ABSTRACT
Inhibition of glycogen phosphorylase (GP) has attracted considerable attention during the last five to 10 years as a means of treating the elevated hepatic glucose production seen in patients with type 2 diabetes. Several different GP inhibitors binding to various binding sites of the GP enzyme have been reported in the literature. In this paper we report on a novel class of compounds that have been identified as potent GP inhibitors. Their synthesis, mode of binding to the allosteric AMP site as well as in vitro data on GP inhibition are shown. The most potent inhibitor was found to be 4-[2,4-bis-(3-nitrobenzoylamino)phenoxy]phthalic acid (4j) with an IC(50) value of 74 nM. This compound together with a closely related analogue was further characterized by enzyme kinetics and in primary rat hepatocytes.
Subject(s)
Benzamides/chemical synthesis , Enzyme Inhibitors/chemical synthesis , Glycogen Phosphorylase/antagonists & inhibitors , Hypoglycemic Agents/chemical synthesis , Phthalic Acids/chemical synthesis , Adenosine Monophosphate/metabolism , Allosteric Site , Animals , Benzamides/chemistry , Benzamides/pharmacology , Cells, Cultured , Crystallography, X-Ray , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Glucose/metabolism , Glycogen Phosphorylase/chemistry , Hypoglycemic Agents/chemistry , Hypoglycemic Agents/pharmacology , Male , Models, Molecular , Phthalic Acids/chemistry , Phthalic Acids/pharmacology , Protein Binding , Rats , Rats, WistarABSTRACT
Protein-tyrosine phosphatases (PTPs) are considered important therapeutic targets because of their pivotal role as regulators of signal transduction and thus their implication in several human diseases such as diabetes, cancer, and autoimmunity. In particular, PTP1B has been the focus of many academic and industrial laboratories because it was found to be an important negative regulator of insulin and leptin signaling, and hence a potential therapeutic target in diabetes and obesity. As a result, significant progress has been achieved in the design of highly selective and potent PTP1B inhibitors. In contrast, little attention has been given to other potential drug targets within the PTP family. Guided by x-ray crystallography, molecular modeling, and enzyme kinetic analyses with wild type and mutant PTPs, we describe the development of a general, low molecular weight, non-peptide, non-phosphorus PTP inhibitor into an inhibitor that displays more than 100-fold selectivity for PTPbeta over PTP1B. Of note, our structure-based design principles, which are based on extensive bioinformatics analyses of the PTP family, are general in nature. Therefore, we anticipate that this strategy, here applied to PTPbeta, in principle can be used in the design and development of selective inhibitors of many, if not most PTPs.
Subject(s)
Enzyme Inhibitors/pharmacology , Protein Tyrosine Phosphatases/antagonists & inhibitors , Cloning, Molecular , Crystallography, X-Ray , Drug Design , Enzyme Inhibitors/chemistry , Hydrogen Bonding , Insulin/metabolism , Kinetics , Leptin/metabolism , Ligands , Models, Chemical , Models, Molecular , Mutation , Phthalimides/chemistry , Protein Conformation , Protein Tyrosine Phosphatase, Non-Receptor Type 1 , Protein Tyrosine Phosphatases/chemistry , Signal Transduction , Structure-Activity Relationship , TemperatureABSTRACT
Protein-tyrosine phosphatase 1B (PTP1B) has recently received much attention as a potential drug target in type 2 diabetes. This has in particular been spurred by the finding that PTP1B knockout mice show increased insulin sensitivity and resistance to diet-induced obesity. Surprisingly, the highly homologous T cell protein-tyrosine phosphatase (TC-PTP) has received much less attention, and no x-ray structure has been provided. We have previously co-crystallized PTP1B with a number of low molecular weight inhibitors that inhibit TC-PTP with similar efficiency. Unexpectedly, we were not able to co-crystallize TC-PTP with the same set of inhibitors. This seems to be due to a multimerization process where residues 130-132, the DDQ loop, from one molecule is inserted into the active site of the neighboring molecule, resulting in a continuous string of interacting TC-PTP molecules. Importantly, despite the high degree of functional and structural similarity between TC-PTP and PTP1B, we have been able to identify areas close to the active site that might be addressed to develop selective inhibitors of each enzyme.
