A 366 is a selective G9a/GLP histone methyltransferase inhibitor that inhibits the activity of G9a (IC50=3.3nM) and GLP (IC50=38nM). A 366 suppresses H3K9me2 methylation and interferes with the binding of Spindlin1 to H3K4me3[1-2]. A 366 can be used in research related to leukemia, prostate cancer, and autism spectrum disorders[3-4].
In vitro, U2OS human osteosarcoma cells were treated with A 366 (10μM) in combination with phleomycin (0.5–1μM) or etoposide (100nM) for 0-180 minutes. A 366 significantly impaired DNA double-strand break repair, enhanced DNA damage accumulation, and synergistically induced tumor cell death[5]. Human monocytes, M1/M2 monocyte-derived macrophages, CD4+ T cells, and B cells were pretreated with A 366 (5–50μM) for 30 minutes, followed by incubation with different inducters for 24 hours to 5 days. A 366 attenuated the M2 polarization of macrophages; A 366 also significantly inhibited the proliferation of B cells, the formation of plasmablasts, and the release of immunoglobulins G and A[6].
In vivo, Balb/c nu/nu mice bearing OPM2 multiple myeloma xenografts were treated with A 366 alone (8mg/kg; i.p.; for 6 days) or in combination with decitabine (0.2mg/kg) for the final 3 days. A 366 significantly inhibited tumor growth in the xenograft model, and the combination therapy demonstrated a synergistic antitumor effect without causing significant toxicity[7]. Wistar rats were treated with A 366 (25mg/kg; a single intraperitoneal injection). A 366 significantly reduced the clonogenic capacity of bone marrow-derived mesenchymal stem cells (BM-MSCs), prolonged their population doubling time, increased their adipogenic differentiation potential, and decreased their osteogenic differentiation potential[8].
References:
[1] Reiner D, Seifert L, Deck C, et al. Epigenetics meets GPCR: inhibition of histone H3 methyltransferase (G9a) and histamine H3 receptor for Prader-Willi Syndrome. Sci Rep. 2020 Aug 11;10(1):13558.
[2] Wagner T, Greschik H, Burgahn T, et al. Identification of a small-molecule ligand of the epigenetic reader protein Spindlin1 via a versatile screening platform. Nucleic Acids Res. 2016 May 19;44(9):e88.
[3] Sweis RF, Pliushchev M, Brown PJ, et al. Discovery and development of potent and selective inhibitors of histone methyltransferase g9a. ACS Med Chem Lett. 2014 Jan 2;5(2):205-9.
[4] Pappano WN, Guo J, He Y, et al. The Histone Methyltransferase Inhibitor A-366 Uncovers a Role for G9a/GLP in the Epigenetics of Leukemia. PLoS One. 2015 Jul 6;10(7):e0131716.
[5] Agarwal P, Jackson SP. G9a inhibition potentiates the anti-tumour activity of DNA double-strand break inducing agents by impairing DNA repair independent of p53 status. Cancer Lett. 2016 Oct 1;380(2):467-475.
[6] Schiffmann S, Henke M, Weber F, et al. Immune-modulatory effects of Spindlin-1 inhibitors. Clin Exp Immunol. 2025 Jan 21;219(1):uxaf013.
[7] Nylund P, Garrido-Zabala B, Tziola SI, et al. Dual targeting of G9a and DNMTs induces antitumor effects in multiple myeloma. Blood Adv. 2025 Oct 14;9(19):4825-4841.
[8] Khanban H, Fattahi E, Talkhabi M. In vivo administration of G9a inhibitor A366 decreases osteogenic potential of bone marrow-derived mesenchymal stem cells. EXCLI J. 2019 Jun 3;18:300-309.