Pharmacokinetics and safety/tolerability of isoniazid, rifampicin and pyrazinamide in children and adolescents treated for tuberculous meningitis.

OBJECTIVE: To assess the pharmacokinetics and safety/tolerability of isoniazid, rifampicin and pyrazinamide in children and adolescents with tuberculous meningitis (TBM). DESIGN: Prospective observational pharmacokinetic study with an exploratory pharmacokinetic/pharmacodynamic analysis. SETTING: Hasan Sadikin Hospital, Bandung, Indonesia. PATIENTS: Individuals aged 0-18 years clinically diagnosed with TBM and receiving first-line anti-tuberculosis drug dosages according to revised WHO-recommended treatment guidelines. INTERVENTIONS: Plasma and cerebrospinal fluid (CSF) concentrations of isoniazid, rifampicin and pyrazinamide were assessed on days 2 and 10 of treatment. MAIN OUTCOME MEASURES: Plasma exposures during the daily dosing interval (AUC0-24), peak plasma concentrations (Cmax) and CSF concentrations. RESULTS: Among 20 eligible patients, geometric mean AUC0-24 of isoniazid, rifampicin and pyrazinamide was 18.5, 66.9 and 315.5 hour∙mg/L on day 2; and 14.5, 71.8 and 328.4 hour∙mg/L on day 10, respectively. Large interindividual variabilities were observed in AUC0-24 and Cmax of all drugs. All patients had suboptimal rifampicin AUC0-24 for TBM treatment indication and very low rifampicin CSF concentrations. Four patients developed grade 2-3 drug-induced liver injury (DILI) within the first 4 weeks of treatment, in whom anti-tuberculosis drugs were temporarily stopped, and no DILI recurred after reintroduction of rifampicin and isoniazid. AUC0-24 of isoniazid, rifampicin and pyrazinamide along with Cmax of isoniazid and pyrazinamide on day 10 were higher in patients who developed DILI than those without DILI (p<0.05). CONCLUSION: Higher rifampicin doses are strongly warranted in treatment of children and adolescents with TBM. The association between higher plasma concentrations of isoniazid, rifampicin and pyrazinamide and the development of DILI needs confirmatory studies.

LC-MS/MS method for simultaneous quantification of the first-line anti-tuberculosis drugs and six primary metabolites in patient plasma: Implications for therapeutic drug monitoring.

The pharmacokinetic profiling of drug substances and corresponding metabolites in the biological matrix is one of the most informative tools for the treatment efficacy assessment. Therefore, to satisfy the need for comprehensive monitoring of anti-tuberculosis drugs in human plasma, a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for simultaneous quantification of first-line anti-tuberculosis drugs (ethambutol, isoniazid, pyrazinamide, and rifampicin) along with their six primary metabolites. Simple single-step protein precipitation with methanol was chosen as the most convenient sample pre-treatment method. Chromatographic separation of the ten analyte mixture was achieved within 10 minutes on a reverse-phase C8 column using mobile phase gradient mode. The multiple reaction monitoring mode (MRM) was used for analyte detection and quantification in patient samples. The chosen quantification ranges fully covered expected plasma concentrations. The method exhibited acceptable selectivity; the within- and between-run accuracy ranged from 87.2 to 113.6%, but within- and between-run precision was between 1.6 and 14.9% (at the LLOQ level CV < 20%). Although the response of the isonicotinic acid varied depending on the matrix source (CV 21.8%), validation results proved that such inconsistency does not affect the accuracy and precision of results. If stored at room temperature plasma samples should be processed within 4 h after collection, temporary storage at -20 °C up to 24 h is acceptable due to stability issues of analytes. The developed method was applied for the patient sample analysis (n = 34) receiving anti-tuberculosis treatment with the first-line drugs.

Population pharmacokinetic model of isoniazid in patients with tuberculosis in Tunisia.

