Mycobacterium abscessus strain variability in preclinical drug development: does it really matter?

BACKGROUND: New treatment options for Mycobacterium abscessus infections are urgently needed. Since a correlation between MICs and clinical outcomes is not clearly established, potency of novel drugs needs to be evaluated using additional in vitro drug activity assays. Preclinical drug activity assays generally use the M. abscessus type strain ATCC 19977. However, M. abscessus complex entails a genetically and morphologically diverse group, and it is questionable whether drug activity observed against ATCC 19977 is representative of drug activity against clinical M. abscessus isolates. OBJECTIVES: To assess whether the relationship between MIC and the quantitative antimycobacterial activity of amikacin, imipenem and clofazimine differs between the ATCC 19977 strain and clinical M. abscessus isolates. METHODS: Experiments were performed with M. abscessus ATCC 19977 and a subset of six clinical isolates covering the three M. abscessus subspecies and the smooth and rough morphotypes. Cultures were exposed to the drugs at 4-fold increasing, MIC-standardized concentrations, and the mycobacterial load was assessed over time. RESULTS: Concentration- and time-dependent activity of amikacin, imipenem and clofazimine against the six clinical isolates was similar. Only slight variations in drug activity were observed between ATCC 19977 and clinical isolates. CONCLUSIONS: Time- and concentration-dependent drug activity against the ATCC 19977 strain seems indicative for in vitro drug behaviour against M. abscessus complex clinical isolates. Including one clinical smooth morphotype isolate alongside ATCC 19977 seems appropriate for reliable interpretation of this particular in vitro drug activity assay as part of the M. abscessus preclinical drug development pipeline.

Relative contributions of the novel diarylquinoline TBAJ-876 and its active metabolite to the bactericidal activity in a murine model of tuberculosis.

BACKGROUND: TBAJ-876 is a next-generation diarylquinoline. In vivo, diarylquinoline metabolites are formed with activity against Mycobacterium tuberculosis. Species-specific differences in parent drug-to-metabolite ratios might impact the translational value of animal model-based predictions. This study investigates the contribution of TBAJ-876 and its major active metabolite, TBAJ-876-M3 (M3), to the total bactericidal activity in a mouse tuberculosis model. METHODS: In vitro activity of TBAJ-876 and M3 was investigated and compared to bedaquiline. Subsequently, a dose-response study was conducted in M. tuberculosis-infected BALB/c mice treated with TBAJ-876 (1.6/6.3/25 mg/kg) or M3 (3.1/12.5/50 mg/kg). Colony-forming units in the lungs and TBAJ-876 and M3 plasma concentrations were determined. M3's contribution to TBAJ-876's bactericidal activity was estimated based on M3-exposure following TBAJ-876 treatment and corresponding M3-activity observed in M3-treated animals. RESULTS: TBAJ-876 and M3 demonstrated profound bactericidal activity. Lungs of mice treated for 4 weeks with 50 mg/kg M3 were culture-negative. Following TBAJ-876 treatment, M3-exposures were 2.2-3.6x higher than for TBAJ-876. TBAJ-876 activity was substantially attributable to M3, given its high exposure and potent activity. CONCLUSION: These findings emphasize the need to consider metabolites and their potentially distinct exposure and activity profiles compared to parent drugs to enhance the translational value of mouse model-driven predictions.

Less Is More: When to Repeat Antimicrobial Susceptibility Testing.

This study investigated the frequency of change of the antimicrobial susceptibility pattern when the same isolate was found in the same patient in various situations. We used laboratory data collected over a period of 8 years (January 2014 to December 2021) at the clinical microbiology laboratory of a tertiary hospital for Escherichia coli, Klebsiella pneumoniae, Enterobacter spp., Pseudomonas aeruginosa, and Staphylococcus aureus. Antimicrobial susceptibility tests (AST) were performed using Vitek 2 automated system. We determined essential agreement and categorical agreement, and introduced the new terms essential MIC increase and change from nonresistant to resistant to present changes in antimicrobial susceptibility over time. During the study period, 18,501 successive AST were included. The risk for S. aureus to be resistant to any antibiotic upon repeated culture was <10% during a follow-up of 30 days. For Enterobacterales, this risk was approximately 10% during a follow-up of 7 days. For P. aeruginosa, this risk was higher. The longer the follow-up period, the higher the risk that the bacteria would show phenotypic resistance. We also found that some drug-bug combinations were more likely to develop phenotypical resistance (i.e., E. coli/amoxicillin-clavulanic acid and E. coli/cefuroxime). A potential consequence of our finding is that if we regard a risk of resistance below 10% as acceptable, it may be feasible to omit follow-up AST within 7 days for the microorganisms investigated in this study. This approach saves money, time, and will reduce laboratory waste. Further studies are needed to determine whether these savings are in balance with the small possibility of treating patients with inadequate antibiotics.

