RESUMO
Tuberculous meningitis (TBM) has a high mortality, possibly due to suboptimal therapy. Drug exposure data of antituberculosis agents in the central nervous system (CNS) are required to develop more effective regimens. Rifabutin is a rifamycin equivalently potent to rifampin in human pulmonary tuberculosis. Here, we show that human-equivalent doses of rifabutin achieved potentially therapeutic exposure in relevant CNS tissues in a rabbit model of TBM, supporting further evaluation in clinical trials.
Assuntos
Modelos Animais de Doenças , Rifabutina , Tuberculose Meníngea , Animais , Coelhos , Rifabutina/uso terapêutico , Rifabutina/farmacologia , Tuberculose Meníngea/tratamento farmacológico , Sistema Nervoso Central/efeitos dos fármacos , Antituberculosos/uso terapêutico , Antituberculosos/farmacologia , Rifampina/uso terapêutico , Rifampina/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Antibióticos Antituberculose/uso terapêutico , Antibióticos Antituberculose/farmacologiaRESUMO
Triaza-coumarin (TA-C) is a Mycobacterium tuberculosis (Mtb) dihydrofolate reductase (DHFR) inhibitor with an IC50 (half maximal inhibitory concentration) of â¼1 µM against the enzyme. Despite this moderate target inhibition, TA-C shows exquisite antimycobacterial activity (MIC50, concentration inhibiting growth by 50% = 10 to 20 nM). Here, we investigated the mechanism underlying this potency disconnect. To confirm that TA-C targets DHFR and investigate its unusual potency pattern, we focused on resistance mechanisms. In Mtb, resistance to DHFR inhibitors is frequently associated with mutations in thymidylate synthase thyA, which sensitizes Mtb to DHFR inhibition, rather than in DHFR itself. We observed thyA mutations, consistent with TA-C interfering with the folate pathway. A second resistance mechanism involved biosynthesis of the redox coenzyme F420 Thus, we hypothesized that TA-C may be metabolized by Mtb F420-dependent oxidoreductases (FDORs). By chemically blocking the putative site of FDOR-mediated reduction in TA-C, we reproduced the F420-dependent resistance phenotype, suggesting that F420H2-dependent reduction is required for TA-C to exert its potent antibacterial activity. Indeed, chemically synthesized TA-C-Acid, the putative product of TA-C reduction, displayed a 100-fold lower IC50 against DHFR. Screening seven recombinant Mtb FDORs revealed that at least two of these enzymes reduce TA-C. This redundancy in activation explains why no mutations in the activating enzymes were identified in the resistance screen. Analysis of the reaction products confirmed that FDORs reduce TA-C at the predicted site, yielding TA-C-Acid. This work demonstrates that intrabacterial metabolism converts TA-C, a moderately active "prodrug," into a 100-fold-more-potent DHFR inhibitor, thus explaining the disconnect between enzymatic and whole-cell activity.
Assuntos
Antagonistas do Ácido Fólico/farmacologia , Complexos Multienzimáticos/metabolismo , Mycobacterium tuberculosis/enzimologia , Oxirredutases/metabolismo , Tetra-Hidrofolato Desidrogenase/metabolismo , Cumarínicos/química , Cumarínicos/farmacologia , Farmacorresistência Bacteriana/efeitos dos fármacos , Ácido Fólico/metabolismo , Antagonistas do Ácido Fólico/química , Genes Bacterianos , Mutação com Perda de Função/genética , Simulação de Acoplamento Molecular , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/genética , Oxirredução , Tetra-Hidrofolato Desidrogenase/genéticaRESUMO
The combination of the ß-lactam tebipenem and the ß-lactamase inhibitor avibactam shows potent bactericidal activity against Mycobacterium abscessus in vitro. Here, we report that the combination of the respective oral prodrugs tebipenem-pivoxil and avibactam ARX-1796 showed efficacy in a mouse model of M. abscessus lung infection. The results suggest that tebipenem-avibactam presents an attractive oral drug candidate pair for the treatment of M. abscessus pulmonary disease and could inform the design of clinical trials.
