Abstract
Despite their proven antidiabetic efficacy, widespread use of peroxisome proliferator-activated receptor (PPAR)γ agonists has been limited by adverse cardiovascular effects. To overcome this shortcoming, selective PPARγ modulators (SPPARγMs) have been identified that have antidiabetic efficacy comparable with full agonists with improved tolerability in preclinical species. The results of structural studies support the proposition that SPPARγMs interact with PPARγ differently from full agonists, thereby providing a physical basis for their novel activities. Herein, we describe a novel PPARγ ligand, SPPARγM2. This compound was a partial agonist in a cell-based transcriptional activity assay, with diminished adipogenic activity and an attenuated gene signature in cultured human adipocytes. X-ray cocrystallography studies demonstrated that, unlike rosiglitazone, SPPARγM2 did not interact with the Tyr473 residue located within helix 12 of the ligand binding domain (LBD). Instead, SPPARγM2 was found to bind to and activate human PPARγ in which the Tyr473 residue had been mutated to alanine (hPPARγY473A), with potencies similar to those observed with the wild-type receptor (hPPARγWT). In additional studies, we found that the intrinsic binding and functional potencies of structurally distinct SPPARγMs were not diminished by the Y473A mutation, whereas those of various thiazolidinedione (TZD) and non-TZD PPARγ full agonists were reduced in a correlative manner. These results directly demonstrate the important role of Tyr473 in mediating the interaction of full agonists but not SPPARγMs with the PPARγ LBD, thereby providing a precise molecular determinant for their differing pharmacologies.
Footnotes
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Use of Industrial Macromolecular Crystallography Association Collaborative Access Team beamline 17-ID (or 17-BM) at the Advanced Photon Source was supported by the companies of the Industrial Macromolecular Crystallography Association through a contract with the Center for Advanced Radiation Sources at the University of Chicago (Chicago, IL). All work presented here was performed by employees of Merck and Co. and was completely funded by Merck and Co.
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M.E. and T.E.A. contributed equally to the research communicated in this article.
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ABBREVIATIONS: PPAR, peroxisome proliferator-activated receptor; TZD, thiazolidinedione; SPPARγM, selective peroxisome proliferator-activated receptor γ modulator; T2DM, type 2 diabetes mellitus; LBD, ligand binding domain; WT, wild type; GST, glutathione transferase; PCR, polymerase chain reaction; nTZDpa, nonthiazolidinedione-selective peroxisome proliferator-activated receptor γ modulator; FABP4, fatty acid-binding protein 4; PEPCK, phosphoenolpyruvate carboxykinase; ADFP, adipose differentiation-related protein; FK-614, 3-(2,4-dichlorobenzyl)-2-methyl-N-(pentylsulfonyl)-3H-benzimidazole-5-carboxamide; GI262570, (2S)-2-[[2-(benzoyl)phenyl]amino]-3-[4-[2-(5-methyl-2-phenyl-1,3-oxazol-4-yl)ethoxy]phenyl]propanoic acid; GW0072, 4-[4-[(2S,5S)-5-[2-(bis(phenylmethyl)amino)-2-oxoethyl]-2-heptyl-4-oxo-1,3-thiazolidin-3-yl]butyl]benzoic acid; PA-082, 1-(3,4-dimethoxy-benzyl)-6,7-dimethoxy-4-[4-(2-methoxy-phenyl)-piperidin-1-ylmethyl]-isoquinoline.
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↵1 Current affiliation: Vitae Pharmaceuticals, Inc., Fort Washington, Pennsylvania.
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↵2 Current affiliation: Pharmasset, Inc., Princeton, New Jersey.
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↵3 Current affiliation: Lilly Research Laboratories, Indianapolis, Indiana.
- Received August 29, 2007.
- Accepted October 16, 2007.
- The American Society for Pharmacology and Experimental Therapeutics
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