff015">
<sup>15</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Hudson</surname>
<given-names>Thomas J.</given-names>
</name>
<xref ref-type="aff" rid="aff016">
<sup>16</sup>
</xref>
<xref ref-type="aff" rid="aff017">
<sup>17</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Jenkins</surname>
<given-names>Mark A.</given-names>
</name>
<xref ref-type="aff" rid="aff015">
<sup>15</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Küry</surname>
<given-names>Sébastien</given-names>
</name>
<xref ref-type="aff" rid="aff006">
<sup>6</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Le Marchand</surname>
<given-names>Loic</given-names>
</name>
<xref ref-type="aff" rid="aff018">
<sup>18</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Leal</surname>
<given-names>Suzanne M.</given-names>
</name>
<xref ref-type="aff" rid="aff019">
<sup>19</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Newcomb</surname>
<given-names>Polly A.</given-names>
</name>
<xref ref-type="aff" rid="aff002">
<sup>2</sup>
</xref>
<xref ref-type="aff" rid="aff003">
<sup>3</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Nickerson</surname>
<given-names>Deborah A.</given-names>
</name>
<xref ref-type="aff" rid="aff020">
<sup>20</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Potter</surname>
<given-names>John D.</given-names>
</name>
<xref ref-type="aff" rid="aff002">
<sup>2</sup>
</xref>
<xref ref-type="aff" rid="aff003">
<sup>3</sup>
</xref>
<xref ref-type="aff" rid="aff021">
<sup>21</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Schoen</surname>
<given-names>Robert E.</given-names>
</name>
<xref ref-type="aff" rid="aff022">
<sup>22</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Schumacher</surname>
<given-names>Fredrick R.</given-names>
</name>
<xref ref-type="aff" rid="aff010">
<sup>10</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Seminara</surname>
<given-names>Daniela</given-names>
</name>
<xref ref-type="aff" rid="aff023">
<sup>23</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Slattery</surname>
<given-names>Martha L.</given-names>
</name>
<xref ref-type="aff" rid="aff024">
<sup>24</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Hsu</surname>
<given-names>Li</given-names>
</name>
<xref ref-type="aff" rid="aff002">
<sup>2</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Chan</surname>
<given-names>Andrew T.</given-names>
</name>
<xref ref-type="aff" rid="aff004">
<sup>4</sup>
</xref>
<xref ref-type="aff" rid="aff025">
<sup>25</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>White</surname>
<given-names>Emily</given-names>
</name>
<xref ref-type="aff" rid="aff002">
<sup>2</sup>
</xref>
<xref ref-type="aff" rid="aff003">
<sup>3</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Berndt</surname>
<given-names>Sonja I.</given-names>
</name>
<xref ref-type="aff" rid="aff026">
<sup>26</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Peters</surname>
<given-names>Ulrike</given-names>
</name>
<xref ref-type="aff" rid="aff002">
<sup>2</sup>
</xref>
<xref ref-type="aff" rid="aff003">
<sup>3</sup>
</xref>
<xref ref-type="corresp" rid="cor001">*</xref>
</contrib>
</contrib-group>
<aff id="aff001">
<label>1</label>
<addr-line>Department of Epidemiology and Biostatistics, Memorial Sloan Kettering Cancer Center, New York, NY, United States of America</addr-line>
</aff>
<aff id="aff002">
<label>2</label>
<addr-line>Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, United States of America</addr-line>
</aff>
<aff id="aff003">
<label>3</label>
<addr-line>School of Public Health, University of Washington, Seattle, WA, United States of America</addr-line>
</aff>
<aff id="aff004">
<label>4</label>
<addr-line>Division of Gastroenterology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, United States of America</addr-line>
</aff>
<aff id="aff005">
<label>5</label>
<addr-line>Department of Biostatistics, University of Michigan School of Public Health, Ann Arbor, MI, United States of America</addr-line>
</aff>
<aff id="aff006">
<label>6</label>
<addr-line>Service de Génétique Médicale, CHU Nantes, Nantes, France</addr-line>
</aff>
<aff id="aff007">
<label>7</label>
<addr-line>Division of Clinical Epidemiology and Aging Research, German Cancer Research Center (DKFZ), Heidelberg, Germany</addr-line>
</aff>
<aff id="aff008">
<label>8</label>
<addr-line>German Cancer Consortium (DKTK), Heidelberg, Germany</addr-line>
</aff>
<aff id="aff009">
<label>9</label>
<addr-line>Division of Research, Kaiser Permanente Medical Care Program of Northern California, Oakland, CA, United States of America</addr-line>
</aff>
<aff id="aff010">
<label>10</label>
<addr-line>Keck School of Medicine, University of Southern California, Los Angeles, CA, United States of America</addr-line>
</aff>
<aff id="aff011">
<label>11</label>
<addr-line>Division of Cancer Epidemiology, German Cancer Research Center (DKFZ), Heidelberg, Germany</addr-line>
</aff>
<aff id="aff012">
<label>12</label>
<addr-line>Translational Genomics Research Institute, Phoenix, Arizona, United States of America</addr-line>
</aff>
<aff id="aff013">
<label>13</label>
<addr-line>Department of Surgery, Mount Sinai Hospital, Toronto, ON, Canada</addr-line>
</aff>
<aff id="aff014">
<label>14</label>
<addr-line>Division of Epidemiology, Department of Population Health, New York University School of Medicine, New York, NY, United States of America</addr-line>
</aff>
<aff id="aff015">
<label>15</label>
<addr-line>Melbourne School of Population and Global Health, The University of Melbourne, Melbourne, Victoria, Australia</addr-line>
</aff>
<aff id="aff016">
<label>16</label>
<addr-line>Ontario Institute for Cancer Research, Toronto, ON, Canada</addr-line>
</aff>
<aff id="aff017">
<label>17</label>
<addr-line>Departments of Medical Biophysics and Molecular Genetics, University of Toronto, Toronto, ON, Canada</addr-line>
</aff>
<aff id="aff018">
<label>18</label>
<addr-line>Epidemiology Program, University of Hawaii Cancer Center, Honolulu, HI, United States of America</addr-line>
</aff>
<aff id="aff019">
<label>19</label>
<addr-line>Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX, United States of America</addr-line>
</aff>
<aff id="aff020">
<label>20</label>
<addr-line>Genome Sciences, University of Washington, Seattle, WA, United States of America</addr-line>
</aff>
<aff id="aff021">
<label>21</label>
<addr-line>Centre for Public Health Research, Massey University, Wellington, New Zealand</addr-line>
</aff>
<aff id="aff022">
<label>22</label>
<addr-line>Department of Medicine and Epidemiology, University of Pittsburgh Medical Center, Pittsburgh, PA, United States of America</addr-line>
</aff>
<aff id="aff023">
<label>23</label>
<addr-line>Division of Cancer Control and Population Sciences, National Cancer Institute, Bethesda, MD, United States of America</addr-line>
</aff>
<aff id="aff024">
<label>24</label>
<addr-line>Department of Internal Medicine, University of Utah Health Sciences Center, Salt Lake City, UT, United States of America</addr-line>
</aff>
<aff id="aff025">
<label>25</label>
<addr-line>Channing Division of Network Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, MA, United States of America</addr-line>
</aff>
<aff id="aff026">
<label>26</label>
<addr-line>Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda, MD, United States of America</addr-line>
</aff>
<contrib-group>
<contrib contrib-type="editor">
<name>
<surname>Wang</surname>
<given-names>Junwen</given-names>
</name>
<role>Editor</role>
<xref ref-type="aff" rid="edit1"></xref>
</contrib>
</contrib-group>
<aff id="edit1">
<addr-line>Mayo Clinic Arizona, UNITED STATES</addr-line>
</aff>
<author-notes>
<fn fn-type="conflict" id="coi001">
<p>
<bold>Competing Interests: </bold>
The authors have declared that no competing interests exist.</p>
</fn>
<fn fn-type="con" id="contrib001">
<p>Conceived and designed the experiments: MD SJ SAB GA SB HB BJC CSC GC JCC DD SG RWH TAH RBH MH JLH TJH MAJ SK LLM SML PAN DAN JDP RES FRS DS MLS LH ATC EW SIB UP. Performed the experiments: DD. Analyzed the data: SJ MD SAB LH UP. Contributed reagents/materials/analysis tools: SJ KRC. Wrote the paper: MD SJ SAB UP. Collected phenotype data and biological samples and contributed these as investigators for their respective study: HB BJC GC JCC SG RWH RBH MH JLH MAJ LLM PAN JDP RES DS MLS ATC EW SIB. Critically reviewed the manuscript drafts and approved the final manuscript: MD SJ SAB MG GA SB HB KB BJC CSC GC JCC DVC KRC DD SG RWH TAR RBH MH JLH TJH MAJ SK LLM SML PAN DAN JDP RES FRS DS MLS LH ATC EW SIB UP</p>
</fn>
<corresp id="cor001">* E-mail:
<email>dumeng@mskcc.org</email>
(MD);
<email>upeters@fhcrc.org</email>
(UP)</corresp>
</author-notes>
<pub-date pub-type="epub">
<day>5</day>
<month>7</month>
<year>2016</year>
</pub-date>
<pub-date pub-type="collection">
<year>2016</year>
</pub-date>
<volume>11</volume>
<issue>7</issue>
<elocation-id>e0157521</elocation-id>
<history>
<date date-type="received">
<day>8</day>
<month>10</month>
<year>2015</year>
</date>
<date date-type="accepted">
<day>1</day>
<month>6</month>
<year>2016</year>
</date>
</history>
<permissions>
<copyright-statement>© 2016 Du et al</copyright-statement>
<copyright-year>2016</copyright-year>
<copyright-holder>Du et al</copyright-holder>
<license xlink:href="http://creativecommons.org/licenses/by/4.0/">
<license-p>This is an open access article distributed under the terms of the
<ext-link ext-link-type="uri" xlink:href="http://creativecommons.org/licenses/by/4.0/">Creative Commons Attribution License</ext-link>
, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.</license-p>
</license>
</permissions>
<self-uri content-type="pdf" xlink:href="pone.0157521.pdf"></self-uri>
<abstract>
<p>Genome-wide association studies (GWAS) have identified many common single nucleotide polymorphisms (SNPs) associated with colorectal cancer risk. These SNPs may tag correlated variants with biological importance. Fine-mapping around GWAS loci can facilitate detection of functional candidates and additional independent risk variants. We analyzed 11,900 cases and 14,311 controls in the Genetics and Epidemiology of Colorectal Cancer Consortium and the Colon Cancer Family Registry. To fine-map genomic regions containing all known common risk variants, we imputed high-density genetic data from the 1000 Genomes Project. We tested single-variant associations with colorectal tumor risk for all variants spanning genomic regions 250-kb upstream or downstream of 31 GWAS-identified SNPs (index SNPs). We queried the University of California, Santa Cruz Genome Browser to examine evidence for biological function. Index SNPs did not show the strongest association signals with colorectal tumor risk in their respective genomic regions. Bioinformatics analysis of SNPs showing smaller
<italic>P</italic>
-values in each region revealed 21 functional candidates in 12 loci (5q31.1, 8q24, 11q13.4, 11q23, 12p13.32, 12q24.21, 14q22.2, 15q13, 18q21, 19q13.1, 20p12.3, and 20q13.33). We did not observe evidence of additional independent association signals in GWAS-identified regions. Our results support the utility of integrating data from comprehensive fine-mapping with expanding publicly available genomic databases to help clarify GWAS associations and identify functional candidates that warrant more onerous laboratory follow-up. Such efforts may aid the eventual discovery of disease-causing variant(s).</p>
</abstract>
<funding-group>
