A derivative of PD156707 selectively inhibits NLRP3 inflammasome activation by directly binding to NLRP3

The NLRP3 inflammasome is a central driver of inflammatory pathology, yet clinical translation of small-molecule inhibitors remains limited by selectivity and safety. Through a 3000-compound screen, we identified PD-156707 as an apparent inhibitor of NLRP3 signaling; however, activity was not reproduced with freshly purchased material. Analytical profiling of the archived sample revealed a previously unrecognized storage-derived derivative, 3-(4-methoxyphenyl)-1-(3,4-methylenedioxyphenyl)-2-(3,4,5-trimethoxybenzylidene)propane-1,3-dione (DPD, 1). DPD, synthesized in a two-step sequence and confirmed to be identical to the degradation product derived from PD-156707, selectively suppressed NLRP3 inflammasome activation in LPS-primed macrophages stimulated with nigericin, ATP, silica, or imiquimod, while sparing AIM2 and NLRC4 pathways. DPD reduced caspase-1 processing and downstream maturation of IL-1β and GSDMD, and impeded ASC oligomerization and speck formation without affecting ASC self-oligomerization in an overexpression system. A DARTS assay indicated direct target engagement of NLRP3, whereas caspase-1 protection was not observed. In silico docking positioned DPD in the NACHT domain (PDB: 7ALV) with a predicted binding affinity of − 9.2 kcal/mol—comparable to MCC950 (– 9.5 kcal/mol)—and suggested interactions with Arg351 within the ATP-binding region. In vivo, DPD reduced IL-1β—but not IL-6—in LPS-induced sepsis and lowered peritoneal IL-1β in MSU-induced peritonitis. Collectively, these data identify DPD as a selective, direct-binding modulator of NLRP3 that disrupts ATP-dependent conformational activation, establishing DPD as a promising lead for inflammasome-targeted therapeutics.