High‐Throughput Screening Reveals That CeeNU Acts as a New NLRP3 Inflammasome Inhibitor

Pyroptosis is a special form of cell death that often occurs during excessive inflammation and injury, leading to tissue damage, disease progression, and other related issues. The Nod-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome is an important regulatory factor in cellular pyroptosis that promotes the inflammatory response. Inhibitors targeting the NLRP3 inflammasome have emerged as promising potential therapeutic agents for inflammatory diseases. Through large-scale screening, we found that the FDA-approved drug CeeNU strongly inhibited NLRP3-mediated pyroptosis. CeeNU exhibited dose-dependent suppression of NLRP3 inflammasome activation and effectively mitigated inflammasome-driven pyroptotic cell death in both human and murine macrophages/microglia. Mechanistically, we further demonstrated that CeeNU specifically binds to arginine 335 within the NACHT domain of NLRP3, abrogating NLRP3 inflammasome activation by blocking its assembly. Importantly, CeeNU showed remarkable protective effects in multiple mouse models of NLRP3 inflammasome-mediated diseases, including experimental autoimmune encephalomyelitis (EAE) induced by myelin oligodendrocyte glycoprotein (MOG), lipopolysaccharide (LPS)-induced septic shock, monosodium urate (MSU)-induced peritonitis, and MSU-induced gouty arthritis. Our results demonstrate that CeeNU, a clinically available drug, acts as an NLRP3 inhibitor and holds therapeutic potential for NLRP3 inflammasome-mediated pyroptotic diseases.