Site-specific Production of Hydroxyl Radicals and Synergistic DNA Damage Induced by the Non-enzymatic Activation of the Anti-tuberculosis Drug Isoniazid by Cu(II)

摘要

Isoniazid (INH) has been recognized as a frontline anti-tuberculosis drug against Mycobacterium tuberculosis (Mtb), which is readily activated by Mtb catalase peroxidase enzyme to produce the reactive isonicotinic acyl radical intermediate responsible for its anti-tuberculosis activity. However, it is not clear whether INH can be activated non-enzymatically by Cu(II). Here we found that INH and Cu(II) together could induce synergistic DNA damage, including DNA strand breakage and 8-oxodG formation, while neither of them alone has any effect. DNA damage induced by INH/Cu(II) could be inhibited by Cu(I)-specific chelator and catalase, but not by SOD and the typical ? OH radical scavengers. Interestingly, ? OH, ? H, C- and N-centered radicals were found to be produced during Cu(II)-catalyzed oxidation of INH by ESR spin-trapping method. The C-centered radical was further unequivocally identified by ESR and ESI-Q-TOF-MS as isonicotinic acyl radical, which can react with nicotinamide coenzyme NADH to form the critical isonicotinic acyl-NAD adducts. Low temperature ESR studies showed that Cu(II) was reduced to Cu(I) by INH. We proposed that the synergistic DNA damage induced by INH/Cu(II) might be due to the synergistic and site-specific production of ? OH near the binding site of copper and DNA. This study provided a new insight on non-enzymatic activation of INH by Cu(II), which may have important biological implications for future research on INH.