<funding-statement>This work was supported by the following: GECCO: National Cancer Institute, National Institutes of Health, U.S. Department of Health and Human Services (U01 CA137088; R01 CA059045; U01 CA164930), M.D. is supported by grants R25 CA94880 and P30 CA008748 from the National Cancer Institute. ASTERISK: Hospital Clinical Research Program (PHRC) and supported by the Regional Council of Pays de la Loire, the Groupement des Entreprises Françaises dans la Lutte contre le Cancer (GEFLUC), the Association Anne de Bretagne Génétique and the Ligue Régionale Contre le Cancer (LRCC). COLO2&3: National Institutes of Health (R01 CA60987). CCFR: National Institutes of Health (RFA # CA-95-011) and through cooperative agreements with members of the Colon Cancer Family Registry and P.I.s. This genome wide scan was supported by the National Cancer Institute, National Institutes of Health by U01 CA122839. 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 CFRs, nor does mention of trade names, commercial products, or organizations imply endorsement by the US Government or the CFR. The following Colon CFR centers contributed data to this manuscript and were supported by National Institutes of Health: Australasian Colorectal Cancer Family Registry (U01 CA097735), Ontario Registry for Studies of Familial Colorectal Cancer (U01 CA074783), and Seattle Colorectal Cancer Family Registry (U01 CA074794). DACHS: German Research Council (Deutsche Forschungsgemeinschaft, BR 1704/6-1, BR 1704/6-3, BR 1704/6-4 and CH 117/1-1), and the German Federal Ministry of Education and Research (01KH0404 and 01ER0814). DALS: National Institutes of Health (R01 CA48998 to M.L.S.); HPFS, NHS, and PHS: HPFS by the National Institutes of Health (P01 CA 055075, UM1 CA167552, R01 137178, and P50 CA 127003), NHS by the National Institutes of Health (R01 CA137178, P01 CA 087969 and P50 CA 127003,) and PHS by the National Institutes of Health (CA42182). A.T.C. is also supported by a Damon Runyon Clinical investigator Award and K24 DK098311. MEC: National Institutes of Health (R37 CA54281, P01 CA033619, and R01 CA63464). OFCCR: National Institutes of Health, through funding allocated to the Ontario Registry for Studies of Familial Colorectal Cancer (U01 CA074783); see CCFR section above. As subset of ARCTIC, OFCCR is supported by a GL2 grant from the Ontario Research Fund, the Canadian Institutes of Health Research, and the Cancer Risk Evaluation (CaRE) Program grant from the Canadian Cancer Society Research Institute. T.J.H. is a recipient of Senior Investigator Awards from the Ontario Institute for Cancer Research, through generous support from the Ontario Ministry of Research and Innovation. PLCO: Intramural Research Program of the Division of Cancer Epidemiology and Genetics and supported by contracts from the Division of Cancer Prevention, National Cancer Institute, NIH, DHHS. Additionally, a subset of control samples were genotyped as part of the Cancer Genetic Markers of Susceptibility (CGEMS) Prostate Cancer GWAS (Yeager, M et al. Nat Genet 2007 May;39(5):645-9), Colon CGEMS pancreatic cancer scan (PanScan) (Amundadottir, L et al. Nat Genet. 2009 Sep;41(9):986-90 and Petersen, GM et al Nat Genet. 2010 Mar;42(3):224-8), and the Lung Cancer and Smoking study. The prostate and PanScan study datasets were accessed with appropriate approval through the dbGaP online resource (
<ext-link ext-link-type="uri" xlink:href="http://cgems.cancer.gov/data/">http://cgems.cancer.gov/data/</ext-link>
) accession numbers phs000207v.1p1 and phs000206.v3.p2, respectively, and the lung datasets were accessed from the dbGaP website (
<ext-link ext-link-type="uri" xlink:href="http://www.ncbi.nlm.nih.gov/gap">http://www.ncbi.nlm.nih.gov/gap</ext-link>
) through accession number phs000093 v2.p2. Funding for the Lung Cancer and Smoking study was provided by National Institutes of Health (NIH), Genes, Environment and Health Initiative (GEI) Z01 CP 010200, NIH U01 HG004446, and NIH GEI U01 HG 004438. For the lung study, the GENEVA Coordinating Center provided assistance with genotype cleaning and general study coordination, and the Johns Hopkins University Center for Inherited Disease Research conducted genotyping. PMH-CCFR: National Institutes of Health (R01 CA076366 to P.A.N.). VITAL: National Institutes of Health (K05 CA154337). WHI: The WHI program is funded by the National Heart, Lung, and Blood Institute, National Institutes of Health, U.S. Department of Health and Human Services through contracts HHSN268201100046C, HHSN268201100001C, HHSN268201100002C, HHSN268201100003C, HHSN268201100004C, and HHSN271201100004C. CORECT: National Cancer Institute, National Institutes of Health under RFA # CA-09-002 (U19 CA148107). 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 CORECT, nor does mention of trade names, commercial products, or organizations imply endorsement by the US Government or CORECT. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.</funding-statement>
</funding-group>
<counts>
<fig-count count="1"></fig-count>
<table-count count="2"></table-count>
<page-count count="18"></page-count>
</counts>
<custom-meta-group>
<custom-meta id="data-availability">
<meta-name>Data Availability</meta-name>
<meta-value>For plco genetic data, the prostate and panscan study datasets can be accessed with appropriate approval through the dbgap online resource (
<ext-link ext-link-type="uri" xlink:href="http://cgems.cancer.gov/data/">http://cgems.cancer.gov/data/</ext-link>
) accession numbers phs000207v.1p1 and phs000206.v3.p2, respectively, and the lung datasets from the dbgap website (
<ext-link ext-link-type="uri" xlink:href="http://www.ncbi.nlm.nih.gov/gap">http://www.ncbi.nlm.nih.gov/gap</ext-link>
) through accession number phs000093 v2.p2. For the remaining gecco studies, genetic data is in the process of being uploaded to dbgap and will be available soon (accession number phs001078.v1.p1). While the data release is being finalized, the gecco coordinating center can assist with any data requests (contact:
<email>gecco@fredhutch.org</email>
).</meta-value>
</custom-meta>
</custom-meta-group>
</article-meta>
<notes>
<title>Data Availability</title>
<p>For plco genetic data, the prostate and panscan study datasets can be accessed with appropriate approval through the dbgap online resource (
<ext-link ext-link-type="uri" xlink:href="http://cgems.cancer.gov/data/">http://cgems.cancer.gov/data/</ext-link>
) accession numbers phs000207v.1p1 and phs000206.v3.p2, respectively, and the lung datasets from the dbgap website (
<ext-link ext-link-type="uri" xlink:href="http://www.ncbi.nlm.nih.gov/gap">http://www.ncbi.nlm.nih.gov/gap</ext-link>
) through accession number phs000093 v2.p2. For the remaining gecco studies, genetic data is in the process of being uploaded to dbgap and will be available soon (accession number phs001078.v1.p1). While the data release is being finalized, the gecco coordinating center can assist with any data requests (contact:
<email>gecco@fredhutch.org</email>
).</p>
</notes>
</front>
<body>
<sec sec-type="intro" id="sec001">
<title>Introduction</title>
<p>Genetics play a key role in colorectal cancer (CRC) development [
<xref rid="pone.0157521.ref001" ref-type="bibr">1</xref>
,
<xref rid="pone.0157521.ref002" ref-type="bibr">2</xref>
]; genome-wide association studies (GWAS) have successfully identified many common genetic variants that predict risk [
<xref rid="pone.0157521.ref003" ref-type="bibr">3</xref>
<xref rid="pone.0157521.ref019" ref-type="bibr">19</xref>
]. Although these variants have modest associations (i.e., per-allele odds ratio less than 1.3), their discovery has reinforced the importance of suspected disease pathways as well as suggested novel ones [
<xref rid="pone.0157521.ref020" ref-type="bibr">20</xref>
].</p>
<p>An important next step is to characterize the biological importance of these loci. However, single nucleotide polymorphisms (SNPs) identified by GWAS (i.e., index SNPs) are themselves unlikely to be the underlying disease-causing variants; instead, they are expected to tag genomic regions containing correlated SNPs, which may have functional consequences [
<xref rid="pone.0157521.ref021" ref-type="bibr">21</xref>
,
<xref rid="pone.0157521.ref022" ref-type="bibr">22</xref>
]. Laboratory evaluation of all these variants is prohibitively cost- and labor-intensive. Fine-mapping efforts can help inform these experiments by narrowing the size of associated genomic regions likely to contain functional variation [
<xref rid="pone.0157521.ref022" ref-type="bibr">22</xref>
,
<xref rid="pone.0157521.ref023" ref-type="bibr">23</xref>
]. Several recent studies have shown the utility of this approach to refine regions of interest and propose promising functional candidates [
<xref rid="pone.0157521.ref014" ref-type="bibr">14</xref>
,
<xref rid="pone.0157521.ref017" ref-type="bibr">17</xref>
,
<xref rid="pone.0157521.ref024" ref-type="bibr">24</xref>
<xref rid="pone.0157521.ref031" ref-type="bibr">31</xref>
].</p>
<p>In addition, some loci may harbor multiple independent risk variants, rather than a single variant. As genomic regions surrounding index SNPs may span more than one linkage disequilibrium block, it is possible these loci harbor additional variants that predict risk independent of the index SNPs. Fine-mapping studies, when conducted within a broader region, can help identify these novel independent risk variants for cancer [
<xref rid="pone.0157521.ref014" ref-type="bibr">14</xref>
,
<xref rid="pone.0157521.ref031" ref-type="bibr">31</xref>
,
<xref rid="pone.0157521.ref032" ref-type="bibr">32</xref>
].</p>
<p>In this study of 11,900 colorectal tumor cases and 14,311 controls of European ancestry, we fine-mapped genomic regions harboring 31 known CRC risk variants using both genotyped data and data imputed from the 1000 Genomes Project [
<xref rid="pone.0157521.ref033" ref-type="bibr">33</xref>
]. This high-density genetic data allowed us to comprehensively examine common (>5%) as well as less common or rare (<5%) genetic variation in these regions. We aimed to narrow the likely region containing the functional variant(s) based on results from association testing, as well as search for novel risk alleles independent of the index SNP. Further, to help inform follow-up laboratory studies, we used a comprehensive bioinformatics-based approach to annotate potential functional candidates.</p>
</sec>
<sec sec-type="materials|methods" id="sec002">
<title>Materials and Methods</title>
<sec id="sec003">
<title>Ethics statement</title>
<p>All participants gave written informed consent and this study has been approved by the Fred Hutchinson Cancer Research Center (FHCRC) Institutional Review Board.</p>
</sec>
<sec id="sec004">
<title>Study population</title>
<p>Details of this study population have been described previously [
<xref rid="pone.0157521.ref003" ref-type="bibr">3</xref>
,
<xref rid="pone.0157521.ref034" ref-type="bibr">34</xref>
] and study-specific descriptions are provided in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
. The study population was derived from studies in the Genetics and Epidemiology of Colorectal Cancer Consortium (GECCO) (13 total: 7 case-control studies nested in prospective cohorts and 6 case-control studies) and the Colon Cancer Family Registry (CCFR) [
<xref rid="pone.0157521.ref003" ref-type="bibr">3</xref>
,
<xref rid="pone.0157521.ref034" ref-type="bibr">34</xref>
]. Study case, control, age, and sex distributions are listed in Table A in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
. We excluded participants of non-European ancestry as determined by principal component analysis [
<xref rid="pone.0157521.ref035" ref-type="bibr">35</xref>
]. The final study population comprised 11,900 cases (11,074 colorectal cancers, 826 advanced colorectal adenomas) and 14,311 controls.</p>
</sec>
<sec id="sec005">
<title>Colorectal tumor case definition</title>
<p>Detailed information on case and control definitions is provided in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
. Colorectal cancer cases were defined as colorectal adenocarcinoma confirmed by medical records, pathologic reports, or death certificates. Controls for colorectal cancer cases were population-based or selected from cohort participants who provided a blood sample and had no previous diagnosis of colorectal cancer. Advanced colorectal adenoma cases in the Nurses' Health Study and Health Professionals Follow-Up Study were confirmed by medical records, histopathology, or pathologic reports. Controls for advanced adenoma cases had a negative colonoscopy (except for controls matched to cases with distal adenoma, which either had a negative sigmoidoscopy or colonoscopy exam).</p>
</sec>
<sec id="sec006">
<title>Genotyping and quality control</title>
<p>Detailed information on genotyping and quality control procedures has been described [
<xref rid="pone.0157521.ref003" ref-type="bibr">3</xref>
,
<xref rid="pone.0157521.ref034" ref-type="bibr">34</xref>
] and are available in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
. Briefly, DNA from blood or buccal samples was genotyped using either Affymetrix (Gene Chip 10K, Mendel) (Affymetrix, Santa Clara, CA) or Illumina arrays (HumanHap550K, 610K, combined 300K and 240K, Human1M, HumanCytoSNP, HumanOmniExpress) (Illumina, Inc., San Diego, CA). Genotyped SNPs were excluded based on call rate (<98%), lack of Hardy-Weinberg Equilibrium in controls (
