Triage Performance of <i>FAM19A4/miR124‐2</i> Methylation: 18‐Month Follow‐Up Results From a Prospective Observational Study Within the Dutch Primary <scp>HPV</scp> ‐Based Cervical Screening Program

DNA methylation analysis of the genes FAM19A4 and miR124-2 has emerged as a promising triage strategy for high-risk (hr) human papillomavirus (HPV)-positive women in cervical screening. This study reports a prospective evaluation of the diagnostic accuracy of FAM19A4/miR124-2 methylation in a large population-based primary HPV-based screening cohort with 18 months of follow-up. In this prospective observational study, clinician-collected cervical samples from 3848 consecutive hrHPV-positive women who participated in the Dutch national primary HPV-based screening program were tested with the QIAsure Methylation Test, a quantitative methylation-specific PCR assessing FAM19A4 and miR124-2 methylation. Through linkage to the Dutch Nationwide Pathology Databank (Palga), 11 cases of cervical carcinoma, 293 cervical intraepithelial neoplasia grade 3 (CIN3), and 29 adenocarcinoma in situ (AIS) were identified during 18 months of follow-up. Clinical performance for CIN3, AIS, and cancer (CIN3+) detection was assessed based on (I) the threshold of the QIAsure Methylation Test, and (II) thresholds corresponding with clinical specificities of 70% and 80% in this hrHPV-positive study population. The sensitivity and specificity of FAM19A4/miR124-2 methylation for CIN3+ detection using the QIAsure Methylation Test threshold (I) were 57.4% (95% CI: 52.0-62.7) and 88.3% (95% CI: 87.2-89.5), respectively. At predefined specificities of 70% and 80%, FAM19A4/miR124-2 methylation (II) demonstrated a CIN3+ sensitivity of 75.4% (95% CI: 70.7-80.0) and 66.1% (95% CI: 61.0-71.2), respectively. In conclusion, FAM19A4/miR124-2 methylation analysis showed good triage performance in a primary HPV-based screening setting. These findings support the use of methylation-based testing as a triage strategy in primary HPV-based screening programs, with an appropriate follow-up policy for hrHPV-positive women who test methylation-negative.