Characterization of Clofazimine Metabolism in Human Liver Microsomal Incubation In Vitro

Clofazimine [N,5-bis(4-chlorophenyl)-3-[(propane-2-yl)rimino]-3,5-dihydrophenazin-2-amine] is an antimycobacterial agent used as a second-line antituberculosis (anti-TB) drug. Nonetheless, little information is known about the metabolic routes of clofazimine, and the enzymes involved in metabolism. This study aimed to characterize the metabolic pathways and enzymes responsible for the metabolism of clofazimine in human liver microsomes. Eight metabolites, including four oxidative metabolites, three glucuronide conjugates, and one sulfate conjugate were identified, and their structures were deduced based on tandem mass spectrometry (MS/MS) spectra. Hydroxylated clofazimine and hydrated clofazimine was generated even in the absence of the NADPH generating system presumably via a nonenzymatic pathway. Hydrolytic-dehalogenated clofazimine was catalyzed mainly by CYP1A2 whereas hydrolytic-deaminated clofazimine was formed by CYP3A4/A5. In case of glucuronide conjugates, UGT1A1, UGT1A3, and UGT1A9 showed catalytic activity toward hydroxylated and hydrated clofazimine glucuronide whereas hydrolytic-deaminated clofazimine glucuronide was catalyzed by UGT1A4, UGT1A9, UGT1A3, and UGT2B4. Our results suggested that CYP1A2 and CYP3A are involved in the formation of oxidative metabolites while UGT1A1, 1A3, 1A4, 1A9, and 2B4 are involved in the formation of glucuronide conjugates of oxidative metabolites of clofazimine.