Assuntos
Infecções por Mycobacterium não Tuberculosas , Mycobacterium abscessus , Animais , Camundongos , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Infecções por Mycobacterium não Tuberculosas/tratamento farmacológico , Infecções por Mycobacterium não Tuberculosas/microbiologia , Modelos Animais de Doenças , Pulmão , Testes de Sensibilidade MicrobianaRESUMO
Tuberculosis meningitis (TBM) is essentially treated with the first-line regimen used against pulmonary tuberculosis, with a prolonged continuation phase. However, clinical outcomes are poor in comparison, for reasons that are only partially understood, highlighting the need for improved preclinical tools to measure drug distribution and activity at the site of disease. A predictive animal model of TBM would also be of great value to prioritize promising drug regimens to be tested in clinical trials, given the healthy state of the development pipeline for the first time in decades. Here, we report the optimization of a rabbit model of TBM disease induced via inoculation of Mycobacterium tuberculosis into the cisterna magna, recapitulating features typical of clinical TBM: neurological deterioration within months post-infection, acid-fast bacilli in necrotic lesions in the brain and spinal cord, and elevated lactate levels in cerebrospinal fluid (CSF). None of the infected rabbits recovered or controlled the disease. We used young adult rabbits, the size of which allows for spatial drug quantitation in critical compartments of the central nervous system that cannot be collected in clinical studies. To illustrate the translational value of the model, we report the penetration of linezolid from plasma into the CSF, meninges, anatomically distinct brain areas, cervical spine, and lumbar spine. Across animals, we measured the bacterial burden concomitant with neurological deterioration, offering a useful readout for drug efficacy studies. The model thus forms the basis for building a preclinical platform to identify improved regimens and inform clinical trial design.
Assuntos
Mycobacterium tuberculosis , Tuberculose Meníngea , Animais , Coelhos , Antituberculosos/farmacologia , Sistema Nervoso Central , Tuberculose Meníngea/tratamento farmacológicoRESUMO
Global infections by non-tuberculous mycobacteria (NTM) are steadily rising. New drugs are needed to treat NTM infections, but the NTM drug pipeline remains poorly populated and focused on repurposing or reformulating approved antibiotics. We sought to accelerate de novo NTM drug discovery by testing advanced compounds with established activity against Mycobacterium tuberculosis 3-aminomethyl 4-halogen benzoxaboroles, a novel class of leucyl-tRNA synthetase inhibitors, were recently discovered as active against M. tuberculosis Here, we report that the benzoxaborole EC/11770 is not only a potent anti-tubercular agent but is active against the M. abscessus and M. avium complexes. Focusing on M. abscessus, which causes the most difficult-to-cure NTM disease, we show that EC/11770 retained potency against drug-tolerant biofilms in vitro and was effective in a mouse lung infection model. Resistant mutant selection experiments showed a low frequency of resistance and confirmed leucyl-tRNA synthetase as the target. This work establishes the benzoxaborole EC/11770 as a novel preclinical candidate for the treatment of NTM lung disease and tuberculosis and validates leucyl-tRNA synthetase as an attractive target for the development of broad-spectrum anti-mycobacterials.
RESUMO
The antituberculosis candidate OPC-167832, an inhibitor of DprE1, was active against Mycobacterium abscessus. Resistance mapped to M. abscessus dprE1, suggesting target retention. OPC-167832 was bactericidal and did not antagonize activity of clinical anti-M. abscessus antibiotics. Due to its moderate potency compared to that against Mycobacterium tuberculosis, the compound lacked efficacy in a mouse model and is thus not a repurposing candidate. These results identify OPC-167832-DprE1 as a lead-target couple for a M. abscessus-specific optimization program.
Assuntos
Infecções por Mycobacterium não Tuberculosas , Mycobacterium abscessus , Mycobacterium tuberculosis , Animais , Camundongos , Infecções por Mycobacterium não Tuberculosas/microbiologia , Antibacterianos/farmacologia , Testes de Sensibilidade MicrobianaRESUMO
Cyclohexyl-griselimycin is a preclinical candidate for use against tuberculosis (TB). Here, we show that this oral cyclodepsipeptide is also active against the intrinsically drug-resistant nontuberculous mycobacterium Mycobacterium abscessus in vitro and in a mouse model of infection. This adds a novel advanced lead compound to the M. abscessus drug pipeline and supports a strategy of screening chemical matter generated in TB drug discovery efforts to fast-track the discovery of novel antibiotics against M. abscessus.
