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Background The hollow-fibre system model of tuberculosis (HFS-TB) has been endorsed by regulators; however, application of HFS-TB requires a thorough understanding of intra- and inter-team variability, statistical power and quality controls. Methods Three teams evaluated regimens matching those in the Rapid Evaluation of Moxifloxacin in Tuberculosis (REMoxTB) study, plus two high-dose rifampicin/pyrazinamide/moxifloxacin regimens, administered daily for up to 28 or 56 days against Mycobacterium tuberculosis (Mtb) under log-phase growth, intracellular growth or semidormant growth under acidic conditions. Target inoculum and pharmacokinetic parameters were pre-specified, and the accuracy and bias at achieving these calculated using percent coefficient of variation (%CV) at each sampling point and two-way analysis of variance (ANOVA). Results A total of 10 530 individual drug concentrations, and 1026 individual cfu counts were measured. The accuracy in achieving intended inoculum was >98%, and >88% for pharmacokinetic exposures. The 95% CI for the bias crossed zero in all cases. ANOVA revealed that the team effect accounted for 20% difference in slope, with a power of >99%. Conclusions HFS-TB is a highly tractable tool for choosing combination regimens with little variability between teams, and between replicates.

The tuberculosis treatment duration is 4-6 months, but can be 18–20 months with multidrug resistance. Ultrashort regimens that cure regardless of resistance status [“pan-tuberculosis regimens”] would be a major step towards global tuberculosis control. Starting with seven drugs [bedaquiline, delamanid, pretomanid, OPC-167832, sutezolid, moxifloxacin, and pyrazinamide] in clinical testing, we calculated that there were thousands of possible novel combinations to evaluate. We used mathematical and pharmacokinetics/pharmacodynamic-based hollow fiber modeling to reduce this complexity to nine combinations, which we tested. Each regimen’s fast and slow growth bacteria kill-slope trajectories and times-to-extinction were estimated and translated to minimum required therapy duration in patients. Sputum microbial kill trajectories of two combination regimens that have undergone clinical testing were correctly predicted by this approach. Four pan-tuberculosis regimens were predicted to achieve relapse-free cure after 2-3 months therapy in patients. The kill-slopes were used to design expedited clinical trials that minimize patients’ risks.

ObjectivesAnimal models have suggested that the combination of pretomanid with pyrazinamide and moxifloxacin (PaMZ) may shorten TB therapy duration to 3–4 months. Here, we tested that in the hollow-fibre system model of TB (HFS-TB).MethodsA series of HFS-TB experiments were performed to compare the kill rates of the PaMZ regimen with the standard three-drug combination therapy. HFS-TB experiments were performed with bacilli in log-phase growth treated for 28 days, intracellular bacilli treated daily for 28 days and semi-dormant Mycobacterium tuberculosis treated with daily therapy for 56 days for sterilizing effect. Next, time-to-extinction equations were employed, followed by morphism transformation and Latin hypercube sampling, to determine the proportion of patients who achieved a time to extinction of 3, 4 or 6 months with each regimen.ResultsUsing linear regression, the HFS-TB sterilizing effect rates of the PaMZ regimen versus the standard-therapy regimen during the 56 days were 0.18 (95% credible interval=0.13–0.23) versus 0.15 (95% credible interval=0.08–0.21) log10 cfu/mL/day, compared with 0.16 (95% credible interval=0.13–0.18) versus 0.11 (95% credible interval=0.09–0.13) log10 cfu/mL/day in the Phase II clinical trial, respectively. Using time-to-extinction and Latin hypercube sampling modelling, the expected percentages of patients in which the PaMZ regimen would achieve sterilization were 40.37% (95% credible interval=39.1–41.34) and 72.30% (95% credible interval=71.41–73.17) at 3 and 4 months duration of therapy, respectively, versus 93.67% (95% credible interval=93.18–94.13) at 6 months for standard therapy.ConclusionsThe kill rates of the PaMZ regimen were predicted to be insufficient to achieve cure in less than 6 months in most patients.

Standard therapy (isoniazid, rifampin, and ethambutol), with or without a macrolide, for pulmonary Mycobacterium kansasii lasts more than a year. Therefore, shorter treatment duration regimens are required., Standard therapy (isoniazid, rifampin, and ethambutol), with or without a macrolide, for pulmonary Mycobacterium kansasii lasts more than a year. Therefore, shorter treatment duration regimens are required. We used data from 32 Taiwanese patients treated with standard therapy who were followed using repetitive sampling-based sputum M. kansasii time-to-positivity in liquid cultures to calculate kill slopes (γ) based on ordinary differential equations and time-to-extinction of each patient’s bacterial burden. The γ was 0.18 (95% confidence interval [CI] = 0.16 to 0.20) log10 CFU/ml/day on standard therapy. Next, we identified the M. kansasii time to extinction in the hollow-fiber system model of pulmonary M. kansasii disease (HFS-Mkn) treated with standard therapy, which was a γ of 0.60 (95% CI = 0.45 to 0.69) log10 CFU/ml/day. The γ and time-to-extinction values between the two data sets formed structure-preserving maps based on category theory: thus, we could map them from one to the other using morphisms. This mapping identified a multistep nonlinear transformation factor for time to extinction from HFS-Mkn to patients. Next, a head-to-head study in the HFS-Mkn identified median time to extinction for standard therapy of 38.7 (95% CI = 29.1 to 53.2) days, isoniazid-rifampin-ethambutol-moxifloxacin therapy of 21.7 (95% CI = 19.1 to 25) days, isoniazid-rifampin-moxifloxacin therapy of 22 (96% CI = 20.1 to 24.5) days, and rifampin-moxifloxacin-tedizolid therapy of 20.7 (95% CI = 18.5 to 29) days. Our transformation-factor based translation predicted the proportion of patients of 90.7 (88.74 to 92.35)% achieving cure with standard therapy at 12 months, and 6-month cure rates of 99.8 (95% CI = 99.27 to 99.95)% for isoniazid-rifampin-ethambutol-moxifloxacin, 92.2 (90.37 to 93.71)% for isoniazid-rifampin-moxifloxacin, and 99.9 (99.44 to 99.99)% for rifampin-moxifloxacin-tedizolid. Thus, rifampin-moxifloxacin-tedizolid and isoniazid-rifampin-ethambutol-moxifloxacin are predicted to be short-course chemotherapy regimens for pulmonary M. kansasii disease.

Pharmacokinetic (PK) and pharmacodynamic (PD) analyses were conducted to determine the cumulative fraction of response (CFR) for 100 mg twice-daily (BID) and 200 mg once-daily (QD) delamanid in patients with multidrug-resistant tuberculosis (MDR-TB), using a pharmacodynamic target (PDT) that achieves 80% of maximum efficacy. First, in the mouse model of chronic TB, the PK/PD index for delamanid efficacy was determined to be area under the drug concentration-time curve over 24 h divided by MIC (AUC0–24/MIC), with a PDT of 252., Pharmacokinetic (PK) and pharmacodynamic (PD) analyses were conducted to determine the cumulative fraction of response (CFR) for 100 mg twice-daily (BID) and 200 mg once-daily (QD) delamanid in patients with multidrug-resistant tuberculosis (MDR-TB), using a pharmacodynamic target (PDT) that achieves 80% of maximum efficacy. First, in the mouse model of chronic TB, the PK/PD index for delamanid efficacy was determined to be area under the drug concentration-time curve over 24 h divided by MIC (AUC0–24/MIC), with a PDT of 252. Second, in the hollow-fiber system model of tuberculosis, plasma-equivalent PDTs were identified as an AUC0–24/MIC of 195 in log-phase bacteria and 201 in pH 5.8 cultures. Third, delamanid plasma AUC0–24/MIC and sputum bacterial decline data from two early bactericidal activity trials identified a clinical PDT of AUC0–24/MIC of 171. Finally, the CFRs for the currently approved 100-mg BID dose were determined to be above 95% in two MDR-TB clinical trials. The CFR for the 200-mg QD dose, evaluated in a trial in which delamanid was administered as 100 mg BID for 8 weeks plus 200 mg QD for 18 weeks, was 89.3% based on the mouse PDT and >90% on the other PDTs. QTcF (QTc interval corrected for heart rate by Fridericia’s formula) prolongation was approximately 50% lower for the 200 mg QD dose than the 100 mg BID dose. In conclusion, while CFRs of 100 mg BID and 200 mg QD delamanid were close to or above 90% in patients with MDR-TB, more-convenient once-daily dosing of delamanid is feasible and likely to have less effect on QTcF prolongation.

