Development of a Conformational Histamine H3 Receptor Biosensor for the Synchronous Screening of Agonists and Inverse Agonists

The histamine H₃ receptor (H₃R) represents a highly attractive drug target for the treatment of various central nervous system disorders, but the discovery of novel H₃R targeting compounds relies on the assessment of highly amplified intracellular signaling events that do not only reflect H₃R modulation and carry the risk of high false-positive and -negative screening rates. To address these limitations, we designed an intramolecular H₃R biosensor based on the principle of bioluminescence resonance energy transfer (BRET) that reports the receptor's real-time conformational dynamics and provides an advanced tool to screen for both H₃R agonists and inverse agonists in a live cell screening-compatible assay format. This conformational G-protein-coupled receptor (GPCR) sensor allowed us to characterize the pharmacological properties of known and new H₃ receptor ligands with unprecedented accuracy. Interestingly, we found that one newly developed H₃ receptor ligand possesses even stronger inverse agonistic activity than reference H₃R inverse agonists including the current gold standard pitolisant. Taken together, we describe here the design and validation of the first screening-compatible H₃R conformational biosensor that will aid in the discovery of novel H₃R ligands and can be employed to gain deeper insights into the (in-)activation mechanism of this highly attractive drug target.

BRET; GPCR; conformational sensor; drug discovery; histamine receptor; inverse agonist.