Assuntos
Infecções por Mycobacterium não Tuberculosas , Mycobacterium abscessus , Animais , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Camundongos , Testes de Sensibilidade Microbiana , Infecções por Mycobacterium não Tuberculosas/tratamento farmacológico , Infecções por Mycobacterium não Tuberculosas/microbiologia , Micobactérias não Tuberculosas , Peptídeos CíclicosRESUMO
Tricyclic pyrrolopyrimidines (TPPs) are a new class of antibacterials inhibiting the ATPase of DNA gyrase. TPP8, a representative of this class, is active against Mycobacterium abscessus in vitro. Spontaneous TPP8 resistance mutations mapped to the ATPase domain of M. abscessus DNA gyrase, and the compound inhibited DNA supercoiling activity of recombinant M. abscessus enzyme. Further profiling of TPP8 in macrophage and mouse infection studies demonstrated proof-of-concept activity against M. abscessus ex vivo and in vivo.
Assuntos
Infecções por Mycobacterium não Tuberculosas , Mycobacterium abscessus , Adenosina Trifosfatases , Animais , Antibacterianos/farmacologia , DNA Girase/genética , Camundongos , Testes de Sensibilidade Microbiana , Infecções por Mycobacterium não Tuberculosas/tratamento farmacológico , Infecções por Mycobacterium não Tuberculosas/microbiologia , Micobactérias não Tuberculosas , Pirimidinas , PirróisRESUMO
Nontuberculous mycobacterial pulmonary disease (NTM-PD) is a potentially fatal infectious disease requiring long treatment duration with multiple antibiotics and against which there is no reliable cure. Among the factors that have hampered the development of adequate drug regimens is the lack of an animal model that reproduces the NTM lung pathology required for studying antibiotic penetration and efficacy. Given the documented similarities between tuberculosis and NTM immunopathology in patients, we first determined that the rabbit model of active tuberculosis reproduces key features of human NTM-PD and provides an acceptable surrogate model to study lesion penetration. We focused on clarithromycin, a macrolide and pillar of NTM-PD treatment, and explored the underlying causes of the disconnect between its favorable potency and pharmacokinetics and inconsistent clinical outcome. To quantify pharmacokinetic-pharmacodynamic target attainment at the site of disease, we developed a translational model describing clarithromycin distribution from plasma to lung lesions, including the spatial quantitation of clarithromycin and azithromycin in mycobacterial lesions of two patients on long-term macrolide therapy. Through clinical simulations, we visualized the coverage of clarithromycin in plasma and four disease compartments, revealing heterogeneous bacteriostatic and bactericidal target attainment depending on the compartment and the corresponding potency against nontuberculous mycobacteria in clinically relevant assays. Overall, clarithromycin's favorable tissue penetration and lack of bactericidal activity indicated that its clinical activity is limited by pharmacodynamic, rather than pharmacokinetic, factors. Our results pave the way toward the simulation of lesion pharmacokinetic-pharmacodynamic coverage by multidrug combinations to enable the prioritization of promising regimens for clinical trials.
Assuntos
Pneumopatias , Infecções por Mycobacterium não Tuberculosas , Animais , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Humanos , Pneumopatias/tratamento farmacológico , Pneumopatias/microbiologia , Macrolídeos/farmacologia , Macrolídeos/uso terapêutico , Infecções por Mycobacterium não Tuberculosas/tratamento farmacológico , Infecções por Mycobacterium não Tuberculosas/microbiologia , Micobactérias não Tuberculosas , CoelhosRESUMO
Bones are the site of multiple diseases requiring chemotherapy, including cancer, arthritis, osteoporosis and infections. Yet limited methodologies are available to investigate the spatial distribution and quantitation of small molecule drugs in bone compartments, due to the difficulty of sectioning undecalcified bones and the interference of decalcification methods with spatially resolved drug quantitation. To measure drug concentrations in distinct anatomical bone regions, we have developed a workflow that enables spatial quantitation of thin undecalcified bone sections by laser-capture microdissection coupled to HPLC/tandem mass spectrometry, and spatial mapping on adjacent sections by mass spectrometry imaging. The adhesive film and staining methods were optimized to facilitate histology staining on the same sections used for mass spectrometry image acquisition, revealing drug accumulation in the underlying bone tissue architecture, for the first time. Absolute spatial concentrations of rifampicin, bedaquiline, doxycycline, vancomycin and several of their active metabolites are shown for both small rodent bones and larger rabbit bones that more closely resemble human bone density. Overlaid MALDI mass spectrometry images of drugs and histology staining enabled the generation of semi-quantitative data from regions of interest within anatomical bone compartments. These data correlated with absolute drug concentrations determined by HPLC-MS/MS in laser-capture microdissection samples. Collectively, these techniques enable semi- and fully quantitative drug distribution investigations within bone tissue compartments for the first time. Our workflow can be translated to image and quantify not only drugs but also biomarkers of disease to investigate drug penetration as well as mechanisms underlying bone disorders.