AIMS: To develop a pharmacokinetic model of isoniazid (INH) concentration taking into account demographic factors and genetic variables [N-acetyltransferase 2 (NAT2) genotype], and to propose an initial INH dosage that could maximize the probability of achieving the desired INH concentration. METHODS: A retrospective analysis was undertaken of INH concentration data collected from patients with tuberculosis in Tunisia. RESULTS: In total, 118 patients were included in this study. The one-compartment model [volume of distribution (V), elimination rate (Ke)] was found to have good predictive performance. Multi-variate analysis showed that NAT2 affected both V and Ke significantly, but age, gender and weight did not. Internal validation of the final model showed correlation of 0.95 between individual predicted INH concentration 3 h after drug intake (C3) and observed C3. External validation showed that percentage mean absolute prediction error and percentage root mean squared error were 9.11% (range 0.62-35.8%) and 11.6%, respectively. Monte-Carlo simulation showed that doses of at least 225 mg/24 h and at least 450 mg/24 h attained a therapeutic concentration in >80% of patients in the NAT2 slow acetylator group and the NAT2 rapid/intermediate acetylator group, respectively. CONCLUSION: The pharmacokinetic model allowed optimization of individual dosing regimens of INH in patients with tuberculosis in Tunisia. This tool may facilitate improved efficacy of INH and prevent its toxicity in this population.

Quantification of pyrazinamide, isoniazid, acetyl-isoniazid, and rifampicin by a high-performance liquid chromatography method in human plasma from patients with tuberculosis.

The aim of this study was to establish and validate an alternative high-performance liquid chromatography method for simultaneous quantification of pyrazinamide, isoniazid, acetyl-isoniazid and rifampicin in plasma of patients under treatment for tuberculosis. The performed method was lineal (r2  > 0.99) in the range of 2.00-50.00 µg/mL for pyrazinamide, 0.50-20.00 µg/mL for both acetyl-isoniazid and isoniazid, and 1.20-25.00 µg/mL for rifampicin. Precision and trueness were demonstrated with coefficient of variation < 15% and deviations < 15%, respectively, for quality controls samples. The lower limits of quantification were 2.00, 0.50, 0.50, and 1.20 µg/mL for pyrazinamide, isoniazid, acetyl-isoniazid and rifampicin, respectively. The method was applied for the analysis of plasma from patients with tuberculosis. This method allowed ensuring reliable quantification of the target compounds and their pharmacokinetics parameters. In general, the mean values of maximum concentration of each antituberculosis drug were located within their respective reference therapeutic ranges. However, patients with sub-therapeutic plasma concentrations of isoniazid and rifampicin were detected. This is the first analytical technique that simultaneously quantifies isoniazid, acetyl-isoniazid, rifampicin, and pyrazinamide concentrations from plasma samples by high-performance liquid chromatography with ultraviolet/visible. The proposed method could be applied for therapeutic drug monitoring and pharmacokinetics studies of the four compounds throughout the treatment of tuberculosis patients.

Effect of interval between food intake and drug administration at fasting condition on the plasma concentrations of first-line anti-tuberculosis drugs in Chinese population.