Background: Ceftazidime-avibactam is an effective agent for the treatment of tuberculosis (TB) but requires frequent administration because of a short half-life. Due to a longer half-life, ceftriaxone could allow intermittent dosing. Methods: First, we identified the MIC of ceftriaxone with 15 mg/L avibactam in 30 clinical Mycobacterium tuberculosis isolates. Next, 2 ceftriaxone exposure-effect studies in the intracellular hollow fiber model of TB (HFS-TB) that mimics disseminated disease in young children, were performed. Ceftriaxone was administered once or twice daily for 28 days to explore percentage of time that the concentration persisted above MIC (%TMIC) ranging from 0 to 100%. In a third HFS-TB experiment, the “double cephalosporin” regimen of ceftazidime-ceftriaxone-avibactam was examined and analyzed using Bliss Independence. Conclusion: The MIC99 of the clinical strains was 32 mg/L, in the presence of 15 mg/L avibactam. Ceftriaxone %TMIC 54% demonstrated a 4.1 log10 colony-forming units per milliliter M. tuberculosis kill, while %TMIC mediating Emax was 68%. The “double cephalosporin” combination was highly synergistic. Monte Carlo experiments of 10,000 subjects identified the optimal ceftriaxone dose as 100 mg/kg twice a day. Conclusion: The combination of ceftriaxone-avibactam at 100 mg/kg could achieve Emax in >90% of children. The ceftriaxone potent activity M. tuberculosis could potentially shorten therapy in children with disseminated TB.

Background MDR-TB and XDR-TB have poor outcomes. Objectives To examine the efficacy of tigecycline monotherapy in the hollow fibre system model of TB. Methods We performed pharmacokinetic/pharmacodynamic studies using tigecycline human-like concentration-time profiles in the hollow fibre system model of TB in five separate experiments using Mycobacterium tuberculosis in log-phase growth or as semi-dormant or intracellular bacilli, as monotherapy. We also compared efficacy with the isoniazid/rifampicin/pyrazinamide combination (standard therapy). We then applied extinction mathematics, morphisms and Latin hypercube sampling to identify duration of therapy with tigecycline monotherapy. Results The median tigecycline MIC for 30 M. tuberculosis clinical and laboratory isolates (67% MDR/XDR) was 2 mg/L. Tigecycline monotherapy was highly effective in killing M. tuberculosis in log-phase-growth and semi-dormant and intracellular M. tuberculosis. Once-a-week dosing had the same efficacy as daily therapy for the same cumulative dose; thus, tigecycline efficacy was linked to the AUC0-24/MIC ratio. Tigecycline replacement by daily minocycline after 4 weeks of therapy was effective in sterilizing bacilli. The AUC0-24/MIC ratio associated with optimal kill was 42.3. Tigecycline monotherapy had a maximum sterilizing effect (day 0 minus day 28) of 3.06 ± 0.20 log10 cfu/mL (r2 = 0.92) compared with 3.92 ± 0.45 log10 cfu/mL (r2 = 0.80) with optimized standard therapy. In our modelling, at a tigecycline monotherapy duration of 12 months, the proportion of patients with XDR-TB who reached bacterial population extinction was 64.51%. Conclusions Tigecycline could cure patients with XDR-TB or MDR-TB who have failed recommended therapy. Once-a-week tigecycline could also replace second-line injectables in MDR-TB regimens.

Drug-resistant tuberculosis represents a global emergency, requiring new drugs. We found that minocycline was highly potent in laboratory strains of Mycobacterium tuberculosis and that 30 drug-susceptible and multidrug/extensively drug-resistant clinical strains were susceptible to clinically achievable concentrations. In the hollow fiber system model, lung concentration-time profiles of 7 mg/kg/day human-equivalent minocycline dose achieved bacterial kill rates equivalent to those of first-line antituberculosis agents. Minocycline killed extracellular bacilli directly. Minocycline also killed intracellular bacilli indirectly, via concentration-dependent granzyme A-driven apoptosis. Moreover, minocycline demonstrated dose-dependent antiinflammatory activity and downregulation of extracellular matrix-based remodeling pathways and, thus, could protect patients from tuberculosis immunopathology. In RNA sequencing of repetitive samples from the hollow fiber system and in independent protein abundance experiments, minocycline demonstrated dose-dependent inhibition of sonic hedgehog-patched-gli signaling. These findings have implications for improved lung remodeling and for dual immunomodulation and direct microbial kill-based treatment shortening regimens for drug-susceptible and drug-resistant latent and active M. tuberculosis infection.

Background: d-cycloserine is used to treat multidrug-resistant tuberculosis. Its efficacy, contribution in combination therapy, and best clinical dose are unclear, also data on the d-cycloserine minimum inhibitory concentration (MIC) distributions is scant.Methods: We performed a systematic search to identify pharmacokinetic and pharmacodynamic studies performed with d-cycloserine. We then performed a combined exposure-effect and dose fractionation study of d-cycloserine in the hollow fiber system model of tuberculosis (HFS-TB). In parallel, we identified d-cycloserine MICs in 415 clinical Mycobacterium tuberculosis (Mtb) isolates from patients. We utilized these results, including intracavitary concentrations, to identify the clinical dose that would be able to achieve or exceed target exposures in 10000 patients using Monte Carlo experiments (MCEs).Results: There were no published d-cycloserine pharmacokinetics/pharmacodynamics studies identified. Therefore, we performed new HFS-TB experiments. Cyloserine killed 6.3 log10 colony-forming units (CFU)/mL extracellular bacilli over 28 days. Efficacy was driven by the percentage of time concentration persisted above MIC (%TMIC), with 1.0 log10 CFU/mL kill achieved by %TMIC = 30% (target exposure). The tentative epidemiological cutoff value with the Sensititre MYCOTB assay was 64 mg/L. In MCEs, 750 mg twice daily achieved target exposure in lung cavities of 92% of patients whereas 500 mg twice daily achieved target exposure in 85% of patients with meningitis. The proposed MCE-derived clinical susceptibility breakpoint at the proposed doses was 64 mg/L.Conclusions: Cycloserine is cidal against Mtb. The susceptibility breakpoint is 64 mg/L. However, the doses likely to achieve the cidality in patients are high, and could be neurotoxic.

BACKGROUND: Linezolid exhibits remarkable sterilizing effect in tuberculosis; however, a large proportion of patients develop serious adverse events. The congener tedizolid could have a better side-effect profile, but its sterilizing effect potential is unknown. METHODS: We performed a 42-day tedizolid exposure-effect and dose-fractionation study in the hollow fiber system model of tuberculosis for sterilizing effect, using human-like intrapulmonary pharmacokinetics. Bacterial burden was examined using time to positivity (TTP) and colony-forming units (CFUs). Exposure-effect was examined using the inhibitory sigmoid maximal kill model. The exposure mediating 80% of maximal kill (EC(80)) was defined as the target exposure, and the lowest dose to achieve EC(80) was identified in 10000-patient Monte Carlo experiments. The dose was also examined for probability of attaining concentrations associated with mitochondrial enzyme inhibition. RESULTS: At maximal effect, tedizolid monotherapy totally eliminated 7.1 log(10) CFU/mL Mycobacterium tuberculosis over 42 days; however, TTP still demonstrated some growth. Once-weekly tedizolid regimens killed as effectively as daily regimens, with an EC(80) free drug 0- to 24-hour area under the concentration–time curve-to-minimum inhibitory concentration (MIC) ratio of 200. An oral tedizolid of 200 mg/day achieved the EC(80) in 92% of 10000 patients. The susceptibility breakpoint was an MIC of 0.5 mg/L. The 200 mg/day dose did not achieve concentrations associated with mitochondrial enzyme inhibition. CONCLUSIONS: Tedizolid exhibits dramatic sterilizing effect and should be examined for pulmonary tuberculosis. A tedizolid dose of 200 mg/day or 700 mg twice a week is recommended for testing in patients; the intermittent tedizolid dosing schedule could be much safer than daily linezolid.

BACKGROUND: One approach that could increase the efficacy and reduce the duration of antituberculosis therapy is pharmacokinetics/pharmacodynamics-based optimization of doses. However, this could increase toxicity. METHODS: We mimicked the concentration-time profiles achieved by human equivalent doses of moxifloxacin 800 mg/day, rifampin 1800 mg/day, and pyrazinamide 4000 mg/day (high-dose regimen) vs isoniazid 300 mg/day, rifampin 600 mg/day, and pyrazinamide 2000 mg/day (standard therapy) in bactericidal and sterilizing effect studies in the hollow fiber system model of tuberculosis (HFS-TB). In an intracellular Mycobacterium tuberculosis (Mtb) HFS-TB experiment, we added a 3-dimensional human organotypic liver to determine potential hepatotoxicity of the high-dose regimen, based on lactate dehydrogenase (LDH). Treatment lasted 28 days and Mtb bacterial burden was based on colony counts. We calculated the time to extinction (TTE) of the Mtb population in the HFS-TB and used morphism-based transformation and Latin hypercube sampling to identify the minimum therapy duration in patients. RESULTS: The kill rate of standard therapy in the bactericidal effect and sterilizing effect experiments were 0.97 (95% confidence interval [CI], .91–.99) log(10) colony-forming units (CFU)/mL/day, and 0.56 (95% CI, .49–.59) log(10) CFU/mL/day, respectively. The high-dose regimen’s bactericidal and sterilizing effect kill rates were 0.99 (95% CI, .96–.99) log(10) CFU/mL/day and 0.72 (95% CI, .56–.79) log(10) CFU/mL/day, respectively. The upper confidence bound for TTE in patients was 4.5–5 months for standard therapy vs 3.7 months on the high-dose regimen. There were no differences in LDH concentrations between the 2 regimens at any time point (P > .05). CONCLUSIONS: The high-dose regimen may moderately shorten therapy without increased hepatotoxicity compared to standard therapy.

