Histone modifications are increasingly being recognized as an important epigenetic mechanism that governs chromatin structure and gene expression. EZH2 is the catalytic subunit of the polycomb repessive complex 2 (PRC2), responsible for tri-methylation of lysine 27 residues on type 3 histones (H3K27Me3) that leads to gene silencing. EZH2 is found to be over-expressed in many different types of cancers, where it is postulated to abnormally repress tumour suppressor genes. Activating mutations described within the catalytic SET domain of EZH2 at codon 646 confer its oncogenic activity, as identified in approximately 3% of melanomas.
We sought to assess whether direct inhibition of EZH2’s methyltransferase activity is a suitable strategy for melanoma treatment. This was tested using a small molecule inhibitor known as GSK126, a potent, highly selective drug that competes with S-adenosyl-methionine substrate. We found that both mutant and non-mutant EZH2 melanoma cell lines over-express EZH2 compared to primary cells, via western blot and that levels of H3K27 methylation were reduced following drug treatment. GSK126 significantly decreased proliferation and reduced the percentage of cells in the G1 phase of the cell cycle, most likely attributed to a G2 arrest. Flow cytometry also revealed an increased sub-G1 population, indicative of drug induced apoptosis. This was confirmed by Annexin-V staining. Growth inhibition by GSK126 was particularly effective in melanoma cells harboring EZH2 activating mutations and had little effect on primary cells that displayed low levels of EZH2.
Expression arrays and chromatin immunoprecipitation are being used to determine which PCR2 target genes are being de-repressed and to decipher the mechanisms of GSK126 growth inhibition. Collectively our initial data indicate that EZH2 represents a promising gene target for therapeutic intervention in melanoma in vitro and that in vivo studies are warranted.