<italic>P</italic>
<1x10
<sup>-4</sup>
), and low minor allele frequency (MAF). All analyses were restricted to samples clustering with the Utah residents of Northern and Western European ancestry, using 1000 Genomes populations as reference, from the Centre d’etude du polymorphisme humain (CEPH) collection (CEU) population in principal component analysis [
<xref rid="pone.0157521.ref035" ref-type="bibr">35</xref>
].</p>
</sec>
<sec id="sec007">
<title>Genotype imputation to 1000 Genomes Project</title>
<p>We imputed genotype data to increase the density of genetic variants. As the reference panel we used the haplotypes of 1,092 samples (all populations) from release version 2 of the 1000 Genomes Project Phase I (
<ext-link ext-link-type="ftp" xlink:href="ftp://ftp-trace.ncbi.nih.gov/1000genomes/ftp/release/20110521">ftp://ftp-trace.ncbi.nih.gov/1000genomes/ftp/release/20110521</ext-link>
) [
<xref rid="pone.0157521.ref036" ref-type="bibr">36</xref>
]. Combining reference data from all populations helps improve imputation accuracy of low-frequency variants [
<xref rid="pone.0157521.ref037" ref-type="bibr">37</xref>
]. The target panel comprised genome-wide genotype data obtained using the methods described above. The target panel was phased using Beagle [
<xref rid="pone.0157521.ref038" ref-type="bibr">38</xref>
], and the phased target panel was imputed to the 1000 Genomes reference panel using Minimac [
<xref rid="pone.0157521.ref039" ref-type="bibr">39</xref>
]. We used Rsq as the imputation quality measure for imputed SNPs [
<xref rid="pone.0157521.ref040" ref-type="bibr">40</xref>
]. For imputed SNPs, we required that variants with low MAFs had higher imputation quality: For SNPs with MAF>0.01, we excluded those with Rsq≤0.3; for MAFs of 0.005–0.01, we excluded Rsq<0.5; and for MAF<0.005, we excluded Rsq<0.99.</p>
</sec>
<sec id="sec008">
<title>Statistical analysis</title>
<sec id="sec009">
<title>Variant selection</title>
<p>To determine genomic regions for fine-mapping, we identified 31 autosomal SNPs (index SNPs) in 22 loci previously associated with CRC risk in GWAS conducted in European ancestry individuals (Table B in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
) [
<xref rid="pone.0157521.ref003" ref-type="bibr">3</xref>
<xref rid="pone.0157521.ref018" ref-type="bibr">18</xref>
]. Through fine-mapping, we aimed to (1) refine the location of potential functional candidate(s) tagged by these index SNPs and (2) identify novel or independent signals near these loci, the latter being hypothesis-generating. We thus defined a broad genomic region of interest as that spanning 250-kb upstream and downstream of each index SNP and evaluated all variants in this 500-kb interval.</p>
</sec>
<sec id="sec010">
<title>Association testing</title>
<p>All statistical analyses were conducted centrally at the GECCO coordinating center on individual-level data to ensure a consistent analytical approach. Unless otherwise indicated, as appropriate, we adjusted for age at the reference time, sex, study center, smoking status (PHS only), batch effects (ASTERISK only: upon quality control there were slight variations in genotyping quality across batches, which were not observed in other studies), and the first three principal components from EIGENSTRAT [
<xref rid="pone.0157521.ref035" ref-type="bibr">35</xref>
] to account for population substructure.</p>
<p>For each study, we estimated the association between individual variants and colorectal tumor risk by calculating odds ratios (ORs) and 95% confidence intervals (CIs) using unconditional logistic regression assuming log-additive genetic effects. Each genotyped SNP was coded as 0, 1 or 2 copies of the variant allele. For imputed SNPs, we used the expected number of copies of the variant allele (the dosage), which gives unbiased effect estimates [
<xref rid="pone.0157521.ref041" ref-type="bibr">41</xref>
]. To combine study-specific estimates across studies, we obtained summary estimates using inverse-variance weighted fixed-effects meta-analysis. Colorectal cancer cases and their controls were analyzed separately from advanced adenoma cases and their controls before meta-analysis. We calculated heterogeneity
<italic>P</italic>
-values using Cochran's Q statistics [
<xref rid="pone.0157521.ref042" ref-type="bibr">42</xref>
]. Quantile-quantile (Q-Q) plots were used to assess whether the distribution of
<italic>P</italic>
-values was consistent with the null distribution (except for the extreme tail). In each region, we searched for additional independent association signals by testing each of the other variants conditioned on the top SNP in that region (i.e., 2 variants included in each model); variants are expected to be less correlated after conditioning on the top SNP. When testing for additional independent signals, we determined the
<italic>P</italic>
-value threshold for statistical significance by using the number of SNPs in each 500-kb region as the Bonferroni correction factor (α-level for a region = 0.05/# SNPs in that region). We used this approach to correct for multiple testing while also accounting for the knowledge that genetic variation in these regions is known to influence predisposition to CRC.</p>
<p>We used R (Version 2.15.1, R Foundation for Statistical Computing, Vienna, Austria) to conduct the statistical analysis, and LocusZoom [
<xref rid="pone.0157521.ref043" ref-type="bibr">43</xref>
] to visualize results. To determine the minimum detectable effect estimates in the present analysis, we estimated statistical power using Quanto Version 1.2.4 (
<ext-link ext-link-type="uri" xlink:href="http://hydra.usc.edu/gxe/">http://hydra.usc.edu/gxe/</ext-link>
).</p>
</sec>
</sec>
<sec id="sec011">
<title>Functional annotation using bioinformatics</title>
<p>Detailed information on functional annotation and various databases is provided in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
. In brief, compared with variants that are either non-functional or not in linkage disequilibrium with the underlying functional variant(s), colorectal tumor association signals are expected to be strongest (show the smallest
<italic>P</italic>
-value) for the functional variant(s), or variants in high linkage disequilibrium with the functional variant(s). Thus we selected the following for bioinformatics follow-up: 1) the variant showing the strongest evidence for association (smallest
<italic>P</italic>
-value) in each region (i.e., top SNP), 2) the index SNP in each region, 3) among the top 10 variants with the smallest
<italic>P</italic>
-values in each region, those that were correlated (
<italic>r</italic>
<sup>
<italic>2</italic>
</sup>
>0.5 in 1000 Genomes European populations) with the index SNP, and 4) any SNP completely correlated (
<italic>r</italic>
<sup>
<italic>2</italic>
</sup>
= 1 in 1000 Genomes European populations) with any SNP listed in parts 1–3. In addition, after performing conditional analyses that simultaneously included the index SNP(s) in multivariable models, we annotated SNPs showing
<italic>P</italic>
≤ 5E-05 and any SNPs completely correlated with these.</p>
<p>We annotated the potential function of variants in coding regions using PolyPhen-2 [
<xref rid="pone.0157521.ref044" ref-type="bibr">44</xref>
]. For variants in non-coding regions, we used HaploReg [
<xref rid="pone.0157521.ref045" ref-type="bibr">45</xref>
,
<xref rid="pone.0157521.ref046" ref-type="bibr">46</xref>
] and the University of California, Santa Cruz (UCSC) Genome Browser [
<xref rid="pone.0157521.ref047" ref-type="bibr">47</xref>
] to align each SNP to the reference genome and annotate them with multiple datasets generated from the Encyclopedia of DNA Elements (ENCODE) Project [
<xref rid="pone.0157521.ref048" ref-type="bibr">48</xref>
,
<xref rid="pone.0157521.ref049" ref-type="bibr">49</xref>
] or the NIH Roadmap program on Epigenomics [
<xref rid="pone.0157521.ref050" ref-type="bibr">50</xref>
] as detailed in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
. Annotation using these databases assumes that the disease-causing variant(s) affects disease by altering gene transcription through multiple regulatory mechanisms [
<xref rid="pone.0157521.ref048" ref-type="bibr">48</xref>
,
<xref rid="pone.0157521.ref049" ref-type="bibr">49</xref>
]. Such mechanisms include indicators for regions that may influence transcriptional regulation of target genes, such as chromatin accessibility (open chromatin), histone modification, binding of regulatory proteins, and alteration of regulatory motifs [
<xref rid="pone.0157521.ref045" ref-type="bibr">45</xref>
,
<xref rid="pone.0157521.ref051" ref-type="bibr">51</xref>
,
<xref rid="pone.0157521.ref052" ref-type="bibr">52</xref>
]. Conservation across vertebrates can provide further evidence of biologically important regions [
<xref rid="pone.0157521.ref053" ref-type="bibr">53</xref>
,
<xref rid="pone.0157521.ref054" ref-type="bibr">54</xref>
]. To identify variants showing any of these indicators of functional importance, we first queried HaploReg [
<xref rid="pone.0157521.ref045" ref-type="bibr">45</xref>
,
<xref rid="pone.0157521.ref046" ref-type="bibr">46</xref>
], which provides an overview of available annotations. We further interrogated variants with any functional evidence using the UCSC Genome Browser [
<xref rid="pone.0157521.ref047" ref-type="bibr">47</xref>
] to examine signal enrichment in regions harboring these variants, which helps correct for false positive signals for each assay (
<ext-link ext-link-type="uri" xlink:href="https://sites.google.com/site/anshulkundaje/projects/idr">https://sites.google.com/site/anshulkundaje/projects/idr</ext-link>
). Specifically, we examined whether variants were located in functionally important regions using the following datasets compiled by HaploReg [
<xref rid="pone.0157521.ref046" ref-type="bibr">46</xref>
] or UCSC Genome Browser [
<xref rid="pone.0157521.ref047" ref-type="bibr">47</xref>
]: DNAse I hypersensitivity data in ENCODE cell lines, including two for CRC (HCT-116 and Caco-2), to assess
<italic>open chromatin structure</italic>
; ChIP-seq data in ENCODE cell lines as well as Roadmap data in normal colon and rectal tissues for
<italic>histone enhancer or promoter modifications</italic>
; ChIP-seq data in ENCODE cell lines to determine regions that
<italic>bound to important regulatory proteins</italic>
(e.g., promoters, enhancers, silencers, and insulators); change in log-odds score based on position weight matrices [
<xref rid="pone.0157521.ref045" ref-type="bibr">45</xref>
] to predict whether a sequence harboring either the reference or alternate allele would exhibit
<italic>altered binding affinities</italic>
for regulatory proteins; and PhastCons scores [
<xref rid="pone.0157521.ref053" ref-type="bibr">53</xref>
,
<xref rid="pone.0157521.ref054" ref-type="bibr">54</xref>
] to predict genomic regions
<italic>conserved across vertebrates</italic>
. As intergenic variants often regulate the closest downstream gene [
<xref rid="pone.0157521.ref048" ref-type="bibr">48</xref>
,
<xref rid="pone.0157521.ref049" ref-type="bibr">49</xref>
], we predicted the gene regulated by each variant based on proximity of each variant to a gene as well as the orientation (3’ or 5’) relative to the nearest end of the gene [
<xref rid="pone.0157521.ref045" ref-type="bibr">45</xref>
]. Recognizing that cis-regulatory elements can also skip the closest gene, in exploratory analyses we integrated expression quantitative trail locus (eQTL) analysis to identify other potential tissue-specific target genes from the Genotype-Tissue Expression (GTEx) database [
<xref rid="pone.0157521.ref046" ref-type="bibr">46</xref>
], GEUVADIS [
<xref rid="pone.0157521.ref055" ref-type="bibr">55</xref>
], and other recent studies [
<xref rid="pone.0157521.ref056" ref-type="bibr">56</xref>
<xref rid="pone.0157521.ref060" ref-type="bibr">60</xref>
] using HaploReg and the GTEx Portal. Further, we evaluated variants in potential splice sites using Genie [
<xref rid="pone.0157521.ref061" ref-type="bibr">61</xref>
].</p>
<p>The relative strength of functional candidates was determined based on the accumulation of evidence from each of these datasets.