BACKGROUND: A major challenge in medicine is translation of preclinical model findings to humans, especially therapy duration. One major example is recent shorter-duration therapy regimen failures in tuberculosis. METHODS: We used set theory mapping to develop a computational/modeling framework to map the time it takes to extinguish the Mycobacterium tuberculosis population on chemotherapy from multiple hollow fiber system model of tuberculosis (HFS-TB) experiments to that observed in patients. The predictive accuracy of the derived translation transformations was then tested using data from 108 HFS-TB Rapid Evaluation of Moxifloxacin in Tuberculosis (REMoxTB) units, including 756 colony-forming units (CFU)/mL. Derived transformations, and Latin hypercube sampling–guided simulations were used to predict cure and relapse after 4 and 6 months of therapy. Outcomes were compared to observations, in 1932 patients in the REMoxTB clinical trial. RESULTS: HFS-TB serial bacillary burden and serial sputum data in the derivation dataset formed a structure-preserving map. Bactericidal effect was mapped with a single step transformation, while the sterilizing effect was mapped with a 3-step transformation function. Using the HFS-TB REMoxTB data, we accurately predicted the proportion of patients cured in the 4-month REMoxTB clinical trial. Model-predicted vs clinical trial observations were (i) the ethambutol arm (77.0% [95% confidence interval {CI}, 74.4%–79.6%] vs 77.7% [95% CI, 74.3%–80.9%]) and (ii) the isoniazid arm (76.4% [95% CI, 73.9%–79.0%] vs 79.5% [95% CI, 76.1%–82.5%]). CONCLUSIONS: We developed a method to translate duration of therapy outcomes from preclinical models to tuberculosis patients.

BackgroundIn the experimental arm of the OFLOTUB trial, gatifloxacin replaced ethambutol in the standard 4-month regimen for drug-susceptible pulmonary tuberculosis. The study included a nested pharmacokinetic (PK) study. We sought to determine if PK variability played a role in patient outcomes.MethodsPatients recruited in the trial were followed for 24 months, and relapse ascertained using spoligotyping. Blood was drawn for drug concentrations on 2 separate days during the first 2 months of therapy, and compartmental PK analyses was performed. Failure to attain sustained sputum culture conversion at the end of treatment, relapse, or death during follow-up defined therapy failure. In addition to standard statistical analyses, we utilized an ensemble of machine-learning methods to identify patterns and predictors of therapy failure from among 27 clinical and laboratory features.ResultsOf 126 patients, 95 (75%) had favorable outcomes and 19 (15%) failed therapy, relapsed, or died. Pyrazinamide and rifampicin peak concentrations and area under the concentration-time curves (AUCs) were ranked higher (more important) than gatifloxacin AUCs. The distribution of individual drug concentrations and their ranking varied significantly between South African and West African trial sites; however, drug concentrations still accounted for 31% and 75% of variance of outcomes, respectively. We identified a 3-way antagonistic interaction of pyrazinamide, gatifloxacin, and rifampicin concentrations. These negative interactions disappeared if rifampicin peak concentration was above 7 mg/L.ConclusionsConcentration-dependent antagonism contributed to death, relapse, and therapy failure but was abrogated by high rifampicin concentrations. Therefore, increasing both rifampin and gatifloxacin doses could improve outcomes.Clinical Trials RegistrationNCT002216385.

Objectives To determine if ceftaroline and ceftazidime combined with avibactam are efficacious against pulmonary Mycobacterium avium complex (MAC) disease. Methods First, we performed a concentration-effect study of ceftaroline and ceftaroline/avibactam against extracellular MAC in test tubes. Given the difficulty of obtaining avibactam at the time of experimentation, we used a single concentration of commercial ceftazidime/avibactam, and two sets of non-treated controls, one with ceftazidime/avibactam and the other without. After finding antimicrobial activity with the ceftazidime/avibactam 'control', we performed ceftazidime/avibactam dose-effect studies in test tubes against extracellular MAC and in 24-well plates against intracellular MAC. We then performed a ceftazidime/avibactam exposure-effect and dose-fractionation studies in the hollow-fibre system model of intracellular pulmonary MAC (HFS-MAC). In each experiment, we repetitively sampled each HFS-MAC at specified times to validate ceftazidime/avibactam pharmacokinetics and to quantify bacterial burden. Results Ceftaroline killed extracellular MAC with maximal microbial kill (Emax) of 4.87 ± 0.26 log10 cfu/mL. However, the ceftazidime/avibactam 'control' also killed MAC compared with the non-treated control. Ceftazidime/avibactam Emax was 3.8 log10 cfu/mL against extracellular bacilli and 3.6 log10 cfu/mL against intracellular MAC. In the HFS-MAC, ceftazidime/avibactam achieved a half-life of 2.5-3.3 h and killed MAC 0.61-2.40 log10 cfu/mL below the starting bacterial burden. The ceftazidime/avibactam efficacy was linked to the proportion of the dosing interval for which the concentration persists above the MIC (fT>MIC), with optimal efficacy at free-drug fT>MIC of 52% (r2 = 0.95). Conclusions Ceftazidime/avibactam effectively kills MAC at exposures easily achieved in the lung by clinical doses. Efficacy was higher than with clinically achievable doses of azithromycin and ethambutol.

Objectives To investigate the performance of the two backbone drugs in the standard combination therapy regimen in the hollow-fibre system (HFS) model of pulmonary Mycobacterium avium complex (MAC) infection. Methods Six HFS were inoculated with human-derived monocytes infected with MAC, and treated with 15 mg/kg of ethambutol and 500 mg of azithromycin daily for 28 days to recapitulate the concentration-time profiles seen in the lungs of humans treated with these drugs and doses. The concentration-time profiles achieved were validated by sampling the central compartment at seven timepoints over 24 h. The total MAC burden, as well as the subpopulation resistant to 3 × MIC of each drug, was identified based on sampling the peripheral compartment of each system on days 0, 3, 7, 14, 21 and 28 of therapy. The experiment was performed twice. Results In non-treated control HFS, MAC grew from 5.0 to 8.53 log10 cfu/mL in 28 days. The dual therapy killed a maximum of 1.52 ± 0.43 log10 cfu/mL during the first 7 days, after which it failed. By day 28 there was no difference in MAC burden between the combination-therapy-treated and non-treated systems. Failure arose in parallel with the emergence of acquired ethambutol resistance. By day 28, 100% of the bacterial population was ethambutol resistant in the combination-therapy-treated HFS replicates. Conclusions The backbone combination of macrolide and ethambutol has poor MAC kill rates and is ineffective. Microbial kill is rapidly abrogated by acquired drug resistance. This backbone should be replaced.

Objectives To perform a systematic review and meta-analysis of the level of funding support and the sputum culture conversion rates in pulmonary Mycobacterium avium-intracellulare complex (P-MAC) disease in adult patients without cystic fibrosis or HIV infection, treated with recommended antibiotic regimens. Methods We performed a literature search to identify clinical trials, prospective studies and registries that reported outcomes in P-MAC patients. Studies that reported P-MAC diagnosis and treatments based on established guidelines met the inclusion criteria and were examined for bias and quality. We modified existing quality scales and came up with a 10 star quality score. Outcomes meta-analysed were sputum conversion incidence ratios (IR) and their 95% CI, weighted for study quality. Results Twenty-one studies that examined 28 regimens, including 2534 patients in intent-to-treat analyses and 1968 in per-protocol analyses, were identified. The study quality mean ± SD scores were 5.4 ± 2.2 out of 10 stars. Only two (9.5%) studies received public funding. There was significant heterogeneity of microbial effect among treatment regimens (I2 > 40%; P > 0.001). The pooled IR for sustained sputum conversion was 0.54 (95% CI 0.45-0.63) for macrolide-containing regimens versus 0.38 (0.25-0.52) with macrolide-free regimens. Prolonging therapy duration beyond 12 months was associated with an average decline in sputum conversion to 22% (95% CI 1%-44%). Conclusions Researchers working on P-MAC therapy have received very little public funding support. As a result, the evidence base for treatment guidelines is based on studies of relatively small numbers of patients in low-quality studies. Nevertheless, these studies showed poor sputum conversion rates in patients receiving recommended treatment regimens.

Background. Infants and toddlers often present with disseminated and lymph node tuberculosis, in which Mycobacterium tuberculosis (Mtb) is predominantly intracellular. Linezolid, used to treat tuberculosis in adults, has not been formally studied in infants. Infants clear linezolid 5 times faster than adults and achieve lower 0- to 24-hour area under the concentration-time curves (AUC0–24). Methods. To mimic intracellular disease, we infected human-derived THP-1 macrophages with Mtb and inoculated hollow fiber systems. We performed dose-effect and dose-scheduling studies in which we recapitulated the linezolid half-life of 3 hours encountered in infants. Repetitive sampling for linezolid pharmacokinetics, Mtb intracellular burden, viable monocyte count, and RNA sequencing reads were performed up to 28 days. Results. The linezolid extracellular half-life was 2.64 ± 0.38 hours, whereas intracellular half-life was 8.93 ± 1.30 hours (r2 = 0.89). Linezolid efficacy was linked to the AUC0–24 to minimum inhibitory concentration (MIC) ratio (r2 = 0.98). The exposure associated with maximal Mtb kill was an AUC0–24/MIC of 23.37 ± 1.16. We identified a 414-gene transcript on exposure to toxic linezolid doses. The largest number of genes mapped to ribosomal proteins, a signature hitherto not associated with linezolid toxicity. The second-largest number of differentially expressed genes mapped to mitochondrial enzyme inhibition. Linezolid AUC0–24 best explained the mitochondrial gene inhibition, with 50% inhibition at 94 mg × hour/L (highest r2 = 0.98). Conclusions. We identified the linezolid AUC0–24/MIC target for optimal efficacy against pediatric intracellular tuberculosis, and an AUC0–24 threshold associated with mitochondrial inhibition. These constitute a therapeutic window to be targeted for optimal linezolid doses in children with tuberculosis.

