Analysis of DNA Methylation at Specific Loci in Stool Samples Detects Colorectal Cancer and High-Grade Dysplasia in Patients With Inflammatory Bowel Disease

John B Kisiel, Pasquale Klepp, Hatim T. Allawi, William R. Taylor, Maria Giakoumopoulos, Tamara Sander, Tracy C. Yab, Bjorn A. Moum, Graham P. Lidgard, Stephan Brackmann, Douglas W. Mahoney, Arne Roseth, David A. Ahlquist

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Abstract

Background & Aims: Patients with inflammatory bowel diseases (IBDs), including ulcerative colitis and Crohn's disease, are at increased risk for colorectal cancer (CRC). Analyses of DNA methylation patterns in stool samples have been reported to detect CRC in patients with IBD. We sought to validate these findings in larger cohorts and assess the accuracy of analysis of DNA methylation patterns in stool for detection of CRC and high-grade dysplasia (HGD) normalized to methylation level at ZDHHC1. Methods: We obtained buffered, frozen stool samples from a US case–control study and from 2 European surveillance cohorts (referral or population based) of patients with chronic ulcerative colitis (n = 248), Crohn's disease (n = 82), indeterminate colitis (n = 2), or IBD with primary sclerosing cholangitis (n = 38). Stool samples were collected before bowel preparation for colonoscopy or at least 1 week after colonoscopy. Among the study samples, stools from individuals with IBD but without neoplasia were used as controls (n = 291). DNA was isolated from stool, exposed to bisulfite, and then assayed by multiplex quantitative allele-specific real-time target and signal amplification. We analyzed methylation levels of BMP3, NDRG4, VAV3, and SFMBT2 relative to the methylation level of ZDHHC1, and compared these between patients with CRC or HGD and controls. Results: Levels of methylation at BMP3 and VAV3, relative to ZDHHC1 methylation, identified patients with CRC and HGD with an area under the curve value of 0.91 (95% CI, 0.77–1.00). Methylation levels at specific promotor regions of these genes identified 11 of the 12 patients with CRC and HGD, with 92% sensitivity (95% CI, 60%–100%) and 90% specificity (95% CI, 86%–93%). The proportion of false-positive results did not differ significantly among the case–control, referral cohort, and population cohort studies (P =.60) when the 90% specificity cut-off from the whole sample set was applied. Conclusions: In an analysis of stool samples from 3 independent studies of 332 patients with IBD, we associated levels of methylation at 2 genes (BMP3 and VAV3), relative to level of methylation at ZDHHC1, with detection of CRC and HGD. These methylation patterns identified patients with CRC and HGD with more than 90% specificity, and might be used in CRC surveillance.

Original languageEnglish (US)
JournalClinical Gastroenterology and Hepatology
DOIs
StateAccepted/In press - Jan 1 2019

Fingerprint

DNA Methylation
Inflammatory Bowel Diseases
Methylation
Colorectal Neoplasms
Colonoscopy
Ulcerative Colitis
Crohn Disease
Referral and Consultation
Sclerosing Cholangitis
Colitis
Genetic Promoter Regions
Population
Genes
Area Under Curve
Cohort Studies
Alleles
DNA

Keywords

  • Biomarker
  • Complications
  • Neoplasm
  • Noninvasive Test
  • Prevention

ASJC Scopus subject areas

  • Hepatology
  • Gastroenterology

Cite this

Analysis of DNA Methylation at Specific Loci in Stool Samples Detects Colorectal Cancer and High-Grade Dysplasia in Patients With Inflammatory Bowel Disease. / Kisiel, John B; Klepp, Pasquale; Allawi, Hatim T.; Taylor, William R.; Giakoumopoulos, Maria; Sander, Tamara; Yab, Tracy C.; Moum, Bjorn A.; Lidgard, Graham P.; Brackmann, Stephan; Mahoney, Douglas W.; Roseth, Arne; Ahlquist, David A.

In: Clinical Gastroenterology and Hepatology, 01.01.2019.

