MCC950/CRID3 potently targets the NACHT domain of wild-type NLRP3 but not disease-associated mutants for inflammasome inhibition

Abstract

The nucleotide-binding-domain (NBD)–and leucine-rich repeat (LRR)–containing (NLR) family, pyrin-domain–containing 3 (NLRP3) inflammasome drives pathological inflammation in a suite of autoimmune, metabolic, malignant, and neurodegenerative diseases. Additionally, NLRP3 gain-of-function point mutations cause systemic periodic fever syndromes that are collectively known as cryopyrin-associated periodic syndrome (CAPS). There is significant interest in the discovery and development of diarylsulfonylurea Cytokine Release Inhibitory Drugs (CRIDs) such as MCC950/CRID3, a potent and selective inhibitor of the NLRP3 inflammasome pathway, for the treatment of CAPS and other diseases. However, drug discovery efforts have been constrained by the lack of insight into the molecular target and mechanism by which these CRIDs inhibit the NLRP3 inflammasome pathway. Here, we show that the NAIP, CIITA, HET-E, and TP1 (NACHT) domain of NLRP3 is the molecular target of diarylsulfonylurea inhibitors. Interestingly, we find photoaffinity labeling (PAL) of the NACHT domain requires an intact (d)ATP-binding pocket and is substantially reduced for most CAPS-associated NLRP3 mutants. In concordance with this finding, MCC950/CRID3 failed to inhibit NLRP3-driven inflammatory pathology in two mouse models of CAPS. Moreover, it abolished circulating levels of interleukin (IL)-1β and IL-18 in lipopolysaccharide (LPS)-challenged wild-type mice but not in Nlrp3L351P knock-in mice and ex vivo-stimulated mutant macrophages. These results identify wild-type NLRP3 as the molecular target of MCC950/CRID3 and show that CAPS-related NLRP3 mutants escape efficient MCC950/CRID3 inhibition. Collectively, this work suggests that MCC950/CRID3-based therapies may effectively treat inflammation driven by wild-type NLRP3 but not CAPS-associated mutants.

Therapeutic modulation of inflammasome pathways

“…47,116,[120][121][122][123] Interestingly, MCC950/CRID3 and related diarylsulfonylurea compounds may be substantially less effective in inhibiting CAPS-related NLRP3 mutants because in vivo MCC950/CRID3 concentrations that potently inhibited NLRP3-driven inflammation in wildtype mice were shown to be ineffective in two in vivo mouse models of CAPS disease. 119 Concordantly, MCC950/CRID3-derived photoaffinity probes labeled the NACHT domain of wildtype NLRP3, but not CAPS-associated NLRP3 mutants. 119 These results raise the possibility that MCC950/CRID3-based therapies may need to be dosed at higher concentrations in CAPS patients to effectively treat inflammatory pathology than is needed to curb inflammatory pathology driven by wildtype NLRP3.…”

“…119 Concordantly, MCC950/CRID3-derived photoaffinity probes labeled the NACHT domain of wildtype NLRP3, but not CAPS-associated NLRP3 mutants. 119 These results raise the possibility that MCC950/CRID3-based therapies may need to be dosed at higher concentrations in CAPS patients to effectively treat inflammatory pathology than is needed to curb inflammatory pathology driven by wildtype NLRP3.…”

“…78,116 More recently, MCC950/CRID3 and related tool compounds were shown to directly target the NACHT domain of NLRP3 in the vicinity of its ATP/dATP binding pocket, and they represent the most potent tool compounds that are currently available for selective NLRP3 inflammasome inhibition in human cells and preclinical rodent studies. [117][118][119] Consequently, MCC950/CRID3-mediated inhibition of NLRP3 has shown beneficial effects in mice and other preclinical species that have been subjected to disease models, among which models of atherosclerosis, myocardial infarction, colitis, and skin and airway inflammation. 47,116,[120][121][122][123] Interestingly, MCC950/CRID3 and related diarylsulfonylurea compounds may be substantially less effective in inhibiting CAPS-related NLRP3 mutants because in vivo MCC950/CRID3 concentrations that potently inhibited NLRP3-driven inflammation in wildtype mice were shown to be ineffective in two in vivo mouse models of CAPS disease.…”

Selective inhibition of NLRP3 inflammasome by designed peptide originating from ASC

“…Sulfonylureabased compound MCC950 (CRID3) 20 seems to be the most potent so far identified NLRP3 inflammasome inhibitor 21 which binds to Walker B motif. [67][68][69] MCC950 was also extensively tested in preclinical models and entered phase II clinical trial for rheumatoid arthritis when liver toxicity was observed, however, relatively large dose was applied. 63 Further direct NLRP3 inhibitors include oridonin, which was shown to bind Cys279 in NLRP3 NACHT domain and abrogate NLRP3-NEK7 interaction.…”

Novel Analgesics with Peripheral Targets

“…However, tranilast can induce cystitis [712], which might be partial agonistic through TRPA1 [713]. MCC950 [714][715][716] and dapansutrile have reached phase II [717] and could be a promising new approach.…”