A multidisciplinary investigation towards inhibitors of glycogen phosphorylase designed and evaluated as antidiabetic molecules – GPdia

We recently achieved the synthesis of various structures (N-acyl-N-glucopyranosyl-ureas, isomeric 5-substituted 3-C-glucopyranosyl-1,2,4-oxadiazoles and 3-substituted 5-C-glucopyranosyl-1,2,4-oxadiazoles, glucopyranosyl-spiro-isoxazolines, glusosyl-spiro-oxathiazolines, 2-(C-b-D-glucopyranosyl)-hydro-, and –benzoquinones) derived from glucose. Several compounds studied by enzymology appeared as good reversible inhibitors of glycogen phosphorylase (GP) while crystallography of the enzyme-ligand complexes provided a structural basis of the inhibition, due to binding at the active site. GP, the rate-limiting enzyme of glycogenolysis eg the depolymerization of glycogen to glucose is a major actor for hepatic glucose production (HGP). In type 2 diabetes (T2D) the ability of insulin to suppress HGP is impaired (hepatic insulin resistance). Storage of glucose as glycogen (glycogenogenesis) as catalyzed by unphosphorylated glycogen synthase (GS) limits hyperglycemia. Since kinase GSK3 can inactivate GS, inhibiting GSK3 should favour glycogenogenesis. Simultaneous inhibition of GP and GSK3 should efficiently limit hyperglycemia. In this context, inhibitions measured in vitro for the prepared inhibitors (best Ki 160 nanoM) call for a collaboration towards detailled in vivo studies with cells and animals to probe this approach in the context of T2D. To this end, cellular tests with human hepatocytes will be carried out with active inhibitors available, for measuring the glucose liberated in the presence of the tested products and estimating their effects and stability. Inhibitors displaying a glucose moiety may behave differently depending on their N-, or C-glucosidic linkage, the latter being presumed more stable. These tests will be critical for the choice of the synthetic tasks, also dependent on theoretical predictive studies (Fragment-Based Drug Design) and comparison of chemical libraries. These methods may even suggest other types of promissing molecules. Special attention will be paid to flavopiridol and analogues since they can act simultaneously on GP and GSK3, so as to reduce hyperglycemia. In addition, NMR will provide within a short time a mean for estimating the inhibition potency of newly prepared molecules, as compared to typical enzymatic assays. The active molecules will be used for preparing enzyme-ligand complexes with GP in view of crystallography for explaining on a rational structural basis the inhibitions observed in vitro. Finally, some among the best inhibitors will be tested with animals (mice) so as to determine the in vivo effects and establish whether inhibitors of GP (as well as GSK3) may be used in view of a pharmacological treatment of type 2 diabetes.