Screening for ligands of human retinoid X receptor-[alpha] using ultrafiltration mass spectrometry

Retinoid X receptors (RXRs) function as ligand-activated transcription factors and are obligatory components of a large number of nuclear receptor heterodimers. RXRs help regulate diverse physiological responses including the cancer prevention responses of cell proliferation, inflammation, cell differentiation, and apoptosis. Since RXRs represent important targets for cancer chemoprevention, an ultrafiltration mass spectrometry-based assay was developed to facilitate the discovery of potential chemoprevention agents that bind to human RXR[alpha]. Natural and synthetic ligands for RXR[alpha] including 9-cis-retinoic acid, docosahexaenoic acid, and LG100268 could be detected and identified in DMSO (dimethyl sulfoxide) or even complex matrixes such as extracts of marine bacteria. Specific binding of ligands to RXR[alpha] was demonstrated through competitive binding using ultrafiltration LC-MS/ MS (liquid chromatography-tandem mass spectrometry), and ligands could be ranked in order of affinity for RXR[alpha]. Therefore, ultrafiltration LC-MS/MS is suitable for the screening of complex mixtures such as natural product extracts for the discovery of new ligands to RXR[alpha].