We aimed to investigate the effect of interval between food intake and drug administration at fasting condition on the plasma concentrations of first-line anti- tuberculosis (TB) drugs in Chinese population. Newly diagnosed TB patients administered the anti-TB drugs under fasting conditions orally, and then had prepared breakfast at 30 minutes and 120 min after dosing, respectively. Blood sampling was also performed 120  minutes after dosing for the detection of Cmax purpose. Overall, twenty-five participants were included in our analysis. The Cmaxs of 30  minutes interval and 120  minutes interval were 21.8 ±â€Š2.0 and 19.2 ±â€Š2.0 µg/mL for rifampin, 1.6 ±â€Š0.2 and 2.1 ±â€Š0.2 µg/mL for isoniazid (INH), 1.5 ±â€Š0.1and 1.5 ±â€Š0.2 µg/mL for ethambutol (EMB), and 49.2 ±â€Š3.7 and 41.5 ±â€Š3.9 µg/mL for pyrazinamide, respectively. Statistical analysis revealed that there was no statistical difference between 2 groups. Additionally, 88.0% and 72.0% of the 25 participants at 2-hour interval group had peak concentrations less than the lower limit of the reference range for INH and EMB, respectively. The Cmaxs of INH were 0.9 ±â€Š0.4 µg/ml for rapid acetylator, which was significantly lower than those of intermediate (1.4 ±â€Š1.0 µg/mL), and slow acetylator (2.5 ±â€Š1.0 µg/mL), respectively (P < .01). In conclusion, our data demonstrate that early food intake at 30 minutes after drug administration had no significant influence on the plasma concentrations. In addition, a high proportion of patients receiving first-line anti-TB regimen fail to achieve the expected plasma drug ranges of INH and EMB (P > .05).

LC-QToF-MS method for quantification of ethambutol, isoniazid, pyrazinamide and rifampicin in human plasma and its application.

In this research, we developed and validated a liquid chromatography coupled to mass spectrometry (LC-QToF-MS) method for simultaneous quantification of the anti-tuberculosis drugs ethambutol, isoniazid, pyrazinamide and rifampicin in human plasma. Plasma samples spiked with cimetidine (internal standard) were extracted using protein precipitation with acetonitrile containing 1% formic acid. Separation was performed using a C18 column under flow gradient conditions with water and acetonitrile, both containing 5 mm ammonium formate and 0.1% formic acid. The method was validated according to the ANVISA and US Food and Drug Administration guidelines for bioanalytical method validation. The calibration curve was linear over a concentration range of 0.2-5 µg ml-1 for ethambutol, 0.2-7.5 µg ml-1 for isoniazid, 1-40 µg ml-1 for pyrazinamide and 0.25-2 µg ml-1 for rifampicin, all with adequate precision and accuracy. The method was reproducible, selective and free of carryover and matrix effects. The validated LC-QToF-MS method was successfully applied to real samples and shown to be applicable to future therapeutic and pharmacokinetic monitoring studies.

Encapsulating Rifampicin into SLNs: A Viable Option for Managing its Bioavailability Issues Upon Co-Delivery with Isoniazid.

BACKGROUND: Rifampicin is known to degrade at the acidic pH of the stomach, especially in the presence of isoniazid. Although isoniazid also degrades partially, its degradation is reversible. OBJECTIVE: Presently, we provide a proof of the fact that the simultaneous oral administration of rifampicin (RIF), upon incorporation into solid lipid nanoparticles (RIF-SLNs), with isoniazid (INH) overcomes its INH-induced degradation and improves its oral bioavailability in rats. METHODS: Solid lipid nanoparticles of RIF (RIF-SLNs) were prepared using a novel and patented method. The effect of INH was investigated on in vivo bioavailability of RIF both in its free and encapsulated (RIF-SLNs) form, after oral administration to rats. RESULTS: Cmax and AUC0-∞ of RIF increased 158 % and 125 %, respectively, upon incorporation into SLNs versus free RIF when combined with INH. The Tmax decreased from 5.67 h to 3.3 h, and the plasma concentration of RIF remained above its MIC (8 µg/ml) at all the tested time points starting with 15 min, when administered as RIF-SLNs in combination with INH. CONCLUSION: The results confirm the scope of combining RIF-SLNs with INH to overcome the bioavailability of free RIF when combined with INH, especially in fixed dose combinations.

Rapid and highly sensitive quantification of the anti-tuberculosis agents isoniazid, ethambutol, pyrazinamide, rifampicin and rifabutin in human plasma by UPLC-MS/MS.

