Hollow Fiber System Research Articles

A hollow fiber system for simple generation of human brain organoids.

3D organoids exhibit near-physiological morphogenesis and histology relying on the self-organization of human pluripotent stem cells (hPSCs), representing a new class of in vitro model for studying developmental biology and diseases. An engineered approach is highly desirable to generate sufficient organoids in a simple and efficient manner. Herein, we present a new strategy for the simple formation of massive human brain organoids from hiPSCs within a hollow fiber reactor system by combining fiber materials with the developmental biology principle. A thin and finely adjustable calcium alginate (CaA) core-shell fiber was constructed using a multilayer coaxial laminar flow microfluidic system. The meter-long hollow fibers enabled neural differentiation of hiPSCs and simple formation of abundant brain organoids in a 3D matrix. The generated brain organoids displayed essential features of human brain organogenesis, including polarized neuroepithelium, cell type heterogeneity and discrete brain regions, resembling the early brain development. This approach is simple and easy to operate, which allows for simplified formation of massive brain organoids, overcoming the tedious procedures in conventional methods. In particular, the facile and scalable characteristics of hollow fibers are compatible with real-time observation and monitoring, as well as flexible tissue manipulations for downstream biological analysis. It might also provide a new platform to advance stem cell-derived organoid models and their utility in biomedical applications.

Hollow Fiber System Biomanufacturing

Hollow Fiber System Biomanufacturing

Concentration-Dependent Synergy and Antagonism of Linezolid and Moxifloxacin in the Treatment of Childhood Tuberculosis: The Dynamic Duo.

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.

A Faropenem, Linezolid, and Moxifloxacin Regimen for Both Drug-Susceptible and Multidrug-Resistant Tuberculosis in Children: FLAME Path on the Milky Way.

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.

Linezolid for Infants and Toddlers With Disseminated Tuberculosis: First Steps.

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.

Amikacin Optimal Exposure Targets in the Hollow-Fiber System Model of Tuberculosis.

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 (Emax) 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) log10 CFU/ml, while the concentration mediating 50% of Emax (EC50) 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 (Cmax)-to-MIC ratio (r2 > 0.99), closely followed by the area under the concentration-time curve from 0 to 24 h (AUC0–24)-to-MIC ratio (r2 = 0.98). The EC90 was a Cmax/MIC ratio of 10.13 (95% CI, 7.73 to 12.48). The EC90 is the dosing target for intermittent therapy that optimizes cure in TB programs for MDR-TB patients.

Artificial Intelligence and Amikacin Exposures Predictive of Outcomes in Multidrug-Resistant Tuberculosis Patients.

Aminoglycosides such as amikacin continue to be part of the backbone of treatment of multidrug-resistant tuberculosis (MDR-TB). We measured amikacin concentrations in 28 MDR-TB patients in Botswana receiving amikacin therapy together with oral levofloxacin, ethionamide, cycloserine, and pyrazinamide and calculated areas under the concentration-time curves from 0 to 24 h (AUC0–24). The patients were followed monthly for sputum culture conversion based on liquid cultures. The median duration of amikacin therapy was 184 (range, 28 to 866) days, at a median dose of 17.30 (range 11.11 to 19.23) mg/kg. Only 11 (39%) patients had sputum culture conversion during treatment; the rest failed. We utilized classification and regression tree analyses (CART) to examine all potential predictors of failure, including clinical and demographic features, comorbidities, and amikacin peak concentrations (Cmax), AUC0–24, and trough concentrations. The primary node for failure had two competing variables, Cmax of <67 mg/liter and AUC0–24 of <568.30 mg · h/L; weight of >41 kg was a secondary node with a score of 35% relative to the primary node. The area under the receiver operating characteristic curve for the CART model was an R2 = 0.90 on posttest. In patients weighing >41 kg, sputum conversion was 3/3 (100%) in those with an amikacin Cmax of ≥67 mg/liter versus 3/15 (20%) in those with a Cmax of <67 mg/liter (relative risk [RR] = 5.00; 95% confidence interval [CI], 1.82 to 13.76). In all patients who had both amikacin Cmax and AUC0–24 below the threshold, 7/7 (100%) failed, compared to 7/15 (47%) of those who had these parameters above threshold (RR = 2.14; 95% CI, 1.25 to 43.68). These amikacin dose-schedule patterns and exposures are virtually the same as those identified in the hollow-fiber system model.