Background. The role of drug concentrations in clinical outcomes in children with tuberculosis is unclear. Target concentrations for dose optimization are unknown. Methods. Plasma drug concentrations measured in Indian children with tuberculosis were modeled using compartmental pharmacokinetic analyses. The children were followed until end of therapy to ascertain therapy failure or death. An ensemble of artificial intelligence algorithms, including random forests, was used to identify predictors of clinical outcome from among 30 clinical, laboratory, and pharmacokinetic variables. Results. Among the 143 children with known outcomes, there was high between-child variability of isoniazid, rifampin, and pyrazinamide concentrations: 110 (77%) completed therapy, 24 (17%) failed therapy, and 9 (6%) died. The main predictors of therapy failure or death were a pyrazinamide peak concentration

Aminoglycosides such as amikacin are currently used for the treatment of multidrug-resistant tuberculosis (MDR-TB). However, formal pharmacokinetic/pharmacodynamic (PK/PD) studies to identify amikacin exposures and dosing schedules that optimize Mycobacterium tuberculosis killing have not been performed. It is believed that aminoglycosides do not work well under acidic conditions, which, if true, would mean poor sterilizing activity against semidormant bacilli at low pH. We performed time-kill studies to compare the bactericidal effect of amikacin in log-phase-growth bacilli with the sterilizing effect in semidormant bacilli at pH 5.8 in broth. In log-phase M. tuberculosis at normal pH versus semidormant M. tuberculosis at pH 5.8, the maximal kill ( E max ) estimate and 95% confidence interval (CI) were 5.39 (95% CI, 4.91 to 5.63) versus 4.88 (CI, 4.46 to 5.22) log 10 CFU/ml, while the concentration mediating 50% of E max (EC 50 ) was 1.0 (CI, 0. 0.86 to 1.12) versus 0.60 (CI, 0.50 to 0.66) times the MIC, respectively. Thus, the optimal exposures and kill rates identified for log-phase M. tuberculosis will be optimal even for semidormant bacilli. Next, we performed exposure-response and dose-scheduling studies in the hollow-fiber system model of tuberculosis using log-phase M. tuberculosis . We recapitulated the amikacin concentration-time profiles observed in lungs of patients treated over 28 days. The PK/PD index linked to M. tuberculosis kill was the peak concentration ( C max )-to-MIC ratio ( r 2 > 0.99), closely followed by the area under the concentration-time curve from 0 to 24 h (AUC 0–24 )-to-MIC ratio ( r 2 = 0.98). The EC 90 was a C max /MIC ratio of 10.13 (95% CI, 7.73 to 12.48). The EC 90 is the dosing target for intermittent therapy that optimizes cure in TB programs for MDR-TB patients.

Children with tuberculosis are treated with drug regimens copied from adults despite significant differences in antibiotic pharmacokinetics, pathology, and the microbial burden between childhood and adult tuberculosis. We sought to develop a new and effective oral treatment regimen specific to children of different ages. We investigated and validated the concept that target drug concentrations associated with therapy failure and death in children are different from those of adults. On that basis, we proposed a 4-step program to rapidly develop treatment regimens for children. First, target drug concentrations for optimal efficacy are derived from preclinical models of disseminated tuberculosis that recapitulate pediatric pharmacokinetics, starting with monotherapy. Second, 2-drug combinations were examined for zones of synergy, antagonism, and additivity based on a whole exposure-response surface. Exposures associated with additivity or synergy were then combined and the regimen was compared to standard therapy. Third, several exposures of the third drug were added, and a 3-drug regimen was identified based on kill slopes in comparison to standard therapy. Fourth, computer-aided clinical trial simulations are used to identify clinical doses that achieve these kill rates in children in different age groups. The proposed program led to the development of a 3-drug combination regimen for children from scratch, independent of adult regimens, in

Mycobacterium avium-intracellulare complex (MAC) causes an intractable intracellular infection that presents as chronic pulmonary disease. Currently, therapy consists of ethambutol and macrolides and takes several years to complete. The neuroleptic phenothiazine thioridazine kills mycobacteria by inhibiting the electron transport chain. In several experiments with bacterial populations of up to 10 12 CFU/ml, we failed to isolate any bacteria resistant to 3 times the MIC of thioridazine, suggesting the absence of resistant mutants at bacterial burdens severalfold higher than those encountered in patients. In the hollow-fiber model of intracellular MAC (HFS-MAC), thioridazine achieved an extracellular half-life of 16.8 h and an intracellular half-life of 19.7 h. Thioridazine concentrations were >28,000-fold higher inside infected macrophages than in the HFS-MAC central compartment (equivalent to plasma). Thioridazine maximal kill was 5.20 ± 0.75 log 10 CFU/ml on day 7 ( r 2 = 0.96) and 7.19 ± 0.31 log 10 CFU/ml on day 14 ( r 2 = 0.99), the highest seen with any drug in the system. Dose fractionation studies revealed that thioridazine efficacy and acquired drug resistance were driven by the peak concentation-to-MIC ratio, with a 50% effective concentration (EC 50 ) of 2.78 ± 0.44 for microbial killing. Acquired drug resistance was encountered by day 21 with suboptimal doses, demonstrating that fluctuating drug concentrations drive evolution faster than static concentrations in mutation frequency studies. However, the thioridazine EC 50 changed 16.14-fold when the concentration of fetal bovine serum was changed from 0% to 50%, suggesting that intracellular potency could be heavily curtailed by protein binding. Efficacy in patients will depend on the balance between trapping of the drug in the pulmonary system and the massive intracellular concentrations versus very high protein binding of thioridazine.

Mycobacterium avium complex is now the leading mycobacterial cause of chronic pneumonia in the United States. Macrolides and ethambutol form the backbone of the regimen used in the treatment of pulmonary disease. However, therapy outcomes remain poor, with microbial cure rates of 4% in cavitary disease. The treatment dose of azithromycin has mostly been borrowed from that used to treat other bacterial pneumonias; there are no formal dose-response studies in pulmonary M. avium disease and the optimal dose is unclear. We utilized population pharmacokinetics and pharmacokinetics/pharmacodynamics-derived azithromycin exposures associated with optimal microbial kill or resistance suppression to perform 10,000 patient Monte Carlo simulations of dose effect studies for daily azithromycin doses of 0.5 to 10 g. The currently recommended dose of 500 mg per day achieved the target exposures in 0% of patients. Exposures associated with optimal kill and resistance suppression were achieved in 87 and 54% of patients, respectively, only by the very high dose of 8 g per day. The azithromycin susceptibility breakpoint above which patients failed therapy on the very high doses of 8 g per day was an MIC of 16 mg/liter, suggesting a critical concentration of 32 mg/liter, which is 8-fold lower than the currently used susceptibility breakpoint of 256 mg/liter. If the standard dose of 500 mg a day were used, then the critical concentration would fall to 2 mg/liter, 128-fold lower than 256 mg/liter. The misclassification of resistant isolates as susceptible could explain the high failure rates of current doses.

Background Current therapy for pulmonary Mycobacterium avium-complex [MAC] disease achieves poor sustained sputum conversion rates and is poorly tolerated. SPR719, the active metabolite of SPR720, a novel gyrase inhibitor, has demonstrated low MICs against MAC. SPR720 is being developed as an oral therapy for use in combination with other antibiotics for the treatment of patients with pulmonary disease due to infection with MAC. Our objective was identify SPR719 pharmacokinetic/pharmacodynamic [PK/PD] parameters and optimal SPR720 dose for treatment of pulmonary MAC. Methods SPR719 was administered once daily for 28 days using a simulated human half-life of 3.3 hours in the hollow fiber system model of pulmonary intracellular MAC [HFS-MAC]. Bacterial burden, including for SPR719-resistant subpopulations, and drug concentrations, were measured via repetitive sampling of HFS-MAC units. A separate dose fractionation study in the HFS-MAC was used to identify the PK/PD index linked to effect. MAC burden versus SPR719 exposure was modeled using the inhibitory sigmoid maximal effect [Emax] model and resistance using the “antibiotic resistance arrow of time” model. Finally, we performed Monte Carlo Experiments to identify the optimal clinical dose of SPR720 monotherapy. Results The median HFS-MAC intracellular-to-extracellular SPR719 AUC0-24 ratio was 2300:1. The PK/PD parameter best linked to microbial kill was determined to be AUC0-24/MIC. SPR719 Emax was -1.5 log10 cfu/mL compared to day 0; 1.0 log10 cfu/mL reduction and acquired-resistance suppression were achieved by an AUC0-24/MIC of 2.0 and 11, respectively. SPR720 1,000 mg/day was predicted to achieve 1.0 log10 cfu/mL kill in 95%, and resistance suppression in 43%, of 10,000 simulated subjects. Conclusion SPR720 monotherapy is predicted to achieve exposures associated with bactericidal effect against pulmonary MAC in 95% of patients at doses that have recently been established to be safe and well tolerated. These data support the continued development of SPR720 for the treatment of pulmonary MAC. Disclosures Nicole Cotroneo, Spero Therapeutics (Employee) David Melnick, MD, Spero Therapeutics (Employee)Spero Therapeutics (Employee) Troy Lister, PhD, Spero Therapeutics (Employee) Suzanne Stokes, PhD, Spero Therapeutics (Employee, Shareholder) Tawanda Gumbo, MD, Praedicare Inc (Employee, Shareholder)