<italic>A priori</italic>
, we defined a score to summarize the amount of functional evidence for each variant using the following algorithm: showed (+1) histone modification, (+1) open chromatin, (+1) protein binding, (+1) protein binding in the presence of open chromatin or histone modification, (+1) different patterns of histone modification in cancerous vs. noncancerous cell lines/ tissues, (+1) regulatory evidence in a CRC cell line (e.g. Caco-2 or HCT-116) or normal colon/rectal tissue, (+0.5) altered binding motif, and (+0.5) a conserved region across vertebrates. Thus, variants were assigned a maximum score of 7. Although not observed in our data, any variant in a coding region predicted by PolyPhen to be “possibly damaging” or “probably damaging” would have been assigned a score of 8 or 9, respectively. Variants in coding regions predicted by PolyPhen to be “benign” or “unknown” were scored as a non-coding regulatory variant, as DNA sequences can act as coding exons in one tissue and enhancers of nearby gene(s) in another [
<xref rid="pone.0157521.ref062" ref-type="bibr">62</xref>
]. Caution should be exercised when interpreting these scores as there is a degree of uncertainty when relating annotation data to SNP function. These data are based on transformed cell lines or tissues instead of living organisms, and regulatory mechanisms may vary temporally as well as across different types of cells or tissues. However, bioinformatics analysis is primarily useful for prioritizing a large number of variants for more onerous laboratory follow-up; to this end, we used these scores to create 3 categories that ranked the strength of functional evidence for each variant: score of 3–3.5 =“weak”, 4–4.5 =“moderate”, and ≥5 =“strong”.</p>
</sec>
</sec>
<sec sec-type="results" id="sec012">
<title>Results</title>
<p>The mean age of the 26,211 participants was 64.2 years, ranging from 19 to 99 years (Table A in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
). Two studies (HPFS, PHS) comprised only males, and 3 studies (NHS, PMH, WHI) only females. The proportion of females in the remaining studies ranged from 30.8% to 52.0%.</p>
<p>For the 31 previously reported CRC-related variants (index SNPs), 17 showed
<italic>P</italic>
-values ≤ 0.001, 22 showed
<italic>P</italic>
-values ≤ 0.01, and 27 showed
<italic>P</italic>
-values ≤ 0.05 (Table B in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
). Further, ORs for 30 of 31 SNPs showed directions consistent with previous findings.</p>
<p>Across the 31 genomic regions encompassing index SNPs, there were on average 1,807 SNPs per 500-kb region, ranging from 967 to 2,364 SNPs per region. SNPs with the strongest evidence of CRC-associations may more likely be functional or strong proxies for functional candidates. To help refine regions harboring functional candidates, we identified the SNP showing the smallest
<italic>P</italic>
-value in each region (i.e., top SNP) (
<xref ref-type="table" rid="pone.0157521.t001">Table 1</xref>
). The initial index SNP did not show the strongest association signal in any genomic region (
<xref ref-type="fig" rid="pone.0157521.g001">Fig 1</xref>
). For loci that harbored more than 1 index SNP, the regions encompassing each index SNP sometimes overlapped, yielding regions in which the top SNP was the same (e.g., at 1q41, the top SNP rs143030473 showed the smallest
<italic>P</italic>
-value in 2 regions, defined by index SNPs rs6687758 and rs6691170). This was observed in 1q41, 12p13.32, 14q22.2, and 15q13. Thus for the 31 regions studied there were 25 unique top SNPs (note 12p13.32 and 15q13 each contained 3 index SNPs); of these, 20 had an association with
<italic>P</italic>
-values ≤ 0.001 and all 25 showed
<italic>P</italic>
-values ≤ 0.01. For these 25 variants, the top SNP was correlated with an index SNP in European populations at
<italic>r</italic>
<sup>
<italic>2</italic>
</sup>
≥ 0.8 for 8 SNPs, 0.6 ≤
<italic>r</italic>
<sup>
<italic>2</italic>
</sup>
< 0.8 for 6 SNPs, 0.4 ≤
<italic>r</italic>
<sup>
<italic>2</italic>
</sup>
< 0.6 for 4 SNPs, 0.2 ≤
<italic>r</italic>
<sup>
<italic>2</italic>
</sup>
< 0.4 for 1 SNP, and
<italic>r</italic>
<sup>
<italic>2</italic>
</sup>
< 0.2 for the remaining 6 SNPs.</p>
<fig id="pone.0157521.g001" orientation="portrait" position="float">
<object-id pub-id-type="doi">10.1371/journal.pone.0157521.g001</object-id>
<label>Fig 1</label>
<caption>
<title>Comparison of
<italic>P</italic>
-values for GWAS-identified variants (index SNPs) vs. variants with the smallest
<italic>P</italic>
-values (top SNPs) in 31 regions.</title>
<p>The height of each bar reflects the–log10
<italic>P</italic>
-value of each SNP in our study population. A grey bar indicates the index SNP, and a black bar indicates the top SNP.</p>
</caption>
<graphic xlink:href="pone.0157521.g001"></graphic>
</fig>
<table-wrap id="pone.0157521.t001" orientation="portrait" position="float">
<object-id pub-id-type="doi">10.1371/journal.pone.0157521.t001</object-id>
<label>Table 1</label>
<caption>
<title>Association results for variants showing the smallest
<italic>P</italic>
-values (top SNPs) in 31 regions surrounding previous GWAS-identified variants (index SNPs).</title>
</caption>
<alternatives>
<graphic id="pone.0157521.t001g" xlink:href="pone.0157521.t001"></graphic>
<table frame="hsides" rules="groups">
<colgroup span="1">
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
</colgroup>
<thead>
<tr>
<th align="left" rowspan="1" colspan="1">Locus</th>
<th align="left" rowspan="1" colspan="1">Index SNP</th>
<th align="left" rowspan="1" colspan="1">Level
<xref ref-type="table-fn" rid="t001fn002">
<sup>a</sup>
</xref>
</th>
<th align="left" rowspan="1" colspan="1"># SNPs</th>
<th align="left" rowspan="1" colspan="1">Top SNP</th>
<th align="left" rowspan="1" colspan="1">Level
<xref ref-type="table-fn" rid="t001fn002">
<sup>a</sup>
</xref>
</th>
<th align="left" rowspan="1" colspan="1">Position
<xref ref-type="table-fn" rid="t001fn006">
<sup>b</sup>
</xref>
</th>
<th align="left" rowspan="1" colspan="1">Genetic</th>
<th align="left" rowspan="1" colspan="1">
<italic>r</italic>
<sup>2</sup>
with</th>
<th align="left" colspan="5" rowspan="1">Top SNP results</th>
</tr>
<tr>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1">in</th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1">region</th>
<th align="left" rowspan="1" colspan="1">index</th>
<th align="left" rowspan="1" colspan="1">Ref/</th>
<th align="left" rowspan="1" colspan="1">Ref</th>
<th align="left" rowspan="1" colspan="1">OR (95% CI)
<xref ref-type="table-fn" rid="t001fn008">
<sup>d</sup>
</xref>
<sup>,</sup>
<xref ref-type="table-fn" rid="t001fn009">
<sup>e</sup>
</xref>
</th>
<th align="left" rowspan="1" colspan="1">
<italic>P</italic>
</th>
<th align="left" rowspan="1" colspan="1">
<italic>P</italic>
-het</th>
</tr>
<tr>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1">region</th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1">SNP
<xref ref-type="table-fn" rid="t001fn007">
<sup>c</sup>
</xref>
</th>
<th align="left" rowspan="1" colspan="1">other</th>
<th align="left" rowspan="1" colspan="1">allele</th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
</tr>
<tr>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1">allele</th>
<th align="left" rowspan="1" colspan="1">freq</th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
</tr>
</thead>
<tbody>
<tr>
<td align="left" rowspan="1" colspan="1">1q25.3</td>
<td align="left" rowspan="1" colspan="1">rs10911251</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">1886</td>
<td align="left" rowspan="1" colspan="1">rs6669796</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">183082825</td>
<td align="left" rowspan="1" colspan="1">
<italic>LAMC1</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.87</td>
<td align="left" rowspan="1" colspan="1">G/C</td>
<td align="left" rowspan="1" colspan="1">0.56</td>
<td align="left" rowspan="1" colspan="1">1.11 (1.07, 1.16)</td>
<td align="left" rowspan="1" colspan="1">5.8E-07</td>
<td align="left" rowspan="1" colspan="1">6.0E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">1q41</td>
<td align="left" rowspan="1" colspan="1">rs6687758</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn003">*</xref>
</td>
<td align="left" rowspan="1" colspan="1">1885</td>
<td align="left" rowspan="1" colspan="1">rs143030473</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">222161943</td>
<td align="left" rowspan="1" colspan="1">
<italic>DUSP10/CICP13</italic>
</td>
<td align="left" rowspan="1" colspan="1"><0.2</td>
<td align="left" rowspan="1" colspan="1">C/T</td>
<td align="left" rowspan="1" colspan="1">0.99</td>
<td align="left" rowspan="1" colspan="1">1.97 (1.35, 2.87)</td>
<td align="left" rowspan="1" colspan="1">4.5E-04</td>
<td align="left" rowspan="1" colspan="1">9.2E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs6691170</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">2096</td>
<td align="left" rowspan="1" colspan="1">rs143030473</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">222161943</td>
<td align="left" rowspan="1" colspan="1">
<italic>DUSP10/CICP13</italic>
</td>
<td align="left" rowspan="1" colspan="1"><0.2</td>
<td align="left" rowspan="1" colspan="1">C/T</td>
<td align="left" rowspan="1" colspan="1">0.99</td>
<td align="left" rowspan="1" colspan="1">1.97 (1.35, 2.87)</td>
<td align="left" rowspan="1" colspan="1">4.5E-04</td>
<td align="left" rowspan="1" colspan="1">9.2E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">2q32.3</td>
<td align="left" rowspan="1" colspan="1">rs11903757</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">1536</td>
<td align="left" rowspan="1" colspan="1">rs6731095</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">192589442</td>
<td align="left" rowspan="1" colspan="1">
<italic>NABP1/SDPR</italic>
</td>
<td align="left" rowspan="1" colspan="1">1.00</td>
<td align="left" rowspan="1" colspan="1">A/G</td>
<td align="left" rowspan="1" colspan="1">0.84</td>
<td align="left" rowspan="1" colspan="1">0.89 (0.84, 0.95)</td>
<td align="left" rowspan="1" colspan="1">2.3E-04</td>
<td align="left" rowspan="1" colspan="1">1.1E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">3q26.2</td>
<td align="left" rowspan="1" colspan="1">rs10936599</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">1651</td>
<td align="left" rowspan="1" colspan="1">rs2421771</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn004">**</xref>
</td>
<td align="left" rowspan="1" colspan="1">169411370</td>
<td align="left" rowspan="1" colspan="1">
<italic>MECOM/MYNN</italic>
</td>
<td align="left" rowspan="1" colspan="1"><0.2</td>
<td align="left" rowspan="1" colspan="1">C/T</td>
<td align="left" rowspan="1" colspan="1">0.96</td>
<td align="left" rowspan="1" colspan="1">1.21 (1.05, 1.40)</td>
<td align="left" rowspan="1" colspan="1">7.5E-03</td>
<td align="left" rowspan="1" colspan="1">9.2E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">5q31.1</td>
<td align="left" rowspan="1" colspan="1">rs647161</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn004">**</xref>
</td>
<td align="left" rowspan="1" colspan="1">1499</td>
<td align="left" rowspan="1" colspan="1">rs2193941</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">134469594</td>
<td align="left" rowspan="1" colspan="1">
<italic>PITX1/ H2AFY</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.57</td>
<td align="left" rowspan="1" colspan="1">G/A</td>
<td align="left" rowspan="1" colspan="1">0.72</td>
<td align="left" rowspan="1" colspan="1">1.09 (1.05, 1.14)</td>
<td align="left" rowspan="1" colspan="1">4.0E-05</td>
<td align="left" rowspan="1" colspan="1">9.2E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">6p21</td>
<td align="left" rowspan="1" colspan="1">rs1321311</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">2364</td>
<td align="left" rowspan="1" colspan="1">rs13215272</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">36589502</td>
<td align="left" rowspan="1" colspan="1">
<italic>SRSF3/CDKN1A</italic>
</td>
<td align="left" rowspan="1" colspan="1"><0.2</td>
<td align="left" rowspan="1" colspan="1">C/T</td>
<td align="left" rowspan="1" colspan="1">0.73</td>
<td align="left" rowspan="1" colspan="1">1.10 (1.04, 1.15)</td>
<td align="left" rowspan="1" colspan="1">5.3E-04</td>
<td align="left" rowspan="1" colspan="1">8.0E-02</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">8q23.3</td>
<td align="left" rowspan="1" colspan="1">rs16892766</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">1432</td>
<td align="left" rowspan="1" colspan="1">rs16888589</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">117635602</td>
<td align="left" rowspan="1" colspan="1">
<italic>TRPS1/EIF3H</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.92</td>
<td align="left" rowspan="1" colspan="1">A/G</td>
<td align="left" rowspan="1" colspan="1">0.91</td>
<td align="left" rowspan="1" colspan="1">0.80 (0.75, 0.86)</td>
<td align="left" rowspan="1" colspan="1">3.3E-10</td>
<td align="left" rowspan="1" colspan="1">7.1E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">8q24</td>
<td align="left" rowspan="1" colspan="1">rs6983267</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">2257</td>
<td align="left" rowspan="1" colspan="1">rs7013278</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">128414892</td>
<td align="left" rowspan="1" colspan="1">
<italic>SRRM1P1/POU5F1B/MYC</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.40</td>
<td align="left" rowspan="1" colspan="1">C/T</td>
<td align="left" rowspan="1" colspan="1">0.66</td>
<td align="left" rowspan="1" colspan="1">0.88 (0.85, 0.92)</td>
<td align="left" rowspan="1" colspan="1">7.8E-11</td>
<td align="left" rowspan="1" colspan="1">7.4E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">9p24</td>
<td align="left" rowspan="1" colspan="1">rs719725</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">1907</td>
<td align="left" rowspan="1" colspan="1">rs7875812</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">6364533</td>
<td align="left" rowspan="1" colspan="1">
<italic>TPD52L3/UHRF2</italic>
</td>
<td align="left" rowspan="1" colspan="1">1.00</td>
<td align="left" rowspan="1" colspan="1">A/T</td>
<td align="left" rowspan="1" colspan="1">0.63</td>
<td align="left" rowspan="1" colspan="1">1.09 (1.04, 1.13)</td>
<td align="left" rowspan="1" colspan="1">7.9E-05</td>
<td align="left" rowspan="1" colspan="1">7.4E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">10p14</td>
<td align="left" rowspan="1" colspan="1">rs10795668</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn003">*</xref>
</td>
<td align="left" rowspan="1" colspan="1">2363</td>
<td align="left" rowspan="1" colspan="1">rs1537603</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn004">**</xref>
</td>
<td align="left" rowspan="1" colspan="1">8734295</td>
<td align="left" rowspan="1" colspan="1">
