Quantitative methylation-specific PCR for the detection of aberrant DNA methylation in liquid-based pap tests

Steven L. Kahn, Brigitte Maria Ronnett, Patti E. Gravitt, Karen S. Gustafson

Research output: Contribution to journalArticle

Abstract

BACKGROUND. Aberrant promoter methylation of selective tumor suppressor genes has been detected in squamous intraepithelial lesions (SIL) and invasive cervical cancer. Identification of methylation profiles of genes that can distinguish high-grade SIL (HSIL) from low-grade SIL (LSIL), and cytologically negative for intraepithelial lesion or malignancy (NILM) residual liquid-based Papanicolaou (Pap) tests may be potentially useful as an ancillary test for cervical cancer screening. METHODS. Using real-time quantitative methylation-specific polymerase chain reaction (PCR) (QMSP), the authors analyzed the frequency and relative level of promoter methylation for DAPK1, IGSF4, SPARC, and TFPI2 in biopsy-confirmed HSIL and LSIL, and NILM residual liquid-based Pap tests. The percentage of methylation (%M) for each gene was calculated using the reference gene, ACTB. The cumulative methylation score for each sample, defined as the sum of %M of all 4 genes, was used to analyze the genes in combination. RESULTS. For each gene analyzed the frequency and relative level of methylation were increased in HSIL compared with combined NILM/LSIL samples. The cumulative methylation scores were significantly higher in HSIL samples (P <.0001). Area under the receiver operating characteristic (ROC) curve (AUC) demonstrated that methylation of each gene could distinguish HSIL from NILM/LSIL samples (AUC range, 0.6-0.67; P ≤ .0028). The combination of 4 genes showed improved test performance (AUC = 0.76; P

Original languageEnglish (US)
Pages (from-to)57-64
Number of pages8
JournalCancer
Volume114
Issue number1
DOIs
StatePublished - Feb 25 2008

Fingerprint

Papanicolaou Test
DNA Methylation
Methylation
Polymerase Chain Reaction
Genes
Area Under Curve
Uterine Cervical Neoplasms
Neoplasms
Tumor Suppressor Genes
Early Detection of Cancer
Gene Frequency
ROC Curve
Biopsy

Keywords

  • Biomarker
  • HSIL
  • Methylation
  • Quantitative MSP

ASJC Scopus subject areas

  • Cancer Research
  • Oncology

Cite this

Quantitative methylation-specific PCR for the detection of aberrant DNA methylation in liquid-based pap tests. / Kahn, Steven L.; Ronnett, Brigitte Maria; Gravitt, Patti E.; Gustafson, Karen S.

In: Cancer, Vol. 114, No. 1, 25.02.2008, p. 57-64.

Research output: Contribution to journalArticle

Kahn, Steven L. ; Ronnett, Brigitte Maria ; Gravitt, Patti E. ; Gustafson, Karen S. / Quantitative methylation-specific PCR for the detection of aberrant DNA methylation in liquid-based pap tests. In: Cancer. 2008 ; Vol. 114, No. 1. pp. 57-64.
@article{665bb7ed15dd489ba34a92fec6bff490,
title = "Quantitative methylation-specific PCR for the detection of aberrant DNA methylation in liquid-based pap tests",
abstract = "BACKGROUND. Aberrant promoter methylation of selective tumor suppressor genes has been detected in squamous intraepithelial lesions (SIL) and invasive cervical cancer. Identification of methylation profiles of genes that can distinguish high-grade SIL (HSIL) from low-grade SIL (LSIL), and cytologically negative for intraepithelial lesion or malignancy (NILM) residual liquid-based Papanicolaou (Pap) tests may be potentially useful as an ancillary test for cervical cancer screening. METHODS. Using real-time quantitative methylation-specific polymerase chain reaction (PCR) (QMSP), the authors analyzed the frequency and relative level of promoter methylation for DAPK1, IGSF4, SPARC, and TFPI2 in biopsy-confirmed HSIL and LSIL, and NILM residual liquid-based Pap tests. The percentage of methylation ({\%}M) for each gene was calculated using the reference gene, ACTB. The cumulative methylation score for each sample, defined as the sum of {\%}M of all 4 genes, was used to analyze the genes in combination. RESULTS. For each gene analyzed the frequency and relative level of methylation were increased in HSIL compared with combined NILM/LSIL samples. The cumulative methylation scores were significantly higher in HSIL samples (P <.0001). Area under the receiver operating characteristic (ROC) curve (AUC) demonstrated that methylation of each gene could distinguish HSIL from NILM/LSIL samples (AUC range, 0.6-0.67; P ≤ .0028). The combination of 4 genes showed improved test performance (AUC = 0.76; P",
keywords = "Biomarker, HSIL, Methylation, Quantitative MSP",
author = "Kahn, {Steven L.} and Ronnett, {Brigitte Maria} and Gravitt, {Patti E.} and Gustafson, {Karen S.}",
year = "2008",
month = "2",
day = "25",
doi = "10.1002/cncr.23258",
language = "English (US)",
volume = "114",
pages = "57--64",
journal = "Cancer",
issn = "0008-543X",
publisher = "John Wiley and Sons Inc.",
number = "1",

