Direct Binding to NLRP3 Pyrin Domain as a Novel Strategy to Prevent NLRP3‐Driven Inflammation and Gouty Arthritis

Objective The NLRP 3 inflammasome is closely linked to the pathophysiology of a wide range of inflammatory diseases. This study was undertaken to identify small molecules that directly bind to NLRP 3 in order to develop pharmacologic interventions for NLRP 3‐related diseases. Methods A structure‐based virtual screening analysis was performed with ~62,800 compounds to select efficient NLRP 3 inhibitors. The production of caspase 1‐p10 and interleukin‐1β ( IL ‐1β) was measured by immunoblotting and enzyme‐linked immunosorbent assay to examine NLRP 3 inflammasome activation. Two gouty arthritis models and an air pouch inflammation model induced by monosodium urate monohydrate ( MSU ) crystal injection were used for in vivo experiments. Primary synovial fluid cells from gout patients were used to determine the relevance of NLRP 3 inflammasome inhibition in human gout. Results Beta‐carotene (provitamin A) suppressed the NLRP 3 inflammasome activation induced by various activators, including MSU crystals, in mouse bone marrow–derived primary macrophages ( P < 0.05). Surface plasmon resonance analysis demonstrated the direct binding of β‐carotene to the pyrin domain ( PYD ) of NLRP 3 ( K D = 3.41 × 10 −6 ). Molecular modeling and mutation assays revealed the interaction mode between β‐carotene and the NLRP 3 PYD . Inflammatory symptoms induced by MSU crystals were attenuated by oral administration of β‐carotene in gouty arthritis mouse models ( P < 0.05), correlating with its suppressive effects on the NLRP 3 inflammasome in inflamed tissues. Furthermore, β‐carotene reduced IL ‐1β secretion from human synovial fluid cells isolated from gout patients ( P < 0.05), showing its inhibitory efficacy in human gout. Conclusion Our results present β‐carotene as a selective and direct inhibitor of NLRP 3, and the binding of β‐carotene to NLRP 3 PYD as a novel pharmacologic strategy to combat NLRP 3 inflammasome–driven diseases, including gouty arthritis.