A 5-hydroxymethylcytosine-based noninvasive model for ultrasensitive early detection of colorectal advanced adenoma and adenocarcinoma: The METHOD-2 study.

Abstract

3042 Background: Detection of colorectal tumors at a time when there are more treatment options is associated with better outcomes. This prospective case-control study (NCT03676075) assessed the performance of 5-hydroxymethycytosine (5hmC) biomarkers in circulating cell-free DNA (cfDNA) to detect advanced colorectal adenomas (ADA) or adenocarcinomas (CRC). Methods: This multi-center study enrolled 2,576 participants, including patients with newly diagnosed CRC (n=1,074), ADA (n=356), other solid tumors (n=80), and healthy individuals (HEA) (n=1,066), followed by genome-wide 5hmC profiling using the 5hmC-Seal, a highly sensitive chemical labeling approach. A weighted diagnostic model for CRC (stage I-III) and ADA was developed using the elastic net regularization in a training set of age-, sex-, region-matched samples and validated in independent samples. Results: Distribution of 5hmC in cfDNA reflected gene regulatory relevance and tissue of origin. In an independent validation set of 482 samples, a 96-gene model trained in stage I-III CRC, ADA, and HEA achieved an AUC (area under the curve) of 92.6% (95% CI [confidence interval], 90.7-94.5%) for distinguishing CRC from HEA, regardless of primary location or mutation. This model also showed high capacity for distinguishing ADA from HEA with an AUC of 87.9% as well as cancer-type specificity. Model performance was further confirmed in samples from multiple centers. Functionally, differential 5hmC features associated with CRC and ADA demonstrated relevance to CRC biology, including pathways such as calcium and MAPK signaling. Conclusions: Genome-wide mapping of 5hmC in cfDNA shows promise as a highly sensitive and specific approach to be integrated in a screening program for improving early detection of CRC and high-risk ADA. Clinical trial information: NCT03676075 .