ABSTRACT
Herein, we report a blended ligand and structure-based pharmacophore screening approach to identify new natural leads against the Protein Lysine Methyltransferase 2 (EHMT2/G9a). The EHMT2/G9a has been associated with Cancer, Alzheimer's, and aging and is considered an emerging drug target having no clinically passed inhibitor. Purposefully, we developed the ligand-based pharmacophore (Pharmacophore-L) based on the common features of known inhibitors and the structure-based pharmacophore (Pharmacophore-S) based on the interaction profile of available crystal structures. The Pharmacophore-L and Pharmacophore-S were subjected to multiple tiers of validations and utilized in combination for the screening of total 741543 compounds coming from multiple databases. Additional layers of stringency were applied in the screening process to test drug-likeness (using Lipinski's rule, Veber's rule, SMARTS and ADMET filtration), to rule out any toxicity (TOPKAT analysis). The interaction profiles, stabilities, and comparative analysis against the reference were carried out by flexible docking, MD simulation, and MM-GBSA analysis, which finally led to three leads as potential inhibitors of G9a.Communicated by Ramaswamy H. Sarma.
Subject(s)
Histone-Lysine N-Methyltransferase , Pharmacophore , Molecular Docking Simulation , Ligands , Molecular Dynamics Simulation , Quantitative Structure-Activity RelationshipABSTRACT
Herein, we report for the first time the G9a/EHMT2 inhibition and anti-Alzheimer's activities of the drug raltitrexed. G9a is a lysine methyltransferase that mainly dimethylates the H3K9 of chromatin, which triggers the repression of genes epigenetically, leading to various diseased conditions, including Alzheimer's disease (AD). First, we demonstrate that raltitrexed inhibits G9a at 120 nM. Moreover, raltitrexed lowers the total H3K9me2/H3K9 levels in AD transgenic C. elegans CL2006 worms, indicating that raltitrexed targets G9a directly. As toxicity is the bottleneck in G9a drug discovery, we conducted detailed in silico toxicity (TOPKAT) analyses of raltitrexed and measured the food consumption by C. elegans, demonstrating that raltitrexed's toxicity/function range is safe for the worm's growth. Moreover, we demonstrate that raltitrexed enhances the locomotive function of worms dose-dependently. Finally, we show that raltitrexed reduced the Aß aggregates in worms up to 47%, highlighting the potential of raltitrexed in AD treatment.
ABSTRACT
The G9a, Lysine Methyltransferase that methylates the histone 3 lysine 9 (H3K9) of the nucleosome, is an excellent epigenetic target having no clinically passed inhibitor currently owing to adverse in vivo ADMET toxicities. In this work, we have carried out detailed computational investigations to find novel and safer lead against the target using advanced 3 D QSAR pharmacophore screening of databases containing more than 400000 entrees of natural compounds. The screening was conducted at different levels at increasing stringencies by employing pharmacophore mapping, druglikenesses and interaction profiles of the selected to identify potential hit compounds. The potential hits were further screened by advanced flexible docking, ADME and toxicity analysis to eight hit compounds. Based on the comparative analysis of the hits with the reference inhibitor, we identified one lead inhibitor against the G9a, having better binding efficacy and a safer ADMET profile than the reference inhibitor. Finally, the results were further verified using robust molecular dynamics simulation and MM-GBSA binding energy calculation. The natural compounds are generally considered benign due to their long human uses and this is the first attempt of in silico screening of a large natural compound library against G9a to our best knowledge. Therefore, the finding of this study may add value towards the development of epigenetic therapeutics against the G9a.Communicated by Ramaswamy H. Sarma.