BACKGROUND: Ethionamide is used to treat multidrug-resistant tuberculosis (MDR-TB). The antimicrobial pharmacokinetics/pharmacodynamics, the contribution of ethionamide to the multidrug regimen, and events that lead to acquired drug resistance (ADR) are unclear. METHODS: We performed a multidose hollow fiber system model of tuberculosis (HFS-TB) study to identify the 0–24 hour area under the concentration-time curve (AUC(0-24)) to minimum inhibitory concentration (MIC) ratios that achieved maximal kill and ADR suppression, defined as target exposures. Ethionamide-resistant isolates underwent whole-genome and targeted Sanger sequencing. We utilized Monte Carlo experiments (MCEs) to identify ethionamide doses that would achieve the target exposures in 10000 patients with pulmonary tuberculosis. We also identified predictors of time-to-sputum conversion in Tanzanian patients on ethionamide- and levofloxacin-based regimens using multivariate adaptive regression splines (MARS). RESULTS: An AUC(0-24)/MIC >56.2 was identified as the target exposure in the HFS-TB. Early efflux pump induction to ethionamide monotherapy led to simultaneous ethambutol and isoniazid ADR, which abrogated microbial kill of an isoniazid-ethambutol-ethionamide regimen. Genome sequencing of isolates that arose during ethionamide monotherapy revealed mutations in both ethA and embA. In MCEs, 20 mg/kg/day achieved the AUC(0-24)/MIC >56.2 in >95% of patients, provided the Sensititre assay MIC was

BACKGROUND: Children are often neglected during early development of antituberculosis agents, and most receive treatment after it is first tested in adults. However, very young children have tuberculosis that differs in many respects from adult cavitary pneumonia and could have different toxicity profiles to drugs. Linezolid is effective against intracellular tuberculosis, a common manifestation in young children. However, linezolid has considerable toxicity due to inhibition of mitochondrial enzymes. Tedizolid could be a replacement if it shows equal efficacy and reduced toxicity. METHODS: We performed tedizolid dose-effect studies in the hollow fiber system model of intracellular tuberculosis. We measured linezolid concentrations, colony-forming units (CFU), time-to-positivity, and monocyte viability and performed RNA sequencing on infected cells collected from repetitive sampling of each system. We also compared efficacy of tedizolid vs linezolid and vs tedizolid-moxifloxacin combination. RESULTS: There was no downregulation of mitochondrial enzyme genes, with a tedizolid 0–24 hour area under the concentration-time curve (AUC(0-24)) of up to 90 mg*h/L. Instead, high exposures led to increased mitochondrial gene expression and monocyte survival. The AUC(0-24) to minimum inhibitory concentration ratio associated with 80% of maximal bacterial kill (EC(80)) was 184 by CFU/mL (r(2) = 0.96) and 189 by time-to-positivity (r(2) = 0.99). Tedizolid EC(80) killed 4.0 log(10) CFU/mL higher than linezolid EC(80). The tedizolid-moxifloxacin combination had a bacterial burden elimination rate constant of 0.27 ± 0.05 per day. CONCLUSIONS: Tedizolid demonstrated better efficacy than linezolid, without the mitochondrial toxicity gene or cytotoxicity signatures encountered with linezolid. Tedizolid-moxifloxacin combination had a high bacterial elimination rate.

Background Gatifloxacin is used for the treatment of multidrug-resistant tuberculosis (MDR-TB). The optimal dose is unknown. Methods We performed a 28-day gatifloxacin hollow-fiber system model of tuberculosis (HFS-TB) study in order to identify the target exposures associated with optimal kill rates and resistance suppression. Monte Carlo experiments (MCE) were used to identify the dose that would achieve the target exposure in 10000 adult patients with meningeal or pulmonary MDR-TB. The optimal doses identified were validated using probit analyses of clinical data from 2 prospective clinical trials of patients with pulmonary and meningeal tuberculosis. Classification and regression-tree (CART) analyses were used to identify the gatifloxacin minimum inhibitory concentration (MIC) below which patients failed or relapsed on combination therapy. Results The target exposure associated with optimal microbial kill rates and resistance suppression in the HFS-TB was a 0–24 hour area under the concentration-time curve-to-MIC of 184. MCE identified an optimal gatifloxacin dose of 800 mg/day for pulmonary and 1200 mg/day for meningeal MDR-TB, and a clinical susceptibility breakpoint of MIC ≤ 0.5 mg/L. In clinical trials, CART identified that 79% patients failed therapy if MIC was >2 mg/L, but 98% were cured if MIC was ≤0.5 mg/L. Probit analysis of clinical data demonstrated a >90% probability of a cure in patients if treated with 800 mg/day for pulmonary tuberculosis and 1200 mg/day for meningeal tuberculosis. Doses ≤400 mg/day were suboptimal. Conclusions Gatifloxacin doses of 800 mg/day and 1200 mg/day are recommended for pulmonary and meningeal MDR-TB treatment, respectively. Gatifloxacin has a susceptible dose-dependent zone at MICs 0.5–2 mg/L.

Background Levofloxacin is used for the treatment of multidrug-resistant tuberculosis; however the optimal dose is unknown. Methods We used the hollow fiber system model of tuberculosis (HFS-TB) to identify 0–24 hour area under the concentration-time curve (AUC0-24) to minimum inhibitory concentration (MIC) ratios associated with maximal microbial kill and suppression of acquired drug resistance (ADR) of Mycobacterium tuberculosis (Mtb). Levofloxacin-resistant isolates underwent whole-genome sequencing. Ten thousands patient Monte Carlo experiments (MCEs) were used to identify doses best able to achieve the HFS-TB–derived target exposures in cavitary tuberculosis and tuberculous meningitis. Next, we used an ensemble of artificial intelligence (AI) algorithms to identify the most important predictors of sputum conversion, ADR, and death in Tanzanian patients with pulmonary multidrug-resistant tuberculosis treated with a levofloxacin-containing regimen. We also performed probit regression to identify optimal levofloxacin doses in Vietnamese tuberculous meningitis patients. Results In the HFS-TB, the AUC0-24/MIC associated with maximal Mtb kill was 146, while that associated with suppression of resistance was 360. The most common gyrA mutations in resistant Mtb were Asp94Gly, Asp94Asn, and Asp94Tyr. The minimum dose to achieve target exposures in MCEs was 1500 mg/day. AI algorithms identified an AUC0-24/MIC of 160 as predictive of microbiologic cure, followed by levofloxacin 2-hour peak concentration and body weight. Probit regression identified an optimal dose of 25 mg/kg as associated with >90% favorable response in adults with pulmonary tuberculosis. Conclusions The levofloxacin dose of 25 mg/kg or 1500 mg/day was adequate for replacement of high-dose moxifloxacin in treatment of multidrug-resistant tuberculosis.

Item does not contain fulltext OBJECTIVES: Our aim was to identify the pharmacokinetic/pharmacodynamic parameters of minocycline in the hollow-fibre system (HFS) model of pulmonary Mycobacterium avium complex (MAC) and to identify the optimal clinical dose. METHODS: Minocycline MICs for 55 MAC clinical isolates from the Netherlands were determined. We also co-incubated primary isolated macrophages infected with MAC with minocycline. Next, we performed a 28 day HFS-MAC model dose-response study in which we mimicked pulmonary concentration-time profiles achieved in patients. The HFS-MAC model was sampled at intervals to determine the minocycline pharmacokinetics and MAC burden. We identified the AUC0-24/MIC ratios associated with 1.0 log10 cfu/mL kill below day 0 (stasis), defined as a bactericidal effect. We then performed 10000 Monte Carlo experiments to identify the optimal dose for a bactericidal effect in patients. RESULTS: The MIC for 50% and 90% of cumulative clinical isolates was 8 and 64 mg/L, respectively. Minocycline decreased MAC bacterial burden below stasis in primary isolated macrophages. In the HFS-MAC model, minocycline achieved a microbial kill of 3.6 log10 cfu/mL below stasis. The AUC0-24/MIC exposure associated with a bactericidal effect was 59. Monte Carlo experiments identified a minocycline susceptibility MIC breakpoint of 16 mg/L. At this proposed breakpoint, the clinical dose of 200 mg/day achieved the bactericidal effect exposure target in approximately 50% of patients, while 400 mg/day achieved this in 73.6% of patients, in Monte Carlo experiments. CONCLUSIONS: Minocycline at a dose of 400 mg/day is expected to be bactericidal. We propose a clinical trial for validation.

