Discovery of a 2,4-Diamino-7-aminoalkoxyquinazoline as a Potent and Selective Inhibitor of Histone Lysine Methyltransferase G9a

Abstract

SAR exploration of the 2,4-diamino-6,7-dimethoxyquinazoline template led to the discovery of 8 (UNC0224) as a potent and selective G9a inhibitor. A high resolution X-ray crystal structure of the G9a-8 complex, the first co-crystal structure of G9a with a small molecule inhibitor, was obtained. The co-crystal structure validated our binding hypothesis and will enable structure-based design of novel inhibitors. 8 is a useful tool for investigating the biology of G9a and its roles in chromatin remodeling.Multicellular organisms have evolved elaborate mechanisms to enable differential and celltype specific expression of genes. Epigenetics refers to these heritable changes in how the genome is accessed in different cell-types and during development and differentiation. This capability permits specialization of function between cells even though each cell contains the same genome. Over the last decade, the cellular machinery that creates these heritable changes has been the subject of intense scientific investigation as there is no area of biology or for that matter no area of human health, where epigenetics may not play a fundamental role. 1 The template upon which the epigenome is written is chromatin -the complex of histone proteins, RNA and DNA that efficiently package the genome in an appropriately accessible state within each cell. The state of chromatin, and therefore access to the genetic code, is mainly regulated by covalent and reversible PTMs to histone proteins and DNA, and the recognition of these marks by other proteins and protein complexes. The PTMs of histones and DNA include: histone lysine methylation, arginine methylation, lysine acetylation, sumoylation, † The coordinates and structure factors of UNC0224 co-crystallized with G9a have been deposited in the Protein Data Bank (www.pdb.org, PDB code 3K5K). ubiquitination, glycosylation and phosphorylation, and DNA methylation. 2 Given the wide-spread importance of chromatin regulation to cell biology, the enzymes that produce these modifications (the 'writers'), the proteins that recognize them (the 'readers'), and the enzymes that remove them (the 'erasers') are critical targets for manipulation in order to further understand the histone code 3, 4 and its role in human disease. Indeed, small molecule histone de-acetylase inhibitors5 and DNA methyltransferase inhibitors6 have already proven useful in the treatment of cancer.Histone lysine methylation refers to covalent methylation of histone lysine tails to produce mono-,di-, or trimethylated states. Among a myriad of PTMs, histone lysine methylation catalyzed by histone lysine methyltransferases (HMTs) has received great attention because of its essential function in many biological processes including gene expression and transcriptional regulation, heterochromatin formation, and X-chromosome inactivation. 7 It is therefore considered to be one of the most significant PTMs of histones. Since the first HMT was characterized in 20008, more than 50 human histone methyltransferases have been ...

Small-molecule histone methyltransferase inhibitors display rapid antimalarial activity against all blood stage forms inPlasmodium 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.…”

“…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.…”

Molecular insights on basal-like breast cancer

“…BC development depends on both genetic alterations and epigenetic changes involving DNA methylation and histone modifications [35]. Roll et al [36] reported a methylation signature in BLBC.…”

“…Moderate to low levels of lysine acetylation (H3K9ac, H3K18ac, and K4K12ac), lysine (H3K4me2 and H4K20me3) and arginine methylation (H4R3me2) were observed in BLBC and HER2-positive tumors and were related with adverse prognosis [42]. Alterations in histone methylation and demethylation are likely critical steps in neoplastic progression by disrupting the normal stem-or progenitor-cell program [35]. Further studies are needed involving BLBC and DNA methylation machinery to fully understand the clinicopahtological implications of the hypermehtylator phenotype in primary BC and subtypes for better diagnosis and improved treatment strategies.…”

Chemical and biochemical approaches in the study of histone methylation and demethylation

“…A few small molecule compounds such as chaetocin, 254 BIX-01294, 217 BIX-01338, 217 UNC0224, 255 DZNep 256 ( Figure 15) have been identified with anti-HKMT activity by random or virtual screening approaches. The first inhibitor of HKMTs is a natural fungal substance named chaetocin.…”