title: Identification, Synthesis, and Biological Evaluation of the Major Human Metabolite of NLRP3 Inflammasome Inhibitor MCC950.
abstract: MCC950 is an orally bioavailable small molecule inhibitor of the NOD-like receptor pyrin domain-containing protein 3 (NLRP3) inflammasome that exhibits remarkable activity in multiple models of inflammatory disease. Incubation of MCC950 with human liver microsomes, and subsequent analysis by HPLC-MS/MS, revealed a major metabolite, where hydroxylation of MCC950 had occurred on the 1,2,3,5,6,7-hexahydro-s-indacene moiety. Three possible regioisomers were synthesized, and coelution using HPLC-MS/MS confirmed the structure of the metabolite. Further synthesis of individual enantiomers and coelution studies using a chiral column in HPLC-MS/MS showed the metabolite wasR-(+)-N-((1-hydroxy-1,2,3,5,6,7-hexahydro-s-indacen-4-yl)carbamoyl)-4-(2-hydroxypropan-2-yl)furan-2-sulfonamide (2a). Incubation of MCC950 with a panel of cytochrome P450 enzymes showed P450s 2A6, 2C9, 2C18, 2C19, 2J2, and 3A4 catalyze the formation of the major metabolite2a, with a lower level of activity shown by P450s 1A2 and 2B6. All of the synthesized compounds were tested for inhibition of NLRP3-induced production of the pro-inflammatory cytokine IL-1β from human monocyte derived macrophages. The identified metabolite2awas 170-fold less potent than MCC950, while one regioisomer had nanomolar inhibitory activity. These findings also give first insight into the SAR of the hexahydroindacene moiety.
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland , Brisbane, Queensland 4072, Australia.
authors.forename: Manohar
authors.initials: M
authors.lastname: Salla
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland , Brisbane, Queensland 4072, Australia.
authors.forename: Mark S
authors.initials: MS
authors.lastname: Butler
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland , Brisbane, Queensland 4072, Australia.
authors.forename: Ruby
authors.initials: R
authors.lastname: Pelingon
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland , Brisbane, Queensland 4072, Australia.
authors.forename: Geraldine
authors.initials: G
authors.lastname: Kaeslin
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland , Brisbane, Queensland 4072, Australia.
authors.forename: Daniel E
authors.initials: DE
authors.lastname: Croker
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland , Brisbane, Queensland 4072, Australia.
authors.forename: Janet C
authors.initials: JC
authors.lastname: Reid
authors.affiliation: School of Chemistry and Molecular Biosciences, The University of Queensland , Brisbane, Queensland 4072, Australia.
authors.forename: Jong Min
authors.initials: JM
authors.lastname: Baek
authors.affiliation: School of Chemistry and Molecular Biosciences, The University of Queensland , Brisbane, Queensland 4072, Australia.
authors.forename: Paul V
authors.initials: PV
authors.lastname: Bernhardt
authors.affiliation: School of Chemistry and Molecular Biosciences, The University of Queensland , Brisbane, Queensland 4072, Australia.
authors.forename: Elizabeth M J
authors.initials: EM
authors.lastname: Gillam
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland , Brisbane, Queensland 4072, Australia.
authors.forename: Matthew A
authors.initials: MA
authors.lastname: Cooper
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland , Brisbane, Queensland 4072, Australia.
authors.forename: Avril A B
authors.initials: AA
authors.lastname: Robertson
keywords: MCC950; NLRP3; cytochrome P450; inflammasome; metabolite; microsome