Treatment of disseminated tuberculosis in children ≤ 6 years has not been optimized. The pyrazinamide-containing combination regimen used to treat disseminated tuberculosis in babies and toddlers was extrapolated from adult pulmonary tuberculosis. Due to hepatotoxicity worries, there are no dose–response studies in children. We designed a hollow fiber system model of disseminated intracellular tuberculosis with co-perfused three-dimensional organotypic liver modules to simultaneously test for efficacy and toxicity. We utilized pediatric pharmacokinetics of pyrazinamide and acetaminophen to determine dose-dependent pyrazinamide efficacy and hepatotoxicity. Acetaminophen concentrations that cause hepatotoxicity in children led to elevated liver function tests, while 100 mg/kg pyrazinamide did not. Surprisingly, pyrazinamide did not kill intracellular Mycobacterium tuberculosis up to fourfold the standard dose as monotherapy or as combination therapy, despite achieving high intracellular concentrations. Host-pathogen RNA-sequencing revealed lack of a pyrazinamide exposure transcript signature in intracellular bacteria or of phagolysosome acidification on pH imaging. Artificial intelligence algorithms confirmed that pyrazinamide was not predictive of good clinical outcomes in children ≤ 6 years who had extrapulmonary tuberculosis. Thus, adding a drug that works inside macrophages could benefit children with disseminated tuberculosis. Our in vitro model can be used to identify such new regimens that could accelerate cure while minimizing toxicity., Highlights • We designed a pre-clinical of disseminated for simultaneous identification of toxicity and efficacy in children. • The system is a co-culture of infected monocytes and 3 dimensional organotypic liver recapitulating children pharmacokinetics. • Pyrazinamide, central drug in treatment regimen, had no effect as monotherapy or contribute to the combination therapy. Due to fear of toxicity children are often not involved in clinical trials, and as a result the optimal treatment regimens are often lacking. As an example, toddlers and babies develop disseminated tuberculosis but are treated with regimens designed for adults with lung cavity disease. We designed a “glass-mouse” model of disseminated tuberculosis that simultaneously tests for the efficacy and toxicity of the anti-tuberculosis drugs for children with disseminated disease. We found that while not causing dose-dependent liver toxicity, one of the central drugs used to treat this children is likely not efficacious.

Mycobacterium abscessus causes chronic pulmonary infections that are extremely difficult to cure. The currently recommended combination therapy is associated with high failure rates and relapse. Tigecycline has been explored in salvage regimens, with a response rate of 43% in those who received at least a month of therapy. We performed a dose-response study in a hollow-fiber system model of pulmonary M. abscessus infection in which we recapitulated tigecycline human pulmonary concentration-time profiles of 8 different doses for 21 days. We identified the maximal kill or efficacy in CFU per milliliter and the ratio of the 0- to 24-h area under the concentration-time curve to MIC (AUC/MIC) associated with 80% efficacy (EC 80 ). The tigecycline efficacy was 5.38 ± 2.35 log 10 CFU/ml, and the drug achieved the unprecedented feat of a bacterial level of 1.0 log 10 CFU/ml below the pretreatment inoculum (1-log kill) of M. abscessus in the hollow-fiber system. The EC 80 AUC/MIC ratio was 36.65, while that for a 1-log kill was 44.6. Monte Carlo experiments with 10,000 patients were used to identify the clinical dose best able to achieve the EC 80 or 1-log kill. The standard dose of 100 mg/day had a cumulative fraction of response of 51% for the EC 80 and 46% for 1-log kill. For both the EC 80 target and 1-log kill, the optimal tigecycline clinical dose was identified as 200 mg/day. The susceptibility breakpoint was ≤0.5 mg/liter. Tigecycline is the most active single agent evaluated to date, and we propose that 200 mg/day be examined as the backbone of new combination therapy regimens to replace current treatment.

The modern chemotherapy era started with Fleming's discovery of benzylpenicillin. He demonstrated that benzylpenicillin did not kill Mycobacterium tuberculosis . In this study, we found that >64 mg/liter of static benzylpenicillin concentrations killed 1.16 to 1.43 log 10 CFU/ml below starting inoculum of extracellular and intracellular M. tuberculosis over 7 days. When we added the β-lactamase inhibitor avibactam, benzylpenicillin maximal kill ( E max ) of extracellular log-phase-growth M. tuberculosis was 6.80 ± 0.45 log 10 CFU/ml at a 50% effective concentration (EC 50 ) of 15.11 ± 2.31 mg/liter, while for intracellular M. tuberculosis it was 2.42 ± 0.14 log 10 CFU/ml at an EC 50 of 6.70 ± 0.56 mg/liter. The median penicillin (plus avibactam) MIC against South African clinical M. tuberculosis strains (80% either multidrug or extensively drug resistant) was 2 mg/liter. We mimicked human-like benzylpenicillin and avibactam concentration-time profiles in the hollow-fiber model of tuberculosis (HFS-TB). The percent time above the MIC was linked to effect, with an optimal exposure of ≥65%. At optimal exposure in the HFS-TB, the bactericidal activity in log-phase-growth M. tuberculosis was 1.44 log 10 CFU/ml/day, while 3.28 log 10 CFU/ml of intracellular M. tuberculosis was killed over 3 weeks. In an 8-week HFS-TB study of nonreplicating persistent M. tuberculosis , penicillin-avibactam alone and the drug combination of isoniazid, rifampin, and pyrazinamide both killed >7.0 log 10 CFU/ml. Monte Carlo simulations of 10,000 preterm infants with disseminated disease identified an optimal dose of 10,000 U/kg (of body weight)/h, while for pregnant women or nonpregnant adults with pulmonary tuberculosis the optimal dose was 25,000 U/kg/h, by continuous intravenous infusion. Penicillin-avibactam should be examined for effect in pregnant women and infants with drug-resistant tuberculosis, to replace injectable ototoxic and teratogenic second-line drugs.

In pharmacokinetic/pharmacodynamic models of pulmonary Mycobacterium abscessus complex, the recommended macrolide-containing combination therapy has poor kill rates. However, clinical outcomes are unknown. We searched the literature for studies published between 1990 and 2017 that reported microbial outcomes in patients treated for pulmonary M. abscessus disease. A good outcome was defined as sustained sputum culture conversion (SSCC) without relapse. Random effects models were used to pool studies and estimate proportions of patients with good outcomes. Odds ratios (OR) and 95% confidence intervals (CI) were computed. Sensitivity analyses and metaregression were used to assess the robustness of findings. In 19 studies of 1,533 patients, combination therapy was administered to 508 patients with M. abscessus subsp. abscessus , 204 with M. abscessus subsp. massiliense , and 301 with M. abscessus with no subspecies specified. Macrolide-containing regimens achieved SSCC in only 77/233 (34%) new M. abscessus subsp. abscessus patients versus 117/141 (54%) M. abscessus subsp. massiliense patients (OR, 0.108 [95% CI, 0.066 to 0.181]). In refractory disease, SSCC was achieved in 20% (95% CI, 7 to 36%) of patients, which was not significantly different across subspecies. The estimated recurrent rates per month were 1.835% (range, 1.667 to 3.196%) for M. abscessus subsp. abscessus versus 0.683% (range, 0.229 to 1.136%) for M. abscessus subsp. massiliense (OR, 6.189 [95% CI, 2.896 to 13.650]). The proportion of patients with good outcomes was 52/223 (23%) with M. abscessus subsp. abscessus versus 118/141 (84%) with M. abscessus subsp. massiliense disease (OR, 0.059 [95% CI, 0.034 to 0.101]). M. abscessus subsp. abscessus pulmonary disease outcomes with the currently recommended regimens are atrocious, with outcomes similar to those for extensively drug-resistant tuberculosis. Therapeutically, the concept of nontuberculous mycobacteria is misguided. There is an urgent need to craft entirely new treatment regimens.

Objectives To determine if tedizolid is effective for pulmonary Mycobacterium avium complex (MAC) disease, and to use pharmacokinetics/pharmacodynamics to design optimal doses. Methods We performed an exposure-response experiment in the hollow-fibre system model of intracellular MAC (HFS-MAC). We mimicked the tedizolid concentration-time profiles achieved in the lungs of patients treated once daily for 28 days. The HFS-MAC was sampled at intervals to determine the tedizolid pharmacokinetics and MAC intracellular burden. We identified the 0-24 h area under the concentration-time curves to MIC (AUC0-24/MIC) ratios associated with the following targets: 80% of maximal kill (EC80), bacteriostasis, and 1.0 and 2.0 log10 cfu/mL kill. We then performed 10 000 patient Monte Carlo simulations to identify the optimal dose for each of the exposure targets. Results Tedizolid achieved the feat of 2.0 log10 cfu/mL kill below initial bacterial burden, an effect not seen before in this model with other antibiotics. The tedizolid exposure associated with 1.0 log10 cfu/mL kill was a non-protein bound AUC0-24/MIC ratio of 23.46, while that associated with 2.0 log10 cfu/mL kill was 37.50, and the EC80 was 21.71. The clinical dose of 200 mg achieved each of these targets in ∼100% of the 10 000 patients, except the 2.0 log10 cfu/mL kill which required 300 mg/day. A tedizolid susceptibility MIC breakpoint of 1 mg/L is proposed. Conclusions Tedizolid, at standard clinical doses, is expected to be bactericidal, and even achieved an unprecedented 2.0 log10 cfu/mL kill of MAC as monotherapy. We propose it as the backbone of short-course anti-MAC chemotherapy.