With the increased cases of multidrug- or rifampicin-resistant tuberculosis and co-infection with HIV globally, it is difficult to achieve ideal clinical responses because of poor drug absorption and drug-drug interactions. Herein, a bioanalytical UPLC-MS/MS method was developed and validated to quantify five anti-TB agents in human plasma samples for detecting blood drug concentrations to improve therapeutic effects. To overcome the matrix effects, stable isotope labeled analogue of each analyte was used for internal standardization. A simple single-step protein precipitation by acetonitrile was employed for the sample preparation, then the analytes including rifampicin, rifabutin, pyrazinamid, ethambutol, isoniazid and their isotope labeled internal standards (ILISs) were implemented on an HILIC silica column with a gradient mode. The linear range for each analyte was covering the peak drug concentration (Cmax) in the 20 times diluted plasma samples. The coefficient of variation of intra- and inter-day precision was less than 17.0 %, and the accuracy ranged between 91.5 and 110.0 %. The extraction recoveries of all agents were ≥90.2 %, and the matrix effects with internal standard-normalization for all agents were 97.1-110.0 %. The optimal blood sampling time was designed basing on the results of stability validation. This UPLC-MS/MS method with a run time of 3.5 min was successfully applied to routine therapeutic monitoring of the five anti-TB agents in patient plasma.

Population Pharmacokinetic Analysis of Isoniazid among Pulmonary Tuberculosis Patients from China.

The blood concentration of isoniazid (INH) is evidently affected by polymorphisms in N-acetyltransferase 2 (NAT2), an enzyme that is primarily responsible for the trimodal (i.e., fast, intermediate, and slow) INH elimination. The pharmacokinetic (PK) variability, driven largely by NAT2 activity, creates a challenge for the deployment of a uniform INH dosage in tuberculosis (TB) patients. Although acetylator-specific INH dosing has long been suggested, well-recognized dosages according to acetylator status remain elusive. In this study, 175 blood samples were collected from 89 pulmonary TB patients within 0.5 to 6 h after morning INH administration. According to their NAT2 genotypes, 32 (36.0%), 38 (42.7%), and 19 (21.3%) were fast, intermediate, and slow acetylators, respectively. The plasma INH concentration was detected by liquid chromatography-tandem mass spectrometry. Population pharmacokinetic (PPK) analysis was conducted using NONMEM and R software. A two-compartment model with first-order absorption and elimination well described the PK parameters of isoniazid. Body weight and acetylator status significantly affected the INH clearance rate. The dosage simulation targeting three indicators, including the well-recognized efficacy-safety indicator maximum concentration in serum (Cmax; 3 to 6 µg/ml), the reported area under the concentration-time curve from 0 h to infinity (AUC0-∞; ≥10.52 µg·h/ml), and the 2-h INH serum concentrations (≥2.19 µg/ml), was associated with the strongest early bactericidal activity. The optimal dosages targeting the different indicators varied from 700 to 900 mg/day, 500 to 600 mg/day, and 300 mg/day for the rapid, intermediate, and slow acetylators, respectively. Furthermore, a PPK model for isoniazid among Chinese tuberculosis patients was established for the first time and suggested doses of approximately 800 mg/day, 500 mg/day, and 300 mg/day for fast, intermediate, and slow acetylators, respectively, after a trade-off between efficacy and the occurrence of side effects.

Pharmacokinetics of isoniazid in children with tuberculosis-A comparative study at two doses.

