Contribution of direct InhA inhibitors to novel drug regimens in a mouse model of tuberculosis.

Isoniazid is an important first-line medicine to treat tuberculosis (TB). Isoniazid resistance increases the risk of poor treatment outcomes and development of multidrug resistance, and is driven primarily by mutations involving katG, encoding the prodrug-activating enzyme, rather than its validated target, InhA. The chemical tractability of InhA has fostered efforts to discover direct inhibitors of InhA (DIIs). In this study, we bridge the gap in understanding the potential contribution of DIIs to novel combination regimens and demonstrate a clear distinction of DIIs, like GSK693 and the newly described GSK138, from isoniazid, based on activity against clinical isolates and contribution to novel drug regimens. The results suggest that DIIs, specifically GSK138 and GSK693, could be promising partners in novel drug regimens, including those used against isoniazid-resistant TB, potentially enhancing their efficacy and/or preventing the selection of resistant mutants and supporting the continued exploration of InhA as a promising target for TB drug development.

Efficacy of Long-Acting Bedaquiline Regimens in a Mouse Model of Tuberculosis Preventive Therapy.

Rationale: Completion of preventive therapy is a major bottleneck in global tuberculosis control. Long-acting injectable drug formulations would shorten therapy administration and may thereby improve completion rates. Recently, a long-acting formulation of bedaquiline demonstrated antituberculosis activity for up to 12 weeks after injection in a validated mouse model of preventive therapy. Objectives: The objectives of this study were to 1) determine the total duration of activity after an injection of long-acting bedaquiline and 2) evaluate the activity of regimens comprised of long-acting bedaquiline plus short (2-4 wk) oral companion courses of bedaquiline, with or without rifapentine, using the validated mouse model of tuberculosis preventive therapy. Methods: After the establishment of a stable Mycobacterium tuberculosis lung infection in bacillus Calmette-Guérin (BCG)-immunized BALB/c mice, treatment was initiated with 1 of 12 randomly assigned regimens. In addition to positive and negative controls, six regimens included one or two injections of long-acting bedaquiline (alone or with oral bedaquiline with or without rifapentine), and four comparator regimens consisted of oral agents only. Lung bacterial burden was measured monthly for up to 28 weeks. Measurements and Main Results: One injection of long-acting bedaquiline at 160 mg/kg exerted antituberculosis activity for 12 weeks. Compared with the positive control (daily isoniazid-rifapentine for 4 wk), six regimens had equivalent bactericidal activity (including two all-oral comparator regimens), and two regimens had superior sterilizing activity: one injection with 2 weeks of oral bedaquiline and high-dose rifapentine; and two injections with 4 weeks of oral bedaquiline. Conclusions: Long-acting injectable bedaquiline has significant potential for shortening tuberculosis preventive therapy.

Novel Regimens of Bedaquiline-Pyrazinamide Combined with Moxifloxacin, Rifabutin, Delamanid and/or OPC-167832 in Murine Tuberculosis Models.

A recent landmark trial showed a 4-month regimen of rifapentine, pyrazinamide, moxifloxacin, and isoniazid (PZMH) to be noninferior to the 6-month standard of care. Here, two murine models of tuberculosis were used to test whether novel regimens replacing rifapentine and isoniazid with bedaquiline and another drug would maintain or increase the sterilizing activity of the regimen. In BALB/c mice, replacing rifapentine in the PZM backbone with bedaquiline (i.e., BZM) significantly reduced both lung CFU counts after 1 month and the proportion of mice relapsing within 3 months after completing 1.5 months of treatment. The addition of rifabutin to BZM (BZMRb) further increased the sterilizing activity. In the C3HeB/FeJ mouse model characterized by caseating lung lesions, treatment with BZMRb resulted in significantly fewer relapses than PZMH after 2 months of treatment. A regimen combining the new DprE1 inhibitor OPC-167832 and delamanid (BZOD) also had superior bactericidal and sterilizing activity compared to PZM in BALB/c mice and was similar in efficacy to PZMH in C3HeB/FeJ mice. Thus, BZM represents a promising backbone for treatment-shortening regimens. Given the prohibitive drug-drug interactions between bedaquiline and rifampin or rifapentine, the BZMRb regimen represents the best opportunity to combine, in one regimen, the treatment-shortening potential of the rifamycin class with that of BZM and deserves high priority for evaluation in clinical trials. Other 4-drug BZM-based regimens and BZOD represent promising opportunities for extending the spectrum of treatment-shortening regimens to rifamycin- and fluoroquinolone-resistant tuberculosis.

