DNA methylation-based testing in peripheral blood mononuclear cells enables accurate and early detection of colorectal cancer.

Abstract

An effective blood-based method for the diagnosis of colorectal cancer (CRC) has not yet been developed. Molecular alterations of immune cells occur early in tumorigenesis, providing the theoretical underpinning for early cancer diagnosis based on immune cell profiling. Therefore, we aimed to develop an effective detection method based on peripheral blood mononuclear cells (PBMCs) to improve the diagnosis of CRC. Analysis of the genome-wide methylation landscape of PBMCs from CRC patients and healthy controls by microarray, pyrosequencing, and targeted bisulfite sequencing revealed five DNA methylation markers for CRC diagnosis, especially early-stage CRC (eCRC). A single-tube multiple methylation-specific quantitative PCR assay (multi-msqPCR) for simultaneous detection of five methylation markers was established, which allowed quantitative analysis of samples with as little as 0.1% PBMC DNA and had better discriminative performance than single-molecule detection. Then, a CRC diagnostic model (CDM) based on methylation markers and the multi-msqPCR method was constructed that achieved high accuracy for eCRC (AUC=0.91; sensitivity=81.18%; specificity=89.39%), which was improved compared to CEA (AUC=0.79). The CDM also enabled a high degree of discrimination for advanced adenoma (AA) cases (AUC=0.85, sensitivity=63.04%). Follow-up data also demonstrated that the CDM could identify CRC potential up to 2 years before currently used diagnostic methods. In conclusion, the approach constructed in this study based on PBMC-derived DNA methylation markers and a multi-msqPCR method is a promising and easily implementable diagnostic method for eCRC.