AIM: To compare the pharmacokinetics of isoniazid (INH) at doses 5 and 10 mg/kg/day. METHODS: INH concentrations were estimated by high-performance liquid chromatography in 24 Indian children aged 1 to 15 years on antituberculosis therapy. Blood samples were collected at 0, 2, 4, 6, 8 hours after administration of INH. Patients were randomly given INH at 5 or 10 mg/kg/day and maximum concentrations (Cmax ) and area under the curve (AUC(0-8) ) were determined in each group. The 2-hour concentration of INH was used as Cmax for this study. RESULTS: Mean (standard deviation) Cmax was reached in 2 hours and was 2.68 ± 1.19 µg/mL in 5 mg/kg/day group and 8.86 ± 3.94 µg/mL in 10 mg/kg/day group (P < .05). The normal therapeutic range at 2-hour concentrations for INH in adults achieving good clinical response is between 3 and 5 µg/mL. Among 5 mg/kg/day, only 4 (33%) patients had INH concentrations within the 2-hour concentrations therapeutic range whereas in 10 mg/kg/day group, 11 (91%) patients achieved Cmax higher than the 2-hour concentrations therapeutic range and 1 (9%) patient had Cmax within the 2-hour concentrations therapeutic range. The mean AUC(0-8) in 5 mg/kg/day group was 10.04 ± 6.12 and 35.93 ± 25.37 µg·h/mL in 10 mg/kg/day group (P = .0001). CONCLUSION: Children on daily INH 10 mg/kg/day have higher AUC and Cmax than the required therapeutic range whereas over 65% of children with daily 5 mg/kg/day INH therapy failed to achieve the optimal therapeutic range.

Nonparametric Population Pharmacokinetic Modeling of Isoniazid in Colombian Patients With Tuberculosis.

BACKGROUND: Isoniazid (INH) is a first-line antituberculosis (TB) agent with a pharmacokinetic profile characterized by high interindividual variation; however, population pharmacokinetic studies in patients with TB are scarce. The aim was to develop a population model for INH in Colombian patients with TB suitable for predicting drug exposure and assessing the probability of target attainment of pharmacodynamic goals. METHODS: Ten hospitalized adult patients with TB undergoing INH treatment were recruited. After an 8-hour fasting, subjects took 300 mg of INH, and 10 samples were taken from 0 to 12 hours. INH was quantified by high-performance liquid chromatography-UV, and data were analyzed with the Pmetrics R package software. A Monte Carlo simulation with the model parameters was run to determine the probability of target attainment for optimal efficacy. RESULTS: The best model included 2 compartments, first-order absorption (Ka), delayed absorption (Tlag), and linear clearance (CL). Median Tlag was 0.25 hours, 5.54 hour for Ka, (Equation is included in full-text article.)for CL, (Equation is included in full-text article.)for the volume of the central compartment (Vc), 1.04 L/h for intercompartmental clearance (Q), and 788 L for the volume of the peripheral compartment (Vp). CL and Vc were allometrically scaled on basis of the normalized body weight. CONCLUSIONS: The Monte Carlo simulation indicated that 300 mg of INH per day is appropriate for Mycobacterium tuberculosis strains with minimal inhibitory concentration (MIC) up to 0.03 mg/L (target: area under the concentration-time curve/MIC >597); however, to cover strains with MIC up to 0.125 mg/L (80% of clinical isolates), a dose of 900 mg per day would be required.

Determinants of serum concentration of first-line anti-tuberculosis drugs from China.

Therapeutic drug monitoring has been employed in anti-tuberculosis (TB) drugs to assess optimal dose for maximum therapeutic effects and minimal toxicity. But the determinants of serum concentration need further evidences.In a retrospective case-control study, clinical and laboratory data were collected from 717 in-patients with TB at Xi'an Chest Hospital, China. Two hours serum concentrations of isoniazid, rifampicin, pyrazinamide as well as ethambutol were obtained and analyzed by liquid chromatography-tandem mass spectrometry.The month 2 culture conversion group had lower concentration of isoniazid, pyrazinamide, and ethambutol than month 1 group. Statistical analysis showed that serum concentrations of isoniazid, rifampicin, pyrazinamide, and ethambutol revealed a positive relationship with dose (mg/kg) (P < .001, P < .001, P < .001, and P = .003, respectively). Furthermore, isoniazid concentration was related to smoking (P = .009) and prior TB (P = .011), while rifampicin and pyrazinamide concentrations were correlated to sex (P = .004 and 0.025, respectively). Ethambutol concentration was associated with creatinine clearance (Ccr, P = .002).It is necessary to optimize drug doses using therapeutic drug monitoring while considering the following determinants: weight, smoking status, prior TB, sex, and Ccr. Furthermore, low 2 hours serum concentrations can be associated with longer culture conversion.

