Novel Epigenetic 12-gene Signature Predictive of Poor Prognosis and MSI-like Phenotype in Human Metastatic Colorectal Carcinomas

Background. Epigenetic remodeling is responsible for tumor progression and drug resistance in human colorectal carcinoma (CRC). A subgroup of human CRCs exhibits the CIMP status, with extensive hypermethylation events in promoter regions of several genes, even though the prognostic significance of CIMP is controversial. This study addressed the hypothesis that DNA methylation profiling may identify metastatic CRC (mCRC) subtypes with different clinical behavior. Methods. Global methylation profile was comparatively analyzed between 24 first-line primary-resistant and 12 drug-sensitive mCRCs (in-house cohort), two subgroups of tumors with significantly different outcome. Methylation and gene expression data from 33 mCRCs of the TCGA COAD dataset (TCGA COAD cohort) were used to identify, among differentially methylated genes, a prognostic signature of functionally methylated genes. Clusters of mCRCs with different methylation patterns were further characterized for DNA mutational load, gene copy number and gene expression profiles. Human CRC HT29 and HCT116 cell lines were adapted to growth in presence of oxaliplatin and irinotecan and used as in vitro model to validate gene expression data.Results. Twelve functionally methylated genes yielded a hierarchical clustering of patients in two well-defined clusters with hypermethylated tumors characterized by a significantly worse relapse-free and overall survival compared to hypomethylated cancers and this was reproduced in both the in-house and the TCGA COAD cohorts. Interestingly, the hypermethylated poor prognosis cluster was enriched of CIMP-high and MSI-like cases. Furthermore, methylation events were enriched in genes located on q-arm of chromosomes 13 and 20, two chromosomal regions with gain/loss alterations strongly associated with adenoma-to-carcinoma progression. Finally, the expression of the 12-genes signature and MSI-enriching genes was confirmed in two independent oxaliplatin- and irinotecan-resistant CRC cell lines. Conclusions. These data represent the proof of concept that the hypermethylation of specific sets of genes may provide prognostic information being able to identify a subgroup of mCRCs with poor prognosis.