308. Ultracentrifugation-Free Chromatography-Mediated Large-Scale Purification of Recombinant Adeno-Associated Virus Serotype 1 (rAAV1) and rAAV9 from the Serum-Free Culture Supernatant

[Background]The current production of rAAV from the transfected cell lysate and purification based on CsCl or iodixanol density ultracentrifugation are not suitable for large-scale processing. Although rAAV1 and rAAV9 are promising therapeutic vectors for genetic neuromuscular disorders, the large-scale purification method for those vectors has not yet been established. In this study, we elaborate the novel chromatography-mediated methods for purification of rAAV1 and rAAV9 from the serum-free culture supernatant with ultracentrifugation-free technique towards large-scale and GMP production.[Methods]rAAV1 (scAAV1-CBA-EGFP) and rAAV9 (scAAV9-CBA-EGFP) were produced by the triple-transfection to HEK293 or HEK293EB cells in serum-free medium with polyethyleneimine (PEI). Five days later, the culture supernatant was tangential flow-filtrated (TFF) and concentrated by the Hollow Fiber system. After reducing protein debris by 1/3-saturated ammonium sulfate (1/3 AS) precipitation, rAAV1 or rAAV9 was precipitated in 1/2 AS solution.Subsequently, the precipitated rAAV1 was dissolved in 10 ml of PBS containing 3 mM MgCl2. After the sample was dialyzed against a buffer containing 5 mM NaCl, the dialysate was diluted with dH2O and loaded to tandem quintet cation-exchange membranes (Mustang SXT) and quintet anion-exchange membranes (Mustang QXT) with a column volume of 0.86 ml. After rAAV1 was eluted from Mustang QXT, it was finally purified by gel-filtrated chromatography (Superdex 200 10/300 GL).The precipitated rAAV9 was dissolved in 22 ml of 3.3 mM MES, 3.3 mM HEPES, 3.3 mM sodium acetate (MHN) buffer (pH8.0) containing 50 mM NaCl and 0.01% Pluronic F-68. After the resultant sample was diluted with MHN buffer, it was loaded to tertiary amine charged anion-exchange column. The passed through fraction containing rAAV9 was finally purified by gel-filtrated chromatography.The physiological and biological properties of the purified rAAV1 and rAAV9 were characterized by qPCR, electron micrograph, FACS, western blot and SDS-PAGE.[Results]The purified rAAV1 and rAAV9 displayed three major bands (VP1, VP2, and VP3) on SDS-PAGE and more than 90% of rAAV1 particles or 95% of rAAV9 particles was contained fully packaged viral genomes according to electron micrographic analysis. We confirmed increasing infectivity of rAAV1 products depending on chromatographic purification step and that of rAAV9 is under evaluation. The resultant genomic titer of the purified rAAV1 was 4.17 × 1013 v.g. (3.63 × 1013 v.g./ml) from the 4 × 109 HEK293 cells and the purified rAAV9 was 1.49 × 1015 v.g. (1.14 × 1014 v.g./ml) from the 5.1 × 109 HEK293EB cells.[Conclusion]Chromatography-mediated purification from the culture supernatant is a major breakthrough, especially for the production of rAAV9. We obtained rAAV1 and rAAV9 by this protocol with high titer, high purity and minimum contamination of empty particles. This novel chromatography-based method would be consistent with GMP production and facilitate clinical studies in the future.

Moxifloxacin's Limited Efficacy in the Hollow-Fiber Model of Mycobacterium abscessus Disease.

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 (kel) of 0.11 ± 0.05 h(-1) (mean ± standard deviation) (half-life of 9.8 h). Inhibitory sigmoid maximal effect (Emax) modeling revealed that the highest Emax was 3.15 ± 1.84 log10 CFU/ml on day 3, and the exposure mediating 50% of Emax (EC50) was a 0- to 24-h area under the concentration time curve (AUC0-24)-to-MIC ratio of 41.99 ± 31.78 (r(2) = 0.99). The EC80 was an AUC0-24/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 AUC0-24/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 EC80 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.

Tigecycline Is Highly Efficacious against Mycobacterium abscessus Pulmonary Disease.

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 (EC80). The tigecycline efficacy was 5.38 ± 2.35 log10 CFU/ml, and the drug achieved the unprecedented feat of a bacterial level of 1.0 log10 CFU/ml below the pretreatment inoculum (1-log kill) of M. abscessus in the hollow-fiber system. The EC80 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 EC80 or 1-log kill. The standard dose of 100 mg/day had a cumulative fraction of response of 51% for the EC80 and 46% for 1-log kill. For both the EC80 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.

A Long-term Co-perfused Disseminated Tuberculosis-3D Liver Hollow Fiber Model for Both Drug Efficacy and Hepatotoxicity in Babies

Treatment of disseminated tuberculosis in children≤6years 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 100mg/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≤6years 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.

