DNA methylation profiling of cervical squamous intraepithelial lesions using liquid‐based cytology specimens

BACKGROUND Cervical carcinoma is a common malignancy among women worldwide, and its pathogenesis is related causally to human papillomavirus infection. The progression from precursor squamous intraepithelial lesions to cervical carcinoma requires additional genetic and epigenetic alterations that have not been characterized fully. The authors examined aberrant promoter methylation of multiple tumor suppressor genes in precursor squamous intraepithelial lesions. METHODS A multiplex, nested, methylation‐specific polymerase chain reaction approach was used to examine promoter methylation of 15 tumor suppressor genes in high‐grade squamous intraepithelial lesions (HSIL, n = 11), low‐grade squamous intraepithelial lesions (LSIL, n = 17), and negative tissues (n = 11) from liquid‐based cytology samples. The area under the receiver‐operating characteristic (ROC) curve was determined for individual methylated tumor suppressor genes and for gene combinations to evaluate test performance for the ability of methylation profiles to distinguish HSIL cytology samples from combined LSIL/negative cytology samples. RESULTS Aberrant promoter methylation of DAPK1 and IGSF4 occurred at a high frequency in HSIL samples and was absent in LSIL and negative samples. There was a significant trend toward increased methylation with the increased severity of lesions, and the mean number of methylated genes was significantly higher in HSIL samples compared with LSIL and negative samples. Using the area under the ROC curve as a measure of test performance, the methylation of IGSF4 and DAPK1 had areas that were significantly greater than 0.5; thus, each had the ability to distinguish HSIL samples from combined LSIL/negative samples. The areas under the curve for the best two‐gene combination (IGSF4/DAPK1) and the best three‐gene combination (IGSF4/DAPK1/HIC1) were not statistically different from the best individual tumor suppressor gene (IGSF4) in distinguishing HSIL samples from combined LSIL/negative samples. CONCLUSIONS Aberrant promoter methylation of tumor suppressor genes is an epigenetic alteration that occurs during neoplastic progression to cervical carcinoma. The methylation status of multiple tumor suppressor genes can be evaluated using ROC analysis to determine methylation profiles that can distinguish HSIL samples from combined LSIL/negative samples. Cancer (Cancer Cytopathol) 2004. © 2004 American Cancer Society. DNA methylation profiling of multiple tumor suppressor gen