Effect of diabetes mellitus on TB drug concentrations in Tanzanian patients.

BACKGROUND: Diabetes mellitus (DM) is associated with poor TB treatment outcome. Previous studies examining the effect of DM on TB drug concentrations yielded conflicting results. No studies have been conducted to date in an African population. OBJECTIVES: To compare exposure to TB drugs in Tanzanian TB patients with and without DM. PATIENTS AND METHODS: A prospective pharmacokinetic study was performed among 20 diabetic and 20 non-diabetic Tanzanian TB patients during the intensive phase of TB treatment. Plasma pharmacokinetic parameters of isoniazid, rifampicin, pyrazinamide and ethambutol were compared using an independent-sample t-test on log-transformed data. Multiple linear regression analysis was performed to assess the effects of DM, gender, age, weight, HIV status and acetylator status on exposure to TB drugs. RESULTS: A trend was shown for 25% lower total exposure (AUC0-24) to rifampicin among diabetics versus non-diabetics (29.9 versus 39.9 mg·h/L, P=0.052). The AUC0-24 and peak concentration (Cmax) of isoniazid were also lower in diabetic TB patients (5.4 versus 10.6 mg·h/L, P=0.015 and 1.6 versus 2.8 mg/L, P=0.013). Pyrazinamide AUC0-24 and Cmax values were non-significantly lower among diabetics (P=0.08 and 0.09). In multivariate analyses, DM remained an independent predictor of exposure to isoniazid and rifampicin, next to acetylator status for isoniazid. CONCLUSIONS: There is a need for individualized dosing of isoniazid and rifampicin based on plasma concentration measurements (therapeutic drug monitoring) and for clinical trials on higher doses of these TB drugs in patients with TB and DM.

Rod-like Co based metal-organic framework embedded into mesoporous carbon composite modified glassy carbon electrode for effective detection of pyrazinamide and isonicotinyl hydrazide in biological samples.

Herein, we report a novel composite fabricated via embedding rod-like Co based metal-organic framework (Co-MOF-74) crystals into MC matrix for the first time. The introduction of MC astricts the size of Co-MOF-74 crystals, enlarges the pore size and improves the electrical conductivity, which lead to the good electrochemical properties of the composite. The fabricated sensor based on Co-MOF-74@MC exhibits superior electrocatalytic activity toward the reduction of pyrazinamide (PZA) and the oxidation of isonicotinyl hydrazide (INZ). Under optimized conditions, the sensor shows two linear ranges from 0.3 to 46.5 µM and 46.5-166.5 µM with a high sensitivity of 7.2 µA µM-1 cm-2 and a detection limit of 0.21 µM for the determination of PZA. The electroanalytical sensing of INZ also gives two linear ranges of 0.15-1.55 µM and 1.55-592.55 µM with a detection limit of 0.094 µM. The mechanism involved was also discussed, briefly. The sensor is assessed toward the detection of PZA and INZ in human serum and urine samples. Recovery values varied from 97.08 to 103.20% for PZA sensing and 96.67-102.90% for INZ sensing, revealing the promising practicality of sensor for PZA and INZ detection.

Anthropometric and Genetic Factors Associated With the Exposure of Rifampicin and Isoniazid in Mexican Patients With Tuberculosis.

