Robert T. Gampe

Asymmetry in the PPARγ/RXRα Crystal Structure Reveals the Molecular Basis of Heterodimerization among Nuclear Receptors

Abstract The nuclear receptor PPARγ/RXRα heterodimer regulates glucose and lipid homeostasis and is the target for the antidiabetic drugs GI262570 and the thiazolidinediones (TZDs). We report the crystal structures of the PPARγ and RXRα LBDs complexed to the RXR ligand 9- cis -retinoic acid (9cRA), the PPARγ agonist rosiglitazone or GI262570, and coactivator peptides. The PPARγ/RXRα heterodimer is asymmetric, with each LBD deviated ∼10° from the C2 symmetry, allowing the PPARγ AF-2 helix to interact with helices 7 and 10 of RXRα. The heterodimer interface is composed of conserved motifs in PPARγ and RXRα that form a coiled coil along helix 10 with additional charge interactions from helices 7 and 9. The structures provide a molecular understanding of the ability of RXR to heterodimerize with many nuclear receptors and of the permissive activation of the PPARγ/RXRα heterodimer by 9cRA.

Discovery of 7-Methyl-5-(1-{[3-(trifluoromethyl)phenyl]acetyl}-2,3-dihydro-1H-indol-5-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine (GSK2606414), a Potent and Selective First-in-Class Inhibitor of Protein Kinase R (PKR)-like Endoplasmic Reticulum Kinase (PERK).

Protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK) is activated in response to a variety of endoplasmic reticulum stresses implicated in numerous disease states. Evidence that PERK is implicated in tumorigenesis and cancer cell survival stimulated our search for small molecule inhibitors. Through screening and lead optimization using the human PERK crystal structure, we discovered compound 38 (GSK2606414), an orally available, potent, and selective PERK inhibitor. Compound 38 inhibits PERK activation in cells and inhibits the growth of a human tumor xenograft in mice.

Crystallization of protein-ligand complexes.

Methods presented for growing protein–ligand complexes fall into the categories of co-expression of the protein with the ligands of interest, use of the ligands during protein purification, cocrystallization and soaking the ligands into existing crystals.

Discovery of GSK2656157: An Optimized PERK Inhibitor Selected for Preclinical Development.

We recently reported the discovery of GSK2606414 (1), a selective first in class inhibitor of protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK), which inhibited PERK activation in cells and demonstrated tumor growth inhibition in a human tumor xenograft in mice. In continuation of our drug discovery program, we applied a strategy to decrease inhibitor lipophilicity as a means to improve physical properties and pharmacokinetics. This report describes our medicinal chemistry optimization culminating in the discovery of the PERK inhibitor GSK2656157 (6), which was selected for advancement to preclinical development.

Synthesis and biological activities of novel indole derivatives as potent and selective PPARγ modulators

Starting from the structure of Telmisartan, a new series of potent and selective PPARgamma modulators was identified. The synthesis, in vitro and in vivo evaluation of the most potent compounds are reported and the X-ray structure of compound 7b bound to the PPARgamma ligand binding domain is described.

Structural Analysis of Rev-erbα Bound to NCoR Reveals a Unique Mechanism of Nuclear Receptor-Corepressor Interaction

Repression of gene transcription by the nuclear receptor Rev-erbα plays an integral role in the core molecular circadian clock. We report the crystal structure of a nuclear receptor–co-repressor (N-CoR) interaction domain 1 (ID1) peptide bound to truncated human Rev-erbα ligand-binding domain (LBD). The ID1 peptide forms an unprecedented antiparallel β-sheet with Rev-erbα, as well as an α-helix similar to that seen in nuclear receptor ID2 crystal structures but out of register by four residues. Comparison with the structure of Rev-erbβ bound to heme indicates that ID1 peptide and heme induce substantially different conformational changes in the LBD. Although heme is involved in Rev-erb repression, the structure suggests that Rev-erbα could also mediate repression via ID1 binding in the absence of heme. The previously uncharacterized secondary structure induced by ID1 peptide binding advances our understanding of nuclear receptor–co-repressor interactions.

Reverse Hydroxamate Inhibitors of Bone Morphogenetic Protein 1

Bone Morphogenetic Protein 1 (BMP1) inhibition is a potential method for treating fibrosis because BMP1, a member of the zinc metalloprotease family, is required to convert pro-collagen to collagen. A novel class of reverse hydroxamate BMP1 inhibitors was discovered, and cocrystal structures with BMP1 were obtained. The observed binding mode is unique in that the small molecule occupies the nonprime side of the metalloprotease pocket providing an opportunity to build in metalloprotease selectivity. Structure-guided modification of the initial hit led to the identification of an oral in vivo tool compound with selectivity over other metalloproteases. Due to irreversible inhibition of cytochrome P450 3A4 for this chemical class, the risk of potential drug-drug interactions was managed by optimizing the series for subcutaneous injection.