Objectives To identify the pharmacokinetic/pharmacodynamic parameters and exposures of linezolid in the treatment of pulmonary Mycobacterium avium complex (MAC) disease. Methods Human-derived monocytes infected with MAC were inoculated into hollow-fibre systems for dose-effect and dose-scheduling studies. We mimicked linezolid concentration-time profiles achieved in adult human lungs treated for 28 days. Sampling to confirm that the intended linezolid pharmacokinetics had been achieved, and for enumeration of MAC colony-forming units, was performed based on repetitive sampling from each system over the 28 days. We then performed 10 000 patient Monte Carlo simulations to identify doses associated with optimal effect in the clinic. Results Linezolid achieved a hitherto unprecedented feat of at least 1.0 log10 cfu/mL reduction. Efficacy was most closely linked to the AUC0-24/MIC ratio. The AUC0-24/MIC ratio associated with no change in bacterial burden or bacteriostasis was 7.82, while that associated with 1.0 log10 cfu/mL kill was 42.06. The clinical dose of 600 mg/day achieved or exceeded the bacteriostasis exposure in 98.73% of patients. The proportion of 10 000 patients treated with the standard 1200 mg/day who achieved the exposure for 1.0 log10 cfu/mL kill was 70.64%, but was 90% for 1800 mg/day. The proposed MIC breakpoint for linezolid is 16 mg/L, with which 49%-80% of clinical isolates would be considered resistant. Conclusions Linezolid is associated with a bactericidal effect in pulmonary MAC that is greater than that seen with other recommended drugs. However, because of the MIC distribution, doses that would optimize the bactericidal effect would be associated with a high adverse event rate.

Objectives Pulmonary Mycobacterium avium complex (MAC) prevalence is on the rise worldwide. The average therapy duration is 1.5 years, which is associated with poor cure rates. Our objective was to develop a programme to design a combination therapy regimen for pulmonary MAC to be administered for 6 months or less with efficacy in > 90% of patients. Methods We performed a literature search for the following MeSH headings 'Mycobacterium avium' AND 'pharmacokinetics/pharmacodynamics' in PubMed up to 2016. The findings were then used to identify steps in the programme to design new regimens with faster microbial kill rates than the current standard regimen. Results First, we designed a strategy for rapid in vitro screening of all antibiotic classes for repurposing against pulmonary MAC. Secondly, we identified and compared maximal microbial kill rates (Emax), and optimal exposures of eight different antibiotics. These studies had all been performed in the hollow-fibre system model of pulmonary MAC (HFS-MAC). Thirdly, all drugs with a high Emax at clinically achievable optimal exposures will be chosen, and exposures associated with synergy or additivity for two/three drugs identified based on Bliss independence. Fourthly, the time-kill slopes and resistance suppression of the chosen combinations will be compared with those of standard combination therapy in the HFS-MAC. Finally, we will identify the clinical doses best able to achieve synergistic or additive combination exposures by taking into account pharmacokinetic variability. Conclusions Our stepwise pharmacokinetics/pharmacodynamics approach provides a scientific rationale and a strategy for achieving short-course chemotherapy for pulmonary MAC disease within a few years.

Objectives To develop a thioridazine/moxifloxacin-based combination regimen for treatment of pulmonary infection due to Mycobacterium avium-intracellulare complex (MAC) that kills bacteria faster than the standard treatment regimen. Methods Monocytes were infected with MAC and inoculated into the hollow-fibre system model for pulmonary MAC disease (HFS-MAC). We co-administered ethambutol plus azithromycin daily for 28 days, to achieve the same human concentration-time profiles that result from standard doses, in three HFS-MAC systems. Two experimental regimens consisted of thioridazine at an exposure associated with optimal kill, given intermittently on days 0, 3, 7 and 10. Regimen A consisted of thioridazine in combination with standard dose azithromycin for the entire study duration. Regimen B was thioridazine plus moxifloxacin at concentration-time profiles achieved by the standard daily dose administered for 14 days, followed by daily azithromycin. Each HFS-MAC was sampled for bacterial burden every 7 days. Results The bacteria in the non-treated HFS-MAC grew at a rate of 0.11 ± 0.01 log10 cfu/mL/day. The azithromycin/ethambutol regimen decreased bacterial burden by 1.21 ± 0.74 log10 cfu/mL below baseline during the first 7 days, after which it failed. Regimen A killed 3.28 ± 0.32 log10 cfu/mL below baseline up to day 14, after which regrowth occurred once thioridazine treatment stopped. Regimen B killed bacteria to below the limits of detection in 7 days (≥5.0 log10 cfu/mL kill), with rebound in the azithromycin continuation phase. Conclusions The thioridazine/moxifloxacin regimen demonstrated that rapid microbial kill could be achieved within 7 days. This is a proof of principle that short-course chemotherapy for pulmonary MAC is possible.

Ceftazidime-avibactam is highly efficacious against extensive- and multidrug-resistant strains of Mycobacterium tuberculosis., There are currently many patients with multidrug-resistant and extensively drug-resistant tuberculosis. Ongoing transmission of the highly drug-resistant strains and high mortality despite treatment remain problematic. The current strategy of drug discovery and development takes up to a decade to bring a new drug to clinical use. We embarked on a strategy to screen all antibiotics in current use and examined them for use in tuberculosis. We found that ceftazidime-avibactam, which is already used in the clinic for multidrug-resistant Gram-negative bacillary infections, markedly killed rapidly growing, intracellular, and semidormant Mycobacterium tuberculosis in the hollow fiber system model. Moreover, multidrug-resistant and extensively drug-resistant clinical isolates demonstrated good ceftazidime-avibactam susceptibility profiles and were inhibited by clinically achievable concentrations. Resistance arose because of mutations in the transpeptidase domain of the penicillin-binding protein PonA1, suggesting that the drug kills M. tuberculosis bacilli via interference with cell wall remodeling. We identified concentrations (exposure targets) for optimal effect in tuberculosis, which we used with susceptibility results in computer-aided clinical trial simulations to identify doses for immediate clinical use as salvage therapy for adults and young children. Moreover, this work provides a roadmap for efficient and timely evaluation of antibiotics and optimization of clinically relevant dosing regimens.

Linezolid has an excellent sterilizing effect in tuberculosis patients but high adverse event rates. The dose that would maximize efficacy and minimize toxicity is unknown. We performed linezolid dose-effect and dose-scheduling studies in the hollow fiber system model of tuberculosis (HFS-TB) for sterilizing effect. HFS-TB units were treated with several doses to mimic human-like linezolid intrapulmonary pharmacokinetics and repetitively sampled for drug concentration, total bacterial burden, linezolid-resistant subpopulations, and RNA sequencing over 2 months. Linezolid-resistant isolates underwent whole-genome sequencing. The expression of genes encoding efflux pumps in the first 1 to 2 weeks revealed the same exposure-response patterns as the linezolid-resistant subpopulation. Linezolid-resistant isolates from the 2nd month of therapy revealed mutations in several efflux pump/transporter genes and a LuxR-family transcriptional regulator. Linezolid sterilizing effect was linked to the ratio of unbound 0- to 24-h area under the concentration-time curve (AUC 0–24 ) to MIC. Optimal microbial kill was achieved at an AUC 0–24 /MIC ratio of 119. The optimal sterilizing effect dose for clinical use was identified using Monte Carlo simulations. Clinical doses of 300 and 600 mg/day (or double the dose every other day) achieved this target in 87% and >99% of 10,000 patients, respectively. The susceptibility breakpoint identified was 2 mg/liter. The simulations identified that a 300-mg/day dose did not achieve AUC 0–24 s associated with linezolid toxicity, while 600 mg/day achieved those AUC 0–24 s in

Background. The regimen of linezolid and moxifloxacin was found to be efficacious in the hollow fiber system model of pediatric intracellular tuberculosis. However, its kill rate was slower than the standard 3-drug regimen of isoniazid, rifampin, and pyrazinamide. We wanted to examine the effect of adding a third oral agent, faropenem, to this dual combination. Methods. We performed a series of studies in the hollow fiber system model of intracellular Mycobacterium tuberculosis, by mimicking pediatric pharmacokinetics of each antibiotic. First, we varied the percentage of time that faropenem persisted above minimum inhibitory concentration (TMIC) on the moxifloxacin-linezolid regimen. After choosing the best faropenem exposure, we performed experiments in which we varied the moxifloxacin and linezolid doses in the triple regimen. Finally, we performed longer-duration therapy validation experiments. Bacterial burden was quantified using both colony-forming units per milliliter (CFU/mL) and time to positivity (TTP). Kill slopes were modeled using exponential regression. Results. TTP was a more sensitive measure of bacterial burden than CFU/mL. A faropenem TMIC > 62% was associated with steepest microbial kill slope. Regimens of standard linezolid and moxifloxacin plus faropenem TMIC > 60%, as well as higher-dose moxifloxacin, achieved slopes equivalent to those of the standard regimen based by both TTP and CFU/mL over 28 days of treatment. Conclusions. We have developed an oral faropenem-linezolid-moxifloxacin (FLAME) regimen that is free of first-line drugs. The regimen could be effective against both multidrug-resistant and drug-susceptible tuberculosis in children.