Combination of Mycobacterium tuberculosis RS Ratio and CFU Improves the Ability of Murine Efficacy Experiments to Distinguish between Drug Treatments.

Murine tuberculosis drug efficacy studies have historically monitored bacterial burden based on CFU of Mycobacterium tuberculosis in lung homogenate. In an alternative approach, a recently described molecular pharmacodynamic marker called the RS ratio quantifies drug effect on a fundamental cellular process, ongoing rRNA synthesis. Here, we evaluated the ability of different pharmacodynamic markers to distinguish between treatments in three BALB/c mouse experiments at two institutions. We confirmed that different pharmacodynamic markers measure distinct biological responses. We found that a combination of pharmacodynamic markers distinguishes between treatments better than any single marker. The combination of the RS ratio with CFU showed the greatest ability to recapitulate the rank order of regimen treatment-shortening activity, providing proof of concept that simultaneous assessment of pharmacodynamic markers measuring different properties will enhance insight gained from animal models and accelerate development of new combination regimens. These results suggest potential for a new era in which antimicrobial therapies are evaluated not only on culture-based measures of bacterial burden but also on molecular assays that indicate how drugs impact the physiological state of the pathogen.

GSK2556286 Is a Novel Antitubercular Drug Candidate Effective In Vivo with the Potential To Shorten Tuberculosis Treatment.

As a result of a high-throughput compound screening campaign using Mycobacterium tuberculosis-infected macrophages, a new drug candidate for the treatment of tuberculosis has been identified. GSK2556286 inhibits growth within human macrophages (50% inhibitory concentration [IC50] = 0.07 µM), is active against extracellular bacteria in cholesterol-containing culture medium, and exhibits no cross-resistance with known antitubercular drugs. In addition, it has shown efficacy in different mouse models of tuberculosis (TB) and has an adequate safety profile in two preclinical species. These features indicate a compound with a novel mode of action, although still not fully defined, that is effective against both multidrug-resistant (MDR) or extensively drug-resistant (XDR) and drug-sensitive (DS) M. tuberculosis with the potential to shorten the duration of treatment in novel combination drug regimens. (This study has been registered at ClinicalTrials.gov under identifier NCT04472897).

Dual mTORC1/mTORC2 Inhibition as a Host-Directed Therapeutic Target in Pathologically Distinct Mouse Models of Tuberculosis.

Efforts to develop more effective and shorter-course therapies for tuberculosis have included a focus on host-directed therapy (HDT). The goal of HDT is to modulate the host response to infection, thereby improving immune defenses to reduce the duration of antibacterial therapy and/or the amount of lung damage. As a mediator of innate and adaptive immune responses involved in eliminating intracellular pathogens, autophagy is a potential target for HDT in tuberculosis. Because Mycobacterium tuberculosis modulates mammalian target of rapamycin (mTOR) signaling to impede autophagy, pharmacologic mTOR inhibition could provide effective HDT. mTOR exists within two distinct multiprotein complexes, mTOR complex-1 (mTORC1) and mTOR complex-2 (mTORC2). Rapamycin and its analogs only partially inhibit mTORC1. We hypothesized that novel mTOR kinase inhibitors blocking both complexes would have expanded therapeutic potential. We compared the effects of two mTOR inhibitors, rapamycin and the orally available mTOR kinase domain inhibitor CC214-2, which blocks both mTORC1 and mTORC2, as adjunctive therapies against murine TB when added to the first-line regimen (isoniazid, rifampin, pyrazinamide, and ethambutol [RHZE]) or the novel bedaquiline-pretomanid-linezolid (BPaL) regimen. Neither mTOR inhibitor affected lung CFU counts after 4 to 8 weeks of treatment when combined with BPaL or RHZE. However, addition of CC214-2 to BPaL and RHZE was associated with significantly fewer relapses in C3HeB/FeJ mice compared to addition of rapamycin and, in RHZE-treated mice, resulted in fewer relapses than RHZE alone. Therefore, CC214-2 and related mTOR kinase inhibitors may be more effective candidates for HDT than rapamycin analogs and may have the potential to shorten the duration of TB treatment.