Hollow fiber vs. flat sheet MBR for the treatment of high strength stabilized landfill leachate

The Membrane Bioreactor (MBR) technology is increasingly becoming a prominent process in the treatment of high-strength wastewater such as leachate resulting from the decomposition of waste in landfills. This study presents a performance comparative assessment of flat sheet and hollow fiber membranes in bioreactors for the treatment of relatively stable landfill leachate with the objective of defining guidelines for pilot/full scale plants. For this purpose, a laboratory scale MBR system was constructed and operated to treat a leachate with Chemical Oxygen Demand (COD) (3900–7800mg/L), Biochemical Oxygen Demand (BOD5) (∼440–1537mg/L), Total Phosphorus (TP) (∼10–59mg/L), Phosphate (PO43−) (5–58mg/L), Total Nitrogen (TN) (1500–5200mg/L), and ammonium (NH4+) (1770–4410mg/L). Both membranes achieved comparable BOD (92.2% vs. 93.2%) and TP (79.4% vs. 78.5%) removals. Higher PO43− removal efficiency or percentage (87.3% vs. 81.3%) and slightly higher, but not statistically significant, COD removal efficiency were obtained with the hollow fiber membrane (71.4% vs. 68.5%). On the other hand, the flat sheet membrane achieved significantly higher TN and NH4+ removal efficiencies (61.2% vs. 49.4% and 63.4% vs. 47.8%, respectively), which may be attributed to the less frequent addition of NaOCl compared to the hollow fiber system.

Amikacin Pharmacokinetics/Pharmacodynamics in a Novel Hollow-Fiber Mycobacterium abscessus Disease Model.

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 (EC80) 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 EC80. 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 EC80 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.

Strategic Regulatory Evaluation and Endorsement of the Hollow Fiber Tuberculosis System as a Novel Drug Development Tool.

The first nonclinical drug development tool (DDT) advanced by the Critical Path to TB Drug Regimens (CPTR) Initiative through a regulatory review process has been endorsed by leading global regulatory authorities. DDTs with demonstrated predictive accuracy for clinical and microbiological outcomes are needed to support decision making. Regulatory endorsement of these DDTs is critical for drug developers, as it promotes confidence in their use in Investigational New Drug and New Drug Application filings. The in vitro hollow fiber system model of tuberculosis (HFS-TB) is able to recapitulate concentration-time profiles (exposure) observed in patients for single drugs and combinations, by evaluating exposure measures for the ability to kill tuberculosis in different physiologic conditions. Monte Carlo simulations make this quantitative output useful to inform susceptibility breakpoints, dosage, and optimal combination regimens in patients, and to design nonclinical experiments in animal models. The Pre-Clinical and Clinical Sciences Working Group within CPTR executed an evidence-based evaluation of the HFS-TB for predictive accuracy. This extensive effort was enabled through the collaboration of subject matter experts representing the pharmaceutical industry, academia, product development partnerships, and regulatory authorities including the Food and Drug Administration (FDA) and the European Medicines Agency (EMA). A comprehensive analysis plan following the regulatory guidance documents for DDT qualification was developed, followed by individual discussions with the FDA and the EMA. The results from the quantitative analyses were submitted to both agencies, pursuing regulatory DDT endorsement. The EMA Qualification Opinion for the HFS-TB DDT was published 26 January 2015 (available at: http://www.ema.europa.eu/ema/index.jsp?curl=pages/regulation/document_listing/document_listing_000319.jsp).

The Hollow Fiber System Model in the Nonclinical Evaluation of Antituberculosis Drug Regimens.

The Hollow Fiber System Model in the Nonclinical Evaluation of Antituberculosis Drug Regimens.

Forecasting Accuracy of the Hollow Fiber Model of Tuberculosis for Clinical Therapeutic Outcomes.

The hollow fiber system model of tuberculosis (HFS-TB), in tandem with Monte Carlo experiments, represents a drug development tool (DDT) with the potential for use to develop tuberculosis treatment regimens. However, the predictive accuracy of the HFS-TB, or any other nonclinical DDT such as an animal model, has yet to be robustly evaluated. To avoid hindsight bias, a literature search was performed to identify clinical studies published at least 6 months after HFS-TB experiments' quantitative predictions. Steps to minimize bias and for reporting systematic reviews were applied as outlined in the Preferred Reporting Items for Systematic Reviews and Meta-Analyses. Publications were scored for quality of evidence. Accuracy was calculated using the mean absolute percentage error, then summated with weighting assigned by sample size and quality-of-evidence score. Given the lack of a gold-standard tuberculosis DDT, the forecasting accuracy of a completely unreliable tool was also calculated from 1000 simulated experiments for a random or "total guesswork" model. The quantitative forecasting accuracy (95% confidence interval [CI]) for the "total guesswork" model was 15.6% (95% CI, 8.7%-22.5%); bias was -0.1% (95% CI, -2.5% to 2.2%). Twenty clinical studies were published after HFS-TB experiments predicted optimal drug exposures and doses, susceptibility breakpoints, and optimal combination regimens. Based on these clinical studies, the predictive accuracy of the HFS-TB was 94.4% (95% CI, 84.3%-99.9%), and bias was 1.8% (95% CI, -13.7% to 6.2%). The HFS-TB model is highly accurate at forecasting optimal drug exposures, doses, and dosing schedules for use in the clinic.