Assuntos
Antibacterianos , Espectrometria de Massas em Tandem , Animais , Osso e Ossos , Cromatografia Líquida de Alta Pressão/métodos , Humanos , Microdissecção e Captura a Laser/métodos , Lasers , Coelhos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Espectrometria de Massas em Tandem/métodosRESUMO
Rifamycin antibiotics are a valuable class of antimicrobials for treating infections by mycobacteria and other persistent bacteria owing to their potent bactericidal activity against replicating and non-replicating pathogens. However, the clinical utility of rifamycins against Mycobacterium abscessus is seriously compromised by a novel resistance mechanism, namely, rifamycin inactivation by ADP-ribosylation. Using a structure-based approach, we rationally redesign rifamycins through strategic modification of the ansa-chain to block ADP-ribosylation while preserving on-target activity. Validated by a combination of biochemical, structural, and microbiological studies, the most potent analogs overcome ADP-ribosylation, restored their intrinsic low nanomolar activity and demonstrated significant in vivo antibacterial efficacy. Further optimization by tuning drug disposition properties afforded a preclinical candidate with remarkable potency and an outstanding pharmacokinetic profile.
Assuntos
Mycobacterium , Rifamicinas , Antibacterianos/farmacologia , Antibacterianos/química , Testes de Sensibilidade Microbiana , Rifamicinas/farmacologia , Rifamicinas/química , ADP-RibosilaçãoRESUMO
Benzoxaboroles are a new class of leucyl-tRNA synthetase inhibitors. We recently reported that the antitubercular 4-halogenated benzoxaboroles are active against Mycobacterium abscessus. Here, we find that the nonhalogenated benzoxaborole epetraborole, a clinical candidate developed for Gram-negative infections, is also active against M. abscessus in vitro and in a mouse model of infection. This expands the repertoire of advanced lead compounds for the discovery of a benzoxaborole-based candidate to treat M. abscessus lung disease.
Assuntos
Pneumopatias , Infecções por Mycobacterium não Tuberculosas , Mycobacterium abscessus , Animais , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Antituberculosos , Pneumopatias/tratamento farmacológico , Camundongos , Testes de Sensibilidade Microbiana , Infecções por Mycobacterium não Tuberculosas/tratamento farmacológico , Micobactérias não TuberculosasRESUMO
New, more-effective drugs for the treatment of lung disease caused by nontuberculous mycobacteria (NTM) are needed. Among NTM opportunistic pathogens, Mycobacterium abscessus is the most difficult to cure and intrinsically multidrug resistant. In a whole-cell screen of a compound collection active against Mycobacterium tuberculosis, we previously identified the piperidine-4-carboxamide (P4C) MMV688844 (844) as a hit against M. abscessus. Here, we identified a more potent analog of 844 and showed that both the parent and improved analog retain activity against strains representing all three subspecies of the M. abscessus complex. Furthermore, P4Cs showed bactericidal and antibiofilm activity. Spontaneous resistance against the P4Cs emerged at a frequency of 10-8/CFU and mapped to gyrA and gyrB encoding the subunits of DNA gyrase. Biochemical studies with recombinant M. abscessus DNA gyrase showed that P4Cs inhibit the wild-type enzyme but not the P4C-resistant mutant. P4C-resistant strains showed limited cross-resistance to the fluoroquinolone moxifloxacin, which is in clinical use for the treatment of macrolide-resistant M. abscessus disease, and no cross-resistance to the benzimidazole SPR719, a novel DNA gyrase inhibitor in clinical development for the treatment of mycobacterial diseases. Analyses of P4Cs in recA promoter-based DNA damage reporter strains showed induction of recA promoter activity in the wild type but not in the P4C-resistant mutant background. This indicates that P4Cs, similar to fluoroquinolones, cause DNA gyrase-mediated DNA damage. Together, our results show that P4Cs present a novel class of mycobacterial DNA gyrase inhibitors with attractive antimicrobial activities against the M. abscessus complex.
