Combined Promoter Methylation Analysis of CADM1 and MAL: An Objective Triage Tool for High-Risk Human Papillomavirus DNA–Positive Women

PURPOSE: Screening women for high-grade cervical intraepithelial neoplasia or cervical cancer (CIN3(+)) by high-risk human papillomavirus (hrHPV) testing has as side-effect the detection of hrHPV-positive women without clinically relevant lesions. Here, we developed an objective assay assessing the methylation status of the promoter regions of CADM1 and MAL to triage hrHPV-positive women for CIN3(+). EXPERIMENTAL DESIGN: In a training set (51 women with CIN3(+) and 224 without CIN2(+)), panels consisting of one to four quantitative methylation-specific PCR (qMSP) assays (CADM1-m12,CADM1-m18,MAL-m1,MAL-m2) were analyzed. Cross-validated receiver-operating characteristics (ROC) curves were constructed and the panel with highest partial cross-validated area under the curve (AUC) was used for validation in an independent set of 236 consecutive hrHPV-positive women from a screening cohort. In the validation set, the ROC curve of the panel was compared with CIN3(+) sensitivity and specificity of cytology and of cytology combined with HPV16/18 genotyping. RESULTS: In the training set, CADM1-m18 combined with MAL-m1 was the best panel (cross-validated partial AUC = 0.719). In the validation set, this panel revealed CIN3(+) sensitivities ranging from 100% (95% CI: 92.4-100) to 60.5% (95% CI: 47.1-74.6), with corresponding specificities ranging from 22.7% (95% CI: 20.2-25.2) to 83.3% (95% CI: 78.4-87.4). For cytology these were 65.8% (95% CI: 52.3-79.0) and 78.8% (95% CI: 73.7-83.1) and for cytology/HPV16/18, these were 84.2% (95% CI: 72.0-92.7) and 54.0% (95% CI: 49.2-58.7), respectively. The point estimates of both cytology and cytology/HPV16/18 were equal to the values of the ROC curve of CADM1-m18/MAL-m1. CONCLUSIONS: We developed an objective methylation marker panel that was equally discriminatory for CIN3(+) as cytology or cytology with HPV16/18 genotyping in hrHPV-positive women. This opens the possibility for complete cervical screening by objective, nonmorphological molecular methods.