Design, Synthesis and Biological Evaluation of Heterocycles Targeting Dipeptidyl Peptidase-4 (DPP-4) for Management of Type 2 Diabetes

Abstract

The research work titled “Design, Synthesis & Biological Evaluation of Heterocycles Targeting Dipeptidyl Peptidase-4 (DPP-4) for management of Type 2 Diabetes” is divided into 7 chapters mentioned in brief as follows. Chapter 1 outlines historical aspects of diabetes, epidemiology, prevalence, classification and current therapies for management of Type 2 Diabetes (T2D). Novel therapies under development for management of T2D encompassing targets and related New Chemical Entities (NCEs) are described. Chapter 2 begins with role of incretin hormones in blood sugar control, effect of Dipetidyl Peptidase-4 (DPP-4) and impact of DPP-4 inhibition on blood sugar levels. It is followed by classification systems of DPP-4 inhibitors, “Gliptins” and discussion about peptidomimetic and non-peptidomimetic inhibitors with importance on diversity in their structure. Chapter is concluded with advantages and limitations of Gliptins. Aim of present work was envisaged to exploit computer-aided drug design approaches to identify new small molecules as DPP-4 inhibitors. Aim and objectives of the research work are briefed in chapter 3. Chapter 4 is detailed on various computational tools to identify new molecules that can inhibit DPP-4. In present work, various X-ray crystal structures of DPP-4 enzymes were analyzed to study their active site and interaction pattern. Following the virtual screening, methodologies were used for identification of new synthetic small molecules having the affinity for DPP-4 inhibitors. Common feature pharmacophore approach was used in combination with docking to screen the commercial compound library for searching new templates that may inhibit DPP-4, and few selected hits were subjected to in-vitro DPP-4 enzyme inhibition assay. Docking poses obtained for few hit compounds were confirmed by molecular dynamics simulation. In second computational approach molecular dynamics structure-based pharmacophore approach was used to generate receptor-based pharmacophore using multiple DPP-4 protein structure. The commercial compound library was pre-filtered employing receptor-based pharmacophores followed by docking into the respective crystal structure of DPP-4 followed by molecular dynamics simulation and in-vitro activity. Few compounds were designed from pharmacophore queries and docking studies encompassing essential molecular recognition feature to interact in DPP-4 catalytic site. Drug-like properties of virtual hits and designed compounds were computed according to Lipinski’s rule of five.In chapter 5, synthesis of eighteen compounds bearing glycine substituted with aromatic and hydrophobic substituent at N-terminus and C-terminus and five compounds having substituted thiazole nucleus bearing L- phenylalanine and their characterization is described in stepwise manner. In the first step of the synthesis amidation of chloroacetyl chloride was performed with variably substituted heterocyclic amine. Subsequent amination using substituted 2-phenylethylamines yielded glycine substituted amine. For synthesis of thiazole derivatives, Boc-protected phenylalanine was coupled with substituted 2-amino-1, 3-thiazoles to provide amide intermediate in first step. Boc-deprotection of intermediates using 4M HCl in dioxane yielded phenylalanine derivatives coupled with thiazoles. Progress of the reactions was monitored by thin layer chromatography (TLC) using silica-gel precoated TLC plates from Merck. Compounds were characterized by 1H NMR, 13C NMR Mass and FT-IR. Chapter 6 encompasses biological activity of 22 virtual hits and 23 synthesized compounds. Virtual hits procured from Specs Database and synthesized compounds were subjected to in vitro DPP 4 enzyme inhibition assay by fluorometric method. Based on the in vitro DPP-4 enzyme inhibition assay results, compounds JAG-A2 (2-(phenethylamino)-1-phenylethanone) and JAG-D2 (2-(4-fluorophenethylamino)-1-(4-(6-fluorobenzo[d]isoxazol-3-yl)piperidin-1-yl)ethanone) were subjected to in vivo OGTT assay in streptozotocin induced diabetic rats. Both compounds produced significant reduction in blood glucose levels at 20 mg/ kg and 40 mg/kg dose when compared with 1mg/kg dose of standard DPP-4 inhibitor sitagliptin. Maximum glucose lowering effect was observed at 40 mg/kg for both compounds. Chapter 7 describes summary of the research work.