Optimization of novel TRPV1 antagonists bearing 1-((2,2-difluorobenzo[d][1,3] dioxol-5-yl) methyl) piperazine for pain management: Mechanistic insights into NLRP3 Inflammasome modulation.

Emerging evidence underscores the pivotal role of the NLRP3 inflammasome in the pathogenesis of inflammatory pain, positioning it as a promising therapeutic target. Herein, we reported the structure-based design, synthesis, and optimization of novel transient receptor potential vanilloid 1 (TRPV1) antagonist bearing a 1-((2,2-difluorobenzo[d][1,3] dioxol-5-yl) methyl) piperazine scaffold, aiming to simultaneously address TRPV1-mediated neurogenic inflammation and NLRP3-dependent inflammatory signaling. A total of 48 compounds were synthesized and lead compound 3q demonstrated potent in vitro TRPV1 antagonism (IC50 = 63.1 ± 9.6 nM) and inhibition of NLRP3 inflammasome activation in THP-1 cells, as evidenced by reduced IL-1β secretion (IC50 = 348.9 ± 69.62 nM). In vivo, 3q significantly alleviated formalin-induced inflammation pain and LPS/ATP-induced acute peritonitis in mice, markedly reducing IL-1β by 51 %. Furthermore, compound 3q exhibited superior efficacy in DSS-induced colitis, resulting in a lower DAI and histological score. Preliminary pharmacokinetic profiling revealed favorable oral bioavailability (F = 34.4 %) and a half-life of 11.04 h in mice. These findings highlight the potential of 3q as an alternative anti-inflammatory analgesic, advancing the development of TRPV1/NLRP3 dual modulatory therapies for inflammatory pain management.