title: Development of time resolved fluorescence resonance energy transfer-based assay for FXR antagonist discovery.
abstract: FXR (farnesoid X receptor, NRIH4), a nuclear receptor, plays a major role in the control of cholesterol metabolism. FXR ligands have been investigated in preclinical studies for targeted therapy against metabolic diseases, but have shown limitations. Therefore, there is a need for new agonist or antagonist ligands of FXR, both for potential clinical applications, as well as to further elucidate its biological functions. Here we describe the use of the X-ray crystal structure of FXR complexed with the potent small molecule agonist GW4064 to design and synthesize a novel fluorescent, high-affinity probe (DY246) for time resolved fluorescence resonance energy transfer (TR-FRET) assays. We then used the TR-FRET assay for high throughput screening of a library of over 5000 bioactive compounds. From this library, we identified 13 compounds that act as putative FXR transcriptional antagonists.
authors.affiliation: Department of Diabetes, Endocrinology and Metabolism, Beckman Research Institute, City of Hope National Medical Center, Duarte, CA 91010, USA. dyu@coh.org
authors.forename: Donna D
authors.initials: DD
authors.lastname: Yu
authors.forename: Wenwei
authors.initials: W
authors.lastname: Lin
authors.forename: Taosheng
authors.initials: T
authors.lastname: Chen
authors.forename: Barry M
authors.initials: BM
authors.lastname: Forman
mesh_terms: D001681:Biological Assay / Q000379:methods*; D055808:Drug Discovery; D005452:Fluoresceins / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D007555:Isoxazoles / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology; D015394:Molecular Structure; D011485:Protein Binding / Q000187:drug effects; D018160:Receptors, Cytoplasmic and Nuclear / Q000819:agonists / Q000037:antagonists & inhibitors*; D000099150:Receptor, Farnesoid X-Activated
chemical_list: D005452:Fluoresceins; D005456:Fluorescent Dyes; D007555:Isoxazoles; D018160:Receptors, Cytoplasmic and Nuclear; D000099150:Receptor, Farnesoid X-Activated; C585737:DY246 compound; C412815:GW 4064