Systematic Analysis of Hollow Fiber Model of Tuberculosis Experiments.

The in vitro hollow fiber system model of tuberculosis (HFS-TB), in tandem with Monte Carlo experiments, was introduced more than a decade ago. Since then, it has been used to perform a large number of tuberculosis pharmacokinetics/pharmacodynamics (PK/PD) studies that have not been subjected to systematic analysis. We performed a literature search to identify all HFS-TB experiments published between 1 January 2000 and 31 December 2012. There was no exclusion of articles by language. Bias minimization was according to Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA). Steps for reporting systematic reviews were followed. There were 22 HFS-TB studies published, of which 12 were combination therapy studies and 10 were monotherapy studies. There were 4 stand-alone Monte Carlo experiments that utilized quantitative output from the HFS-TB. All experiments reported drug pharmacokinetics, which recapitulated those encountered in humans. HFS-TB studies included log-phase growth studies under ambient air, semidormant bacteria at pH 5.8, and nonreplicating persisters at low oxygen tension of ≤ 10 parts per billion. The studies identified antibiotic exposures associated with optimal kill of Mycobacterium tuberculosis and suppression of acquired drug resistance (ADR) and informed predictions about optimal clinical doses, expected performance of standard doses and regimens in patients, and expected rates of ADR, as well as a proposal of new susceptibility breakpoints. The HFS-TB model offers the ability to perform PK/PD studies including humanlike drug exposures, to identify bactericidal and sterilizing effect rates, and to identify exposures associated with suppression of drug resistance. Because of the ability to perform repetitive sampling from the same unit over time, the HFS-TB vastly improves statistical power and facilitates the execution of time-to-event analyses and repeated event analyses, as well as dynamic system pharmacology mathematical models.

Hollow Fiber System Model for Tuberculosis: The European Medicines Agency Experience.

The in vitro hollow fiber system model has been qualified by the European Medicines Agency as a methodology for use in support of selection and development of antituberculosis regimens. More data are expected to be generated in the future to further characterize its value.

Pharmacokinetic-pharmacodynamic and dose-response relationships of antituberculosis drugs: recommendations and standards for industry and academia.

Antimicrobial pharmacokinetic-pharmacodynamic (PK/PD) science is vital to early antibiotic drug development to enable more efficient dose-effect study designs, identification of doses that may suppress drug resistance and choice of susceptibility breakpoints. Proper conduct of such studies is essential in the field of tuberculosis. We conducted an exhaustive review of literature on the hollow fiber system (HFS) model, murine model, and guinea pig model of tuberculosis as well as clinical studies to identify PK/PD studies that have been applied to antituberculosis therapy. Lessons learned are presented as recommendations and standards for both industry and academia in the field of antituberculosis drug development. PK/PD studies have been performed for both first-line and experimental antituberculosis agents. When properly designed exposure-effect and dose-fractionation studies have been performed in preclinical models, optimal drug exposures, and PK/PD parameters identified in these models have been found to be similar to clinical studies. Susceptibility breakpoints identified using these methods differed from previous concentrations in the literature but were found to be similar to those in prospective clinical studies. Preclinical PK/PD studies are essential value added in the development of antituberculosis agents. We provide 8 recommendations and standards for the proper conduct of such studies.

Nonclinical models for antituberculosis drug development: a landscape analysis.

Several nonclinical drug-development tools (DDTs) have been used for antituberculosis drug development over several decades. The role of the DDTs used for evaluating the efficacy of antituberculosis drug combinations and the gaps in the evidence base for which new tools or approaches are needed are as yet undefined. We performed a landscape analysis based on a comprehensive literature review to create evidence based guidelines. There are 3 important questions that a DDT should answer with regard to antituberculosis drugs: What combination(s) of drugs will be most effective? What dose(s) and schedule(s) of each drug should be administered? and What duration(s) of treatment will be efficacious? Four DDTs were identified as having a track record to answer these questions: in vitro susceptibility tests, the hollow fiber system model of tuberculosis, mice, and guinea pigs. No single nonclinical in vitro or animal model recapitulates all aspects of human tuberculosis. Therefore, a combination of models is recommended for drug development. Gaps identified include the need for standardization of nonclinical model experiments, evaluation of animal models with pathology more similar to that in humans, and identification of experimental quantitative output in the DDTs that correlates with sterilizing effect in humans. There is a need for formal quantitative analyses of how well DDTs forecast clinical outcomes.