Research output: Contribution to journalArticle

Kisiel, John B ; Klepp, Pasquale ; Allawi, Hatim T. ; Taylor, William R. ; Giakoumopoulos, Maria ; Sander, Tamara ; Yab, Tracy C. ; Moum, Bjorn A. ; Lidgard, Graham P. ; Brackmann, Stephan ; Mahoney, Douglas W. ; Roseth, Arne ; Ahlquist, David A. / Analysis of DNA Methylation at Specific Loci in Stool Samples Detects Colorectal Cancer and High-Grade Dysplasia in Patients With Inflammatory Bowel Disease. In: Clinical Gastroenterology and Hepatology. 2019.
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title = "Analysis of DNA Methylation at Specific Loci in Stool Samples Detects Colorectal Cancer and High-Grade Dysplasia in Patients With Inflammatory Bowel Disease",
abstract = "Background & Aims: Patients with inflammatory bowel diseases (IBDs), including ulcerative colitis and Crohn's disease, are at increased risk for colorectal cancer (CRC). Analyses of DNA methylation patterns in stool samples have been reported to detect CRC in patients with IBD. We sought to validate these findings in larger cohorts and assess the accuracy of analysis of DNA methylation patterns in stool for detection of CRC and high-grade dysplasia (HGD) normalized to methylation level at ZDHHC1. Methods: We obtained buffered, frozen stool samples from a US case–control study and from 2 European surveillance cohorts (referral or population based) of patients with chronic ulcerative colitis (n = 248), Crohn's disease (n = 82), indeterminate colitis (n = 2), or IBD with primary sclerosing cholangitis (n = 38). Stool samples were collected before bowel preparation for colonoscopy or at least 1 week after colonoscopy. Among the study samples, stools from individuals with IBD but without neoplasia were used as controls (n = 291). DNA was isolated from stool, exposed to bisulfite, and then assayed by multiplex quantitative allele-specific real-time target and signal amplification. We analyzed methylation levels of BMP3, NDRG4, VAV3, and SFMBT2 relative to the methylation level of ZDHHC1, and compared these between patients with CRC or HGD and controls. Results: Levels of methylation at BMP3 and VAV3, relative to ZDHHC1 methylation, identified patients with CRC and HGD with an area under the curve value of 0.91 (95{\%} CI, 0.77–1.00). Methylation levels at specific promotor regions of these genes identified 11 of the 12 patients with CRC and HGD, with 92{\%} sensitivity (95{\%} CI, 60{\%}–100{\%}) and 90{\%} specificity (95{\%} CI, 86{\%}–93{\%}). The proportion of false-positive results did not differ significantly among the case–control, referral cohort, and population cohort studies (P =.60) when the 90{\%} specificity cut-off from the whole sample set was applied. Conclusions: In an analysis of stool samples from 3 independent studies of 332 patients with IBD, we associated levels of methylation at 2 genes (BMP3 and VAV3), relative to level of methylation at ZDHHC1, with detection of CRC and HGD. These methylation patterns identified patients with CRC and HGD with more than 90{\%} specificity, and might be used in CRC surveillance.",
keywords = "Biomarker, Complications, Neoplasm, Noninvasive Test, Prevention",
author = "Kisiel, {John B} and Pasquale Klepp and Allawi, {Hatim T.} and Taylor, {William R.} and Maria Giakoumopoulos and Tamara Sander and Yab, {Tracy C.} and Moum, {Bjorn A.} and Lidgard, {Graham P.} and Stephan Brackmann and Mahoney, {Douglas W.} and Arne Roseth and Ahlquist, {David A.}",
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month = "1",
day = "1",
doi = "10.1016/j.cgh.2018.05.004",
language = "English (US)",
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TY - JOUR

T1 - Analysis of DNA Methylation at Specific Loci in Stool Samples Detects Colorectal Cancer and High-Grade Dysplasia in Patients With Inflammatory Bowel Disease

AU - Kisiel, John B

AU - Klepp, Pasquale

AU - Allawi, Hatim T.

AU - Taylor, William R.

AU - Giakoumopoulos, Maria

AU - Sander, Tamara

AU - Yab, Tracy C.

AU - Moum, Bjorn A.

AU - Lidgard, Graham P.

AU - Brackmann, Stephan

AU - Mahoney, Douglas W.

AU - Roseth, Arne

AU - Ahlquist, David A.