Assuntos
Infecções por Mycobacterium não Tuberculosas , Mycobacterium abscessus , Antibacterianos/farmacologia , DNA Girase/genética , Humanos , Testes de Sensibilidade Microbiana , Infecções por Mycobacterium não Tuberculosas/tratamento farmacológico , Mycobacterium abscessus/genética , Micobactérias não Tuberculosas , Piperidinas/farmacologiaRESUMO
Fluoroquinolones-the only clinically used DNA gyrase inhibitors-are effective against tuberculosis (TB) but are in limited clinical use for nontuberculous mycobacteria (NTM) lung infections due to intrinsic drug resistance. We sought to test alternative DNA gyrase inhibitors for anti-NTM activity. Mycobacterium tuberculosis gyrase inhibitors (MGIs), a subclass of novel bacterial topoisomerase inhibitors (NBTIs), were recently shown to be active against the tubercle bacillus. Here, we show that the MGI EC/11716 not only has potent anti-tubercular activity but is active against M. abscessus and M. avium in vitro. Focusing on M. abscessus, which causes the most difficult to cure NTM disease, we show that EC/11716 is bactericidal, active against drug-tolerant biofilms, and efficacious in a murine model of M. abscessus lung infection. Based on resistant mutant selection experiments, we report a low frequency of resistance to EC/11716 and confirm DNA gyrase as its target. Our findings demonstrate the potential of NBTIs as anti-M. abscessus and possibly broad-spectrum anti-mycobacterial agents.
Assuntos
Infecções por Mycobacterium não Tuberculosas , Mycobacterium abscessus , Mycobacterium tuberculosis , Animais , Camundongos , Testes de Sensibilidade Microbiana , Infecções por Mycobacterium não Tuberculosas/tratamento farmacológico , Micobactérias não Tuberculosas , Tioinosina/análogos & derivados , Inibidores da Topoisomerase II/farmacologiaRESUMO
SQ109 is a novel well-tolerated drug candidate in clinical development for the treatment of drug-resistant tuberculosis (TB). It is the only inhibitor of the MmpL3 mycolic acid transporter in clinical development. No SQ109-resistant mutant has been directly isolated thus far in vitro, in mice, or in patients, which is tentatively attributed to its multiple targets. It is considered a potential replacement for poorly tolerated components of multidrug-resistant TB regimens. To prioritize SQ109-containing combinations with the best potential for cure and treatment shortening, one must understand its contribution against different bacterial populations in pulmonary lesions. Here, we have characterized the pharmacokinetics of SQ109 in the rabbit model of active TB and its penetration at the sites of disease-lung tissue, cellular and necrotic lesions, and caseum. A two-compartment model with first-order absorption and elimination described the plasma pharmacokinetics. At the human-equivalent dose, parameter estimates fell within the ranges published for preclinical species. Tissue concentrations were modeled using an "effect" compartment, showing high accumulation in lung and cellular lesion areas with penetration coefficients in excess of 1,000 and lower passive diffusion in caseum after 7 daily doses. These results, together with the hydrophobic nature and high nonspecific caseum binding of SQ109, suggest that multiweek dosing would be required to reach steady state in caseum and poorly vascularized compartments, similar to bedaquiline. Linking lesion pharmacokinetics to SQ109 potency in assays against replicating, nonreplicating, and intracellular M. tuberculosis showed SQ109 concentrations markedly above pharmacokinetic-pharmacodynamic targets in lung and cellular lesions throughout the dosing interval.
