Metabolic Activation of Thiacetazone by the Flavin Monooxygenase EtaA of Mycobacterium tuberculosis

Abstract

Thiacetazone (TAZ) and ethionamide (ETA) are thioamide-containing second line antitubercular prodrugs for which there is an impressive clinical history of cross-resistance in M. tuberculosis and M. leprae. The recently identified FAD-containing monooxygenase EtaA has previously been shown to oxidize ETA to a sulfenic acid (-SOH) and eventually to the amide. EtaA is shown here to also be responsible for activation of TAZ. The two metabolites identified in the in vitro oxidation of TAZ by heterologously expressed EtaA are a sulfinic acid (-SO2H) and a carbodiimide (-N=C=NH), both of which apparently derive from an initially formed but not detected sulfenic acid intermediate. The pH dependence of the oxidation of TAZ by EtaA demonstrated that under basic conditions formation of the carbodiimide metabolite is favored, whereas under neutral or acidic conditions the sulfinic acid is preferred. The sulfenic acid intermediate undergoes redox cycling with glutathione whereas the carbodiimide metabolite forms a covalent adduct with glutathione. Analogous reactions are observed with the human flavin monooxygenases FMO1 and FMO2. The metabolites formed provide a possible rationale for the cytotoxicity of TAZ. This work was supported by NIH grant GM56531.