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Preclinical treatment outcome evaluation of tuberculosis (TB) occurs primarily in mice. Current designs compare relapse rates of different regimens at selected time points, but lack information about the correlation between treatment length and treatment outcome, which is required to efficiently estimate a regimens' treatment-shortening potential. Therefore we developed a new approach. BALB/c mice were infected with a Mycobacterium tuberculosis Beijing genotype strain and were treated with rifapentine-pyrazinamide-isoniazid-ethambutol (R p ZHE), rifampicin-pyrazinamide-moxifloxacin-ethambutol (RZME) or rifampicin-pyrazinamide-moxifloxacin-isoniazid (RZMH). Treatment outcome was assessed in n = 3 mice after 9 different treatment lengths between 2-6 months. Next, we created a mathematical model that best fitted the observational data and used this for inter-regimen comparison. The observed data were best described by a sigmoidal E max model in favor over linear or conventional E max models. Estimating regimen-specific parameters showed significantly higher curative potentials for RZME and R p ZHE compared to RZMH. In conclusion, we provide a new design for treatment outcome evaluation in a mouse TB model, which (i) provides accurate tools for assessment of the relationship between treatment length and predicted cure, (ii) allows for efficient comparison between regimens and (iii) adheres to the reduction and refinement principles of laboratory animal use.

Insight into drug transport mechanisms is highly relevant to the efficacious treatment of tuberculosis (TB). Major problems in TB treatment are related to the transport of antituberculosis (anti-TB) drugs across human and mycobacterial membranes, affecting the concentrations of these drugs systemically and locally. Firstly, transporters located in the intestines, liver, and kidneys all determine the pharmacokinetics and pharmacodynamics of anti-TB drugs, with a high risk of drug-drug interactions in the setting of concurrent use of antimycobacterial, antiretroviral, and antidiabetic agents. Secondly, human efflux transporters limit the penetration of anti-TB drugs into the brain and cerebrospinal fluid, which is especially important in the treatment of TB meningitis. Finally, efflux transporters located in the macrophage and Mycobacterium tuberculosis cell membranes play a pivotal role in the emergence of phenotypic tolerance and drug resistance, respectively. We review the role of efflux transporters in TB drug disposition and evaluate the promise of efflux pump inhibition from a novel holistic perspective.

The aim of this study was to investigate pharmacodynamic (PD) interactions in mice infected with Mycobacterium tuberculosis using population pharmacokinetics (PKs), the Multistate Tuberculosis Pharmacometric (MTP) model, and the General Pharmacodynamic Interaction (GPDI) model. Rifampicin, isoniazid, ethambutol, or pyrazinamide were administered in monotherapy for 4 weeks. Rifampicin and isoniazid showed effects in monotherapy, whereas the animals became moribund after 7 days with ethambutol or pyrazinamide alone. No PD interactions were observed against fast-multiplying bacteria. Interactions between rifampicin and isoniazid on killing slow and non-multiplying bacteria were identified, which led to an increase of 0.86 log 10 colony-forming unit (CFU)/lungs at 28 days after treatment compared to expected additivity (i.e., antagonism). An interaction between rifampicin and ethambutol on killing non-multiplying bacteria was quantified, which led to a decrease of 2.84 log 10 CFU/lungs at 28 days after treatment (i.e., synergism). These results show the value of pharmacometrics to quantitatively assess PD interactions in preclinical tuberculosis drug development., (© 2017 The Authors CPT: Pharmacometrics & Systems Pharmacology published by Wiley Periodicals, Inc. on behalf of American Society for Clinical Pharmacology and Therapeutics.)