PY - 2019/1/1

Y1 - 2019/1/1

N2 - Background & Aims: Patients with inflammatory bowel diseases (IBDs), including ulcerative colitis and Crohn's disease, are at increased risk for colorectal cancer (CRC). Analyses of DNA methylation patterns in stool samples have been reported to detect CRC in patients with IBD. We sought to validate these findings in larger cohorts and assess the accuracy of analysis of DNA methylation patterns in stool for detection of CRC and high-grade dysplasia (HGD) normalized to methylation level at ZDHHC1. Methods: We obtained buffered, frozen stool samples from a US case–control study and from 2 European surveillance cohorts (referral or population based) of patients with chronic ulcerative colitis (n = 248), Crohn's disease (n = 82), indeterminate colitis (n = 2), or IBD with primary sclerosing cholangitis (n = 38). Stool samples were collected before bowel preparation for colonoscopy or at least 1 week after colonoscopy. Among the study samples, stools from individuals with IBD but without neoplasia were used as controls (n = 291). DNA was isolated from stool, exposed to bisulfite, and then assayed by multiplex quantitative allele-specific real-time target and signal amplification. We analyzed methylation levels of BMP3, NDRG4, VAV3, and SFMBT2 relative to the methylation level of ZDHHC1, and compared these between patients with CRC or HGD and controls. Results: Levels of methylation at BMP3 and VAV3, relative to ZDHHC1 methylation, identified patients with CRC and HGD with an area under the curve value of 0.91 (95% CI, 0.77–1.00). Methylation levels at specific promotor regions of these genes identified 11 of the 12 patients with CRC and HGD, with 92% sensitivity (95% CI, 60%–100%) and 90% specificity (95% CI, 86%–93%). The proportion of false-positive results did not differ significantly among the case–control, referral cohort, and population cohort studies (P =.60) when the 90% specificity cut-off from the whole sample set was applied. Conclusions: In an analysis of stool samples from 3 independent studies of 332 patients with IBD, we associated levels of methylation at 2 genes (BMP3 and VAV3), relative to level of methylation at ZDHHC1, with detection of CRC and HGD. These methylation patterns identified patients with CRC and HGD with more than 90% specificity, and might be used in CRC surveillance.

AB - Background & Aims: Patients with inflammatory bowel diseases (IBDs), including ulcerative colitis and Crohn's disease, are at increased risk for colorectal cancer (CRC). Analyses of DNA methylation patterns in stool samples have been reported to detect CRC in patients with IBD. We sought to validate these findings in larger cohorts and assess the accuracy of analysis of DNA methylation patterns in stool for detection of CRC and high-grade dysplasia (HGD) normalized to methylation level at ZDHHC1. Methods: We obtained buffered, frozen stool samples from a US case–control study and from 2 European surveillance cohorts (referral or population based) of patients with chronic ulcerative colitis (n = 248), Crohn's disease (n = 82), indeterminate colitis (n = 2), or IBD with primary sclerosing cholangitis (n = 38). Stool samples were collected before bowel preparation for colonoscopy or at least 1 week after colonoscopy. Among the study samples, stools from individuals with IBD but without neoplasia were used as controls (n = 291). DNA was isolated from stool, exposed to bisulfite, and then assayed by multiplex quantitative allele-specific real-time target and signal amplification. We analyzed methylation levels of BMP3, NDRG4, VAV3, and SFMBT2 relative to the methylation level of ZDHHC1, and compared these between patients with CRC or HGD and controls. Results: Levels of methylation at BMP3 and VAV3, relative to ZDHHC1 methylation, identified patients with CRC and HGD with an area under the curve value of 0.91 (95% CI, 0.77–1.00). Methylation levels at specific promotor regions of these genes identified 11 of the 12 patients with CRC and HGD, with 92% sensitivity (95% CI, 60%–100%) and 90% specificity (95% CI, 86%–93%). The proportion of false-positive results did not differ significantly among the case–control, referral cohort, and population cohort studies (P =.60) when the 90% specificity cut-off from the whole sample set was applied. Conclusions: In an analysis of stool samples from 3 independent studies of 332 patients with IBD, we associated levels of methylation at 2 genes (BMP3 and VAV3), relative to level of methylation at ZDHHC1, with detection of CRC and HGD. These methylation patterns identified patients with CRC and HGD with more than 90% specificity, and might be used in CRC surveillance.

KW - Biomarker

KW - Complications

KW - Neoplasm

KW - Noninvasive Test

KW - Prevention

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U2 - 10.1016/j.cgh.2018.05.004

DO - 10.1016/j.cgh.2018.05.004

M3 - Article

C2 - 29775793

AN - SCOPUS:85052205637

JO - Clinical Gastroenterology and Hepatology

JF - Clinical Gastroenterology and Hepatology

SN - 1542-3565

ER -