A chemical probe selectively inhibits G9a and GLP methyltransferase activity in cells

Abstract

Protein lysine methyltransferases G9a and GLP modulate the transcriptional repression of a variety of genes via dimethylation of Lys9 on histone H3 (H3K9me2) as well as dimethylation of non-histone targets. Here we report the discovery of UNC0638, an inhibitor of G9a and GLP with excellent potency and selectivity over a wide range of epigenetic and non-epigenetic targets. UNC0638 treatment of a variety of cell lines resulted in lower global H3K9me2 levels, equivalent to levels observed for small hairpin RNA knockdown of G9a and GLP with the functional potency of UNC0638 being well separated from its toxicity. UNC0638 markedly reduced the clonogenicity of MCF7 cells, reduced the abundance of H3K9me2 marks at promoters of known G9a-regulated endogenous genes and disproportionately affected several genomic loci encoding microRNAs. In mouse embryonic stem cells, UNC0638 reactivated G9a-silenced genes and a retroviral reporter gene in a concentration-dependent manner without promoting differentiation.

Small-molecule histone methyltransferase inhibitors display rapid antimalarial activity against all blood stage forms in Plasmodium falciparum

“…These results indicate that BIX-01294 does indeed exert an effect on parasite histone methylation levels, and this effect is not simply a consequence of parasites dying. Together with its significant precedence as an HKMT inhibitor (33)(34)(35)(36)(37)(38), this result is highly suggestive of inhibition of parasite histone methyltransferase activity.…”

“…BIX-01294 was shown to be noncompetitive for the methyl donor S-adenosylmethionine and instead competitive for the histone substrate (30,37). In light of the high precedence of this class of compounds to serve as mammalian HKMT inhibitors (33)(34)(35)(36)(37)(38), focused BIX-01294 derivatives were explored for the development of parasitespecific histone methyltransferase inhibitors in our study. Our discovery that BIX-01294-treated and TM2-115-treated P. falciparum parasites exhibit greatly reduced H3K4me3 levels in BIX-01294-treated parasites, while maintaining a distinct profile compared with human cell lines treated with BIX-01294, is highly supportive of our compounds acting as parasite histone methyltransferase inhibitors.…”

“…1) was discovered in a high-throughput screen and was shown to be an inhibitor of the HKMTs G9a/GLP (30). BIX-01294 has also been used successfully in stem cell modulation (31,32), and subsequent medicinal chemistry studies have shown the potential of this scaffold in the discovery of compounds with increased potency, selectivity, and cellular permeability (33)(34)(35)(36)(37)(38). In this work, we identified two compounds that target histone methylation in P. falciparum.…”

The methyltransferase G9a regulates HoxA9-dependent transcription in AML

“…These results suggested that catalytic inhibition of G9a should mimic the phenotype observed in G9a À/À (Vav) cells. Indeed, treatment with the G9a/GLP inhibitor UNC0638 (Vedadi et al 2011) inhibited the growth of G9a +/À (Vav) ; A9M but not G9a À/À (Vav) ; A9M cells in vitro (Fig. 3H).…”

“…Furthermore, LSD1/KDM1A inhibition suppresses acute myeloid leukemia (AML) stem cell activity (Harris et al 2012;Schenk et al 2012), and disruption of the chromatin binding of Brd4 by a BET bromodomain inhibitor blocks c-Myc expression and the proliferation of leukemic cells (Delmore et al 2011;Zuber et al 2011). To date, several inhibitors of histone methyltransferases and demethylases have been reported, including those that target G9a and GLP with high specificity (Kubicek et al 2007;Vedadi et al 2011;Yuan et al 2012). G9a/GLP uniquely catalyze mono-and dimethylation of histone 3 on Lys9 (H3K9me1/2) (Tachibana et al 2002(Tachibana et al , 2005, a highly abundant chromatin mark in mammalian cells.…”