Abstract
ABSTRACTActive tuberculosis (TB) and latent Mycobacterium tuberculosis infection both require lengthy treatments to achieve durable cures. This problem has partly been attributable to the existence of nonreplicating M. tuberculosis “persisters” that are difficult to kill using conventional anti-TB treatments. Compounds that target the respiratory pathway have the potential to kill both replicating and persistent M. tuberculosis and shorten TB treatment, as this pathway is essential in both metabolic states. We developed a novel respiratory pathway-specific whole-cell screen to identify new respiration inhibitors. This screen identified the biphenyl amide GSK1733953A (DG70) as a likely respiration inhibitor. DG70 inhibited both clinical drug-susceptible and drug-resistant M. tuberculosis strains. Whole-genome sequencing of DG70-resistant colonies identified mutations in menG (rv0558), which is responsible for the final step in menaquinone biosynthesis and required for respiration. Overexpression of menG from wild-type and DG70-resistant isolates increased the DG70 MIC by 4× and 8× to 30×, respectively. Radiolabeling and high-resolution mass spectrometry studies confirmed that DG70 inhibited the final step in menaquinone biosynthesis. DG70 also inhibited oxygen utilization and ATP biosynthesis, which was reversed by external menaquinone supplementation. DG70 was bactericidal in actively replicating cultures and in a nutritionally deprived persistence model. DG70 was synergistic with the first-line TB drugs isoniazid, rifampin, and the respiratory inhibitor bedaquiline. The combination of DG70 and isoniazid completely sterilized cultures in the persistence model by day 10. These results suggest that MenG is a good therapeutic target and that compounds targeting MenG along with standard TB therapy have the potential to shorten TB treatment duration.
Highlights
IMPORTANCE This study shows that MenG, which is responsible for the last enzymatic step in menaquinone biosynthesis, may be a good drug target for improving TB treatments
To identify novel inhibitors of mycobacterial respiration, we developed a whole-cell respiratory pathway-specific screen that combined the advantages of a whole-cell screen with those of a target-based approach [18]
A Novel Inhibitor of MenG in M. tuberculosis the transition to anaerobiosis in M. tuberculosis [19, 20]. We used this reporter strain to detect possible respiratory inhibitors that are able to penetrate into live mycobacterial cells
Summary
IMPORTANCE This study shows that MenG, which is responsible for the last enzymatic step in menaquinone biosynthesis, may be a good drug target for improving TB treatments. New anti-TB drugs with the ability to kill M. tuberculosis in both replicating and nonreplicating states could be used to treat drug-resistant TB and potentially to reduce the required duration of treatment. M. tuberculosis cannot support its energy needs through substrate-level phosphorylation Instead, both actively growing and nonreplicating persistent M. tuberculosis bacteria are dependent on respiration to synthesize adequate amounts of ATP [12]. The M. tuberculosis respiratory chain consists of various electron donors that transfer two electrons to lipoquinone with the help of corresponding dehydrogenases. We report a novel anti-TB chemotype, GSK1733953A (here renamed DG70), which was discovered through this screen; show that DG70 acts through inhibition of MenG; and investigate the effects and potential therapeutic utility of MenG inhibition
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