BACKGROUND: Tuberculosis (TB) remains a critical infectious, contagious disease worldwide with high prevalence and mortality rate. The directly observed treatment short-course therapy includes rifampicin (RMP) and isoniazid (INH) for at least 6 months. The purposes of this scheme are to interrupt the transmissibility of the Mycobacterium tuberculosis complex and to avoid secondary complications. Low plasma concentrations of these anti-TB drugs have been associated with extended treatment duration, therapeutic failure, and relapse. The determination of anthropometric, genetic, and clinical variables that may affect plasma concentrations of RMP and INH might facilitate the detection of patients at increased risk of therapeutic failure. METHODS: A prospective observational study was performed in patients with TB diagnosis. A fixed-dose combined formulation was administered following clinical guidelines, and 12 venous blood samples were collected within 24 hours after dose for the quantification of plasma levels of RMP and INH by high-performance liquid chromatography-ultraviolet. The plasma concentrations versus time for each drug in each patient were assessed by a noncompartmental approach to obtain Cmax, and the area under the concentration-time curve to the last observation point (AUC0-24 h) was calculated by the linear trapezoidal rule. Genetic polymorphisms of the enzyme involved in INH metabolism (NAT2) and proteins involved in RMP transport (glycoprotein-P and OATP1B1) were determined. RESULTS: A total of 34 patients aged between 18 and 72 years with the diagnosis of TB were included in the current study. A multivariate analysis was performed to determine the anthropometric and genetic characteristics that modified the Cmax and AUC0-24 h of RMP and INH. Results indicated that RMP Cmax and AUC0-24 h were affected by sex, dose/weight, and single nucleotide polymorphism of MDR1. In addition, age, body mass index, and NAT2 acetylator genotype were shown to determine the Cmax and AUC0-24 h for INH. CONCLUSIONS: Anthropometric, genetic, and dosage characteristics of Mexican patients with TB are an important source of risk for subtherapeutic plasma concentrations of anti-TB drugs. Factors such as lower-than-recommended RMP dose, male patients with TB, and MDR1 3435 genotype, in addition to age group, body mass index, and INH acetylator phenotype based on NAT2 genotype, should be considered during treatment.

Optimal Sampling Strategies for Therapeutic Drug Monitoring of First-Line Tuberculosis Drugs in Patients with Tuberculosis.

BACKGROUND: The 24-h area under the concentration-time curve (AUC24)/minimal inhibitory concentration ratio is the best predictive pharmacokinetic/pharmacodynamic (PK/PD) parameter of the efficacy of first-line anti-tuberculosis (TB) drugs. An optimal sampling strategy (OSS) is useful for accurately estimating AUC24; however, OSS has not been developed in the fed state or in the early phase of treatment for first-line anti-TB drugs. METHODS: An OSS for the prediction of AUC24 of isoniazid, rifampicin, ethambutol and pyrazinamide was developed for TB patients starting treatment. A prospective, randomized, crossover trial was performed during the first 3 days of treatment in which first-line anti-TB drugs were administered either intravenously or in fasting or fed conditions. The PK data were used to develop OSS with best subset selection multiple linear regression. The OSS was internally validated using a jackknife analysis and externally validated with other patients from different ethnicities and in a steady state of treatment. RESULTS: OSS using time points of 2, 4 and 8 h post-dose performed best. Bias was < 5% and imprecision was < 15% for all drugs except ethambutol in the fed condition. External validation showed that OSS2-4-8 cannot be used for rifampicin in steady state conditions. CONCLUSION: OSS at 2, 4 and 8 h post-dose enabled an accurate and precise prediction of AUC24 values of first-line anti-TB drugs in this population. TRIAL REGISTRATION: ClinicalTrials.gov (NCT02121314).

Eosinophil peroxidase oxidizes isoniazid to form the active metabolite against M. tuberculosis, isoniazid-NAD.

