Inducing vulnerability to InhA inhibition restores isoniazid susceptibility in drug resistantMycobacterium tuberculosis

Abstract

AbstractOf the approximately 10 million cases ofMycobacterium tuberculosis(Mtb) infections each year, over 10% are resistant to the frontline antibiotic isoniazid (INH). INH resistance is predominantly caused by mutations that decrease the activity of the bacterial enzyme KatG, which mediates conversion of the pro-drug INH to its active form INH-NAD. We previously discovered an inhibitor ofMtbrespiration, C10, that enhances the bactericidal activity of INH, prevents the emergence of INH-resistant mutants, and re-sensitizes a collection of INH-resistant mutants to INH through an unknown mechanism. To investigate the mechanism of action of C10, we exploited the toxicity of high concentrations of C10 to select for resistant mutants. We discovered two mutations that confer resistance to the disruption of energy metabolism and allow for growth ofMtbin high C10 concentrations, indicating that growth inhibition by C10 is associated with inhibition of respiration. Using these mutants as well as direct inhibitors of theMtbelectron transport chain, we provide evidence that inhibition of energy metabolism by C10 is neither sufficient nor necessary to potentiate killing by INH. Instead, we find that C10 acts downstream of INH-NAD synthesis, causingMtbto become particularly sensitive to inhibition of the INH-NAD target, InhA, without changing the concentration of INH-NAD or the activity of InhA, the two predominant mechanisms of potentiating INH. Our studies revealed that there exists a vulnerability inMtbthat can be exploited to renderMtbsensitive to otherwise subinhibitory concentrations of InhA inhibitor.SignificanceIsoniazid (INH) is a critical frontline antibiotic to treatMycobacterium tuberculosis(Mtb) infections. INH efficacy is limited by its suboptimal penetration of theMtb-containing lesion and by the prevalence of clinical INH-resistance. We previously discovered a compound, C10, that enhances the bactericidal activity of INH, prevents the emergence of INH-resistant mutants, and re-sensitizes a set of INH-resistant mutants to INH. Resistance is typically mediated bykatGmutations that decrease the activation of INH, which is required for INH to inhibit the essential enzyme InhA. Our current work demonstrates that C10 re-sensitizes INH-resistantkatG-hypomorphs without enhancing the activation of INH. We furthermore show that C10 causesMtbto become particularly vulnerable to InhA inhibition without compromising InhA activity on its own. Therefore, C10 represents a novel strategy to curtail the development of INH resistance and to sensitizeMtbto sub-lethal doses of INH, such as those achieved at the infection site.