RESUMO
The recent development and spread of extensively drug-resistant and totally drug-resistant resistant (TDR) strains of Mycobacterium tuberculosis highlight the need for new antitubercular drugs. Protein synthesis inhibitors have played an important role in the treatment of tuberculosis (TB) starting with the inclusion of streptomycin in the first combination therapies. Although parenteral aminoglycosides are a key component of therapy for multidrug-resistant TB, the oxazolidinone linezolid is the only orally available protein synthesis inhibitor that is effective against TB. Here, we show that small-molecule inhibitors of aminoacyl-tRNA synthetases (AARSs), which are known to be excellent antibacterial protein synthesis targets, are orally bioavailable and effective against M. tuberculosis in TB mouse infection models. We applied the oxaborole tRNA-trapping (OBORT) mechanism, which was first developed to target fungal cytoplasmic leucyl-tRNA synthetase (LeuRS), to M. tuberculosis LeuRS. X-ray crystallography was used to guide the design of LeuRS inhibitors that have good biochemical potency and excellent whole-cell activity against M. tuberculosis Importantly, their good oral bioavailability translates into in vivo efficacy in both the acute and chronic mouse models of TB with potency comparable to that of the frontline drug isoniazid.
Assuntos
Antituberculosos/farmacologia , Leucina-tRNA Ligase/antagonistas & inibidores , Mycobacterium tuberculosis/efeitos dos fármacos , Inibidores da Síntese de Proteínas/farmacologia , Administração Oral , Animais , Antituberculosos/administração & dosagem , Antituberculosos/química , Antituberculosos/farmacocinética , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos/métodos , Feminino , Humanos , Leucina-tRNA Ligase/química , Leucina-tRNA Ligase/genética , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Testes de Sensibilidade Microbiana , Mycobacterium smegmatis/efeitos dos fármacos , Mycobacterium smegmatis/genética , Mycobacterium tuberculosis/genética , Inibidores da Síntese de Proteínas/administração & dosagem , Inibidores da Síntese de Proteínas/química , Inibidores da Síntese de Proteínas/farmacocinética , Relação Estrutura-Atividade , Tuberculose/tratamento farmacológico , Células VeroRESUMO
New drugs and drugs with a novel mechanism of action are desperately needed to shorten the duration of tuberculosis treatment, to prevent the emergence of drug resistance, and to treat multiple-drug-resistant strains of Mycobacterium tuberculosis. Recently, there has been renewed interest in clofazimine (CFZ). In this study, we utilized the C3HeB/FeJ mouse model, possessing highly organized, hypoxic pulmonary granulomas with caseous necrosis, to evaluate CFZ monotherapy in comparison to results with BALB/c mice, which form only multifocal, coalescing cellular aggregates devoid of caseous necrosis. While CFZ treatment was highly effective in BALB/c mice, its activity was attenuated in the lungs of C3HeB/FeJ mice. This lack of efficacy was directly related to the pathological progression of disease in these mice, since administration of CFZ prior to the formation of hypoxic, necrotic granulomas reconstituted bactericidal activity in this mouse strain. These results support the continued use of mouse models of tuberculosis infection which exhibit a granulomatous response in the lungs that more closely resembles the pathology found in human disease.
Assuntos
Antituberculosos/uso terapêutico , Clofazimina/uso terapêutico , Granuloma/tratamento farmacológico , Tuberculose/tratamento farmacológico , Animais , Carga Bacteriana , Modelos Animais de Doenças , Feminino , Granuloma/patologia , Interferon gama/genética , Interferon gama/fisiologia , Pulmão/microbiologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Knockout , Testes de Sensibilidade Microbiana , Mycobacterium tuberculosis/efeitos dos fármacos , Necrose , Tuberculose/complicaçõesRESUMO
New chemotherapeutics active against multidrug-resistant Mycobacterium tuberculosis are urgently needed. We report on the identification of an adamantyl urea compound that shows potent bactericidal activity against M. tuberculosis and a unique mode of action, namely the abolition of the translocation of mycolic acids from the cytoplasm, where they are synthesized to the periplasmic side of the plasma membrane and are in turn transferred onto cell wall arabinogalactan or used in the formation of virulence-associated, outer membrane, trehalose-containing glycolipids. Whole-genome sequencing of spontaneous-resistant mutants of M. tuberculosis selected in vitro followed by genetic validation experiments revealed that our prototype inhibitor targets the inner membrane transporter MmpL3. Conditional gene expression of mmpL3 in mycobacteria and analysis of inhibitor-treated cells validate MmpL3 as essential for mycobacterial growth and support the involvement of this transporter in the translocation of trehalose monomycolate across the plasma membrane.