Preserved Efficacy and Reduced Toxicity with Intermittent Linezolid Dosing in Combination with Bedaquiline and Pretomanid in a Murine Tuberculosis Model.

The novel regimen of bedaquiline, pretomanid, and linezolid (BPaL) is highly effective against drug-resistant tuberculosis, but linezolid toxicities are frequent. We hypothesized that, for a similar total weekly cumulative dose, thrice-weekly administration of linezolid would preserve efficacy while reducing toxicity compared with daily dosing, in the context of the BPaL regimen. Using C3HeB/FeJ and BALB/c mouse models of tuberculosis disease, thrice-weekly linezolid dosing was compared with daily dosing, with intermittent dosing introduced (i) from treatment initiation or (ii) after an initial period of daily dosing. In all animals, BPa was dosed daily throughout treatment. Blood counts were used to assess hematologic toxicity. After unexpected findings of apparent antagonism, we conducted additional experiments to investigate strain-to-strain differences in the contribution of linezolid to regimen efficacy by comparing each 1- and 2-drug component to the BPaL regimen in BALB/c mice infected with Mycobacterium tuberculosis H37Rv or HN878. Giving linezolid daily for 1 to 2 months achieved the greatest efficacy but, after that, results were similar if the drug was stopped, dosed thrice-weekly, or continued daily. Erythrocyte counts were lower with daily than thrice-weekly dosing. Linezolid had additive effects with BPa against M. tuberculosis H37Rv but antagonistic effects with BPa against M. tuberculosis HN878. However, the overall efficacy of BPaL was high and similar against both strains. Dosing linezolid daily for the first 2 months and then less frequently thereafter may optimize its therapeutic margin. Linezolid's contribution to BPaL regimens may depend on the M. tuberculosis strain.

Mechanistic Modeling of Mycobacterium tuberculosis Infection in Murine Models for Drug and Vaccine Efficacy Studies.

Tuberculosis (TB) drug, regimen, and vaccine development rely heavily on preclinical animal experiments, and quantification of bacterial and immune response dynamics is essential for understanding drug and vaccine efficacy. A mechanism-based model was built to describe Mycobacterium tuberculosis H37Rv infection over time in BALB/c and athymic nude mice, which consisted of bacterial replication, bacterial death, and adaptive immune effects. The adaptive immune effect was best described by a sigmoidal function on both bacterial load and incubation time. Applications to demonstrate the utility of this baseline model showed (i) the important influence of the adaptive immune response on pyrazinamide (PZA) drug efficacy, (ii) a persistent adaptive immune effect in mice relapsing after chemotherapy cessation, and (iii) the protective effect of vaccines after M. tuberculosis challenge. These findings demonstrate the utility of our model for describing M. tuberculosis infection and corresponding adaptive immune dynamics for evaluating the efficacy of TB drugs, regimens, and vaccines.

Telacebec for Ultrashort Treatment of Buruli Ulcer in a Mouse Model.

Telacebec (Q203) is a new antitubercular drug with extremely potent activity against Mycobacterium ulcerans Here, we explored the treatment-shortening potential of Q203 alone or in combination with rifampin (RIF) in a mouse footpad infection model. The first study compared Q203 at 5 and 10 mg/kg doses alone and with rifampin. Q203 alone rendered most mouse footpads culture negative in 2 weeks. Combining Q203 with rifampin resulted in a relapse-free cure 24 weeks after completing 2 weeks of treatment, compared to a 25% relapse rate in mice receiving RIF with clarithromycin, the current standard of care, for 4 weeks. The second study explored the dose-ranging activity of Q203 alone and with RIF, including the extended activity of Q203 after treatment discontinuation. The bactericidal activity of Q203 persisted for ≥ 4 weeks beyond the last dose. All mice receiving just 1 week of Q203 at 2 to 10 mg/kg were culture negative 4 weeks after stopping treatment. Mice receiving 2 weeks of Q203 at 0.5, 2, and 10 mg/kg were culture negative 4 weeks after treatment. RIF did not increase the efficacy of Q203. A pharmacokinetics substudy revealed that Q203 doses of 2 to 10 mg/kg in mice produce plasma concentrations similar to those produced by 100 to 300 mg doses in humans, with no adverse effect of RIF on Q203 concentrations. These results indicate the extraordinary potential of Q203 to reduce the duration of treatment necessary for a cure to ≤ 1 week (or 5 doses of 2 to 10 mg/kg) in our mouse footpad infection model and warrant further evaluation of Q203 in clinical trials.

