RESUMO
BACKGROUND AND PURPOSE: The histamine H4 receptor, originally thought to signal merely through Gαi proteins, has recently been shown to also recruit and signal via ß-arrestin2. Following the discovery that the reference antagonist indolecarboxamide JNJ 7777120 appears to be a partial agonist in ß-arrestin2 recruitment, we have identified additional biased hH4R ligands that preferentially couple to Gαi or ß-arrestin2 proteins. In this study, we explored ligand and receptor regions that are important for biased hH4R signalling. EXPERIMENTAL APPROACH: We evaluated a series of 48 indolecarboxamides with subtle structural differences for their ability to induce hH4R-mediated Gαi protein signalling or ß-arrestin2 recruitment. Subsequently, a Fingerprints for Ligands and Proteins three-dimensional quantitative structure-activity relationship analysis correlated intrinsic activity values with structural ligand requirements. Moreover, a hH4R homology model was used to identify receptor regions important for biased hH4R signalling. KEY RESULTS: One indolecarboxamide (75) with a nitro substituent on position R7 of the aromatic ring displayed an equal preference for the Gαi and ß-arrestin2 pathway and was classified as unbiased hH4R ligand. The other 47 indolecarboxamides were ß-arrestin2-biased agonists. Intrinsic activities of the unbiased as well as ß-arrestin2-biased indolecarboxamides to induce ß-arrestin2 recruitment could be correlated with different ligand features and hH4R regions. CONCLUSION AND IMPLICATIONS: Small structural modifications resulted in diverse intrinsic activities for unbiased (75) and ß-arrestin2-biased indolecarboxamides. Analysis of ligand and receptor features revealed efficacy hotspots responsible for biased-ß-arrestin2 recruitment. This knowledge is useful for the design of hH4R ligands with biased intrinsic activities and aids our understanding of the mechanism of H4R activation.
