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Pharmacological targeting of NLRP3 deubiquitination for treatment of NLRP3-associated inflammatory diseases

Thiolutin, an inhibitor of the BRISC deubiquitinating enzyme complex, specifically and potently inhibits NLRP3 inflammasome activation. DUBbing new inflammasome inhibitors Inflammasome assembly and activation leading to mature IL-1β release is dysregulated in a wide range of inflammatory diseases. Optimal activation of the NLRP3 inflammasome requires the activity of BRISC, a deubiquitinating enzyme (DUB) complex composed of four protein subunits including BRCC3. Ren et al. demonstrate that the compound thiolutin, a zinc chelator that inhibits JAMM domain–containing metalloproteases including BRCC3, can potently inhibit NLRP3 deubiquitination and inflammasome activation. Thiolutin was effective at inhibiting NLRP3 activation and preventing IL-1β production in multiple mouse models of inflammatory disease, including a model of diet-induced nonalcoholic fatty liver disease. Holomycin, a derivative of thiolutin with reduced toxicity, was also effective at inhibiting NLRP3, paving the way to develop novel agents that selectively target deubiquitination of NLRP3 to regulate its activity. Pharmacologically inhibiting nucleotide-binding domain and leucine-rich repeat-containing (NLR) family, pyrin domain–containing protein 3 (NLRP3) inflammasome activation results in potent therapeutic effects in a wide variety of preclinical inflammatory disease models. NLRP3 deubiquitination is essential for efficient NLRP3 inflammasome activity, but it remains unclear whether this process can be harnessed for therapeutic benefit. Here, we show that thiolutin (THL), an inhibitor of the JAB1/MPN/Mov34 (JAMM) domain–containing metalloprotease, blocks NLRP3 inflammasome activation by canonical, noncanonical, alternative, and transcription-independent pathways at nanomolar concentrations. In addition, THL potently inhibited the activation of multiple NLRP3 mutants linked with cryopyrin-associated periodic syndromes (CAPS). Treatment with THL alleviated NLRP3-related diseases in mouse models of lipopolysaccharide-induced sepsis, monosodium urate–induced peritonitis, experimental autoimmune encephalomyelitis, CAPS, and methionine-choline–deficient diet-induced nonalcoholic fatty liver disease. Mechanistic studies revealed that THL inhibits the BRCC3-containing isopeptidase complex (BRISC)–mediated NLRP3 deubiquitination and activation. In addition, we show that holomycin, a natural methyl derivative of THL, displays an even higher inhibitory activity against NLRP3 inflammasome than THL. Our study validates that posttranslational modification of NLRP3 can be pharmacologically targeted to prevent or treat NLRP3-associated inflammatory diseases. Future clinical development of derivatives of THL may provide new therapies for NLRP3-related diseases.