Structural requirements for inverse agonism and neutral antagonism of indole-, benzimidazole-, and thienopyrrole-derived histamine H4 receptor ligands.

The human histamine H(4) receptor (hH(4)R), coexpressed with Galpha(i2) and Gbeta(1)gamma(2) in Sf9 insect cells, is highly constitutively active, and thioperamide [THIO; N-cyclohexyl-4-(imidazol-4-yl)-1-piperidinecarbothioamide] is one of the most efficacious hH(4)R inverse agonists. High constitutive hH(4)R activity may have pathophysiological implications in which case inverse agonists may behave differently than neutral antagonists. To learn more about the structural requirements for hH(4)R inverse agonism, we investigated 25 compounds (indole, benzimidazole, and thienopyrrole derivatives) structurally related to the standard antagonist JNJ-7777120 [1-[(5-chloro-1H-indol-2-yl)carbonyl]-4-methyl-piperazine]. We characterized the compounds in radioligand binding assays by using [(3)H]histamine ([(3)H]HA) and in steady-state GTPase assays in the presence (antagonist mode) and absence (inverse agonist mode) of the agonist HA, yielding the following results: 1) Twenty-two compounds were inverse agonists (efficacy: 15-62% of the THIO effect), and only three compounds (12%) showed neutral antagonism. Thus, inverse agonism is far more common than neutral antagonism. 2) The inverse agonistic efficacy of the R5-monosubstituted indole-derived compounds increased with the volume of R5. R5 may interact with Trp(6.48) of the rotamer toggle switch and stabilize the inactive receptor conformation. 3) A subset of compounds showed large differences between the K(i) value from [(3)H]HA competition binding and the EC(50) value from steady-state GTPase assays, whereas the K(b) values were closer to the K(i) values. Thus, the two-state model should be extended to a model comprising a constitutively active hH(4)R state, which can be discriminated by inverse agonists from a structurally distinct HA-stabilized active state.