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Buruli ulcer (BU), caused by Mycobacterium ulcerans, is currently treated with rifampin–streptomycin or rifampin–clarithromycin daily for 8 weeks recommended by World Health Organization (WHO). These options are lengthy with severe side effects. A new anti-tuberculosis drug, TB47, targeting QcrB in cytochrome bc1:aa3 complex is being developed in China. TB47-containing regimens were evaluated in a well-established murine model using an autoluminescent M. ulcerans strain. High-level TB47-resistant spontaneous M. ulcerans mutants were selected and their qcrB genes were sequenced. The in vivo activities of TB47 against both low-level and high-level TB47-resistant mutants were tested in BU murine model. Here, we show that TB47-containing oral 3-drug regimens can completely cure BU in ≤2 weeks for daily use or in ≤3 weeks given twice per week (6 doses in total). All high-level TB47-resistant mutants could only be selected using the low-level mutants which were still sensitive to TB47 in mice. This is the first report of double mutations in QcrB in mycobacteria. In summary, TB47-containing regimens have promise to cure BU highly effectively and prevent the emergence of drug resistance. Novel QcrB mutations found here may guide the potential clinical molecular diagnosis of resistance and the discovery of new drugs against the high-level resistant mutants.

Pathogenic mycobacteria pose a sustained threat to global human health. Recently, cytochrome bcc complexes have gained interest as targets for antibiotic drug development. However, there is currently no structural information for the cytochrome bcc complex from these pathogenic mycobacteria. Here, we report the structures of Mycobacterium tuberculosis cytochrome bcc alone (2.68 Å resolution) and in complex with clinical drug candidates Q203 (2.67 Å resolution) and TB47 (2.93 Å resolution) determined by single-particle cryo-electron microscopy. M. tuberculosis cytochrome bcc forms a dimeric assembly with endogenous menaquinone/menaquinol bound at the quinone/quinol-binding pockets. We observe Q203 and TB47 bound at the quinol-binding site and stabilized by hydrogen bonds with the side chains of QcrBThr313 and QcrBGlu314, residues that are conserved across pathogenic mycobacteria. These high-resolution images provide a basis for the design of new mycobacterial cytochrome bcc inhibitors that could be developed into broad-spectrum drugs to treat mycobacterial infections.

Buruli ulcer (BU), caused by Mycobacterium ulcerans , is currently treated with rifampin–streptomycin or rifampin–clarithromycin daily for 8 weeks recommended by World Health Organization (WHO). These options are lengthy with severe side effects. A new anti-tuberculosis drug, TB47, targeting QcrB in cytochrome bc1:aa3 complex is being developed in China. TB47-containing regimens were evaluated in a well-established murine model using an autoluminescent M. ulcerans strain. High-level TB47-resistant spontaneous M. ulcerans mutants were selected and their qcrB genes were sequenced. The in vivo activities of TB47 against both low-level and high-level TB47-resistant mutants were tested in BU murine model. Here, we show that TB47-containing oral 3-drug regimens can completely cure BU in ≤2 weeks for daily use or in ≤3 weeks given twice per week (6 doses in total). All high-level TB47-resistant mutants could only be selected using the low-level mutants which were still sensitive to TB47 in mice. This is the first report of double mutations in QcrB in mycobacteria. In summary, TB47-containing regimens have promise to cure BU highly effectively and prevent the emergence of drug resistance. Novel QcrB mutations found here may guide the potential clinical molecular diagnosis of resistance and the discovery of new drugs against the high-level resistant mutants. Novel regimens containing TB47, a QcrB inhibitor, cure Buruli ulcer in a murine model within 14 days or with only 6 doses. TB47 was active against the low-level TB47-resistant mutants but inactive against the high-level ones in which double mutations in QcrB were first found. [Display omitted] [ABSTRACT FROM AUTHOR]

As an obligate aerobe, Mycobacterium tuberculosis relies on its branched electron transport chain (ETC) for energy production through oxidative phosphorylation. Regimens targeting ETC exhibit promising potential to enhance bactericidal activity against M. tuberculosis and hold the prospect of shortening treatment duration. Our previous research demonstrated that the bacteriostatic drug candidate TB47 (T) inhibited the growth of M. tuberculosis by targeting the cytochrome bc1 complex and exhibited synergistic activity with clofazimine (C). Here, we found synergistic activities between C and sudapyridine (S), a structural analog of bedaquiline (B). S has shown similar anti-tuberculosis efficacy and may share a mechanism of action with B, which inhibits ATP synthesis and the energy metabolism of bacteria. We evaluated the efficacy of SCT in combination with linezolid (L) or pyrazinamide (Z) using a well-established murine model of tuberculosis. Compared to the BPa(pretomanid)L regimen, SCT and SCTL demonstrated similar bactericidal and sterilizing activities. There was no significant difference in activity between SCT and SCTL. In contrast, SCZ and SCTZ showed much higher activities, with none of the 15 mice experiencing relapse after 2 months of treatment with either SCZ or SCTZ. However, T did not contribute to the activity of the SCZ. Our findings emphasize the efficacy and the potential clinical significance of combination therapy with ETC inhibitors. Additionally, cross-resistance exists not only between S and B but also between S/B and C. This is supported by our findings, as spontaneous S-resistant mutants exhibited mutations in Rv0678 , which are associated with cross-resistance to B and C., Competing Interests: The authors declare no conflict of interest.

TB47, a new drug candidate targeting QcrB in the electron transport chain, has shown a unique synergistic activity with clofazimine and forms a highly sterilizing combination. Here, we investigated the sterilizing effects of several all-oral regimens containing TB47 plus clofazimine and linezolid as a block and the roles of fluoroquinolones and pyrazinamide in them. All these regimens cured tuberculosis within 4 to 6 months in a well-established mouse model, and adding pyrazinamide showed a significant difference in bactericidal effects.

Mycobacterium abscessus is intrinsically resistant to most antimicrobial agents. The emerging infections caused by M. abscessus and the lack of effective treatment call for rapid attention. Here, we intended to construct a selectable marker-free autoluminescent M. abscessus strain (designated UAlMab) as a real-time reporter strain to facilitate the discovery of effective drugs and regimens for treating M. abscessus The UAlMab strain was constructed using the dif /Xer recombinase system. In vitro and in vivo activities of several drugs, including clofazimine and TB47, a recently reported cytochrome bc inhibitor, were assessed using UAlMab. Furthermore, the efficacy of multiple drug combinations, including the clofazimine and TB47 combination, were tested against 20 clinical 1 inhibitor, were assessed using UAlMab. Furthermore, the efficacy of multiple drug combinations, including the clofazimine and TB47 combination, were tested against 20 clinical M. abscessus and in live BALB/c mice in a real-time, noninvasive fashion. Importantly, although TB47 showed marginal activity either alone or in combination with clarithromycin, amikacin, or roxithromycin, the drug markedly potentiated the activity of clofazimine, both in vitro and in live BALB/c mice in a real-time, noninvasive fashion. Importantly, although TB47 showed marginal activity either alone or in combination with clarithromycin, amikacin, or roxithromycin, the drug markedly potentiated the activity of clofazimine, both in vitro This study demonstrates that the use of the UAlMab strain can significantly facilitate rapid evaluation of new drugs and regimens. The clofazimine and TB47 combination is effective against in vivo This study demonstrates that the use of the UAlMab strain can significantly facilitate rapid evaluation of new drugs and regimens. The clofazimine and TB47 combination is effective against M. abscessus , and dual/triple electron transport chain (ETC) targeting can be an effective therapeutic approach for treating mycobacterial infections., (Copyright © 2020 American Society for Microbiology.)