<italic>KRT8P16/TCEB1P3</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.45</td>
<td align="left" rowspan="1" colspan="1">C/T</td>
<td align="left" rowspan="1" colspan="1">0.50</td>
<td align="left" rowspan="1" colspan="1">1.06 (1.02, 1.10)</td>
<td align="left" rowspan="1" colspan="1">1.2E-03</td>
<td align="left" rowspan="1" colspan="1">7.9E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">11q13.4</td>
<td align="left" rowspan="1" colspan="1">rs3824999</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">1788</td>
<td align="left" rowspan="1" colspan="1">rs72977282</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">74300441</td>
<td align="left" rowspan="1" colspan="1">
<italic>LIPT2/POLD3</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.59</td>
<td align="left" rowspan="1" colspan="1">T/A</td>
<td align="left" rowspan="1" colspan="1">0.59</td>
<td align="left" rowspan="1" colspan="1">0.92 (0.88, 0.95)</td>
<td align="left" rowspan="1" colspan="1">9.8E-06</td>
<td align="left" rowspan="1" colspan="1">8.1E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">11q23</td>
<td align="left" rowspan="1" colspan="1">rs3802842</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">1830</td>
<td align="left" rowspan="1" colspan="1">rs7130173</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">111154072</td>
<td align="left" rowspan="1" colspan="1">
<italic>C11orf93</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.95</td>
<td align="left" rowspan="1" colspan="1">C/A</td>
<td align="left" rowspan="1" colspan="1">0.73</td>
<td align="left" rowspan="1" colspan="1">0.89 (0.85, 0.93)</td>
<td align="left" rowspan="1" colspan="1">2.7E-08</td>
<td align="left" rowspan="1" colspan="1">3.8E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">12p13.32</td>
<td align="left" rowspan="1" colspan="1">rs10774214</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn003">*</xref>
</td>
<td align="left" rowspan="1" colspan="1">1656</td>
<td align="left" rowspan="1" colspan="1">rs3217874</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">4400808</td>
<td align="left" rowspan="1" colspan="1">
<italic>CCND2</italic>
</td>
<td align="left" rowspan="1" colspan="1"><0.2</td>
<td align="left" rowspan="1" colspan="1">C/T</td>
<td align="left" rowspan="1" colspan="1">0.58</td>
<td align="left" rowspan="1" colspan="1">0.89 (0.85, 0.93)</td>
<td align="left" rowspan="1" colspan="1">3.1E-08</td>
<td align="left" rowspan="1" colspan="1">7.8E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs3217810</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">1571</td>
<td align="left" rowspan="1" colspan="1">rs3217874</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">4400808</td>
<td align="left" rowspan="1" colspan="1">
<italic>CCND2</italic>
</td>
<td align="left" rowspan="1" colspan="1"><0.2</td>
<td align="left" rowspan="1" colspan="1">C/T</td>
<td align="left" rowspan="1" colspan="1">0.58</td>
<td align="left" rowspan="1" colspan="1">0.89 (0.85, 0.93)</td>
<td align="left" rowspan="1" colspan="1">3.1E-08</td>
<td align="left" rowspan="1" colspan="1">7.8E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs3217901</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">1539</td>
<td align="left" rowspan="1" colspan="1">rs3217874</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">4400808</td>
<td align="left" rowspan="1" colspan="1">
<italic>CCND2</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.65</td>
<td align="left" rowspan="1" colspan="1">C/T</td>
<td align="left" rowspan="1" colspan="1">0.58</td>
<td align="left" rowspan="1" colspan="1">0.89 (0.85, 0.93)</td>
<td align="left" rowspan="1" colspan="1">3.1E-08</td>
<td align="left" rowspan="1" colspan="1">7.8E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">12q13.13</td>
<td align="left" rowspan="1" colspan="1">rs11169552</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn003">*</xref>
</td>
<td align="left" rowspan="1" colspan="1">1310</td>
<td align="left" rowspan="1" colspan="1">rs7306677</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">51205763</td>
<td align="left" rowspan="1" colspan="1">
<italic>ATF1</italic>
</td>
<td align="left" rowspan="1" colspan="1"><0.2</td>
<td align="left" rowspan="1" colspan="1">C/T</td>
<td align="left" rowspan="1" colspan="1">0.62</td>
<td align="left" rowspan="1" colspan="1">0.92 (0.89, 0.96)</td>
<td align="left" rowspan="1" colspan="1">6.3E-05</td>
<td align="left" rowspan="1" colspan="1">3.4E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs7136702</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">967</td>
<td align="left" rowspan="1" colspan="1">rs11169524</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">51089734</td>
<td align="left" rowspan="1" colspan="1">
<italic>DIP2B</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.67</td>
<td align="left" rowspan="1" colspan="1">T/A</td>
<td align="left" rowspan="1" colspan="1">0.67</td>
<td align="left" rowspan="1" colspan="1">0.92 (0.88, 0.96)</td>
<td align="left" rowspan="1" colspan="1">8.5E-05</td>
<td align="left" rowspan="1" colspan="1">5.3E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">12q24.21</td>
<td align="left" rowspan="1" colspan="1">rs59336</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">2072</td>
<td align="left" rowspan="1" colspan="1">rs1427760</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">115100714</td>
<td align="left" rowspan="1" colspan="1">
<italic>TBX5/TBX3</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.71</td>
<td align="left" rowspan="1" colspan="1">T/C</td>
<td align="left" rowspan="1" colspan="1">0.50</td>
<td align="left" rowspan="1" colspan="1">0.90 (0.87, 0.94)</td>
<td align="left" rowspan="1" colspan="1">5.0E-07</td>
<td align="left" rowspan="1" colspan="1">2.5E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">14q22.2</td>
<td align="left" rowspan="1" colspan="1">rs1957636</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">1613</td>
<td align="left" rowspan="1" colspan="1">rs10130587</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">54419110</td>
<td align="left" rowspan="1" colspan="1">
<italic>BMP4</italic>
</td>
<td align="left" rowspan="1" colspan="1"><0.2</td>
<td align="left" rowspan="1" colspan="1">G/C</td>
<td align="left" rowspan="1" colspan="1">0.63</td>
<td align="left" rowspan="1" colspan="1">0.89 (0.85, 0.93)</td>
<td align="left" rowspan="1" colspan="1">4.1E-08</td>
<td align="left" rowspan="1" colspan="1">9.0E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs4444235</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">1659</td>
<td align="left" rowspan="1" colspan="1">rs10130587</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">54419110</td>
<td align="left" rowspan="1" colspan="1">
<italic>BMP4</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.67</td>
<td align="left" rowspan="1" colspan="1">G/C</td>
<td align="left" rowspan="1" colspan="1">0.63</td>
<td align="left" rowspan="1" colspan="1">0.89 (0.85, 0.93)</td>
<td align="left" rowspan="1" colspan="1">4.1E-08</td>
<td align="left" rowspan="1" colspan="1">9.0E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">15q13</td>
<td align="left" rowspan="1" colspan="1">rs11632715</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn004">**</xref>
</td>
<td align="left" rowspan="1" colspan="1">1735</td>
<td align="left" rowspan="1" colspan="1">rs2293582</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">33010412</td>
<td align="left" rowspan="1" colspan="1">
<italic>GREM1</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.23</td>
<td align="left" rowspan="1" colspan="1">G/A</td>
<td align="left" rowspan="1" colspan="1">0.80</td>
<td align="left" rowspan="1" colspan="1">0.86 (0.82, 0.91)</td>
<td align="left" rowspan="1" colspan="1">1.0E-09</td>
<td align="left" rowspan="1" colspan="1">3.8E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs16969681</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn004">**</xref>
</td>
<td align="left" rowspan="1" colspan="1">1692</td>
<td align="left" rowspan="1" colspan="1">rs2293582</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">33010412</td>
<td align="left" rowspan="1" colspan="1">
<italic>GREM1</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.22</td>
<td align="left" rowspan="1" colspan="1">G/A</td>
<td align="left" rowspan="1" colspan="1">0.80</td>
<td align="left" rowspan="1" colspan="1">0.86 (0.82, 0.91)</td>
<td align="left" rowspan="1" colspan="1">1.0E-09</td>
<td align="left" rowspan="1" colspan="1">3.8E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs4779584</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">1701</td>
<td align="left" rowspan="1" colspan="1">rs2293582</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">33010412</td>
<td align="left" rowspan="1" colspan="1">
<italic>GREM1</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.71</td>
<td align="left" rowspan="1" colspan="1">G/A</td>
<td align="left" rowspan="1" colspan="1">0.80</td>
<td align="left" rowspan="1" colspan="1">0.86 (0.82, 0.91)</td>
<td align="left" rowspan="1" colspan="1">1.0E-09</td>
<td align="left" rowspan="1" colspan="1">3.8E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">16q22.1</td>
<td align="left" rowspan="1" colspan="1">rs9929218</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn004">**</xref>
</td>
<td align="left" rowspan="1" colspan="1">1867</td>
<td align="left" rowspan="1" colspan="1">rs9932005</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn004">**</xref>
</td>
<td align="left" rowspan="1" colspan="1">68822019</td>
<td align="left" rowspan="1" colspan="1">
<italic>CDH1</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.63</td>
<td align="left" rowspan="1" colspan="1">C/T</td>
<td align="left" rowspan="1" colspan="1">0.76</td>
<td align="left" rowspan="1" colspan="1">1.08 (1.03, 1.13)</td>
<td align="left" rowspan="1" colspan="1">1.2E-03</td>
<td align="left" rowspan="1" colspan="1">6.6E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">18q21</td>
<td align="left" rowspan="1" colspan="1">rs4939827</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">1931</td>
<td align="left" rowspan="1" colspan="1">rs2337113</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">46452327</td>
<td align="left" rowspan="1" colspan="1">
<italic>SMAD7</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.91</td>
<td align="left" rowspan="1" colspan="1">A/G</td>
<td align="left" rowspan="1" colspan="1">0.55</td>
<td align="left" rowspan="1" colspan="1">1.13 (1.09, 1.18)</td>
<td align="left" rowspan="1" colspan="1">1.0E-10</td>
<td align="left" rowspan="1" colspan="1">2.5E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">19q13.1</td>
<td align="left" rowspan="1" colspan="1">rs10411210</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn003">*</xref>
</td>
<td align="left" rowspan="1" colspan="1">2307</td>
<td align="left" rowspan="1" colspan="1">rs75414102</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn004">**</xref>
</td>
<td align="left" rowspan="1" colspan="1">33302424</td>
<td align="left" rowspan="1" colspan="1">
<italic>TDRD12/SLC7A9</italic>
</td>
<td align="left" rowspan="1" colspan="1"><0.2</td>
<td align="left" rowspan="1" colspan="1">G/A</td>
<td align="left" rowspan="1" colspan="1">0.99</td>
<td align="left" rowspan="1" colspan="1">0.51 (0.33, 0.79)</td>
<td align="left" rowspan="1" colspan="1">2.6E-03</td>
<td align="left" rowspan="1" colspan="1">9.1E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">20p12.3</td>
<td align="left" rowspan="1" colspan="1">rs2423279</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">1901</td>
<td align="left" rowspan="1" colspan="1">rs118184022</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn004">**</xref>
</td>
<td align="left" rowspan="1" colspan="1">7719449</td>
<td align="left" rowspan="1" colspan="1">
<italic>BMP2/HAO1</italic>
</td>
<td align="left" rowspan="1" colspan="1"><0.2</td>
<td align="left" rowspan="1" colspan="1">C/T</td>
<td align="left" rowspan="1" colspan="1">0.98</td>
<td align="left" rowspan="1" colspan="1">0.58 (0.41, 0.83)</td>
<td align="left" rowspan="1" colspan="1">2.8E-03</td>
<td align="left" rowspan="1" colspan="1">9.4E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs4813802</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">1825</td>
<td align="left" rowspan="1" colspan="1">rs6085662</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">6698372</td>
<td align="left" rowspan="1" colspan="1">
<italic>FERMT1/BMP2</italic>
</td>
<td align="left" rowspan="1" colspan="1">1.00</td>
<td align="left" rowspan="1" colspan="1">G/C</td>
<td align="left" rowspan="1" colspan="1">0.65</td>
<td align="left" rowspan="1" colspan="1">0.91 (0.87, 0.95)</td>
<td align="left" rowspan="1" colspan="1">5.1E-06</td>
<td align="left" rowspan="1" colspan="1">3.0E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs961253</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">1990</td>
<td align="left" rowspan="1" colspan="1">rs56083061</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">6430696</td>
<td align="left" rowspan="1" colspan="1">
<italic>FERMT1/BMP2</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.28</td>
<td align="left" rowspan="1" colspan="1">G/A</td>
<td align="left" rowspan="1" colspan="1">0.79</td>
<td align="left" rowspan="1" colspan="1">0.90 (0.85, 0.94)</td>
<td align="left" rowspan="1" colspan="1">5.6E-06</td>
<td align="left" rowspan="1" colspan="1">7.6E-01</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">20q13.33</td>
<td align="left" rowspan="1" colspan="1">rs4925386</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn004">**</xref>
</td>
<td align="left" rowspan="1" colspan="1">2173</td>
<td align="left" rowspan="1" colspan="1">rs1760073</td>
<td align="left" rowspan="1" colspan="1">
<xref ref-type="table-fn" rid="t001fn005">***</xref>
</td>
<td align="left" rowspan="1" colspan="1">60926106</td>
<td align="left" rowspan="1" colspan="1">
<italic>LAMA5</italic>
</td>
<td align="left" rowspan="1" colspan="1">0.81</td>
<td align="left" rowspan="1" colspan="1">G/A</td>
<td align="left" rowspan="1" colspan="1">0.67</td>
<td align="left" rowspan="1" colspan="1">1.08 (1.04, 1.12)</td>
<td align="left" rowspan="1" colspan="1">1.9E-04</td>
<td align="left" rowspan="1" colspan="1">8.8E-02</td>
</tr>
</tbody>
</table>
</alternatives>
<table-wrap-foot>
<fn id="t001fn001">
<p>Abbreviations: SNP, single nucleotide polymorphism; Ref, reference; Freq, frequency; OR, odds ratio; CI, confidence interval;
<italic>P</italic>
-het,
<italic>P</italic>
value for test of heterogeneity across studies</p>
</fn>
<fn id="t001fn002">
<p>
<sup>a</sup>
Level of statistical evidence:</p>
</fn>
<fn id="t001fn003">
<p>*SNP with
<italic>P</italic>
≤ 0.05</p>
</fn>
<fn id="t001fn004">
<p>**SNP with
<italic>P ≤</italic>
0.01</p>
</fn>
<fn id="t001fn005">
<p>***SNP with
<italic>P</italic>
≤0.001.
