Identification of DNA methylation differences in colon mucosa deriving from healthy controls and patients with colon polyps

Colorectal cancer develops through a multistep adenoma–carcinoma sequence driven by early genetic and epigenetic alterations. While advanced Colorectal cancer has been extensively characterized, less is known about molecular changes occurring in benign colorectal polyps. This study investigated early DNA methylation alterations and oncogenic mutation status in colorectal polyps compared with healthy colon mucosa to identify potential early biomarkers of colorectal tumorigenesis. Colon mucosa samples were collected from 21 patients with colorectal polyps and 20 control individuals undergoing colonoscopy at Sahlgrenska University Hospital. DNA methylation levels of LINE-1, MLH1, MGMT, and CDKN2A (p16INK4a) were quantified using bisulfite pyrosequencing. KRAS and BRAF mutation status were assessed by digital polymerase chain reaction. Associations between methylation patterns, mutation status, and clinical variables were evaluated, and the diagnostic performance of LINE-1 methylation was assessed using receiver operating characteristic analysis. Global LINE-1 methylation was significantly reduced in polyp-adjacent mucosa compared with control mucosa and demonstrated strong discriminatory power between groups (AUC = 0.863). In contrast, no aberrant hypermethylation of MLH1 or MGMT was observed in polyp-adjacent mucosa, and CDKN2A (p16INK4a) methylation was significantly lower in polyps than in controls. MLH1 methylation remained largely absent, consistent with the lack of BRAF mutations and microsatellite instability–associated features in this cohort. No activating KRAS or BRAF mutations were detected. The CDKN2A (p16INK4a) rs3814960 variant showed genotype-dependent methylation patterns but was not an independent predictor after adjustment for clinical variables.These findings indicate that global DNA hypomethylation precedes tumor suppressor gene hypermethylation in early colorectal lesions and highlight LINE-1 hypomethylation as a promising early epigenetic biomarker in colorectal tumorigenesis.