Pharmacokinetics and pharmacodynamics of anti-tuberculosis drugs: An evaluation of in vitro, in vivo methodologies and human studies.

There has been an increased interest in pharmacokinetics and pharmacodynamics (PKPD) of anti-tuberculosis drugs. A better understanding of the relationship between drug exposure, antimicrobial kill and acquired drug resistance is essential not only to optimize current treatment regimens but also to design appropriately dosed regimens with new anti-tuberculosis drugs. Although the interest in PKPD has resulted in an increased number of studies, the actual bench-to-bedside translation is somewhat limited. One of the reasons could be differences in methodologies and outcome assessments that makes it difficult to compare the studies. In this paper we summarize most relevant in vitro, in vivo, in silico and human PKPD studies performed to optimize the drug dose and regimens for treatment of tuberculosis. The in vitro assessment focuses on MIC determination, static time-kill kinetics, and dynamic hollow fibre infection models to investigate acquisition of resistance and killing of Mycobacterium tuberculosis populations in various metabolic states. The in vivo assessment focuses on the various animal models, routes of infection, PK at the site of infection, PD read-outs, biomarkers and differences in treatment outcome evaluation (relapse and death). For human PKPD we focus on early bactericidal activity studies and inclusion of PK and therapeutic drug monitoring in clinical trials. Modelling and simulation approaches that are used to evaluate and link the different data types will be discussed. We also describe the concept of different studies, study design, importance of uniform reporting including microbiological and clinical outcome assessments, and modelling approaches. We aim to encourage researchers to consider methods of assessing and reporting PKPD of anti-tuberculosis drugs when designing studies. This will improve appropriate comparison between studies and accelerate the progress in the field.

Unraveling antibiotic resistance mechanisms in Mycobacterium abscessus: the potential role of efflux pumps.

OBJECTIVES: Mycobacterium abscessus is an opportunistic respiratory pathogen in patients with underlying lung disease. It is infamously known for its low treatment success rates because of its resistance to multiple classes of antibiotics. Further insight into M. abscessus resistance mechanisms is needed to improve treatment options. In this in vitro study, the role of efflux pumps in reaction to antibiotic stress is explored, as well as the ability of the putative efflux inhibitors, thioridazine and verapamil, to potentiate the activity of guideline-recommended antibiotics. METHODS: To evaluate the effects of antibiotic stress on mycobacterial efflux pumps, M. abscessus subspecies abscessus was exposed to amikacin, cefoxitin, clarithromycin, clofazimine, and tigecycline for 24 hours. Transcriptomic responses were measured by RNA sequencing to gain insight into upregulation of efflux pump encoding genes. Subsequently, in time-kill kinetics assays, the above-mentioned antibiotics were combined with thioridazine and verapamil to evaluate their potentiating capacity. RESULTS: All five antibiotics led to a fold change of ≥2 Log2 in expression of one or more genes encoding transporter systems. This effect was most pronounced for the ribosome-targeting antibiotics amikacin, clarithromycin, and tigecycline. Time-kill kinetics assays demonstrated synergy between amikacin, tigecycline, clofazimine, cefoxitin, and both thioridazine and verapamil. CONCLUSION: Antibiotic stressors induce expression of efflux pump encoding genes in M. abscessus, especially antibiotics that target the ribosome. Putative efflux inhibitors thioridazine and verapamil show synergy with various guideline-recommended antibiotics, making them interesting candidates for the improvement of M. abscessus treatment.

Treatment and Outcome of Culture-Confirmed Mycobacterium marinum Disease.

