Prospective evaluation of a multi-gene cfDNA methylation assay for CRC screening in a high-risk cohort

OBJECTIVES: Colorectal cancer (CRC) screening is crucial for early detection, yet the invasiveness of the gold-standard colonoscopy limits compliance. To address the limitation, this study proposes a novel multi-target free DNA methylation model and evaluated its risk stratification performance for CRC in high-risk populations. METHODS: In this prospective study, a cohort of 603 high-risk individuals was enrolled. Following exclusions, 570 participants who underwent successful colonoscopy were analyzed. Their plasma samples were assessed for the methylation status of a six-biomarker panel (Septin9 region 1, BCAT1, IKZF1, BCAN, VAV3, Septin9 region 2). The performance of a model integrating these markers was evaluated against colonoscopy. RESULTS: The combined six-marker model demonstrated superior auxiliary screening performance compared to any single marker. It achieved a high sensitivity of 85.42 % and a negative predictive value (NPV) of 98.56 % for CRC, with a significantly larger AUC of 0.911.The model showed strong performance in early-stage cancers (85.71 % for stage I, 92.86 % for stage II) and for high-grade intraepithelial neoplasia (HGIN) (75.00 %). Specificity exceeded 94 % for distinguishing CRC from non-neoplastic conditions and normal tissue. Compared to conventional tumor markers, the six-marker model achieved significantly higher AUCs (0.911 vs. 0.599-0.711) and superior sensitivity at 90 % fixed specificity (87.5 % vs. 29.2-41.7 %). Multivariable analysis identified the model score as the sole independent predictor of CRC (OR=1.12, p<0.001). CONCLUSIONS: The results demonstrate the clinical potential of a multi-gene cfDNA methylation-based approach to augment CRC screening strategies for high-risk individuals.