Project description:Background: Infectious diseases are still a leading cause of death and, with the emergence of drug resistance, pose a great threat to human health. New drugs are thus continually being developed, without their effects on the immune system being studied in-depth. Here we aim to evaluate the impact of a recently release anti-TB drug, named bedaquiline (BDQ), on the response of human macrophages infected with Mycobacterium tuberculosis (MTB). Results: Here, we used MTB resistant to BDQ as a model of bacterial infection to examine the impact of BDQ on the human macrophage response. Gene expression profiling has revealed about 1,500 genes differentially expressed in MTB infected macrophages incubated with BDQ. The expression of 499 genes was upregulated and that of 996 downregulated upon treatment. The gene set up-regulated by BDQ was significantly enriched for genes involved in glucose/phospholipid metabolism, lysosome and autophagy. We found similar results with uninfected macrophages-treated with BDQ. Conclusions: Our results highlight the importance to study the interactions between antibiotics and host cells at the molecular level to better understand the consequences of antibiotic treatment during infection Overall design: Evaluation of host response to BDQ treatment in infected and uninfected human macrophages.
Project description:Bacteria commonly adapt to stresses by altering gene expression. To understand the response of M. tuberculosis (MTB) to bedaquiline, we performed transcriptomics over a time-course on MTB bacilli exposed to the drug. Overall design: Bacteria were harvested at 1, 6, 24, 48, 96 hrs after being exposed to concentrations of bedaquiline (no drug, 1.5μM, 15μM) during a 7-day drug exposure time course experiment (as indicated in each sample title). RNA was isolated and applied to custom Agilent tiled arrays. Please note that experiments were performed technically as dual channel (eg, Cy3 and Cy5-labeled samples hybridized to the same array) but processed the results as though they are single channel (Cy3 and Cy5 signals are calculated). Therefore, there are two sample records per each raw data file and the raw data file associated with each sample is indicated in the corresponding sample description field.
Project description:Bedaquiline is a recently approved drug for the treatment of multidrug-resistant tuberculosis. Adverse cardiac and hepatic drug reactions to bedaquiline have been noted in clinical practice. The current study investigated bedaquiline metabolism in human hepatocytes using a metabolomic approach. Bedaquiline N-demethylation via CYP3A4 was confirmed as the major pathway in bedaquiline metabolism. In addition to CYP3A4, we found that both CYP2C8 and CYP2C19 contributed to bedaquiline N-demethylation. The Km values of CYP2C8, CYP2C19, and CYP3A4 in bedaquiline N-demethylation were 13.1, 21.3, and 8.5 µM, respectively. We also identified a novel metabolic pathway of bedaquiline that produced an aldehyde intermediate. In summary, this study extended our knowledge of bedaquiline metabolism, which can be applied to predict and prevent drug-drug interactions and adverse drug reactions associated with bedaquiline.
Project description:The ability of Mycobacterium tuberculosis (Mtb) to adopt heterogeneous physiological states, underlies it’s success in evading the immune system and tolerating antibiotic killing. Drug tolerant phenotypes are a major reason why the tuberculosis (TB) mortality rate is so high, with over 1.8 million deaths annually. To develop new TB therapeutics that better treat the infection (faster and more completely), a systems-level approach is needed to reveal the complexity of network-based adaptations of Mtb. Here, we report the transcriptional response of Mtb to the drug Bedaquiline. We performed transcriptomic sequencing (RNA-seq) on Mtb bacilli at 4, 24, 72 h following exposure to the drug. Overall design: Cultures of Mycobacterium tuberculosis were grown in 7H9 media supplemented with ADC, 0.2% glycerol and 0.05% tyloxapol in a 37 degrees C incubator with shaking until mid log phase. Frozen 1 mL stocks of Mtb cells were added to 7H9-rich media and grown until the culture reached an OD600 of ~0.4-0.8. The cells were then diluted to OD600 of 0.05 and added to 7H9-rich media containing Bedaquiline at the predetermined amounts (0, 5.75, 11.5 μg/mL, as indicated in each sample title). Samples were collected at 4 h, 24 h, and 72 h from three biological replicates. The sequence reads that passed quality filters were aligned with Bowtie2 and processed using the R package DuffyNGS.