<italic>P</italic>
-values are not adjusted for multiple testing as these data are intended to help refine previously identified CRC-related regions and highlight sets of variants warranting bioinformatics-based follow-up.</p>
</fn>
<fn id="t001fn006">
<p>
<sup>b</sup>
Based on NCBI build 37 data.</p>
</fn>
<fn id="t001fn007">
<p>
<sup>c</sup>
Based on 1000 Genomes Project data in European populations.</p>
</fn>
<fn id="t001fn008">
<p>
<sup>d</sup>
Adjusted for age in years, sex, first 3 principal components, study center, batch (ASTERISK only), and smoking (PHS only).</p>
</fn>
<fn id="t001fn009">
<p>
<sup>e</sup>
Estimate calculated using the log-additive genetic model for each additional reference allele.</p>
</fn>
</table-wrap-foot>
</table-wrap>
<p>Variants carried forward for functional annotation spanned a median interval of 32-kb. We scored 21 variants in 12 loci as having “strong” functional evidence (
<xref ref-type="table" rid="pone.0157521.t002">Table 2</xref>
, additional details in
<xref ref-type="supplementary-material" rid="pone.0157521.s001">S1 Table</xref>
). At 4 loci (8q24, 11q13.4, 19q13.1, 20p12.3) the index SNP was among the SNPs with the highest functional scores. All 21 candidates were located in regions that were non-coding (15 intronic and 6 intergenic) with open chromatin structure (i.e., accessible to regulatory factors). Twenty of 21 candidates (all except for 18q21/rs34007497) bound to multiple transcription factors. Fifteen variants were predicted to disrupt transcription factor binding. Several candidates showed different patterns of histone enhancer or promoter marks when comparing cancer cells vs. normal cells or tissues. Only 3 variants (8q24/rs6983267, 18q21/rs4939567, 20p12.3/rs4813802) were located in an evolutionarily conserved region, suggesting that most of the predicted regulatory regions may be dynamic through evolution.</p>
<table-wrap id="pone.0157521.t002" orientation="portrait" position="float">
<object-id pub-id-type="doi">10.1371/journal.pone.0157521.t002</object-id>
<label>Table 2</label>
<caption>
<title>SNPs showing strong evidence for functional importance based on bioinformatics.</title>
</caption>
<alternatives>
<graphic id="pone.0157521.t002g" xlink:href="pone.0157521.t002"></graphic>
<table frame="hsides" rules="groups">
<colgroup span="1">
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
<col align="left" valign="middle" span="1"></col>
</colgroup>
<thead>
<tr>
<th align="left" rowspan="1" colspan="1">
<bold>Locus</bold>
</th>
<th align="left" rowspan="1" colspan="1">Index SNP</th>
<th align="left" rowspan="1" colspan="1">Strong</th>
<th align="left" rowspan="1" colspan="1">Mean</th>
<th align="left" rowspan="1" colspan="1">
<italic>r</italic>
<sup>
<italic>2</italic>
</sup>
with</th>
<th align="left" rowspan="1" colspan="1">Predicted</th>
<th align="left" rowspan="1" colspan="1">Genomic</th>
<th align="left" rowspan="1" colspan="1"># Cell lines</th>
<th align="left" colspan="3" rowspan="1">Show histone regulatory marks
<xref ref-type="table-fn" rid="t002fn005">
<sup>e</sup>
</xref>
</th>
<th align="left" rowspan="1" colspan="1">Proteins</th>
<th align="left" rowspan="1" colspan="1">Altered binding</th>
<th align="left" rowspan="1" colspan="1">Evidence</th>
</tr>
<tr>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1">functional</th>
<th align="left" rowspan="1" colspan="1">imput-</th>
<th align="left" rowspan="1" colspan="1">index</th>
<th align="left" rowspan="1" colspan="1">regulated</th>
<th align="left" rowspan="1" colspan="1">location of</th>
<th align="left" rowspan="1" colspan="1">showing</th>
<th align="left" rowspan="1" colspan="1"># Cancer/</th>
<th align="left" rowspan="1" colspan="1"># Normal</th>
<th align="left" rowspan="1" colspan="1">Normal</th>
<th align="left" rowspan="1" colspan="1">bound
<xref ref-type="table-fn" rid="t002fn005">
<sup>e</sup>
</xref>
</th>
<th align="left" rowspan="1" colspan="1">motifs
<xref ref-type="table-fn" rid="t002fn007">
<sup>g</sup>
</xref>
</th>
<th align="left" rowspan="1" colspan="1">for</th>
</tr>
<tr>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1">candidate</th>
<th align="left" rowspan="1" colspan="1">ation</th>
<th align="left" rowspan="1" colspan="1">SNP
<xref ref-type="table-fn" rid="t002fn002">
<sup>b</sup>
</xref>
</th>
<th align="left" rowspan="1" colspan="1">gene
<xref ref-type="table-fn" rid="t002fn003">
<sup>c</sup>
</xref>
</th>
<th align="left" rowspan="1" colspan="1">functional</th>
<th align="left" rowspan="1" colspan="1">open</th>
<th align="left" rowspan="1" colspan="1">progenitor</th>
<th align="left" rowspan="1" colspan="1">cell lines</th>
<th align="left" rowspan="1" colspan="1">colorectal</th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1">conserved</th>
</tr>
<tr>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1">
<italic>r</italic>
<sup>
<italic>2</italic>
</sup>
<xref ref-type="table-fn" rid="t002fn001">
<sup>a</sup>
</xref>
</th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1">candidate</th>
<th align="left" rowspan="1" colspan="1">chromatin
<xref ref-type="table-fn" rid="t002fn004">
<sup>d</sup>
</xref>
</th>
<th align="left" rowspan="1" colspan="1">cell lines</th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1">tissues
<xref ref-type="table-fn" rid="t002fn006">
<sup>f</sup>
</xref>
</th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1"></th>
<th align="left" rowspan="1" colspan="1">region
<xref ref-type="table-fn" rid="t002fn008">
<sup>h</sup>
</xref>
</th>
</tr>
</thead>
<tbody>
<tr>
<td align="left" rowspan="1" colspan="1">5q31.1</td>
<td align="left" rowspan="1" colspan="1">rs647161</td>
<td align="left" rowspan="1" colspan="1">rs1366111</td>
<td align="left" rowspan="1" colspan="1">0.89</td>
<td align="left" rowspan="1" colspan="1">0.57</td>
<td align="left" rowspan="1" colspan="1">
<italic>PITX1</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">1, 2</td>
<td align="left" rowspan="1" colspan="1">POL2,EGR1</td>
<td align="left" rowspan="1" colspan="1">Zfx</td>
<td align="left" rowspan="1" colspan="1">0</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">8q24</td>
<td align="left" rowspan="1" colspan="1">rs6983267</td>
<td align="left" rowspan="1" colspan="1">rs6983267</td>
<td align="left" rowspan="1" colspan="1">0.95</td>
<td align="left" rowspan="1" colspan="1">index</td>
<td align="left" rowspan="1" colspan="1">
<italic>MYC</italic>
</td>
<td align="left" rowspan="1" colspan="1">intergenic</td>
<td align="left" rowspan="1" colspan="1">1 CRC line</td>
<td align="left" rowspan="1" colspan="1">1</td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">TCF4 (CRC line),</td>
<td align="left" rowspan="1" colspan="1">AP-1, Sox, TCF4</td>
<td align="left" rowspan="1" colspan="1">1</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">P300, FOXA1,</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">RXRA, SP1</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">11q13.4</td>
<td align="left" rowspan="1" colspan="1">rs3824999</td>
<td align="left" rowspan="1" colspan="1">rs3824999</td>
<td align="left" rowspan="1" colspan="1">0.99</td>
<td align="left" rowspan="1" colspan="1">index</td>
<td align="left" rowspan="1" colspan="1">
<italic>POLD3</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">4</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">4</td>
<td align="left" rowspan="1" colspan="1">JUND</td>
<td align="left" rowspan="1" colspan="1">AP-3, Evi-1, Mef2</td>
<td align="left" rowspan="1" colspan="1">0</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">11q23</td>
<td align="left" rowspan="1" colspan="1">rs3802842</td>
<td align="left" rowspan="1" colspan="1">rs7130173</td>
<td align="left" rowspan="1" colspan="1">0.95</td>
<td align="left" rowspan="1" colspan="1">0.93</td>
<td align="left" rowspan="1" colspan="1">
<italic>Unknown</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">31 including</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">2, 3</td>
<td align="left" rowspan="1" colspan="1">RAD21, SMC3,</td>
<td align="left" rowspan="1" colspan="1">GSP1, SRF</td>
<td align="left" rowspan="1" colspan="1">0</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">1 CRC line</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">YY1, CTCF</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">12p13.32</td>
<td align="left" rowspan="1" colspan="1">rs3217901</td>
<td align="left" rowspan="1" colspan="1">rs3217827</td>
<td align="left" rowspan="1" colspan="1">0.85</td>
<td align="left" rowspan="1" colspan="1">0.61</td>
<td align="left" rowspan="1" colspan="1">
<italic>CCND2</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">1</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">3</td>
<td align="left" rowspan="1" colspan="1">RAD21, CTCF</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">0</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">12q24.21</td>
<td align="left" rowspan="1" colspan="1">rs59336</td>
<td align="left" rowspan="1" colspan="1">rs71807</td>
<td align="left" rowspan="1" colspan="1">0.89</td>
<td align="left" rowspan="1" colspan="1">0.86</td>
<td align="left" rowspan="1" colspan="1">
<italic>TBX3</italic>
</td>
<td align="left" rowspan="1" colspan="1">intergenic</td>
<td align="left" rowspan="1" colspan="1">1</td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">4</td>
<td align="left" rowspan="1" colspan="1">1, 3</td>
<td align="left" rowspan="1" colspan="1">BAF155, HAE2F1,</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">0</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">CTCF</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs484443</td>
<td align="left" rowspan="1" colspan="1">0.91</td>
<td align="left" rowspan="1" colspan="1">0.90</td>
<td align="left" rowspan="1" colspan="1">
<italic>TBX3</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">8</td>
<td align="left" rowspan="1" colspan="1">1</td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">2, 4</td>
<td align="left" rowspan="1" colspan="1">P300,USF1</td>
<td align="left" rowspan="1" colspan="1">RREB-1, Rad21</td>
<td align="left" rowspan="1" colspan="1">0</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">14q22.2</td>
<td align="left" rowspan="1" colspan="1">rs4444235</td>
<td align="left" rowspan="1" colspan="1">rs2071047</td>
<td align="left" rowspan="1" colspan="1">1.00</td>
<td align="left" rowspan="1" colspan="1">0.75</td>
<td align="left" rowspan="1" colspan="1">
<italic>BMP4</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">9</td>
<td align="left" rowspan="1" colspan="1">3</td>
<td align="left" rowspan="1" colspan="1">3</td>
<td align="left" rowspan="1" colspan="1">1–4</td>
<td align="left" rowspan="1" colspan="1">TCF4 (CRC line)</td>
<td align="left" rowspan="1" colspan="1">BCL, HNF4, RXRA</td>
<td align="left" rowspan="1" colspan="1">0</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs10130587</td>
<td align="left" rowspan="1" colspan="1">0.74</td>
<td align="left" rowspan="1" colspan="1">0.67</td>
<td align="left" rowspan="1" colspan="1">
<italic>BMP4</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">3</td>
<td align="left" rowspan="1" colspan="1">3</td>
<td align="left" rowspan="1" colspan="1">1–4</td>
<td align="left" rowspan="1" colspan="1">GATA3</td>
<td align="left" rowspan="1" colspan="1">5 motifs</td>
<td align="left" rowspan="1" colspan="1">0</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs35107139</td>
<td align="left" rowspan="1" colspan="1">0.82</td>
<td align="left" rowspan="1" colspan="1">0.64</td>
<td align="left" rowspan="1" colspan="1">
<italic>BMP4</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">3</td>
<td align="left" rowspan="1" colspan="1">3</td>
<td align="left" rowspan="1" colspan="1">1, 3, 4</td>
<td align="left" rowspan="1" colspan="1">GATA3</td>
<td align="left" rowspan="1" colspan="1">12 motifs</td>
<td align="left" rowspan="1" colspan="1">0</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">15q13</td>
<td align="left" rowspan="1" colspan="1">rs4779584</td>
<td align="left" rowspan="1" colspan="1">rs2293582</td>
<td align="left" rowspan="1" colspan="1">0.94</td>
<td align="left" rowspan="1" colspan="1">0.71</td>
<td align="left" rowspan="1" colspan="1">
<italic>GREM1</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">4</td>
<td align="left" rowspan="1" colspan="1">3</td>
<td align="left" rowspan="1" colspan="1">5</td>
<td align="left" rowspan="1" colspan="1">1–4</td>
<td align="left" rowspan="1" colspan="1">POL2</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">0</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs2293581</td>
<td align="left" rowspan="1" colspan="1">0.97</td>
<td align="left" rowspan="1" colspan="1">0.66</td>
<td align="left" rowspan="1" colspan="1">
<italic>GREM1</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">36</td>
<td align="left" rowspan="1" colspan="1">4</td>
<td align="left" rowspan="1" colspan="1">5</td>
<td align="left" rowspan="1" colspan="1">1–4</td>
<td align="left" rowspan="1" colspan="1">SUZ12</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">0</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs1406389</td>
<td align="left" rowspan="1" colspan="1">0.97</td>