Background: Mycobacterium marinum is a nontuberculous mycobacterium that causes skin and soft tissue infections. Treatment consists of multiple antibiotics, sometimes combined with surgical debridement. There is little evidence for the choice of antibiotics, the duration of treatment, and the role of susceptibility testing. Methods: We performed a retrospective cohort study of culture-confirmed M. marinum infections in the Netherlands in the 2011-2018 period. Clinical characteristics, in vitro susceptibility, extent of disease, treatment regimens, and outcomes were analyzed. Incidence was assessed from laboratory databases. Results: Forty cases of M. marinum infection could be studied. Antibiotic treatment cured 36/40 patients (90%) after a mean treatment duration of 25 weeks. Failure/relapse occurred in 3 patients, and 1 patient was lost to follow-up. Antibiotic treatment consisted of monotherapy in 35% and 2-drug therapy in 63%. Final treatment contained mostly ethambutol-macrolide combinations (35%). Eleven patients (28%) received additional surgery. We recorded high rates of in vitro resistance to tetracyclines (36% of isolates). Tetracycline resistance seemed correlated with poor response to tetracycline monotherapy. The annual incidence rate was 0.15/100 000/year during the study period. Conclusions: Prolonged and susceptibility-guided treatment results in a 90% cure rate in M. marinum disease. Two-drug regimens of ethambutol and a macrolide are effective for moderately severe infections. Tetracycline monotherapy in limited disease should be used vigilantly, preferably with proven in vitro susceptibility.

Delamanid or pretomanid? A Solomonic judgement!

Given the low treatment success rates of drug-resistant tuberculosis (TB), novel TB drugs are urgently needed. The landscape of TB treatment has changed considerably over the last decade with the approval of three new compounds: bedaquiline, delamanid and pretomanid. Of these, delamanid and pretomanid belong to the same class of drugs, the nitroimidazoles. In order to close the knowledge gap on how delamanid and pretomanid compare with each other, we summarize the main findings from preclinical research on these two compounds. We discuss the compound identification, mechanism of action, drug resistance, in vitro activity, in vivo pharmacokinetic profiles, and preclinical in vivo activity and efficacy. Although delamanid and pretomanid share many similarities, several differences could be identified. One finding of particular interest is that certain Mycobacterium tuberculosis isolates have been described that are resistant to either delamanid or pretomanid, but with preserved susceptibility to the other compound. This might imply that delamanid and pretomanid could replace one another in certain regimens. Regarding bactericidal activity, based on in vitro and preclinical in vivo activity, delamanid has lower MICs and higher mycobacterial load reductions at lower drug concentrations and doses compared with pretomanid. However, when comparing in vivo preclinical bactericidal activity at dose levels equivalent to currently approved clinical doses based on drug exposure, this difference in activity between the two compounds fades. However, it is important to interpret these comparative results with caution knowing the variability inherent in preclinical in vitro and in vivo models.

Predictive Modeling to Study the Treatment-Shortening Potential of Novel Tuberculosis Drug Regimens, Toward Bundling of Preclinical Data.

BACKGROUND: Given the persistently high global burden of tuberculosis, effective and shorter treatment options are needed. We explored the relationship between relapse and treatment length as well as interregimen differences for 2 novel antituberculosis drug regimens using a mouse model of tuberculosis infection and mathematical modeling. METHODS: Mycobacterium tuberculosis-infected mice were treated for up to 13 weeks with bedaquiline and pretomanid combined with moxifloxacin and pyrazinamide (BPaMZ) or linezolid (BPaL). Cure rates were evaluated 12 weeks after treatment completion. The standard regimen of isoniazid, rifampicin, pyrazinamide, and ethambutol (HRZE) was evaluated as a comparator. RESULTS: Six weeks of BPaMZ was sufficient to achieve cure in all mice. In contrast, 13 weeks of BPaL and 24 weeks of HRZE did not achieve 100% cure rates. Based on mathematical model predictions, 95% probability of cure was predicted to occur at 1.6, 4.3, and 7.9 months for BPaMZ, BPaL, and HRZE, respectively. CONCLUSION: This study provides additional evidence for the treatment-shortening capacity of BPaMZ over BPaL and HRZE. To optimally use preclinical data for predicting clinical outcomes, and to overcome the limitations that hamper such extrapolation, we advocate bundling of available published preclinical data into mathematical models.

The association of body temperature with antibiotic therapy and mortality in patients attending the emergency department with suspected infection.