The formation of isonicotinyl-nicotinamide adenine dinucleotide (INH-NAD+) by the mycobacterial catalase-peroxidase enzyme, KatG, was known to be the major component of the mode of action of isoniazid (INH), an anti-tuberculosis drug. However, there are other enzymes that may catalyze this reaction. We have previously reported that neutrophil myeloperoxidase (MPO) is capable of metabolizing INH through the formation of INH-NAD+ adduct, which could be attributed to being a possible mode of action of INH. However, eosinophilic infiltration of the lungs is more pronounced and characteristic of granulomas in Mycobacterium tuberculosis-infected patients. Thus, the aim of the present study is to investigate the role of eosinophil peroxidase (EPO), a key eosinophil enzyme, during INH metabolism and the formation of its active metabolite, INH-NAD+ using purified EPO and eosinophils isolated from asthmatic donors. UV-Vis spectroscopy revealed INH oxidation by EPO led to a new product (λmax = 326 nm) in the presence of NAD+. This adduct was confirmed to be INH-NAD+ using LC-MS analysis where the intact adduct was detected (m/z = 769). Furthermore, EPO catalyzed the oxidation of INH and formed several free radical intermediates as assessed by electron paramagnetic resonance (EPR) spin-trapping; a carbon-centred radical, which is considered to be the reactive metabolite that binds with NAD+, was found when superoxide dismutase was included in the reaction. Our findings suggest that eosinophilic EPO may also play a role in the pharmacological activity of INH through the formation of INH-NAD+ adduct, and supports further evidence that human cells and enzymes are capable of producing the active metabolite involved in tuberculosis treatment.

Pharmacokinetics of First-Line Anti-Tubercular Drugs.

Determining the optimal dosages of isoniazid, rifampicin, pyrazinamide and ethambutol in children is necessary to obtain therapeutic serum concentrations of these drugs. Revised dosages have improved the exposure of 1st line anti-tubercular drugs to some extent; there is still scope for modification of the dosages to achieve exposures which can lead to favourable outcome of the disease. High dose of rifampicin is being investigated in clinical trials in adults with some benefit; studies are required in children. Inter-individual pharmacokinetic variability and the effect of age, nutritional status, Human immunodeficiency virus (HIV) infection, acetylator genotype may need to be accounted for in striving for the dosages best suited for an individual.

Polymorphisms of NAT2, CYP2E1, GST, and HLA related to drug-induced liver injury in indonesian tuberculosis patients.

Background: Gene polymorphisms have been associated with drug-induced liver injury (DILI). This study aimed to elucidate the association between polymorphisms of NAT2, CYP2E1, GSTT1, GSTM1, and HLA genes with isoniazid plasma concentration and DILI. Methods: This study was a prospective cohort study recruiting adult newly diagnosed tuberculosis (TB) patients who met the inclusion criteria from the Public Health Centers in Yogyakarta and Lampung. Defined single-nucleotide polymorphisms were rs1799929, rs1799930, rs1799931, rs1801280, and rs1041983 of NAT2; rs2031920, rs8192775, and rs2515641 of CYP2E1; rs1041981, rs1063355, and rs6906021 of HLA. GSTT1 and GSTM1 were defined as GSTT1, GSTM1, and GSTT1 deletion and GSTM1 deletion. The DNA was taken from the patient saliva. Data of anti-TB drug plasma concentration on the weeks 4-8 of treatment were retrieved from the patients' medical report. Statistical analysis was performed using Chi-square test, Student's t-test, and multinomial logistic regression. Results: Over the 207 patients, up to 1.9% of them experienced DILI. The percentage of slow acetylators of NAT2 was 69.5%. Patients with extensive acetylator phenotype did not experience DILI (odds ratio [OR]: 0.46; 95% confidence interval [CI]: 0.23-0.94). The G carriership of HLA rs1063355 could protect the patients from the DILI (OR: 0.39; 95% CI: 0.14-0.9). Furthermore, the C carriership of HLA rs1041981 can protect the patients from DILI (OR: 0.38; 95% CI: 0.15-0.50). The genotype of HLA-DQB*0302 significantly affects the isoniazid concentration. Conclusion: The NAT2 genotype was significantly associated with DILI. Furthermore, the absence of G carriership of HLA-DQA*0102 could protect the patients from DILI without being associated with an effect on the isoniazid concentration.