Synthesis and structure–activity relationships of pyrazolodiazepine derivatives as human P2X 7 receptor antagonists

Abstract

Screening of library compounds has yielded pyrazolodiazepine derivatives with P2X 7 receptor antagonist activity. To explore the structure–activity relationships (SAR) of these pyrazolodiazepines as human P2X 7 receptor antagonists, derivatives were synthesized by substitutions at positions R 2 and R 3 of the pyrazolodiazepine skeleton. Using a 2′(3′)- O-(4-benzoylbenzoyl)ATP (BzATP)-induced fluorescent ethidium uptake assay, the activities of these derivatives were tested in HEK-293 cells stably expressing human P2X 7 receptors. Moreover, the effect of these derivatives was assessed by measuring their effect on IL-1β release induced by BzATP-induced activation of differentiated THP-1 cells. A 2-phenethyl pyrazolodiazepine derivative with a 1-methyl-1 H-3-indolyl group at position R 2 had fivefold greater activity than the derivative with a 5-isoquinolinyl at R 2. Moreover, a benzyl moiety at R 3 had fivefold greater activity than a bicyclic moiety. The stereochemical effect at C-6 showed a preference for the ( R)-isomer. Among the series of active derivatives, compound 23b, with a phenethyl group at R 1, a 3-methyl indole at R 2, and a benzyl at R 3, exhibited activity similar to that of the positive control, KN-62, as shown by the inhibitory effects of IL-1β release.