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The diarylquinoline R207910 (TMC207) is a promising candidate in clinical development for the treatment of tuberculosis. Though R207910-resistant mycobacteria bear mutations in ATP synthase, the compound's precise target is not known. Here we establish by genetic, biochemical and binding assays that the oligomeric subunit c (AtpE) of ATP synthase is the target of R207910. Thus targeting energy metabolism is a new, promising approach for antibacterial drug discovery.

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The diarylquinoline TMC207 kills Mycobacterium tuberculosis by specifically inhibiting ATP synthase. We show here that human mitochondrial ATP synthase (50% inhibitory concentration [IC 50 ] of >200 M) displayed more than 20,000-fold lower sensitivity for TMC207 compared to that of mycobacterial ATP synthase (IC 50 of 10 nM). Also, oxygen consumption in mouse liver and bovine heart mitochondria showed very low sensitivity for TMC207. These results suggest that TMC207 may not elicit ATP synthesis-related toxicity in mammalian cells. ATP synthase, although highly conserved between prokaryotes and eukaryotes, may still qualify as an attractive antibiotic target.A new series of compounds, the diarylquinolines, was reported to be highly active against Mycobacterium tuberculosis (3). TMC207, the lead compound of the diarylquinoline series, displays MICs of 30 nM for M. tuberculosis and 15 nM for Mycobacterium smegmatis. We recently demonstrated that TMC207 targets ATP synthase, the enzyme responsible for ATP production by oxidative phosphorylation (11). The inhibition of ATP synthase by TMC207 was highly specific, with a 50% inhibitory concentration (IC 50 ) for this enzyme corresponding to the MIC for bacterial growth inhibition (11). This compound, which efficiently kills replicating as well as dormant mycobacteria (12,18), is currently in clinical development in phase IIb trials in patients with multidrug-resistant tuberculosis.An important factor to consider for a new antibacterial drug is the lack of a eukaryotic homologue of the target, as inhibition of a homologous enzyme could lead to toxicity and safety concerns in humans. In the case of TMC207, the target enzyme ATP synthase is essential for survival in higher organisms, as it supplies cells with the bulk of their ATP via oxidative phosphorylation (20). ATP synthase is evolutionarily strongly conserved among prokaryotes and eukaryotes. Universally, ATP synthesis is coupled to the flow of protons from the intercristae region in mitochondria and the periplasmic space in bacteria to the mitochondrial matrix and the bacterial cytoplasm, respectively. Subunit c of ATP synthase, forming a membrane-spanning oligomer, is essential for this proton transport (8).TMC207 binds to subunit c of mycobacterial ATP synthase (11). Several natural compounds, such as oligomycin and venturicidin, are known to block ATP synthase action by interaction with subunit c. However, these compounds are not selective and inhibit ATP synthase not only in bacteria but also in mitochondria (14,15). This lack of selectivity prevents their clinical usage due to toxicity issues and fatality concerns. Mitochondrial toxicity is a major concern in the clinical development of new drugs, as it may lead to disease conditions, such as pancreatitis, peripheral neuropathy, and cardial or skeletal myopathies (1, 21).Hence, it is of key importance to investigate the selectivity of TMC207 towards mycobacterial ATP synthase compared with that towards mitochondrial ATP synthase.Mycobacterium smegmatis mc 2 ...

Targeting respiration and ATP synthesis has received strong interest as a new strategy for combatting drug-resistant Mycobacterium tuberculosis. Mycobacteria employ a respiratory chain terminating with two branches. One of the branches includes a cytochrome bc1 complex and an aa3-type cytochrome c oxidase while the other branch terminates with a cytochrome bd-type quinol oxidase. In this communication we show that genetic inactivation of cytochrome bd, but not of cytochrome bc1, enhances the susceptibility of Mycobacterium smegmatis to hydrogen peroxide and antibiotic-induced stress. The type-II NADH dehydrogenase effector clofazimine and the ATP synthase inhibitor bedaquiline were bacteriostatic against wild-type M. smegmatis, but strongly bactericidal against a cytochrome bd mutant. We also demonstrated that the quinone-analog aurachin D inhibited mycobacterial cytochrome bd at sub-micromolar concentrations. Our results identify cytochrome bd as a key survival factor in M. smegmatis during antibiotic stress. Targeting the cytochrome bd respiratory branch therefore appears to be a promising strategy that may enhance the bactericidal activity of existing tuberculosis drugs.