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Purpose  Colonoscopy and the fecal immunochemical test (FIT) are currently the most widely used screening modalities for colorectal cancer (CRC), however, both with their own limitations. Here we aim to identify and validate stool-based DNA methylation markers for the early detection of CRC and investigate the biological pathways prone to DNA methylation.    Methods  DNA methylation marker discovery was performed using The Cancer Genome Atlas (TCGA) colon adenocarcinoma data set consisting of normal and primary colon adenocarcinoma tissue. The performance of the five best candidate markers and a previously identified marker,  NDRG4 , was evaluated on tissues and whole stool samples of healthy subjects and CRC patients using quantitative MSP assays. The results were compared and combined with FIT data. Finally, pathway and gene ontology enrichment analyses were performed using ToppFun, GOrilla and clusterProfiler.    Results   GDNF, HAND2, SLC35F3, SNAP91  and  SORCS1  were ranked as the best performing markers. Gene combinations of all five markers,  NDRG4  and FIT were evaluated to establish the biomarker panel with the highest diagnostic potential, resulting in the identification of  GDNF/SNAP91/NDRG4/FIT  as the best performing marker panel. Pathway and gene ontology enrichment analyses revealed that genes associated with the nervous system were enriched in the set of best performing CRC-specific biomarkers.    Conclusion  In silico discovery analysis using TCGA-derived data yielded a novel DNA-methylation-based assay for the early detection of CRC, potentially improving current screening modalities. Additionally, nervous system-related pathways were enriched in the identified genes, indicating an epigenetic regulation of neuronal genes in CRC.    Supplementary Information  The online version contains supplementary material available at 10.1186/s13148-021-01067-9.

Identification of DNA methylation markers for early detection of CRC indicates a role for nervous system-related genes in CRC