Assuntos
Adamantano/análogos & derivados , Antibacterianos/química , Antibacterianos/farmacologia , Membrana Celular/metabolismo , Mycobacterium tuberculosis/efeitos dos fármacos , Ácidos Micólicos/metabolismo , Compostos de Fenilureia/farmacologia , Adamantano/química , Adamantano/farmacologia , Antibacterianos/farmacocinética , Proteínas de Bactérias/metabolismo , Transporte Biológico/efeitos dos fármacos , Membrana Celular/efeitos dos fármacos , Fatores Corda , Avaliação Pré-Clínica de Medicamentos/métodos , Farmacorresistência Bacteriana , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Testes de Sensibilidade Microbiana , Mutação , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/metabolismo , Compostos de Fenilureia/química , Bibliotecas de Moléculas Pequenas , Trealose/metabolismoRESUMO
In preclinical testing of antituberculosis drugs, laboratory-adapted strains of Mycobacterium tuberculosis are usually used both for in vitro and in vivo studies. However, it is unknown whether the heterogeneity of M. tuberculosis stocks used by various laboratories can result in different outcomes in tests of antituberculosis drug regimens in animal infection models. In head-to-head studies, we investigated whether bactericidal efficacy results in BALB/c mice infected by inhalation with the laboratory-adapted strains H37Rv and Erdman differ from each other and from those obtained with clinical tuberculosis strains. Treatment of mice consisted of dual and triple drug combinations of isoniazid (H), rifampin (R), and pyrazinamide (Z). The results showed that not all strains gave the same in vivo efficacy results for the drug combinations tested. Moreover, the ranking of HRZ and RZ efficacy results was not the same for the two H37Rv strains evaluated. The magnitude of this strain difference also varied between experiments, emphasizing the risk of drawing firm conclusions for human trials based on single animal studies. The results also confirmed that the antagonism seen within the standard HRZ regimen by some investigators appears to be an M. tuberculosis strain-specific phenomenon. In conclusion, the specific identity of M. tuberculosis strain used was found to be an important variable that can change the apparent outcome of in vivo efficacy studies in mice. We highly recommend confirmation of efficacy results in late preclinical testing against a different M. tuberculosis strain than the one used in the initial mouse efficacy study, thereby increasing confidence to advance potent drug regimens to clinical trials.
Assuntos
Antituberculosos/uso terapêutico , Isoniazida/uso terapêutico , Mycobacterium tuberculosis/classificação , Mycobacterium tuberculosis/efeitos dos fármacos , Pirazinamida/uso terapêutico , Rifampina/uso terapêutico , Tuberculose Pulmonar/tratamento farmacológico , Animais , Antituberculosos/farmacologia , Modelos Animais de Doenças , Quimioterapia Combinada , Feminino , Humanos , Isoniazida/farmacologia , Pulmão/microbiologia , Camundongos , Camundongos Endogâmicos BALB C , Testes de Sensibilidade Microbiana/estatística & dados numéricos , Mycobacterium tuberculosis/genética , Pirazinamida/administração & dosagem , Pirazinamida/farmacologia , Rifampina/administração & dosagem , Rifampina/farmacologia , Resultado do Tratamento , Tuberculose Pulmonar/microbiologiaRESUMO
A novel subclass of quinolones, 2-pyridones, showed potent activity against Mycobacterium tuberculosis, with KRQ-10018 being an early lead. KRQ-10018 showed better activity in vitro against M. tuberculosis versus moxifloxacin. In vivo efficacy of KRQ-10018 at 300 mg/kg of body weight was similar to that of isoniazid at 25 mg/kg, but showed less activity than moxifloxacin at 300 mg/kg.