Immune-modulating drugs that target myeloid-derived suppressor cells or stimulate natural killer T cells have been shown to reduce mycobacterial loads in tuberculosis (TB). We aimed to determine if a combination of these drugs as adjunct immunotherapy to conventional antibiotic treatment could also increase therapeutic efficacy against TB. In our model of pulmonary TB in mice, we applied treatment with isoniazid, rifampicin, and pyrazinamide for 13 weeks alone or combined with immunotherapy consisting of all-trans retinoic acid, 1,25(OH) 2 -vitamin D3, and α-galactosylceramide. Outcome parameters were mycobacterial load during treatment (therapeutic activity) and 13 weeks after termination of treatment (therapeutic efficacy). Moreover, cellular changes were analyzed using flow cytometry and cytokine expression was assessed at the mRNA and protein levels. Addition of immunotherapy was associated with lower mycobacterial loads after 5 weeks of treatment and significantly reduced relapse of disease after a shortened 13-week treatment course compared with antibiotic treatment alone. This was accompanied by reduced accumulation of immature myeloid cells in the lungs at the end of treatment and increased TNF-α protein levels throughout the treatment period. We demonstrate, in a mouse model of pulmonary TB, that immunotherapy consisting of three clinically approved drugs can improve the therapeutic efficacy of standard antibiotic treatment.

Due to the emergence of multidrug-resistant and extensively drug-resistant tuberculosis (TB), there is an urgent need for new TB drugs or for compounds that improve the efficacy of currently used drugs. In this study, time-kill kinetics of SILA-421 as a single drug and in combination with isoniazid (INH), rifampicin (RIF), moxifloxacin (MXF) or amikacin (AMK) against Mycobacterium tuberculosis were assessed. Therapeutic efficacy in vivo in a mouse TB model was also studied. Further in vitro analysis was performed with a RIF-susceptible and RIF-resistant strains of M. tuberculosis. When used as a single drug, SILA-421 in vitro showed concentration-dependent and time-dependent bactericidal activity. SILA-421 also enhanced the activity of INH and RIF, resulting in synergy in the case of INH. Emergence of INH resistance following exposure to INH can be prevented by the addition SILA-421. SILA-421 had no additional value in combination with MXF or AMK. Furthermore, SILA-421 enhanced the activity of RIF towards a RIF-resistant strain and resulted in complete elimination of RIF-resistant mycobacteria. Unfortunately, in mice with TB induced by a Beijing genotype strain, addition of SILA-421 to an isoniazid-rifampicin-pyrazinamide regimen for 13 weeks did not result in enhanced therapeutic efficacy., (Copyright © 2015 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.)

Objectives: Assessment of the activity of thioridazine towards Mycobacterium tuberculosis (Mtb), in vitro and in vivo as a single drug and in combination with tuberculosis (TB) drugs., Methods: The in vitro activity of thioridazine as single drug or in combination with TB drugs was assessed in terms of MIC and by use of the time-kill kinetics assay. Various Mtb strains among which the Beijing genotype strain BE-1585 were included. In vivo, mice with TB induced by BE-1585 were treated with a TB drug regimen with thioridazine during 13 weeks. Therapeutic efficacy was assessed by the change in mycobacterial load in the lung, spleen and liver during treatment and 13 weeks post-treatment., Results: In vitro, thioridazine showed a concentration-dependent and time-dependent bactericidal activity towards both actively-replicating and slowly-replicating Mtb. Thioridazine at high concentrations could enhance the activity of isoniazid and rifampicin, and in case of isoniazid resulted in elimination of mycobacteria and prevention of isoniazid-resistant mutants. Thioridazine had no added value in combination with moxifloxacin or amikacin. In mice with TB, thioridazine was poorly tolerated, limiting the maximum tolerated dose (MTD). The addition of thioridazine at the MTD to an isoniazid-rifampicin-pyrazinamide regimen for 13 weeks did not result in enhanced therapeutic efficacy., Conclusions: Thioridazine is bactericidal towards Mtb in vitro, irrespective the mycobacterial growth rate and results in enhanced activity of the standard regimen. The in vitro activity of thioridazine in potentiating isoniazid and rifampicin is not reflected by improved therapeutic efficacy in a murine TB-model.