Background. When treated with the same antibiotic dose, children achieve different 0- to 24-hour area under the concentration-time curves (AUC0–24) because of maturation and between-child physiological variability on drug clearance. Children are also infected by Mycobacterium tuberculosis isolates with different antibiotic minimum inhibitory concentrations (MICs). Thus, each child will achieve different AUC0–24/MIC ratios when treated with the same dose. Methods. We used 10 000-subject Monte Carlo experiments to identify the oral doses of linezolid, moxifloxacin, and faropenem that would achieve optimal target exposures associated with optimal efficacy in children with disseminated tuberculosis. The linezolid and moxifloxacin exposure targets were AUC0–24/MIC ratios of 62 and 122, and a faropenem percentage of time above MIC >60%, in combination therapy. A linezolid AUC0–24 of 93.4 mg × hour/L was target for toxicity. Population pharmacokinetic parameters of each drug and between-child variability, as well as MIC distribution, were used, and the cumulative fraction of response (CFR) was calculated. We also considered drug penetration indices into meninges, bone, and peritoneum. Results. The linezolid dose of 15 mg/kg in full-term neonates and infants aged up to 3 months and 10 mg/kg in toddlers, administered once daily, achieved CFR ≥ 90%, with 90% in infants, but the optimal dose was 20 mg/kg/day in older children. The faropenem medoxomil optimal dosage was 30 mg/kg 3–4 times daily. Conclusions. The regimen and doses of linezolid, moxifloxacin, and faropenem identified are proposed to be adequate for all disseminated tuberculosis syndromes, whether drug-resistant or -susceptible.

Background. No treatment regimens have been specifically designed for children, in whom tuberculosis is predominantly intracellular. Given their activity as monotherapy and their ability to penetrate many diseased anatomic sites that characterize disseminated tuberculosis, linezolid and moxifloxacin could be combined to form a regimen for this need. Methods. We examined microbial kill of intracellular Mycobacterium tuberculosis (Mtb) by the combination of linezolid and moxifloxacin multiple exposures in a 7-by-7 mathematical matrix. We then used the hollow fiber system (HFS) model of intracellular tuberculosis to identify optimal dose schedules and exposures of moxifloxacin and linezolid in combination. We mimicked pediatric half-lives and concentrations achieved by each drug. We sampled the peripheral compartment on days 0, 7, 14, 21, and 28 for Mtb quantification, and compared the slope of microbial kill of Mtb by these regimens to the standard regimen of isoniazid, rifampin, and pyrazinamide, based on exponential decline regression. Results. The full exposure-response surface identified linezolid-moxifloxacin zones of synergy, antagonism, and additivity. A regimen based on each of these zones was then used in the HFS model, with observed half-lives of 4.08 ± 0.66 for linezolid and 3.80 ± 1.34 hours for moxifloxacin. The kill rate constant was 0.060 ± 0.012 per day with the moxifloxacin-linezolid regimen in the additivity zone vs 0.083 ± 0.011 per day with standard therapy, translating to a bacterial burden half-life of 11.52 days vs 8.53 days, respectively. Conclusions. We identified doses and dose schedules of a linezolid and moxifloxacin backbone regimen that could be highly efficacious in disseminated tuberculosis in children.

Ethambutol, together with a macrolide, is the backbone for treatment of disseminated Mycobacterium avium disease. However, at the standard dose of 15 mg/kg of body weight/day, ethambutol efficacy is limited. In addition, susceptibility breakpoints have consistently failed to predict clinical outcome. We performed dose-effect studies with extracellular M. avium as well as with bacilli within human macrophages. The maximal kill rate ( E max ) for ethambutol against extracellular bacilli was 5.54 log 10 CFU/ml, compared to 0.67 log 10 CFU/ml for intracellular M. avium , after 7 days of exposure. Thus, extracellular assays demonstrated high efficacy. We created a hollow-fiber system model of intracellular M. avium and performed microbial pharmacokinetic-pharmacodynamic studies using pharmacokinetics similar to those of ethambutol for humans. The E max in the systems was 0.79 log 10 CFU/ml with 7 days of daily therapy, so the kill rates approximated those encountered in patients treated with ethambutol monotherapy. Ratio of peak concentration to MIC ( C max /MIC) was linked to microbial kill rate. The C max /MIC ratio needed to achieve the 90% effective concentration (EC 90 ) in serum was 1.23, with a calculated intramacrophage C max /MIC ratio of 13. In 10,000 patient Monte Carlo simulations, doses of 15, 50, and 75 mg/kg achieved the EC 90 in 35.50%, 76.81%, and 86.12% of patients, respectively. Therefore, ethambutol doses of ≥50 mg/kg twice a week would be predicted to be better than current doses of 15 mg/kg for treatment of disseminated M. avium disease. New susceptibility breakpoints and critical concentrations of 1 to 2 mg/liter were identified for the determination of ethambutol-resistant M. avium in Middlebrook broth. Given that the modal MIC of clinical isolates is around 2 mg/liter, most isolates should be considered ethambutol resistant.

Current regimens used to treat pulmonary Mycobacterium abscessus disease have limited efficacy. There is an urgent need for new drugs and optimized combinations and doses. We performed hollow-fiber-system studies in which M. abscessus was exposed to moxifloxacin lung concentration-time profiles similar to human doses of between 0 and 800 mg/day. The minimum bactericidal concentration and MIC were 8 and 2 mg/liter, respectively, in our M. abscessus strain, suggesting bactericidal activity. Measurement of the moxifloxacin concentrations in each hollow-fiber system revealed an elimination rate constant ( k el ) of 0.11 ± 0.05 h −1 (mean ± standard deviation) (half-life of 9.8 h). Inhibitory sigmoid maximal effect ( E max ) modeling revealed that the highest E max was 3.15 ± 1.84 log 10 CFU/ml on day 3, and the exposure mediating 50% of E max (EC 50 ) was a 0- to 24-h area under the concentration time curve (AUC 0–24 )-to-MIC ratio of 41.99 ± 31.78 ( r 2 = 0.99). The EC 80 was an AUC 0–24 /MIC ratio of 102.11. However, no moxifloxacin concentration killed the bacteria to burdens below the starting inoculum. There was regrowth beyond day 3 in all doses, with replacement by a resistant subpopulation that had an MIC of >32 mg/liter by the end of the experiment. A quadratic function best described the relationship between the AUC 0–24 /MIC ratio and the moxifloxacin-resistant subpopulation. Monte Carlo simulations of 10,000 patients revealed that the 400- to 800-mg/day doses would achieve or exceed the EC 80 in ≤12.5% of patients. The moxifloxacin susceptibility breakpoint was 0.25 mg/liter, which means that almost all M. abscessus clinical strains are moxifloxacin resistant by these criteria. While moxifloxacin's efficacy against M. abscessus was poor, formal combination therapy studies with moxifloxacin are still recommended.

The treatment of pulmonary Mycobacterium abscessus disease is associated with very high failure rates and easily acquired drug resistance. Amikacin is the key drug in treatment regimens, but the optimal doses are unknown. No good preclinical model exists to perform formal pharmacokinetics/pharmacodynamics experiments to determine these optimal doses. We developed a hollow-fiber system model of M. abscessus disease and studied amikacin exposure effects and dose scheduling. We mimicked amikacin human pulmonary pharmacokinetics. Both amikacin microbial kill and acquired drug resistance were linked to the peak concentration-to-MIC ratios; the peak/MIC ratio associated with 80% of maximal kill (EC 80 ) was 3.20. However, on the day of the most extensive microbial kill, the bacillary burden did not fall below the starting inoculum. We performed Monte Carlo simulations of 10,000 patients with pulmonary M. abscessus infection and examined the probability that patients treated with one of 6 doses from 750 mg to 4,000 mg would achieve or exceed the EC 80 . We also examined these doses for the ability to achieve a cumulative area under the concentration-time curve of 82,232 mg · h/liter × days, which is associated with ototoxicity. The standard amikacin doses of 750 to 1,500 mg a day achieved the EC 80 in ≤21% of the patients, while a dose of 4 g/day achieved this in 70% of the patients but at the cost of high rates of ototoxicity within a month or two. The susceptibility breakpoint was an MIC of 8 to 16 mg/liter. Thus, amikacin, as currently dosed, has limited efficacy against M. abscessus . It is urgent that different antibiotics be tested using our preclinical model and new regimens developed.

Disseminated Mycobacterium avium complex (MAC) is treated with a macrolide and ethambutol. However, the kill rates are extremely slow so that therapy takes many months to years to achieve and even then more than 40% of patients are not completely cured. Recent studies have demonstrated that assays that detect extracellular MAC have a limited predictive value. Antibiotics kill at a much slower and more disappointing rate against bacilli within macrophages. Use of pharmacodynamic/pharmacokinetic models has resulted in design of new doses and dosing schedules for disseminated MAC, as well as new susceptibility breakpoints for ethambutol and moxifloxacin.