Project description:AIMS:To externally validate an earlier characterized relationship between bedaquiline exposure and decline in bacterial load in a more difficult-to-treat patient population, and to explore the performances of alternative dosing regimens through simulations. METHODS:The bedaquiline exposure-response relationship was validated using time-to-positivity data from 233 newly diagnosed or treatment-experienced patients with drug-resistant tuberculosis from the C209 open-label study. The significance of the exposure-response relationship on the bacterial clearance was compared to a constant drug effect model. Tuberculosis resistance type and the presence and duration of antituberculosis pre-treatment were evaluated as additional covariates. Alternative dosing regimens were simulated for tuberculosis patients with different types of drug resistance. RESULTS:High bedaquiline concentrations were confirmed to be associated with faster bacterial load decline in patients, given that the exposure-effect relationship provided a significantly better fit than the constant drug effect (relative likelihood = 0.0003). The half-life of bacterial clearance was identified to be 22% longer in patients with pre-extensively drug-resistant (pre-XDR) tuberculosis (TB) and 86% longer in patients with extensively drug-resistant (XDR) TB, compared to patients with multidrug-resistant (MDR) TB. Achievement of the same treatment response for (pre-)XDR TB patients as for MDR TB patients would be possible by adjusting the dose and dosing frequency. Furthermore, daily bedaquiline administration as in the ZeNix regimen, was predicted to be as effective as the approved regimen. CONCLUSION:The confirmed bedaquiline exposure-response relationship offers the possibility to predict efficacy under alternative dosing regimens, and provides a useful tool for potential treatment optimization.
Project description:Transcriptional profiling of M. smegmatis mc2155 grown on Hartmans de Bont (HdB) supplemented with 0.2% glycerol and 0.05% Tween80 challenged with 2 μg/ml Bedaquiline and Dimethyl sulfoxide (DMSO) Two-condition experiment. Biological replicates: 5 independently grown and harvested. One replicate per array.
Project description:The main goal of our study was to evaluate the in vitro bedaquiline susceptibility of six prevalent species of pathogenic nontuberculous mycobacteria (NTM) in China. In addition, we investigated the potential molecular mechanisms contributing to bedaquiline resistance in the different NTM species. Among slowly growing mycobacteria (SGM), bedaquiline exhibited the highest activity against Mycobacterium avium; the MIC50 and MIC90 values were 0.03 and 16 mg/liter, respectively. Among rapidly growing mycobacteria (RGM), Mycobacterium abscessus subsp. abscessus (M. abscessus) and Mycobacterium abscessus subsp. massiliense (M. massiliense) seemed more susceptible to bedaquiline than Mycobacterium fortuitum, with MIC50 and MIC90 values of 0.13 and >16 mg/liter, respectively, for both species. On the basis of bimodal distributions of bedaquiline MICs, we proposed the following epidemiological cutoff (ECOFF) values: 1.0 mg/liter for SGM and 2.0 mg/liter for RGM. Among M. avium, Mycobacterium intracellulare, Mycobacterium kansasii, M. abscessus, M. massiliense, and M. fortuitum isolates, 14 (29.8%), 41 (27.2%), 33 (39.3%), 44 (20.2%), 42 (25.8%), and 7 (31.8%), respectively, were resistant to bedaquiline. No significant differences in the proportions of bedaquiline resistance among these species were observed (P > 0.05). Genetic mutations were observed in 74 isolates (10.8%), with all nucleotide substitutions being synonymous. In conclusion, our data demonstrate that bedaquiline shows moderate in vitro activity against NTM species. Using the proposed ECOFF values, we could distinguish between bedaquiline-resistant and -susceptible strains with the broth dilution method. In addition, no nonsynonymous mutations in the atpE gene that conferred bedaquiline resistance in all six NTM species were identified.