BACKGROUND AND IMPORTANCE: Previous studies found that septic patients with normothermia have higher mortality than patients with fever. We hypothesize that antibiotic therapy is less frequently initiated if infectious patients present with normothermia to the emergency department (ED). OBJECTIVES: To examine the association of body temperature with the initiation of antibiotic therapy in patients attending the ED with suspected and proven infection. Additionally, the association of temperature with 30-day mortality was assessed. DESIGN, SETTINGS AND PARTICIPANTS: We conducted a retrospective cohort study between 2012 and 2016 at a tertiary university hospital. Adult patients attending the ED with a blood culture taken (i.e. suspected infection) and a positive blood culture (i.e. proven bacteremia) were included. EXPOSURE: Tympanic temperature at arrival was categorized as hypothermia (<36.1°C), normothermia (36.1-38.0°C) or hyperthermia (>38.0°C). OUTCOME MEASURES AND ANALYSIS: Primary outcome was the initiation of antibiotic therapy. A secondary outcome was 30-day mortality. Multivariable logistic regression was used to control for covariates. MAIN RESULTS: Of 5997 patients with a suspected infection, 45.8% had normothermia, 44.6% hyperthermia and 5.6% hypothermia. Patients with hyperthermia received more often antibiotic therapy (53.5%) compared to normothermic patients (27.6%, adjusted odds ratio [95% confidence interval], 2.59 [2.27-2.95]). Patients with hyperthermia had lower mortality (4.7%) than those with normothermia (7.4%, adjusted odds ratio [95% confidence interval], 0.50 [0.39-0.64]). Sensitivity analyses in patients with proven bacteremia (n = 934) showed similar results. CONCLUSION: Normothermia in patients presenting with infection was associated with receiving less antibiotic therapy in the ED compared to presentations with hyperthermia. Moreover, normothermia was associated with a higher mortality risk than hyperthermia.

Duration and key determinants of infectious virus shedding in hospitalized patients with coronavirus disease-2019 (COVID-19).

Key questions in COVID-19 are the duration and determinants of infectious virus shedding. Here, we report that infectious virus shedding is detected by virus cultures in 23 of the 129 patients (17.8%) hospitalized with COVID-19. The median duration of shedding infectious virus is 8 days post onset of symptoms (IQR 5-11) and drops below 5% after 15.2 days post onset of symptoms (95% confidence interval (CI) 13.4-17.2). Multivariate analyses identify viral loads above 7 log10 RNA copies/mL (odds ratio [OR] of 14.7 (CI 3.57-58.1; p < 0.001) as independently associated with isolation of infectious SARS-CoV-2 from the respiratory tract. A serum neutralizing antibody titre of at least 1:20 (OR of 0.01 (CI 0.003-0.08; p < 0.001) is independently associated with non-infectious SARS-CoV-2. We conclude that quantitative viral RNA load assays and serological assays could be used in test-based strategies to discontinue or de-escalate infection prevention and control precautions.

Superior Efficacy of a Bedaquiline, Delamanid, and Linezolid Combination Regimen in a Mouse Tuberculosis Model.

BACKGROUND: The treatment success rate of drug-resistant (DR) tuberculosis is alarmingly low. Therefore, more effective and less complex regimens are urgently required. METHODS: We compared the efficacy of an all oral DR tuberculosis drug regimen consisting of bedaquiline (25 mg/kg), delamanid (2.5 mg/kg), and linezolid (100 mg/kg) (BDL) on the mycobacterial load in the lungs and spleen of tuberculosis-infected mice during a treatment period of 24 weeks. This treatment was compared with the standard regimen of isoniazid, rifampicin, pyrazinamide, and ethambutol (HRZE). Relapse was assessed 12 weeks after treatment. Two logistic regression models were developed to compare the efficacy of both regimens. RESULTS: Culture negativity in the lungs was achieved at 8 and 20 weeks of treatment with BDL and HRZE, respectively. After 14 weeks of treatment only 1 mouse had relapse in the BDL group, while in the HRZE group relapse was still observed at 24 weeks of treatment. Predictions from the final mathematical models showed that a 95% cure rate was reached after 20.5 and 28.5 weeks of treatment with BDL and HRZE, respectively. CONCLUSION: The BDL regimen was observed to be more effective than HRZE and could be a valuable option for the treatment of DR tuberculosis.

The tough process of unmasking the slow-growing mycobacterium: case report of Mycobacterium microti infection.