Rationale: The dosage of 10 mg/kg/d rifampin, as currently used in the treatment of tuberculosis (TB), is not an optimal dose. Shortening of treatment duration might be achievable using an increased rifampin dose., Objectives: Determination of optimal rifampin dosage in mice, resulting in maximum therapeutic effect and without adverse effects. Assessment of associated pharmacokinetic parameters and pharmacokinetic/pharmacodynamic indices., Methods: A murine TB infection using a Beijing genotype Mycobacterium tuberculosis strain was established by intratracheal bacterial instillation followed by proper inhalation, while keeping mice in a vertical position. We assessed dose-dependent activity of rifampin in single-drug treatment during 3 weeks. The maximum tolerated dosage, pharmacokinetic parameters, and pharmacokinetic/pharmacodynamic index were determined. Therapeutic efficacy of a range of rifampin (R) dosages added to a regimen of isoniazid (H) and pyrazinamide (Z) was assessed., Measurements and Main Results: Maximum tolerated dosage of rifampin in the murine TB was 160 mg/kg/d. Pharmacokinetic measurement in HR(10)Z and HR(160)Z therapy regimens showed for rifampin a C(max) of 16.2 and 157.3 mg/L, an AUC(0-24h) of 132 and 1,782 h·mg/L, and AUC(0-24h)/minimum inhibitory concentration ratios of 528 and 7129, respectively. A clear dose-effect correlation was observed for rifampin after 3-week single-drug treatment. Administration of HR(80)Z allowed 9-week treatment duration to be effective without relapse of infection., Conclusions: Our findings indicate that the currently used rifampin dosage in the therapy of TB is too low. In our murine TB model a rifampin dosage of 80 mg/kg/d enabled a significant reduction in therapy duration without adverse effects.

Relapse of tuberculosis (TB) is defined as re-emergence of clinical symptoms after stopping anti-TB treatment, while this treatment appeared effective initially. Relapse of TB can occur in patients that are therapy-compliant, but the risk of relapse is dramatically increased when patients are non-compliant. Additionally, the probability of antibiotic resistance is higher in those patients who have a relapse of TB and thus longer treatment is recommended. Further insight in the pathogenesis of relapsing TB could provide a basis for future treatment improvement. In the present study, using a murine TB model, we assessed the differences between primary TB and relapse of TB in terms of mycobacterial load in infected organs, (immuno-) histopathology, and plasma cytokine concentrations. Compared to primary TB, in relapse of TB we observed a lower mycobacterial load in lung, spleen and liver at the phase of established infection. Also the levels of TNF-α, IFN-γ, IL-6, MIG/CXCL9, IP-10/CXCL10 and IL-17 were significantly lower. It was observed that in relapse of TB memory Th-1 cells were locally and systemically expanded and congregated in the lung, permitting an efficient control of Mtb growth. Treatment response in relapse of TB is as good as the treatment response in primary TB; thereby no supportive evidence could be given for the recommended longer treatment duration in case of relapse of TB., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Summary Background Multiple-drug therapy for tuberculosis (TB) and TB-associated co-morbidity increase the likelihood of drug–drug interactions (DDIs). Inhibition of membrane transporters is an important mechanism underlying DDIs. In this study, we assessed the in vitro inhibitory potential of currently used first and second-line TB drugs and of proposed mycobacterial efflux pump inhibitors (EPIs) on the major ABC transporters relevant to drug transport, namely P-gp, BCRP, BSEP and MRP1-5. Methods Membrane vesicles isolated from transporter-overexpressing HEK293 cells were used to study the inhibitory action of TB drugs and EPIs on the transport of model substrates [ 3 H]-NMQ (P-gp); [ 3 H]-E 1 S (BCRP); [ 3 H]-TCA (BSEP); [ 3 H]-E217βG (MRP1, 3 and 4) and [ 3 H]-MTX (MRP2 and 5). Results A strong inhibition (IC 50 value <15 μM) was observed for clofazimine (P-gp, BCRP and MRP1), thioridazine (BCRP), timcodar (P-gp, BSEP and MRP1) and SQ109 (P-gp and BCRP). Rifampicin inhibited all transporters, but less potently. Conclusions Co-administration of clofazimine, thioridazine, timcodar, SQ109 and possibly rifampicin with drugs that are substrates for the inhibited transporters may lead to DDIs. The mycobacterial EPIs potently inhibited a wider range of human ABC transporters than previously reported. These vesicular transport data are especially valuable considering the current emphasis on development of TB drug regimens. [ABSTRACT FROM AUTHOR]