Assuntos
Mycobacterium tuberculosis , Preparações Farmacêuticas , Tuberculose Resistente a Múltiplos Medicamentos , Tuberculose , Animais , Antituberculosos/uso terapêutico , Humanos , Camundongos , Coelhos , Tuberculose/tratamento farmacológico , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológicoRESUMO
Amikacin and kanamycin are second-line injectables used in the treatment of multidrug-resistant tuberculosis (MDR-TB) based on the clinical utility of streptomycin, another aminoglycoside and first-line anti-TB drug. While streptomycin was tested as a single agent in the first controlled TB clinical trial, introduction of amikacin and kanamycin into MDR-TB regimens was not preceded by randomized controlled trials. A recent large retrospective meta-analysis revealed that compared with regimens without any injectable drug, amikacin provided modest benefits, and kanamycin was associated with worse outcomes. Although their long-term use can cause irreversible ototoxicity, they remain part of MDR-TB regimens because they have a role in preventing emergence of resistance to other drugs. To quantify the contribution of amikacin and kanamycin to second-line regimens, we applied two-dimensional matrix-assisted laser desorption ionization (MALDI) mass spectrometry imaging in large lung lesions, quantified drug exposure in lung and in lesions of rabbits with active TB, and measured the concentrations required to kill or inhibit growth of the resident bacterial populations. Using these metrics, we applied site-of-action pharmacokinetic and pharmacodynamic (PK-PD) concepts and simulated drug coverage in patients' lung lesions. The results provide a pharmacological explanation for the limited clinical utility of both agents and reveal better PK-PD lesion coverage for amikacin than kanamycin, consistent with retrospective data of contribution to treatment success. Together with recent mechanistic studies dissecting antibacterial activity from aminoglycoside ototoxicity, the limited but rapid penetration of streptomycin, amikacin, and kanamycin to the sites of TB disease supports the development of analogs with improved efficacy and tolerability.
Assuntos
Mycobacterium tuberculosis , Tuberculose Resistente a Múltiplos Medicamentos , Tuberculose Pulmonar , Animais , Antituberculosos/uso terapêutico , Humanos , Canamicina , Coelhos , Ensaios Clínicos Controlados Aleatórios como Assunto , Estudos Retrospectivos , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Tuberculose Pulmonar/tratamento farmacológicoRESUMO
There is no reliable cure for Mycobacterium abscessus lung disease. Rifampin is not used clinically due to poor in vitro potency. In contrast, we have shown that rifabutin, another approved rifamycin used to treat tuberculosis, is potent in vitro against M. abscessus Here, we report that rifabutin is as active as clarithromycin against M. abscessus K21 in NOD.CB17-Prkdcscid/NCrCrl mice. This suggests that rifabutin should be considered a repurposing candidate for patients with M. abscessus disease.
Assuntos
Antibacterianos/farmacologia , Pneumopatias/tratamento farmacológico , Infecções por Mycobacterium não Tuberculosas/tratamento farmacológico , Mycobacterium abscessus/efeitos dos fármacos , Rifabutina/farmacologia , Animais , Antibacterianos/química , Claritromicina/farmacologia , Modelos Animais de Doenças , Reposicionamento de Medicamentos , Feminino , Humanos , Pneumopatias/microbiologia , Camundongos , Camundongos Endogâmicos NOD , Infecções por Mycobacterium não Tuberculosas/microbiologia , Rifabutina/químicaRESUMO
Lung disease caused by Mycobacterium abscessus is very difficult to cure, and treatment failure rates are high. The antituberculosis drug bedaquiline (BDQ) is used as salvage therapy against this dreadful disease. However, BDQ is highly lipophilic, displays a long terminal half-life, and presents a cardiotoxicity liability associated with QT interval prolongation. Recent medicinal chemistry campaigns resulted in the discovery of 3,5-dialkoxypyridine analogues of BDQ which are less lipophilic, have higher clearance, and display lower cardiotoxic potential. TBAJ-876, a clinical development candidate of this series, shows attractive in vitro antitubercular activity and efficacy in a murine tuberculosis model. Here, we asked whether TBAJ-876 is active against M. abscessus TBAJ-876 displayed submicromolar in vitro activity against reference strains representing the three subspecies of M. abscessus and against a collection of clinical isolates. Drug-drug potency interaction studies with commonly used anti-M. abscessus antibiotics showed no antagonistic effects, suggesting that TBAJ-876 could be coadministered with currently used drugs. Efficacy studies, employing a mouse model of M. abscessus infection, demonstrated potent activity in vivo In summary, we demonstrate that TBAJ-876 shows attractive in vitro and in vivo activities against M. abscessus, similar to its BDQ parent. This suggests that next-generation BDQ, with improved tolerability and pharmacological profiles, may be useful for the treatment of M. abscessus lung disease in addition to the treatment of tuberculosis.