Mycobacterium microti belongs to the Mycobacterium tuberculosis complex (MTBC). It can cause pulmonary and extrapulmonary tuberculosis in humans. Compared to M. tuberculosis , which is the most prevalent subspecies of the MTBC, M. microti infection has a different etiology. Moreover, establishing the diagnosis with conventional bacteriology can be difficult. We will illustrate this with a case of an extrapulmonary tuberculosis of the hip caused by M .microti in an immunocompetent patient in The Netherlands.

Non-adherence to antimicrobial guidelines in patients with bloodstream infection visiting the emergency department.

OBJECTIVE: Non-adherence to antimicrobial guidelines in patients with bloodstream infection can result in undertreatment, overtreatment, or equivalent treatment, and could lead to suboptimal care. Our aim was to examine the association between non-adherence and appropriate coverage as well as to assess the impact of non-adherence on 30-day mortality. METHODS: We conducted a retrospective cohort study between 2012 and 2017 at a tertiary university hospital. Adult patients attending the emergency department with a bloodstream infection were included. Adherence was defined as guideline-recommended antibiotic therapy. Non-adherence was either undertreatment (too narrow-spectrum), overtreatment (too broad-spectrum), or equivalent treatment. Outcomes were appropriate coverage (i.e. antibiotic therapy that matches in vitro susceptibility of the isolated bacteria) and 30-day mortality. RESULTS: We included 909 patients of whom 395 (43.5%) were treated adherently, 355 (39.1%) were undertreated, 87 (9.6%) were overtreated, and 72 (7.9%) received an equivalent treatment. Overtreated patients were more severely ill, whilst undertreated patients had more favorable patient characteristics. Overtreatment did not result in higher appropriate coverage, whereas undertreatment was associated with lower coverage (OR[95%CI]: 0.18 [0.12; 0.26]). Overtreatment and undertreatment were not associated with 30-day mortality. CONCLUSIONS: Guideline adherence likely depends on disease severity, because overtreatment was more often observed in patients with high disease severity and undertreatment in less severely ill patients. Undertreatment was associated lower appropriate coverage but not with higher mortality. However, this can be the result of residual confounding . Overtreatment did not result in higher appropriate antibiotic coverage nor a survival benefit . Therefore, overtreatment seems not justifiable.

Mycobacterium tuberculosis clinical isolates of the Beijing and East-African Indian lineage induce fundamentally different host responses in mice compared to H37Rv.

Substantial differences exist in virulence among Mycobacterium tuberculosis strains in preclinical TB models. In this study we show how virulence affects host responses in mice during the first four weeks of infection with a mycobacterial strain belonging to the Beijing, East-African-Indian or Euro-American lineage. BALB/c mice were infected with clinical isolates of the Beijing-1585 strain or the East-African Indian (EAI)-1627 strain and host responses were compared to mice infected with the non-clinical H37Rv strain of the Euro-American lineage. We found that H37Rv induced a 'classical' T-cell influx with high IFN-γ levels, while Beijing-1585 and EAI-1627 induced an influx of B-cells into the lungs together with elevated pulmonary IL-4 protein levels. Myeloid cells in the lungs appeared functionally impaired upon infection with Beijing-1585 and EAI-1627 with reduced iNOS and IL-12 expression levels compared to H37Rv infection. This impairment might be related to significantly reduced expression in the bone marrow of IFN-γ, TNF-α and IFN-ß in mice infected with Beijing-1585 and EAI-1627, which could be detected from the third day post infection onwards. Our findings suggest that increased virulence of two clinical isolates compared to H37Rv is associated with a fundamentally different systemic immune response, which already can be detected early during infection.

Appropriate empirical antibiotic therapy and mortality: Conflicting data explained by residual confounding.

