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 < or = .0028). The combination of 4 genes showed improved test performance (AUC = 0.76; P < .0001). There was no significant difference in cumulative methylation in HSIL cases with histologic outcomes of cervical intraepithelial neoplasia grade 2 (CIN2) versus CIN3. There was no association between the methylation of any gene and the presence of human papillomavirus. CONCLUSIONS: The methylation profile of multiple genes in combination can better distinguish HSIL from combined NILM/LSIL samples. Although aberrant DNA methylation has the potential to function as a molecular biomarker of HSIL in liquid-based Pap tests, additional genes that are selectively methylated in HSIL are needed to improve the clinical performance.