Abstract
Neutrophils ingest Mycobacteria tuberculosis (Mtb) in the lungs of infected individuals. During phagocytosis they use myeloperoxidase (MPO) to catalyze production of hypochlorous acid (HOCl), their most potent antimicrobial agent. Isoniazid (INH), the foremost antibiotic in the treatment of tuberculosis, is oxidized by MPO. It rapidly reduced compound I of MPO [k=(1.22±0.05)×106M−1s−1] but reacted less favorably with compound II [(9.8±0.6)×102M−1s−1]. Oxidation of INH by MPO and hydrogen peroxide was unaffected by chloride, the physiological substrate for compound I, and the enzyme was partially converted to compound III. This indicates that INH is oxidized outside the classical peroxidation cycle. In combination with superoxide dismutase (SOD), MPO oxidized INH without exogenous hydrogen peroxide. SOD must favor reduction of oxygen by the INH radical to give superoxide and ultimately hydrogen peroxide. In both oxidation systems, an adduct with methionine was formed and it was a major product with MPO and SOD. We show that it is a conjugate of an acyldiimide with amines. INH substantially inhibited HOCl production by MPO and neutrophils below pharmacological concentrations. The reversible inhibition is explained by diversion of MPO to its ferrous and compound III forms during oxidation of INH. MPO, along with SOD released by Mtb, will oxidize INH at sites of infection and their interactions are likely to limit the efficacy of the drug, promote adverse drug reactions via formation of protein adducts, and impair a major bacterial killing mechanism of neutrophils.
Highlights
Isonicotinic acid hydrazide, or isoniazid (INH), remains an important drug in the fight against tuberculosis 60 years after its discovery [1,2]
INH, isonicotinic acid, isonicotinamide, methionine and lysine compounds, taurine, diethylenetriamine penta-acetic acid, tetramethylbenzidine, phorbyl myristate acetate, superoxide dismutase, catalase, and Extra-avidin alkaline phosphatase were all from Sigma–Aldrich (USA). 4-Aminobenzoic acid hydrazide (ABAH) was from Fluka Chemicals (Germany)
It is becoming increasingly apparent that neutrophils play a major role in host defense against Mycobacterium tuberculosis (Mtb) [39]
Summary
Isonicotinic acid hydrazide, or isoniazid (INH), remains an important drug in the fight against tuberculosis 60 years after its discovery [1,2]. Resistance of Mycobacterium tuberculosis (Mtb) to INH, is an increasing clinical problem in the management of this disease worldwide [3]. Given that 30% of the world’s population is infected with Mtb and 10 million new cases occur annually, it is critically important to fully understand the molecular and cellular basis of the action of INH. INH is a prodrug that undergoes oxidation in Mtb by a catalase-peroxidase, KatG [4]. Mutations in KatG that make Mtb resistant to INH blunt superoxide-dependent oxidation of INH but have little impact on peroxide-dependent oxidation [5]. These findings suggest that KatG uses superoxide to oxidize INH within Mtb
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