Establishment of a Methodology for <scp>PAX1</scp> and <scp>CADM1</scp> Methylation Detection and Preliminary Assessment of Its Clinical Value for Cervical Cancer Diagnosis

Cervical cancer is a common malignant tumor that poses a serious threat to women's health. Early diagnosis of cervical cancer can effectively improve the prognosis. However, traditional cytological screening methods exhibit certain limitations. Utilizing bisulfite conversion and Sanger sequencing techniques, we analyzed the promoter regions and identified specific methylated sites of the PAX1 and CADM1 genes as diagnostic sites. Subsequently, a multiplex fluorescent PCR system for these sites was established. The assay's sensitivity and specificity were assessed in a cohort of participants with confirmed pathological diagnoses. Systematic screening identified candidate diagnostic sites at position 5 of the PAX1 promoter and position 3 of the CADM1 promoter. The PAX1 and CADM1 methylation-specific quantitative PCR assay demonstrated detection sensitivities of 82 copies/μL and 134 copies/μL of methylated DNA, respectively. Validation in a cohort of 144 cervical samples revealed that PAX1 had a sensitivity of 60.9% and a specificity of 79.6%, whereas CADM1 demonstrated a sensitivity of 67.4% and a specificity of 88.8%. Under the combined testing strategy, parallel testing (defined as a positive result for either gene) significantly improved the sensitivity to 80.4%, while serial testing (defined as positive results for both genes) increased the specificity to 94.9%. The specific methylated sites within the PAX1 and CADM1 promoter regions were associated with cervical cancer. The combined PAX-CADM1 dual-gene methylation assay enhances diagnostic sensitivity and specificity, offering a novel strategy for the clinical diagnosis of cervical cancer.