Abstract
Activated immune cells generate reactive oxygen species as part of the immune defence against infectious agents or irritants. When the rate of oxidant production exceeds the cell’s antioxidant systems, this can lead to oxidative stress and chronic inflammation. Under these conditions, oxidants can damage DNA, proteins, and lipids, thereby disrupting critical cellular processes. Chronic inflammation is a well-established risk factor for colorectal cancer, but the underlying biological mechanisms remain poorly defined. One potential pathway is through the disruption of epigenetic regulation, particularly DNA methylation, which plays a key role in maintaining gene expression patterns. Dysregulated methylation can drive carcinogenesis by silencing tumour suppressor genes and activating oncogenes.
Our research group has previously shown that the immune cell-derived oxidant, glycine chloramine, causes global and site-specific DNA hypomethylation in Jurkat T-lymphoma and HCT116 colorectal cancer cell lines. Oxidative stress has also been linked to increased cell surface expression of the immune checkpoint protein Programmed Death Ligand 1 in other cancer types. Therefore, understanding whether glycine chloramine exposure induces similar effects in colorectal cancer may provide insight into how inflammation-driven oxidative stress promotes tumour immune evasion and progression.
In this study, we re-optimised previous experimental conditions for the HT29 colorectal cancer cell line and developed an alternative method to quantify site-specific DNA methylation using fluorescent probe hybridisation and qPCR. The assay was validated against stored HCT116 samples by comparison with top-ranked loci previously identified by the Illumina MethylationEPIC array.
The technique produced a trend congruent with the results from the EPIC array data, however, further optimisation and refinement are required to confirm its robustness. No significant methylation changes were observed at the examined CpG sites in HT29 cells following glycine chloramine treatment. Interestingly, Programmed Death Ligand 1 expression was significantly increased in HT29 cells following glycine chloramine exposure, suggesting that oxidative stress may promote immune-evasive phenotypes. Further investigation is required to determine whether changes in DNA methylation at the Programmed Death Ligand 1 promoter contribute to this outcome. Together, these findings indicate that oxidants generated during inflammation can drive epigenetic remodelling and immune-evasive phenotypes in colorectal cancer.
However, this effect may not be conserved across different types of cancer. This research adds to the growing body of evidence linking oxidative stress to cancer development, with important implications for understanding tumour progression and informing therapeutic strategies that combine redox modulation with immunotherapy.