59P - Unraveling methylation signatures in RAS/BRAF wild-type colorectal cancer patients to identify predictive biomarkers for anti-epidermal growth factor receptor therapy

Background

Although anti-epidermal growth factor receptor (EGFR) therapy has established efficacy in RAS/BRAF wild type (WT) mCRC, 10-20% of WT patients exhibit primary resistance to this treatment. Biomarker discovery has mainly focused on tumor-specific DNA mutations, which suffer from variable mutation frequencies among patients. Epigenetic abnormalities, such as promoter hypermethylation, represent a common and early event in carcinogenesis, making it promising for resistance detection. In fact, several studies have shown an association between DNA methylation patterns and resistance to anti-EGFR therapy in CRC. To validate and build upon these findings, we investigated the methylomes of primary tumors with resistant and responsive phenotypes.

Methods

We isolated DNA from 15 primary CRC tumors. These DNA samples were subjected to Enzymatic Methyl-Sequencing (EMSeq), creating a genome-wide methylation dataset. We calculated methylation levels at CpGs and evaluated their correlation with resistance status. Based on the methylation values, a differential methylation analysis was performed using a modified linear mixed regression model. The CpGs were ranked based on the largest Δmethylation values and most significant p-values across the three patient groups. We performed a false discovery rate (FDR) evaluation, which led to the selection of differentially methylated CpGs (DMCs) with the highest likelihood of being genuine associations. Additionally, the DMCs were used as input for gene set enrichment analysis (GSEA).

Results

The GSEA revealed significant enrichment in 1 KEGG and 8 Reactome pathways, including MAPK-related, Akt and Wnt signaling. A gene ontology (GO) overrepresentation analysis was conducted on the significant genes. Notably, the most prominent GO categories were GTPase activity for molecular function and response to oxidative stress for biological processes.

Conclusions

We have created a comprehensive dataset of methylation sites with a substantial effect on conferring resistance to anti-EGFR therapy in CRC patients. In addition to identifying critical DMCs, the pathways hold potential as a rich source of predictive biomarkers.

Editorial acknowledgement

Clinical trial identification

Legal entity responsible for the study

The authors.

Funding

University of Antwerp.

Disclosure

All authors have declared no conflicts of interest.