Discovery of circulating cell-free DNA 5hmC  biomarkers for peritoneal metastasis in colorectal and appendiceal cancer
Peritoneal metastasis (PM) is associated with a poor prognosis in patients with colorectal cancer (CRC) or appendiceal adenocarcinoma (AA). Detection of PM remains challenging due to the low sensitivity of current imaging modalities and the invasiveness of tissue pathology-based diagnostics. Here, we demonstrate for the first time the feasibility of using 5-hydroxymethylcytosine (5hmC) signatures derived from circulating cell-free DNA (cfDNA) as minimally biomarker s for PM. Using nano-hmC-Seal on plasma-derived cfDNA and next-generation sequencing, we obtained genome-wide 5hmC profiles from 112 CRC or AA patients   (n = 71 PM+ and n = 41 PM–) and 73 non-cancer controls. Predictive models trained on genomic region  5hmC levels distinguished PM status with an area under the curve of 0.827  (92.4% sensitivity, 46.1% specificity). Pathway enrichment analysis identified epigenetically dysregulated cancer, cell migration, adhesion and immune-related pathways in PM. This study lays a foundation for future clinical assay development for PM. Overall design: Case-control study: Differential 5hmC features were identified by comparing PM+ patients, PM- patients, and non-cancer controls Access to demographic information requires contacting the corresponding author and will only be provided upon reasonable request.