Treatment-Shortening Effect of a Novel Regimen Combining Clofazimine and High-Dose Rifapentine in Pathologically Distinct Mouse Models of Tuberculosis.

Clofazimine and high-dose rifapentine have each separately been associated with treatment-shortening activity when incorporated into tuberculosis (TB) treatment regimens. We hypothesized that both modifications, i.e., the addition of clofazimine and the replacement of rifampin with high-dose rifapentine, in the first-line regimen for drug-susceptible TB would significantly shorten the duration of treatment necessary for cure. We tested this hypothesis in a well-established BALB/c mouse model of TB chemotherapy and also in a C3HeB/FeJ mouse model in which mice can develop caseous necrotic lesions, an environment where rifapentine and clofazimine may individually be less effective. In both mouse models, replacing rifampin with high-dose rifapentine and adding clofazimine in the first-line regimen resulted in greater bactericidal and sterilizing activity than either modification alone, suggesting that a rifapentine- and clofazimine-containing regimen may have the potential to significantly shorten the treatment duration for drug-susceptible TB. These data provide preclinical evidence supporting the evaluation of regimens combining high-dose rifapentine and clofazimine in clinical trials.

Contribution of Pretomanid to Novel Regimens Containing Bedaquiline with either Linezolid or Moxifloxacin and Pyrazinamide in Murine Models of Tuberculosis.

Novel regimens combining bedaquiline and pretomanid with either linezolid (BPaL regimen) or moxifloxacin and pyrazinamide (BPaMZ regimen) shorten the treatment duration needed to cure tuberculosis (TB) in BALB/c mice compared to that of the first-line regimen and have yielded promising results in initial clinical trials. However, the independent contribution of the investigational new drug pretomanid to the efficacy of BPaMZ has not been examined, and its contribution to BPaL has been examined only over the first 2 months of treatment. In the present study, the addition of pretomanid to BL increased bactericidal activity, prevented emergence of bedaquiline resistance, and shortened the duration needed to prevent relapse with drug-susceptible isolates by at least 2 months in BALB/c mice. Addition of pretomanid to bedaquiline, moxifloxacin, and pyrazinamide (BMZ) resulted in a 1-log10 greater CFU reduction after 1 month of treatment and/or reduced the number of mice relapsing in each of 2 experiments in BALB/c mice and in immunocompromised nude mice. Bedaquiline-resistant isolates were found at relapse in only one BMZ-treated nude mouse. Treatment of infection with a pyrazinamide-resistant mutant in BALB/c mice with BPaMZ prevented selection of bedaquiline-resistant mutants and reduced the proportion of mice relapsing compared to that for BMZ treatment alone. Among severely ill C3HeB/FeJ mice with caseous pneumonia and cavitation, BPaMZ increased median survival (≥60 versus 21 days) and reduced median lung CFU by 2.4 log10 at 1 month compared to the level for BMZ. In conclusion, in 3 different mouse models, pretomanid contributed significantly to the efficacy of the BPaMZ and BPaL regimens, including restricting the selection of bedaquiline-resistant mutants.

Advancing the Therapeutic Potential of Indoleamides for Tuberculosis.

Indole-2-carboxamide derivatives are inhibitors of MmpL3, the cell wall-associated mycolic acid transporter of Mycobacterium tuberculosis In the present study, we characterized indoleamide effects on bacterial cell morphology and reevaluated pharmacokinetics and in vivo efficacy using an optimized oral formulation. Morphologically, indoleamide-treated M. tuberculosis cells demonstrated significantly higher numbers of dimples near the poles or septum, which may serve as the mechanism of cell death for this bactericidal scaffold. Using the optimized formulation, an expanded-spectrum indoleamide, compound 2, showed significantly improved pharmacokinetic (PK) parameters and in vivo efficacy in mouse infection models. In a comparative study, compound 2 showed superior efficacy over compound 3 (NITD-304) in a high-dose aerosol mouse infection model. Since indoleamides are equally active on drug-resistant M. tuberculosis, these findings demonstrate the therapeutic potential of this novel scaffold for the treatment of both drug-susceptible and drug-resistant tuberculosis.

