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Publication Abstract from PubMed

BACKGROUND: The nuclear hormone receptor RORgamma regulates transcriptional genes involved in the production of the pro-inflammatory interleukin IL-17 which has been linked to autoimmune diseases such as rheumatoid arthritis, multiple sclerosis and inflammatory bowel disease. This transcriptional activity of RORgamma is modulated through a protein-protein interaction involving the activation function 2 (AF2) helix on the ligand binding domain of RORgamma and a conserved LXXLL helix motif on coactivator proteins. Our goal was to develop a RORgamma specific inverse agonist that would help down regulate pro-inflammatory gene transcription by disrupting the protein protein interaction with coactivator proteins as a therapeutic agent. RESULTS: We identified a novel series of synthetic benzoxazinone ligands having an agonist (BIO592) and inverse agonist (BIO399) mode of action in a FRET based assay. We show that the AF2 helix of RORgamma is proteolytically sensitive when inverse agonist BIO399 binds. Using x-ray crystallography we show how small modifications on the benzoxazinone agonist BIO592 trigger inverse agonism of RORgamma. Using an in vivo reporter assay, we show that the inverse agonist BIO399 displayed specificity for RORgamma over ROR sub-family members alpha and beta. CONCLUSION: The synthetic benzoxazinone ligands identified in our FRET assay have an agonist (BIO592) or inverse agonist (BIO399) effect by stabilizing or destabilizing the agonist conformation of RORgamma. The proteolytic sensitivity of the AF2 helix of RORgamma demonstrates that it destabilizes upon BIO399 inverse agonist binding perturbing the coactivator protein binding site. Our structural investigation of the BIO592 agonist and BIO399 inverse agonist structures identified residue Met358 on RORgamma as the trigger for RORgamma specific inverse agonism.

Structural determinant for inducing RORgamma specific inverse agonism triggered by a synthetic benzoxazinone ligand.,Marcotte DJ, Liu Y, Little K, Jones JH, Powell NA, Wildes CP, Silvian LF, Chodaparambil JV BMC Struct Biol. 2016 Jun 1;16(1):7. doi: 10.1186/s12900-016-0059-3. PMID:27246200^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.