}

TY - JOUR

T1 - Quantitative methylation-specific PCR for the detection of aberrant DNA methylation in liquid-based pap tests

AU - Kahn, Steven L.

AU - Ronnett, Brigitte Maria

AU - Gravitt, Patti E.

AU - Gustafson, Karen S.

PY - 2008/2/25

Y1 - 2008/2/25

N2 - BACKGROUND. Aberrant promoter methylation of selective tumor suppressor genes has been detected in squamous intraepithelial lesions (SIL) and invasive cervical cancer. Identification of methylation profiles of genes that can distinguish high-grade SIL (HSIL) from low-grade SIL (LSIL), and cytologically negative for intraepithelial lesion or malignancy (NILM) residual liquid-based Papanicolaou (Pap) tests may be potentially useful as an ancillary test for cervical cancer screening. METHODS. Using real-time quantitative methylation-specific polymerase chain reaction (PCR) (QMSP), the authors analyzed the frequency and relative level of promoter methylation for DAPK1, IGSF4, SPARC, and TFPI2 in biopsy-confirmed HSIL and LSIL, and NILM residual liquid-based Pap tests. The percentage of methylation (%M) for each gene was calculated using the reference gene, ACTB. The cumulative methylation score for each sample, defined as the sum of %M of all 4 genes, was used to analyze the genes in combination. RESULTS. For each gene analyzed the frequency and relative level of methylation were increased in HSIL compared with combined NILM/LSIL samples. The cumulative methylation scores were significantly higher in HSIL samples (P <.0001). Area under the receiver operating characteristic (ROC) curve (AUC) demonstrated that methylation of each gene could distinguish HSIL from NILM/LSIL samples (AUC range, 0.6-0.67; P ≤ .0028). The combination of 4 genes showed improved test performance (AUC = 0.76; P

AB - BACKGROUND. Aberrant promoter methylation of selective tumor suppressor genes has been detected in squamous intraepithelial lesions (SIL) and invasive cervical cancer. Identification of methylation profiles of genes that can distinguish high-grade SIL (HSIL) from low-grade SIL (LSIL), and cytologically negative for intraepithelial lesion or malignancy (NILM) residual liquid-based Papanicolaou (Pap) tests may be potentially useful as an ancillary test for cervical cancer screening. METHODS. Using real-time quantitative methylation-specific polymerase chain reaction (PCR) (QMSP), the authors analyzed the frequency and relative level of promoter methylation for DAPK1, IGSF4, SPARC, and TFPI2 in biopsy-confirmed HSIL and LSIL, and NILM residual liquid-based Pap tests. The percentage of methylation (%M) for each gene was calculated using the reference gene, ACTB. The cumulative methylation score for each sample, defined as the sum of %M of all 4 genes, was used to analyze the genes in combination. RESULTS. For each gene analyzed the frequency and relative level of methylation were increased in HSIL compared with combined NILM/LSIL samples. The cumulative methylation scores were significantly higher in HSIL samples (P <.0001). Area under the receiver operating characteristic (ROC) curve (AUC) demonstrated that methylation of each gene could distinguish HSIL from NILM/LSIL samples (AUC range, 0.6-0.67; P ≤ .0028). The combination of 4 genes showed improved test performance (AUC = 0.76; P

KW - Biomarker

KW - HSIL

KW - Methylation

KW - Quantitative MSP

UR - http://www.scopus.com/inward/record.url?scp=39749114948&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=39749114948&partnerID=8YFLogxK

U2 - 10.1002/cncr.23258

DO - 10.1002/cncr.23258

M3 - Article

VL - 114

SP - 57

EP - 64

JO - Cancer

JF - Cancer

SN - 0008-543X

IS - 1

ER -