Eosinophil peroxidase oxidizes isoniazid to form the active metabolite against M. tuberculosis, isoniazid-NAD

Chem Biol Interact. 2019 May 25;305:48-53. doi: 10.1016/j.cbi.2019.03.019. Epub 2019 Mar 25.

Abstract

The formation of isonicotinyl-nicotinamide adenine dinucleotide (INH-NAD+) by the mycobacterial catalase-peroxidase enzyme, KatG, was known to be the major component of the mode of action of isoniazid (INH), an anti-tuberculosis drug. However, there are other enzymes that may catalyze this reaction. We have previously reported that neutrophil myeloperoxidase (MPO) is capable of metabolizing INH through the formation of INH-NAD+ adduct, which could be attributed to being a possible mode of action of INH. However, eosinophilic infiltration of the lungs is more pronounced and characteristic of granulomas in Mycobacterium tuberculosis-infected patients. Thus, the aim of the present study is to investigate the role of eosinophil peroxidase (EPO), a key eosinophil enzyme, during INH metabolism and the formation of its active metabolite, INH-NAD+ using purified EPO and eosinophils isolated from asthmatic donors. UV-Vis spectroscopy revealed INH oxidation by EPO led to a new product (λmax = 326 nm) in the presence of NAD+. This adduct was confirmed to be INH-NAD+ using LC-MS analysis where the intact adduct was detected (m/z = 769). Furthermore, EPO catalyzed the oxidation of INH and formed several free radical intermediates as assessed by electron paramagnetic resonance (EPR) spin-trapping; a carbon-centred radical, which is considered to be the reactive metabolite that binds with NAD+, was found when superoxide dismutase was included in the reaction. Our findings suggest that eosinophilic EPO may also play a role in the pharmacological activity of INH through the formation of INH-NAD+ adduct, and supports further evidence that human cells and enzymes are capable of producing the active metabolite involved in tuberculosis treatment.

Keywords: Eosinophil; Free radicals; INH-NAD(+); Isoniazid; Peroxidase.

MeSH terms

  • Asthma / metabolism
  • Asthma / pathology
  • Chromatography, High Pressure Liquid
  • Electron Spin Resonance Spectroscopy
  • Eosinophil Peroxidase / metabolism*
  • Eosinophils / chemistry
  • Eosinophils / drug effects
  • Eosinophils / enzymology*
  • Humans
  • Isoniazid / analogs & derivatives*
  • Isoniazid / blood
  • Isoniazid / chemistry
  • Isoniazid / metabolism*
  • Isoniazid / pharmacology
  • Mass Spectrometry
  • Mycobacterium tuberculosis / drug effects
  • Mycobacterium tuberculosis / pathogenicity
  • NAD / analogs & derivatives*
  • NAD / blood
  • NAD / chemistry
  • NAD / metabolism*
  • Oxidation-Reduction
  • Platelet Activating Factor / pharmacology
  • Superoxide Dismutase / metabolism

Substances

  • Platelet Activating Factor
  • isonicotinyl-NAD
  • NAD
  • Eosinophil Peroxidase
  • Superoxide Dismutase
  • Isoniazid