Abstract 5532: Discovery of A-366, a novel small molecule inhibitor that uncovers an unappreciated role for G9a/GLP in the epigenetics of leukemia
Authors: Guo Jun , Pliushchev Marina , He Yupeng , Ferguson Debra , Jagadeeswaran Sujatha , Petros Andrew , Sun Chaohong , Soni Niru B. , Shaw Bailin , Korepanova Alla , Maag David , Sweis Ramzi , Buchanan Fritz G. , Michaelides Michael , Shoemaker Alex , Tse Chris , Chiang Gary G. , Pappano William N. 
Published: May 2015

Abstract
Understanding the roles of epigenetic alterations in cancer development and maintenance holds great promise for cancer prevention, detection, and therapy. Cancer can be considered as a pathogenic state where cellular differentiation is suppressed (i.e. stem cell-like) and aberrant epigenetic patterning is commonly observed in tumors. Histone methyltransferases play a key role in epigenetics by modifying key lysine and arginine residues on histones and thereby influencing biological processes. Previous studies have suggested that the histone lysine methyltransferase G9a (EHMT2) is required to perpetuate malignant phenotypes through over-expression in a variety of cancer types. These reports have shown that pharmacologic inhibition or genetic ablation of G9a leads to retardation of tumor cell growth and cellular invasion in vitro as well as inhibition of metastasis in vivo. To further elucidate the enzymatic role of G9a in cancer, we describe herein the discovery of a novel histone methyltransferase inhibitor, A-366, that selectively inhibits G9a and the closely related GLP (EHMT1). A-366 is a peptide competitive inhibitor of G9a/GLP with in vitro enzymatic IC50 of ∼ 3 nM and cellular activity of ∼ 100 nM and &gt; 100-fold selectivity over other methyltransferases and other non-epigenetic targets. A-366 has significantly less cytotoxic effects on the growth of solid tumor cell lines compared to other known G9a/GLP small molecule inhibitors despite roughly equivalent cellular activity on methylation of H3K9me2. However, the excellent selectivity profile of A-366 has aided in the discovery of an important role for G9a/GLP in lineage maintenance of a subset of leukemias. Treatment of various leukemia cell lines in vitro resulted in marked differentiation and morphological changes of these tumors in the absence of cytotoxicity resulting in cytostasis. Furthermore, treament of MV4;11 flank xenografts with A-366 resulted in growth inhibition in vivo consistent with the profile of H3K9me2 reduction observed. In summary, A-366 is a novel and highly selective peptide-competitive inhibitor of G9a/GLP that has enabled the discovery of a role for G9a/GLP enzymatic activity in the epigenetic maintenance of a subset of leukemia cells.
Citation Format: Jun Guo, Marina Pliushchev, Yupeng He, Debra Ferguson, Sujatha Jagadeeswaran, Andrew Petros, Chaohong Sun, Niru B. Soni, Bailin Shaw, Alla Korepanova, David Maag, Ramzi Sweis, Fritz G. Buchanan, Michael Michaelides, Alex Shoemaker, Chris Tse, Gary G. Chiang, William N. Pappano. Discovery of A-366, a novel small molecule inhibitor that uncovers an unappreciated role for G9a/GLP in the epigenetics of leukemia. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5532. doi:10.1158/1538-7445.AM2014-5532