Mechanistic Studies of Retinoic Acid Receptor-Related Orphan Receptor Modulators by HDX Mass Spectrometry

摘要

Utilizing HDX in the drug discovery process requires a panel of ligands, ideally containing a mixture of full-agonists, partial agonists, and inverse agonists. The array of cellular and biochemical assays which monitor NR activity can be costly, difficult to reproduce, and sometimes produce conflicting results. One of our goals is to correlate cellular and biochemical results with specific HDX profiles such that the drug discovery process can be driven, in part, by mass spectrometry. Current advancements in robotics, MS instrumentation, and software tools for data analysis allow us to generate and analyze HDX data for each ligand in approximately 10 hours of hands-on time and 12 hours of instrument time. In the absence crystallography data, HDX can drive mutational analysis to localize ligand binding sites. For example, Leu287 as well as the region 491-485 in helix 11 show moderate protection by a sub-set of the ligands studied herein. A closer look at the crystal structure of ROR(gamma) suggests the protection in helix 11 is likely due to Arg481 or Leu482.