Background: Many cancers show aberrant silencing of gene expression and
overexpression of histone methyltransferases. The histone methyltransferases (HKMT)
EZH2 and EHMT2 maintain the repressive chromatin histone marks H3K27 and H3K9
methylation respectively, which are associated with transcriptional silencing. Although
selective HKMT inhibitors reduce levels of individual repressive marks, removal of
H3K27me3 by specific EZH2 inhibitors, for instance, may not be sufficient for inducing
expression of genes with multiple repressive marks.
Results: We report that gene expression and inhibition of triple negative breast cancer cell
growth (MDA-MB-231) are markedly increased when targeting both EZH2 and EHMT2,
either by siRNA knockdown or pharmacological inhibition, rather than independently. Indeed,
expression of certain genes is only induced upon dual inhibition. We sought to identify
compounds which showed evidence of dual EZH2 and EHMT2 inhibition. Using a cell-based
assay, based on the substrate-competitive EHMT2 inhibitor BIX01294, we have identified
proof-of-concept compounds that induce re-expression of a subset of genes consistent with
dual HKMT inhibition. Chromatin immunoprecipitation verified a decrease in silencing marks
and an increase in permissive marks at the promoter and transcription start site of reexpressed
genes, while Western analysis showed reduction in global levels of H3K27me3
and H3K9me3. The compounds inhibit growth in a panel of breast cancer and lymphoma cell
lines with low to sub-micromolar IC50s. Biochemically, the compounds are substrate
competitive inhibitors against both EZH2 and EHMT1/2.
Conclusions: We have demonstrated that dual inhibition of EZH2 and EHMT2 is more
effective at eliciting biological responses of gene transcription and cancer cell growth
inhibition compared to inhibition of single HKMTs, and we report the first dual EZH2-
EHMT1/2 substrate competitive inhibitors that are functional in cells.