<td align="left" rowspan="1" colspan="1">0.66</td>
<td align="left" rowspan="1" colspan="1">
<italic>GREM1</italic>
</td>
<td align="left" rowspan="1" colspan="1">intergenic</td>
<td align="left" rowspan="1" colspan="1">12</td>
<td align="left" rowspan="1" colspan="1">3</td>
<td align="left" rowspan="1" colspan="1">4</td>
<td align="left" rowspan="1" colspan="1">1, 2, 4</td>
<td align="left" rowspan="1" colspan="1">SUZ12</td>
<td align="left" rowspan="1" colspan="1">Irf, SIX5</td>
<td align="left" rowspan="1" colspan="1">0</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">18q21</td>
<td align="left" rowspan="1" colspan="1">rs4939827</td>
<td align="left" rowspan="1" colspan="1">rs11874392</td>
<td align="left" rowspan="1" colspan="1">0.93</td>
<td align="left" rowspan="1" colspan="1">1.00</td>
<td align="left" rowspan="1" colspan="1">
<italic>SMAD7</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">4</td>
<td align="left" rowspan="1" colspan="1">3</td>
<td align="left" rowspan="1" colspan="1">1</td>
<td align="left" rowspan="1" colspan="1">1, 2, 4</td>
<td align="left" rowspan="1" colspan="1">11 proteins</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">1</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs4939567</td>
<td align="left" rowspan="1" colspan="1">0.91</td>
<td align="left" rowspan="1" colspan="1">1.00</td>
<td align="left" rowspan="1" colspan="1">
<italic>SMAD7</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">1</td>
<td align="left" rowspan="1" colspan="1">4</td>
<td align="left" rowspan="1" colspan="1">3</td>
<td align="left" rowspan="1" colspan="1">1–4</td>
<td align="left" rowspan="1" colspan="1">MAFK</td>
<td align="left" rowspan="1" colspan="1">RREB-1, VDR_2</td>
<td align="left" rowspan="1" colspan="1">1</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs34007497</td>
<td align="left" rowspan="1" colspan="1">0.91</td>
<td align="left" rowspan="1" colspan="1">0.57</td>
<td align="left" rowspan="1" colspan="1">
<italic>SMAD7</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">22</td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">4</td>
<td align="left" rowspan="1" colspan="1">1–4</td>
<td align="left" rowspan="1" colspan="1">CTCF,STAT1</td>
<td align="left" rowspan="1" colspan="1">4 motifs</td>
<td align="left" rowspan="1" colspan="1">0</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">19q13.1</td>
<td align="left" rowspan="1" colspan="1">rs10411210</td>
<td align="left" rowspan="1" colspan="1">rs10411210</td>
<td align="left" rowspan="1" colspan="1">0.94</td>
<td align="left" rowspan="1" colspan="1">index</td>
<td align="left" rowspan="1" colspan="1">
<italic>RHPN2</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">48 including</td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">1</td>
<td align="left" rowspan="1" colspan="1">1, 2</td>
<td align="left" rowspan="1" colspan="1">17 proteins</td>
<td align="left" rowspan="1" colspan="1">Mef2, STAT</td>
<td align="left" rowspan="1" colspan="1">1</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">1 CRC line</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">20p12.3</td>
<td align="left" rowspan="1" colspan="1">rs961253</td>
<td align="left" rowspan="1" colspan="1">rs966817</td>
<td align="left" rowspan="1" colspan="1">1.00</td>
<td align="left" rowspan="1" colspan="1">0.87</td>
<td align="left" rowspan="1" colspan="1">
<italic>BMP2</italic>
</td>
<td align="left" rowspan="1" colspan="1">intergenic</td>
<td align="left" rowspan="1" colspan="1">18</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">1</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">CJUN</td>
<td align="left" rowspan="1" colspan="1">Evi-1, FAC1, GLI</td>
<td align="left" rowspan="1" colspan="1">1</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs4813802</td>
<td align="left" rowspan="1" colspan="1">rs4813802</td>
<td align="left" rowspan="1" colspan="1">0.86</td>
<td align="left" rowspan="1" colspan="1">index</td>
<td align="left" rowspan="1" colspan="1">
<italic>BMP2</italic>
</td>
<td align="left" rowspan="1" colspan="1">intergenic</td>
<td align="left" rowspan="1" colspan="1">10</td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">6 proteins</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">1</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">rs6085661</td>
<td align="left" rowspan="1" colspan="1">0.89</td>
<td align="left" rowspan="1" colspan="1">0.76</td>
<td align="left" rowspan="1" colspan="1">
<italic>BMP2</italic>
</td>
<td align="left" rowspan="1" colspan="1">intergenic</td>
<td align="left" rowspan="1" colspan="1">19</td>
<td align="left" rowspan="1" colspan="1">1</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">7 proteins</td>
<td align="left" rowspan="1" colspan="1">BCL, NRSF, VDR</td>
<td align="left" rowspan="1" colspan="1">1</td>
</tr>
<tr>
<td align="left" rowspan="1" colspan="1">20q13.33</td>
<td align="left" rowspan="1" colspan="1">rs4925386</td>
<td align="left" rowspan="1" colspan="1">rs1741634</td>
<td align="left" rowspan="1" colspan="1">0.96</td>
<td align="left" rowspan="1" colspan="1">0.80</td>
<td align="left" rowspan="1" colspan="1">
<italic>LAMA5</italic>
</td>
<td align="left" rowspan="1" colspan="1">intronic</td>
<td align="left" rowspan="1" colspan="1">7</td>
<td align="left" rowspan="1" colspan="1"></td>
<td align="left" rowspan="1" colspan="1">2</td>
<td align="left" rowspan="1" colspan="1">1, 2</td>
<td align="left" rowspan="1" colspan="1">GR</td>
<td align="left" rowspan="1" colspan="1">EWSR1-FLI1</td>
<td align="left" rowspan="1" colspan="1">0</td>
</tr>
</tbody>
</table>
</alternatives>
<table-wrap-foot>
<fn id="t002fn001">
<p>
<sup>a</sup>
Estimated value of the squared correlation between imputed genotypes and true (unobserved) genotypes for the strong functional candidate, averaged across studies.</p>
</fn>
<fn id="t002fn002">
<p>
<sup>b</sup>
Based on data from 1000 Genomes Pilot 1 CEU.</p>
</fn>
<fn id="t002fn003">
<p>
<sup>c</sup>
Based on HaploReg data using proximity of each variant to a gene as well as the orientation (3’ or 5’) relative to the nearest end of the gene.</p>
</fn>
<fn id="t002fn004">
<p>
<sup>d</sup>
Based on data from DNAse I hypersensitivity assays.</p>
</fn>
<fn id="t002fn005">
<p>
<sup>e</sup>
ENCODE ChIP-seq assay.</p>
</fn>
<fn id="t002fn006">
<p>
<sup>f</sup>
Based on data from ChIP-seq assays in the Roadmap Epigenomics Project in the following normal (non-cancerous) tissues: (1) colon mucosa, (2) rectal mucosa, (3) colon smooth muscle, and (4) rectal smooth muscle.</p>
</fn>
<fn id="t002fn007">
<p>
<sup>g</sup>
Position weight matrix score between SNP alleles confers a change in log-odds (LOD) score > 5.</p>
</fn>
<fn id="t002fn008">
<p>
<sup>h</sup>
Based on PhastCons scores: 1 = strong evidence of conserved element; 0 = no evidence of conserved element.</p>
</fn>
</table-wrap-foot>
</table-wrap>
<p>In each region, after conditioning on the top SNP and accounting for the number of tests, we did not observe any statistically significant SNPs.</p>
</sec>
<sec sec-type="conclusions" id="sec013">
<title>Discussion</title>
<p>In this large study population of over 26,000 participants of European ancestry, we used high-density genetic data imputed from the 1000 Genomes Project to comprehensively fine-map genomic regions harboring 31 GWAS-identified CRC risk variants. In association tests, the index SNP did not show the smallest
<italic>P</italic>
-value in any genomic region. Using bioinformatics-based annotation to follow-up variants with the strongest association signals, we showed strong evidence for 21 functional candidates in 12 CRC-related loci. We observed limited evidence of additional independent CRC association signals within GWAS-identified regions.</p>
<p>Although the index SNP did not show the smallest
<italic>P</italic>
-value in any genomic region, all functional candidates were correlated with the index SNP (
<italic>r</italic>
<sup>
<italic>2</italic>
</sup>
of at least 0.57). Interestingly, however, the index SNP was a strong functional candidate in only 4 of the 12 loci harboring a strong functional candidate. Combined, these data from our association testing and functional annotation support the hypothesis that most GWAS-identified index SNPs are not the underlying functional variant, but may instead act as proxies of correlated variants with biological importance.</p>
<p>Eight previous studies have fine-mapped a limited number of GWAS-identified CRC loci in individuals of European ancestry [
<xref rid="pone.0157521.ref014" ref-type="bibr">14</xref>
,
<xref rid="pone.0157521.ref017" ref-type="bibr">17</xref>
,
<xref rid="pone.0157521.ref024" ref-type="bibr">24</xref>
<xref rid="pone.0157521.ref029" ref-type="bibr">29</xref>
]; these studies have reported 34 candidate SNPs showing functional evidence (summarized in Table C in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
). In addition, 2 recent studies have comprehensively fine-mapped known CRC loci: Whiffin et al. [
<xref rid="pone.0157521.ref031" ref-type="bibr">31</xref>
] identified 4 additional candidates in 1q41, 15q13, 18q21, and 20q13.33 in European ancestry individuals (5,626 cases; 7,817 controls); Wang et al. [
<xref rid="pone.0157521.ref030" ref-type="bibr">30</xref>
] identified 1 additional candidate in 1q41 in African Americans (1,894 cases; 4,703 controls). Of these 39 reported candidates in 11 loci, 36 passed genotyping quality control in our study. In the present analysis, 16 of these SNPs had
<italic>P</italic>
-values ≤ 0.001, 5 had
<italic>P</italic>
-values >0.001 and ≤ 0.01, and 7 had
<italic>P</italic>
-values >0.01 and ≤ 0.05. Similar to our findings, only 5 of 39 previously reported functional candidates were GWAS-identified SNPs. We observed 3 exonic candidates out of an expanded list of 51 variants showing “weak”, “moderate”, or “strong” functional evidence (see
<xref ref-type="supplementary-material" rid="pone.0157521.s001">S1 Table</xref>
for an expanded list of functional candidates); similarly, only 2 previously reported candidates (rs706793, rs28626308) were in coding regions [
<xref rid="pone.0157521.ref024" ref-type="bibr">24</xref>
,
<xref rid="pone.0157521.ref025" ref-type="bibr">25</xref>
]—highlighting the importance of non-coding effects on CRC [
<xref rid="pone.0157521.ref063" ref-type="bibr">63</xref>
].</p>
<p>To identify potential variants for laboratory follow-up, we compared all previously reported candidates (in 11 loci) with variants in the present analysis that showed “moderate” or “strong” functional evidence (Table C in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
). In 5 loci (8q23.3, 8q24, 15q13, 18q21, and 19q13.1) our data confirmed previously reported functional candidates [
<xref rid="pone.0157521.ref025" ref-type="bibr">25</xref>
,
<xref rid="pone.0157521.ref027" ref-type="bibr">27</xref>
,
<xref rid="pone.0157521.ref029" ref-type="bibr">29</xref>
,
<xref rid="pone.0157521.ref031" ref-type="bibr">31</xref>
]. In addition, in 11q23 and 16q22.1 we observed candidate(s) that highly correlated and were within 5-kb of a previously reported candidate variant [
<xref rid="pone.0157521.ref025" ref-type="bibr">25</xref>
,
<xref rid="pone.0157521.ref028" ref-type="bibr">28</xref>
]. Fine-mapping can be limited in distinguishing which of several highly correlated SNPs located very close together is the true causal variant. Our candidates in these 2 loci, 11q23/rs7130173 and 16q22.1/rs9929218, showed stronger functional evidence compared with reported candidates, which either were not selected for functional annotation or showed less than “weak” functional evidence (scored less than 3). These data show that to avoid missing functional variation, laboratory studies should follow-up not just the strongest candidates, but also variants showing any evidence of biological importance that are very close and highly correlated. Our data did not show functional evidence for reported candidates in the remaining 4 loci. In 1q41 we did not identify a functional candidate; in 12q13.13 and 14q22.2 we predicted functional candidates that were >150-kb from those previously reported [
<xref rid="pone.0157521.ref014" ref-type="bibr">14</xref>
,
<xref rid="pone.0157521.ref024" ref-type="bibr">24</xref>
]; and in 20q13.33 our candidates were >5-kb away and did not show high correlation with those previously reported (
<italic>r</italic>
<sup>
<italic>2</italic>
</sup>
= 0.59–0.60) [
<xref rid="pone.0157521.ref031" ref-type="bibr">31</xref>
]. In 3 of these loci (1q41, 12q13.13, 14q22.2) only 1 of 7 previously reported candidates showed
<italic>P</italic>
<0.05 in our study population, suggesting they may be false positives. In 20q13.33 the reported candidates, rs1741640 and rs2236202, were not selected for functional annotation in our study based on their
<italic>P</italic>