Verapamil Increases the Bioavailability and Efficacy of Bedaquiline but Not Clofazimine in a Murine Model of Tuberculosis.

Drug efflux pumps play important roles in intrinsic and acquired drug resistance. Verapamil, an efflux inhibitor that enhances the activity of bedaquiline, clofazimine, and other drugs against Mycobacterium tuberculosis, has been proposed as a potential adjunctive agent for treatment of tuberculosis (TB). However, the extent to which verapamil enhances in vivo efficacy by inhibiting bacterial efflux pumps versus inhibiting mammalian drug transporters to improve oral bioavailability has not been delineated. We found that verapamil potentiated the in vitro activity of bedaquiline and clofazimine against M. tuberculosis clinical isolates, including those harboring rv0678 mutations. Verapamil increased the efficacy of bedaquiline in a murine TB model by the same extent to which it increased systemic bedaquiline exposure. However, verapamil showed no effect on the oral bioavailability or efficacy of clofazimine in mice. The addition of verapamil increased the sterilizing activity of a regimen composed of bedaquiline, clofazimine, and pyrazinamide. These results confirm that verapamil has adjunctive activity in vivo, but they also demonstrate that the adjunctive effect is likely due to enhanced systemic exposure to companion drugs via effects on mammalian transporters, rather than inhibition of bacterial pumps. Therefore, there may be no advantage to administering verapamil versus increasing the doses of companion drugs.

Impact of Clofazimine Dosing on Treatment Shortening of the First-Line Regimen in a Mouse Model of Tuberculosis.

The antileprosy drug clofazimine was recently repurposed as part of a newly endorsed short-course regimen for multidrug-resistant tuberculosis. It also enables significant treatment shortening when added to the first-line regimen for drug-susceptible tuberculosis in a mouse model. However, clofazimine causes dose- and duration-dependent skin discoloration in patients, and the optimal clofazimine dosing strategy in the context of the first-line regimen is unknown. We utilized a well-established mouse model to systematically address the impacts of duration, dose, and companion drugs on the treatment-shortening activity of clofazimine in the first-line regimen. In all studies, the primary outcome was relapse-free cure (culture-negative lungs) 6 months after stopping treatment, and the secondary outcome was bactericidal activity, i.e., the decline in the lung bacterial burden during treatment. Our findings indicate that clofazimine activity is most potent when coadministered with first-line drugs continuously throughout treatment and that equivalent treatment-shortening results are obtained with half the dose commonly used in mice. However, our studies also suggest that clofazimine at low exposures may have negative impacts on treatment outcomes, an effect that was evident only after the first 3 months of treatment. These data provide a sound evidence base to inform clofazimine dosing strategies to optimize the antituberculosis effect while minimizing skin discoloration. The results also underscore the importance of conducting long-term studies to allow the full evaluation of drugs administered in combination over long durations.

Immunodeficiency and Intermittent Dosing Promote Acquired Rifamycin Monoresistance in Murine Tuberculosis.

More-permissive preclinical models may be useful in evaluating antituberculosis regimens for their propensity to select drug-resistant mutants. To evaluate whether acquired rifamycin monoresistance could be recapitulated in mice and, if so, to evaluate the effects of immunodeficiency, intermittent dosing, and drug exposures, athymic nude and BALB/c mice were infected. Controls received daily rifapentine alone or 2 months of rifampin, isoniazid, pyrazinamide, and ethambutol, followed by 4 months of rifampin/isoniazid, either daily or twice weekly with one of two isoniazid doses. Test groups received the same intensive regimen followed by once-weekly rifapentine or isoniazid/rifapentine with rifapentine doses of 10, 15, or 20 mg/kg of body weight plus one of two isoniazid doses. All combination regimens rendered BALB/c mouse cultures negative but selected mutants resistant to isoniazid (8.5%, 12/140) or rifampin (3.5%, 5/140) in nude mice (P < 0.001). Intermittently dosed intensive-phase therapy selected isoniazid and rifampin resistance in 10% (8/80, P < 0.001) and 20% (16/80, P = 0.009) of nude mice, respectively, compared to 0% treated with a daily regimen. Once-weekly rifapentine-containing continuation-phase regimens selected rifampin-resistant mutants at a rate of 18.0% (18/100, P = 0.035 compared to rifampin/isoniazid regimens). Higher isoniazid doses in the intermittent-treatment control regimen and higher rifapentine doses in once-weekly regimens were associated with less selection of isoniazid resistance. Acquired resistance, including rifamycin monoresistance, was more likely to occur in nude mice despite administration of combination therapy. These results recapitulate clinical outcomes and indicate that nude mice may be useful for evaluating the ability of novel regimens to prevent the selection of resistance.