Assuntos
Antituberculosos/farmacologia , Antituberculosos/uso terapêutico , Mycobacterium tuberculosis/efeitos dos fármacos , Piridonas/farmacologia , Piridonas/uso terapêutico , Tuberculose Pulmonar/tratamento farmacológico , Animais , Antituberculosos/administração & dosagem , Disponibilidade Biológica , Interferon gama/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Testes de Sensibilidade Microbiana , Piridonas/administração & dosagem , Quinolonas/administração & dosagem , Quinolonas/farmacologia , Quinolonas/uso terapêutico , Resultado do Tratamento , Tuberculose Pulmonar/microbiologiaRESUMO
This study extends earlier reports regarding the in vitro and in vivo efficacies of the nitroimidazopyran PA-824 against Mycobacterium tuberculosis. PA-824 was tested in vitro against a broad panel of multidrug-resistant clinical isolates and was found to be highly active against all isolates (MIC<1 microg/ml). The activity of PA-824 against M. tuberculosis was also assessed grown under conditions of oxygen depletion. PA-824 showed significant activity at 2, 10, and 50 microg/ml, similar to that of metronidazole, in a dose-dependent manner. In a short-course mouse infection model, the efficacy of PA-824 at 50, 100, and 300 mg/kg of body weight formulated in methylcellulose or cyclodextrin/lecithin after nine oral treatments was compared with those of isoniazid, rifampin, and moxifloxacin. PA-824 at 100 mg/kg in cyclodextrin/lecithin was as active as moxifloxacin at 100 mg/kg and isoniazid at 25 mg/kg and was slightly more active than rifampin at 20 mg/kg. Long-term treatment with PA-824 at 100 mg/kg in cyclodextrin/lecithin reduced the bacterial load below 500 CFU in the lungs and spleen. No significant differences in activity between PA-824 and the other single drug treatments tested (isoniazid at 25 mg/kg, rifampin at 10 mg/kg, gatifloxacin at 100 mg/kg, and moxifloxacin at 100 mg/kg) could be observed. In summary, its good activity in in vivo models, as well as its activity against multidrug-resistant M. tuberculosis and against M. tuberculosis isolates in a potentially latent state, makes PA-824 an attractive drug candidate for the therapy of tuberculosis. These data indicate that there is significant potential for effective oral delivery of PA-824 for the treatment of tuberculosis.
Assuntos
Antituberculosos , Mycobacterium tuberculosis/efeitos dos fármacos , Nitroimidazóis , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Tuberculose Resistente a Múltiplos Medicamentos/microbiologia , Animais , Antituberculosos/administração & dosagem , Antituberculosos/farmacologia , Antituberculosos/uso terapêutico , Contagem de Colônia Microbiana , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Feminino , Humanos , Pulmão/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Testes de Sensibilidade Microbiana , Mycobacterium tuberculosis/crescimento & desenvolvimento , Mycobacterium tuberculosis/isolamento & purificação , Nitroimidazóis/administração & dosagem , Nitroimidazóis/farmacologia , Nitroimidazóis/uso terapêutico , Organismos Livres de Patógenos Específicos , Baço/microbiologia , Tuberculose Pulmonar/tratamento farmacológico , Tuberculose Pulmonar/microbiologiaRESUMO
We have developed a rapid new in vivo method for screening experimental drugs for their activity against Mycobacterium tuberculosis by using the gamma interferon gene-disrupted (GKO) C57BL/6 mouse. Due to the rapid growth of the infection, statistical differences indicating positive efficacy of active compounds can be seen after only 8 days of treatment. To validate this model, several fluoroquinolones, including ciprofloxacin, levofloxacin, moxifloxacin, and gatifloxacin, were tested in parallel.