OBJECTIVE: Clinical practice universally assumes that appropriate empirical antibiotic therapy improves survival in patients with bloodstream infection. However, this is not generally supported by previous studies. We examined the association between appropriate therapy and 30-day mortality, while minimizing bias due to confounding by indication. METHODS: We conducted a retrospective cohort study between 2012 and 2017 at a tertiary university hospital in the Netherlands. Adult patients with bloodstream infection attending the emergency department were included. Based on in vitro susceptibility, antibiotic therapy was scored as appropriate or inappropriate. Primary outcome was 30-day mortality. To control for confounding, we performed conventional multivariable logistic regression and propensity score methods. Additionally, we performed an analysis in a more homogeneous subgroup (i.e. antibiotic monotherapy). RESULTS: We included 1.039 patients, 729 (70.2%) received appropriate therapy. Overall 30-day mortality was 10.4%. Appropriately treated patients had more unfavorable characteristics, indicating more severe illness. Despite adjustments, we found no association between appropriate therapy and mortality. For the antibiotic monotherapy subgroup (n = 449), patient characteristics were more homogeneous. Within this subgroup, appropriate therapy was associated with lower mortality (Odds Ratios [95% Confidence Intervals] ranging from: 0.31 [0.14; 0.67] to 0.40 [0.19; 0.85]). CONCLUSIONS: Comparing heterogeneous treatment groups distorts associations despite use of common methods to prevent bias. Consequently, conclusions of such observational studies should be interpreted with care. If possible, future investigators should use our method of attempting to identify and analyze the most homogeneous treatment groups nested within their study objective, because this minimizes residual confounding.

Omadacycline as a promising new agent for the treatment of infections with Mycobacterium abscessus.

BACKGROUND: Despite intensive treatment regimens, the outcome of Mycobacterium abscessus infections is extremely poor and thus novel treatment regimens are needed. Although tigecycline seems to be one of the best options currently available, its long-term use is hampered by severe toxic side effects as well as the need for intravenous administration and the relatively high concentrations required for efficacy. OBJECTIVES: To assess the in vitro activity of omadacycline against M. abscessus and compare it with the activity of tigecycline. METHODS: The concentration- and time-dependent killing capacities of omadacycline and tigecycline against M. abscessus subspecies abscessus were determined using a time-kill kinetics assay. Time-kill curves as well as concentration-effect curves were generated. RESULTS: Time-kill curves showed strong concentration-dependent antimicrobial activity for both omadacycline and tigecycline. Omadacycline showed inhibition of mycobacterial growth at 4 mg/L and mycobacterial killing at concentrations ≥16 mg/L. Tigecycline showed mycobacterial killing at concentrations ≥4 mg/L, achieving elimination at concentrations ≥16 mg/L. The concentration-effect curves after 7 days of exposure showed stasis, 1 log mycobacterial killing and 2 log mycobacterial killing at 3.3, 4.0 and 4.8 mg/L for omadacycline and 2.2, 2.7 and 3.4 mg/L for tigecycline, respectively. CONCLUSIONS: The results of this in vitro study on omadacycline activity, together with its favourable (pharmacokinetic) properties, suggest that omadacycline is a potential new agent for the treatment of M. abscessus infections.

Assessment of the Additional Value of Verapamil to a Moxifloxacin and Linezolid Combination Regimen in a Murine Tuberculosis Model.

The favorable treatment outcome rate for multidrug-resistant tuberculosis (MDR-TB) is only 54%, and therefore new drug regimens are urgently needed. In this study, we evaluated the activity of the combination of moxifloxacin and linezolid as a possible new MDR-TB regimen in a murine TB model and the value of the addition of the efflux pump inhibitor verapamil to this backbone. BALB/c mice were infected with drug-sensitive Mycobacterium tuberculosis and were treated with human-equivalent doses of moxifloxacin (200 mg/kg of body weight) and linezolid (100 mg/kg) with or without verapamil (12.5 mg/kg) for 12 weeks. Pharmacokinetic parameters were collected during treatment at the steady state. After 12 weeks of treatment, a statistically significant decline in mycobacterial load in the lungs was observed with the moxifloxacin-linezolid regimen with and without verapamil (5.9 and 5.0 log CFU, respectively), but sterilization was not achieved yet. The spleens of all mice were culture negative after 12 weeks of treatment with both treatment modalities, and the addition of verapamil caused a significant reduction in relapse (14/14 positive spleens without versus 9/15 with verapamil, P = 0.017). In conclusion, treatment with a combination regimen of moxifloxacin and linezolid showed a strong decline in mycobacterial load in the mice. The addition of verapamil to this backbone had a modest additional effect in terms of reducing mycobacterial load in the lung as well as reducing the spleen relapse rate. These results warrant further studies on the role of efflux pump inhibition in improving the efficacy of MDR-TB backbone regimens.

Improving treatment outcome assessment in a mouse tuberculosis model.

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 (RpZHE), 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 Emax model in favor over linear or conventional Emax models. Estimating regimen-specific parameters showed significantly higher curative potentials for RZME and RpZHE 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.