Bactericidal and Sterilizing Activity of a Novel Regimen with Bedaquiline, Pretomanid, Moxifloxacin, and Pyrazinamide in a Murine Model of Tuberculosis.

New regimens based on 2 or more novel agents are sought to shorten or to simplify treatment of tuberculosis (TB), including drug-resistant forms. Prior studies showed that the novel combinations of bedaquiline (BDQ) plus pretomanid (PMD) plus pyrazinamide (PZA) and PMD plus moxifloxacin (MXF) plus PZA shortened the treatment duration necessary to prevent relapse by 2 to 3 months and 1 to 2 months, respectively, compared with the current first-line regimen, in a murine TB model. These 3-drug combinations are now being studied in clinical trials. Here, the 4-drug combination of BDQ+PMD+MXF+PZA was compared to its 3-drug component regimens and different treatment durations of PZA and MXF were explored, to identify the optimal regimens and treatment times and to estimate the likelihood of success against drug-resistant strains. BDQ+PMD+MXF+PZA rendered all mice relapse-free after 2 months of treatment. PZA administration could be discontinued after the first month of treatment without worsening outcomes, whereas the absence of MXF, PZA, or BDQ administration from the beginning necessitated approximately 0.5, 1, or 2 months, respectively, of additional treatment to attain the same outcome.

In vitro and in vivo activity of biapenem against drug-susceptible and rifampicin-resistant Mycobacterium tuberculosis.

Background: Biapenem, a carbapenem antibiotic, has been shown to have synergistic bactericidal anti-TB activity when combined with rifampicin both in vitro and in the mouse model of TB chemotherapy. We hypothesized that this synergy would result in biapenem/rifampicin activity against rifampicin-resistant Mycobacterium tuberculosis . Objectives: Our objective was to evaluate the synergy of biapenem/rifampicin against both low- and high-level rifampicin-resistant strains of M. tuberculosis , in vitro and in the mouse model. Methods: Biapenem/rifampicin activity was evaluated using three strains of M. tuberculosis : strain 115R (low-level rifampicin resistance); strain 124R (high-level rifampicin resistance); and the drug-susceptible H37Rv parent strain. Biapenem/rifampicin synergy was evaluated in vitro by chequerboard titration. In vivo , we first conducted a dose-ranging experiment with biapenem against H37Rv in the mouse model. We then evaluated biapenem/rifampicin activity in mice infected with each M. tuberculosis strain. Results: In vitro , synergy was observed between biapenem and rifampicin against H37Rv and strain 115R. In vivo , biapenem exhibited clear dose-dependent activity against H37Rv, with all biapenem doses as active or more active than rifampicin alone. Biapenem and rifampicin had synergistic bactericidal activity against H37Rv in the mouse model; no synergy was observed in mice infected with either of the rifampicin-resistant strains. Biapenem alone was active against all three strains. Conclusions: Our preclinical experiments indicate that biapenem has potential for use as an anti-TB drug, including for use against rifampicin-resistant TB. Thus, biapenem has promise for repurposing as a 'new' - and desperately needed - drug for the treatment of drug-resistant TB.

Contribution of Oxazolidinones to the Efficacy of Novel Regimens Containing Bedaquiline and Pretomanid in a Mouse Model of Tuberculosis.