Assuntos
Antibacterianos/farmacologia , Diarilquinolinas/farmacologia , Mycobacterium abscessus/efeitos dos fármacos , Animais , Modelos Animais de Doenças , Feminino , Humanos , Camundongos SCID , Testes de Sensibilidade Microbiana , Infecções por Mycobacterium não Tuberculosas/microbiologia , Mycobacterium abscessus/isolamento & purificaçãoRESUMO
Doxycycline, an FDA-approved tetracycline, is used in tuberculosis in vivo models for the temporal control of mycobacterial gene expression. In these models, animals are infected with recombinant Mycobacterium tuberculosis carrying genes of interest under transcriptional control of the doxycycline-responsive TetR-tetO unit. To minimize fluctuations of plasma levels, doxycycline is usually administered in the diet. However, tissue penetration studies to identify the minimum doxycycline content in food achieving complete repression of TetR-controlled genes in tuberculosis (TB)-infected organs and lesions have not been conducted. Here, we first determined the tetracycline concentrations required to achieve silencing of M. tuberculosis target genes in vitro Next, we measured doxycycline concentrations in plasma, major organs, and lung lesions in TB-infected mice and rabbits and compared these values to silencing concentrations measured in vitro We found that 2,000 ppm doxycycline supplemented in mouse and rabbit feed is sufficient to reach target concentrations in TB lesions. In rabbit chow, the calcium content had to be reduced 5-fold to minimize chelation of doxycycline and deliver adequate oral bioavailability. Clearance kinetics from major organs and lung lesions revealed that doxycycline levels fall below concentrations that repress tet promoters within 7 to 14 days after doxycycline is removed from the diet. In summary, we have shown that 2,000 ppm doxycycline supplemented in standard mouse diet and in low-calcium rabbit diet delivers concentrations adequate to achieve full repression of tet promoters in infected tissues of mice and rabbits.
Assuntos
Antibacterianos/farmacocinética , Doxiciclina/farmacocinética , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/genética , Tuberculose/metabolismo , Ração Animal , Animais , Antibacterianos/administração & dosagem , Antibacterianos/uso terapêutico , Disponibilidade Biológica , Cálcio/farmacologia , Modelos Animais de Doenças , Doxiciclina/administração & dosagem , Doxiciclina/uso terapêutico , Feminino , Inativação Gênica , Pulmão/metabolismo , Camundongos , Coelhos , Resistência a Tetraciclina , Distribuição Tecidual/genética , TransgenesRESUMO
Bacteria use various endogenous antioxidants for protection against oxidative stress associated with environmental survival or host infection. Although glutathione (GSH) is the most abundant and widely used antioxidant in Proteobacteria, ergothioneine (EGT) is another microbial antioxidant, mainly produced by fungi and Actinobacteria. The Burkholderia genus is found in diverse environmental niches. We observed that gene homologs required for the synthesis of EGT are widely distributed throughout the genus. By generating gene-deletion mutants and monitoring production with isotope-labeled substrates, we show that pathogenic Burkholderia pseudomallei and environmental B. thailandensis are able to synthesize EGT de novo. Unlike most other bacterial EGT synthesis pathways described, Burkholderia spp. use cysteine rather than γ-glutamyl cysteine as the thiol donor. Analysis of recombinant EgtB indicated that it is a proficient sulfoxide synthase, despite divergence in the active site architecture from that of mycobacteria. The absence of GSH, but not EGT, increased bacterial susceptibility to oxidative stresses in vitro. However, deletion of EGT synthesis conferred a reduced fitness to B. pseudomallei, with a delay in organ colonization and time to death during mouse infection. Therefore, despite the lack of an apparent antioxidant role in vitro, EGT is important for optimal bacterial pathogenesis in the mammalian host.-Gamage, A. M., Liao, C., Cheah, I. K., Chen, Y., Lim, D. R. X., Ku, J. W. K., Chee, R. S. L., Gengenbacher, M., Seebeck, F. P., Halliwell, B., Gan, Y.-H. The proteobacterial species Burkholderia pseudomallei produces ergothioneine, which enhances virulence in mammalian infection.