-values relative to other variants in the region. Taken together, these data support the utility of fine-mapping to reveal potential functional variation, but also highlight that these studies only serve as an initial step toward determining the underlying causal variant(s) that lead to disease. Results from bioinformatics-based annotation depend on various factors (e.g., queried variants, queried databases, choice of cell lines and tissues, uncertainty in interpreting data from qualitative assays, among others), which vary between studies. It is likely these differences in methodology and interpretation when annotating variants account in part for inconsistencies in results. Consequently, replication of fine-mapping findings is useful, and only targeted functional studies can provide more definitive evidence of SNP function [
<xref rid="pone.0157521.ref022" ref-type="bibr">22</xref>
,
<xref rid="pone.0157521.ref023" ref-type="bibr">23</xref>
].</p>
<p>In our study, for instance, the 500-kb region containing rs6983267 (8q24) harbored 2,257 SNPs. Based on association testing, we narrowed this region to a 13-kb interval that included 7 correlated SNPs showing stronger association signals (Figure A panel A in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
). After bioinformatics analysis, the best functional candidate was the index SNP rs6983267, which was predicted to alter the binding of TCF4 transcription factor. Consistent with this, Tuupanen et al. [
<xref rid="pone.0157521.ref027" ref-type="bibr">27</xref>
] showed
<italic>in vitro</italic>
and
<italic>in vivo</italic>
that rs6983267 resulted in differential TCF4 binding, which may result in enhanced responsiveness to Wnt signaling and a subsequent increase in risk. Further, several other laboratory experiments support the biological importance of this variant in CRC [
<xref rid="pone.0157521.ref064" ref-type="bibr">64</xref>
<xref rid="pone.0157521.ref066" ref-type="bibr">66</xref>
]. Similarly, the 500-kb region containing rs3802842 (11q23) harbored 1,830 SNPs. Association tests narrowed this region to an 18-kb interval that included 9 correlated SNPs for which we performed bioinformatics follow-up (Figure A panel B in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
). Among these, rs7130173 showed strong regulatory evidence in our study. Consistent with these findings, Biancolella et al. [
<xref rid="pone.0157521.ref067" ref-type="bibr">67</xref>
] recently showed that the risk allele of rs7130173 reduced enhancer activity and resulted in reduced transcription factor binding affinity in CRC cells. A combination of fine-mapping and laboratory functional follow-up has also shown similar successes for other cancers and chronic diseases [
<xref rid="pone.0157521.ref023" ref-type="bibr">23</xref>
,
<xref rid="pone.0157521.ref068" ref-type="bibr">68</xref>
,
<xref rid="pone.0157521.ref069" ref-type="bibr">69</xref>
]. Taken together, these data suggest that by combining association testing and bioinformatics analysis, fine-mapping can reduce the size of relevant genomic regions and successfully prioritize candidates for molecular characterization, which greatly reduces the cost, time, and labor associated with testing a large number of variants in the laboratory.</p>
<p>In addition to confirming previous candidates, we suggest several novel candidates with strong functional evidence. These, located in 4 loci with previously reported functional candidates (12q13.13, 14q22.2, 15q13, 20q13.33) and 5 loci without any previously reported candidates (5q31.1, 11q13.4, 12p13.32, 12q24.21, 20p12.3), implicated genes expected to be involved in CRC development as well as those that were unexpected. For instance, duplication in the
<italic>GREM1</italic>
(gremlin 1) promoter has been linked to hereditary mixed polyposis syndrome [
<xref rid="pone.0157521.ref070" ref-type="bibr">70</xref>
], suggesting it is a candidate gene for colorectal tumorigenesis. In our analysis rs2293582, an intronic SNP in
<italic>GREM1</italic>
(15q13), showed the smallest
<italic>P</italic>
-value in this region and was among our best functional candidates. The region containing rs2293582 exhibited open chromatin and bound RNA Polymerase 2
<italic>in vivo</italic>
(ENCODE tracks shown in Figure B in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
). This region also showed strong promoter marks in colon cancer cell lines, but greatly reduced signals in normal colon and rectal tissues. These data suggest rs2293582 warrants experimental follow-up, along with two highly correlated variants within 1-kb, rs2293581 (
<italic>r</italic>
<sup>
<italic>2</italic>
</sup>
= 0.94) and a previously reported candidate rs1406389 (
<italic>r</italic>
<sup>
<italic>2</italic>
</sup>
= 0.94) [
<xref rid="pone.0157521.ref031" ref-type="bibr">31</xref>
], located in regions showing histone marks, open chromatin, and binding to the repressive transcription factor SUZ12 [
<xref rid="pone.0157521.ref071" ref-type="bibr">71</xref>
]. Fine-mapping can also help identify functional candidates that implicate unexpected genes for further functional study.
<italic>LAMA5</italic>
(laminin, alpha 5), for instance, is involved in maintaining the extracellular matrix [
<xref rid="pone.0157521.ref072" ref-type="bibr">72</xref>
], which may not be expected to predict cancer risk. The SNP showing the smallest
<italic>P</italic>
-value in the 20q13.33/
<italic>LAMA5</italic>
region, rs1760073, was completely correlated (
<italic>r</italic>
<sup>
<italic>2</italic>
</sup>
= 1) with the best functional candidate, rs1741634, which was located in an intron of
<italic>LAMA5</italic>
. The region containing rs1741634 exhibited open chromatin, bound the glucocorticoid receptor transcription factor, which has been implicated in cancer [
<xref rid="pone.0157521.ref073" ref-type="bibr">73</xref>
], and interestingly, was located in a region showing different enhancer marks in CRC cell lines vs. normal colon and rectal tissues (Figure C in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
). In addition, Whiffin et al. [
<xref rid="pone.0157521.ref031" ref-type="bibr">31</xref>
] recently reported other functional candidates in this region. Thus, although unexpected, these data, along with those from GWAS showing associations with a variant in another laminin gene,
<italic>LAMC1</italic>
(laminin, gamma 1) [
<xref rid="pone.0157521.ref003" ref-type="bibr">3</xref>
,
<xref rid="pone.0157521.ref019" ref-type="bibr">19</xref>
], support the role of laminin proteins in colorectal carcinogenesis.</p>
<p>Particular strengths of this study included the large study population, high-density genetic data, as well as systematic approach to fine-mapping all GWAS-identified CRC risk variants; however, limitations should be noted. As we aimed to comprehensively investigate both common and less common genetic variation, we examined directly genotyped SNPs as well as SNPs imputed from the 1000 Genomes Project. Imputed genotypes can be called with varying accuracy, and we accounted for this using the genotype dosage, which have been shown to yield unbiased estimates [
<xref rid="pone.0157521.ref041" ref-type="bibr">41</xref>
]. However, lower imputation accuracy may attenuate the estimated significance of association signals [
<xref rid="pone.0157521.ref074" ref-type="bibr">74</xref>
,
<xref rid="pone.0157521.ref075" ref-type="bibr">75</xref>
], and thus relative
<italic>P</italic>
-values for individual variants may not necessarily correspond to their functional importance. Accordingly, rather than only assessing the SNP showing the smallest
<italic>P</italic>
-value in each region we identified a set of SNPs showing stronger association signals for bioinformatics analysis, which enabled us to reduce considerably the number of potential functional SNPs per region and still be able to identify promising functional candidates. Even within our large study, limited statistical power may have accounted for the absence of less common independent association signals at known CRC susceptibility loci, particularly for regions where the initial GWAS showed weak effects. For common genetic variants (allele frequency = 20%), the present analysis had 80% power to detect a per-allele OR of 1.12; for less common variants (allele frequency = 1%), there was 80% power to detect a per-allele OR of 1.51 (Figure D in
<xref ref-type="supplementary-material" rid="pone.0157521.s002">S1 Text</xref>
). These estimates suggest that although larger populations are likely needed to detect weaker associations with less common variants, our data provided sufficient statistical power to detect less common SNPs with larger effect sizes.</p>
<p>In this large population, we comprehensively fine-mapped known common variants that predict CRC risk. We refined genomic regions harboring risk variants and proposed novel functional candidates, as well as confirmed several regions previously reported to contain functional variation. These findings support the utility of a systematic fine-mapping approach that integrates information from expanding publicly available databases to help refine regions surrounding GWAS-identified risk variants and identify a limited number of functional candidates. These insights may help establish a framework for follow-up laboratory studies, which are necessary to yield definitive evidence of functional SNPs that drive common genetic predisposition to CRC.</p>
</sec>
<sec sec-type="supplementary-material" id="sec014">
<title>Supporting Information</title>
<supplementary-material content-type="local-data" id="pone.0157521.s001">
<label>S1 Table</label>
<caption>
<title>Expanded list of SNPs with weak, moderate, or strong evidence of biological function based on bioinformatics (including those in
<xref ref-type="table" rid="pone.0157521.t002">Table 2</xref>
).</title>
<p>(XLSX)</p>
</caption>
<media xlink:href="pone.0157521.s001.xlsx">
<caption>
<p>Click here for additional data file.</p>
</caption>
</media>
</supplementary-material>
<supplementary-material content-type="local-data" id="pone.0157521.s002">
<label>S1 Text</label>
<caption>
<title>Supplementary materials.</title>
<p>Describes in detail the study population and case/control definition; genotyping and quality control; as well as functional annotation using bioinformatics. Also includes Supplementary Tables A-C and Supplementary Figures A-D.</p>
<p>(DOCX)</p>
</caption>
<media xlink:href="pone.0157521.s002.docx">
<caption>
<p>Click here for additional data file.</p>
</caption>
</media>
</supplementary-material>
</sec>
</body>
<back>
<ack>
<p>
<bold>ASTERISK:</bold>
We are very grateful to Dr. Bruno Buecher without whom this project would not have existed. We also thank all those who agreed to participate in this study, including the patients and the healthy control persons, as well as all the physicians, technicians and students.</p>
<p>
<bold>DACHS</bold>
: We thank all participants and cooperating clinicians, and Ute Handte-Daub, Renate Hettler-Jensen, Utz Benscheid, Muhabbet Celik and Ursula Eilber for excellent technical assistance.</p>
<p>
<bold>GECCO</bold>
: The authors would like to thank all those at the GECCO Coordinating Center for helping bring together the data and making this project possible.</p>
<p>
<bold>HPFS, NHS and PHS</bold>
: We would like to acknowledge Patrice Soule and Hardeep Ranu of the Dana Farber Harvard Cancer Center High-Throughput Polymorphism Core who assisted in the genotyping for NHS, HPFS, and PHS under the supervision of Dr. Immaculata De Vivo and Dr. David Hunter, Qin (Carolyn) Guo and Lixue Zhu who assisted in programming for NHS and HPFS, and Haiyan Zhang who assisted in programming for the PHS. We would like to thank the participants and staff of the Nurses' Health Study and the Health Professionals Follow-Up Study, for their valuable contributions as well as the following state cancer registries for their help: AL, AZ, AR, CA, CO, CT, DE, FL, GA, ID, IL, IN, IA, KY, LA, ME, MD, MA, MI, NE, NH, NJ, NY, NC, ND, OH, OK, OR, PA, RI, SC, TN, TX, VA, WA, WY. In addition, this study was approved by the Connecticut Department of Public Health (DPH) Human Investigations Committee. Certain data used in this publication were obtained from the DPH. The authors assume full responsibility for analyses and interpretation of these data.</p>
<p>
<bold>PLCO</bold>
: The authors thank Drs. Christine Berg and Philip Prorok, Division of Cancer Prevention, National Cancer Institute, the Screening Center investigators and staff or the Prostate, Lung, Colorectal, and Ovarian (PLCO) Cancer Screening Trial, Mr. Tom Riley and staff, Information Management Services, Inc., Ms. Barbara O’Brien and staff, Westat, Inc., and Drs. Bill Kopp, Wen Shao, and staff, SAIC-Frederick. Most importantly, we acknowledge the study participants for their contributions to making this study possible.</p>
<p>
<bold>PMH-CCFR</bold>
: The authors would like to thank the study participants and staff of the Hormones and Colon Cancer study.</p>
<p>
<bold>WHI</bold>
: The authors thank the WHI investigators and staff for their dedication, and the study participants for making the program possible. A full listing of WHI investigators can be found at:
<ext-link ext-link-type="uri" xlink:href="https://cleo.whi.org/researchers/Documents%20%20Write%20a%20Paper/WHI%20Investigator%20Short%20List.pdf">https://cleo.whi.org/researchers/Documents%20%20Write%20a%20Paper/WHI%20Investigator%20Short%20List.pdf</ext-link>
</p>
</ack>
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