New regimens based on two or more novel agents are sought to shorten or simplify treatment of tuberculosis (TB). Pretomanid (PMD) is a nitroimidazole in phase 3 trials that has significant bactericidal activity alone and in combination with bedaquiline (BDQ) and/or pyrazinamide (PZA). We previously showed that the novel combination of BDQ+PMD plus the oxazolidinone sutezolid (SZD) had sterilizing activity superior to that of the first-line regimen in a murine model of TB. The present experiments compared the activity of different oxazolidinones in combination with BDQ+PMD with or without PZA in the same model. The 3-drug regimen of BDQ+PMD plus linezolid (LZD) had sterilizing activity approaching that of BDQ+PMD+SZD and superior to that of the first-line regimen. The addition of PZA further enhanced activity. Reducing the duration of LZD to 1 month did not significantly affect the activity of the regimen. Halving the LZD dose or replacing LZD with RWJ-416457 modestly reduced activity over the first month but not after 2 months. AZD5847 and tedizolid also increased the bactericidal activity of BDQ+PMD, but they were less effective than the other oxazolidinones. These results provide optimism for safe, short-course oral regimens for drug-resistant TB that may also be superior to the current first-line regimen for drug-susceptible TB.

High Systemic Exposure of Pyrazinoic Acid Has Limited Antituberculosis Activity in Murine and Rabbit Models of Tuberculosis.

Pyrazinamide (PZA) is a prodrug requiring conversion to pyrazinoic acid (POA) by an amidase encoded by pncA for in vitro activity. Mutation of pncA is the most common cause of PZA resistance in clinical isolates. To determine whether the systemic delivery of POA or host-mediated conversion of PZA to POA could circumvent such resistance, we evaluated the efficacy of orally administered and host-derived POA in vivo Dose-ranging plasma and intrapulmonary POA pharmacokinetics and the efficacy of oral POA or PZA treatment against PZA-susceptible tuberculosis were determined in BALB/c and C3HeB/FeJ mice. The activity of host-derived POA was assessed in rabbits infected with a pncA-null mutant and treated with PZA. Median plasma POA values for the area under the concentration-time curve from 0 h to infinity (AUC0-∞) were 139 to 222 µg·h/ml and 178 to 287 µg·h/ml after doses of PZA and POA of 150 mg/kg of body weight, respectively, in mice. Epithelial lining fluid POA concentrations in infected mice were comparable after POA and PZA administration. In chronically infected BALB/c mice, PZA at 150 mg/kg reduced lung CFU counts by >2 log10 after 4 weeks. POA was effective only at 450 mg/kg, which reduced lung CFU counts by ∼0.7 log10 POA had no demonstrable bactericidal activity in C3HeB/FeJ mice, nor did PZA administered to rabbits infected with a PZA-resistant mutant. Oral POA administration and host-mediated conversion of PZA to POA producing plasma POA exposures comparable to PZA administration was significantly less effective than PZA. These results suggest that the intrabacillary delivery of POA and that producing higher POA concentrations at the site of infection will be more effective strategies for maximizing POA efficacy.

Contribution of the nitroimidazoles PA-824 and TBA-354 to the activity of novel regimens in murine models of tuberculosis.

New regimens based on two or more novel agents are sought in order to shorten or simplify the treatment of both drug-susceptible and drug-resistant forms of tuberculosis. PA-824 is a nitroimidazo-oxazine now in phase II trials and has shown significant early bactericidal activity alone and in combination with the newly approved agent bedaquiline or with pyrazinamide with or without moxifloxacin. While the development of PA-824 continues, a potential next-generation derivative, TBA-354, has been discovered to have in vitro potency superior to that of PA-824 and greater metabolic stability than that of the other nitroimidazole derivative in clinical development, delamanid. In the present study, we compared the activities of PA-824 and TBA-354 as monotherapies in murine models of the initial intensive and continuation phases of treatment, as well as in combination with bedaquiline plus pyrazinamide, sutezolid, and/or clofazimine. The monotherapy studies demonstrated that TBA-354 is 5 to 10 times more potent than PA-824, but selected mutants are cross-resistant to PA-824 and delamanid. The combination studies revealed that TBA-354 is 2 to 4 times more potent than PA-824 when combined with bedaquiline, and when administered at a dose equivalent to that of PA-824, TBA-354 demonstrated superior sterilizing efficacy. Perhaps most importantly, the addition of either nitroimidazole significantly improved the sterilizing activities of bedaquiline and sutezolid, with or without pyrazinamide, confirming the value of each agent in this potentially universally active short-course regimen.