Progressive methylation changes in colorectal cancers on gaining chemoresistance

Abstract

The modes by which cancer cells deal with and escape chemotherapeutics remains an area researchers are trying to understand. Protein lysine methylation has shown to be involved in chemoresistance, but its exact role is still being understood. This project aims to monitor any changes in mono-, di- or tri- methylated lysine residues in colorectal cancer cells upon gaining doxorubicin (DOX) resistance. HCT116 and Caco-2 cell lines were cultured and treated with increasing doses of DOX (0.25, 0.50, 1.00 and 2.00 μg/ml) to see if the cells gain chemoresistance. Throughout the treatment, pictures of the cells were recorded to monitor any progressive changes in confluency and cell morphology. MTS assay was performed to monitor cell viability in sensitive (untreated) and DOX posttreatment cells. Resistant cells were collected at each DOX concentration for protein extraction. The proteins extracted were separated using SDS-PAGE and transferred onto a nitrocellulose membrane for Western blotting. The blots were exposed to primary antibodies which targeted the different degrees of lysine methylation. Both cell lines became DOX resistant by the end of the treatment, with several characteristic features of chemoresistance being observed in both cell lines. The global lysine methylation patterns detected in both cell lines were altered as the cells gained chemoresistance, with lysine mono- and tri- methylation being the most dominant, while lysine di-methylation was the least detected during this study. This project presents a better understanding of global protein lysine methylation upon gain of chemoresistance and provides a good foundation for further studies.