A battery of tandem mass spectrometry assays with stable isotope-dilution for the quantification of 15 anti-tuberculosis drugs and two metabolites in patients with susceptible-, multidrug-resistant- and extensively drug-resistant tuberculosis.

OBJECTIVE: Anti-tuberculosis (antiTB) drugs are characterized by an important inter-interindividual pharmacokinetic variability poorly predictable from individual patients' characteristics. Therapeutic drug monitoring (TDM) may therefore be beneficial for patients with Mycobacterium tuberculosis infection, especially for the management of multidrug/extensively drug resistant- (MDR/XDR)-TB. Our objective was to develop robust HPLC-MS/MS methods for plasma quantification of 15 antiTB drugs and 2 metabolites, namely rifampicin, isoniazid plus N-acetyl-isoniazid, pyrazinamide, ethambutol (the conventional quadritherapy for susceptible TB) as well as combination of agents against MDR/XDR-TB: bedaquiline, clofazimine, delamanid and its metabolite M1, levofloxacin, linezolid, moxifloxacin, pretomanid, rifabutin, rifapentine, sutezolid, and cycloserine. METHODS: Plasma protein precipitation was used for all analytes except cycloserine, which was analyzed separately after derivatization with benzoyl chloride. AntiTB quadritherapy drugs (Pool1) were separated by Hydrophilic Interaction Liquid Chromatography (column Xbridge BEH Amide, 2.1 × 150 mm, 2.5 µm, Waters®) while MDR/XDR-TB agents (Pool 2) and cycloserine (as benzoyl derivative) were analyzed by reverse phase chromatography on a column XSelect HSS T3, 2.1 × 75 mm, 3.5 µm (Waters®). All runs last <7 min. Quantification was performed by selected reaction monitoring electrospray tandem mass spectrometry, using stable isotopically labelled internal standards. RESULTS: The method covers the clinically relevant plasma levels and was extensively validated based on FDA recommendations, with intra- and inter-assay precision (CV) < 15% over the validated ranges. Application of the method is illustrated by examples of TDM for two patients treated for drug-susceptible- and MDR-TB. CONCLUSION: Such convenient extraction methods and the use of stable isotope-labelled drugs as internal standards provide an accurate and precise quantification of plasma concentrations of all major clinically-used antiTB drugs regimens and is optimally suited for clinically efficient TDM against tuberculosis.

Recent Discoveries of Nitrogen-Containing Heterocyclic Compounds as InhA Inhibitors against Mycobacterium tuberculosis: An Overview.

It is a formidable challenge to treat tuberculosis as there are increasing cases of multidrugresistant tuberculosis (MDR-TB) and extensively drug-resistant tuberculosis (XDR-TB) cases. Moreover, the emergence of totally drug-resistant tuberculosis (TDR-TB) makes it vital and imperative to develop a new generation of anti-tubercular drugs to have control over resistance. The nitrogencontaining heterocyclic class of compounds is being studied extensively to ascertain their anti-TB potentials. Nitrogen-containing compounds have a broad range of targets; wherein, InhA is the most important one. Hence, the primary focus of this review is to summarize the recent developments in the discovery of nitrogen-containing heterocyclic compounds as InhA inhibitors to combat tuberculosis.

Assessment of the efficacy of clofazimine alone and in combination with primary agents against Mycobacterium tuberculosis in vitro.

OBJECTIVES: The chemotherapeutic regimens of patients with drug-susceptible (DS)- tuberculosis (TB) comprise four primary anti-TB drugs: rifampicin (RMP), isoniazid (INH), ethambutol (EMB) and pyrazinamide (PZA), administered for six-to-nine months. These drug regimens target the various microbial populations that include actively replicating (AR), slow-replicating (SR) and non-replicating (NR) organisms. Clofazimine (CFZ) has showed benefit in shortening DS-TB treatment in vivo from six to four months when used in combination with this regimen in murine models of experimental infection. However, its antimicrobial efficacy when used in combination with the primary drugs against the various microbial populations of Mycobacterium tuberculosis has not been demonstrated. METHODS: In the current in vitro study, the inhibitory and bactericidal activities of CFZ in combination with the primary anti-TB drugs, RMP, INH and EMB against the AR and SR organisms in planktonic and biofilm-forming cultures, respectively, were evaluated by fractional inhibitory concentration index (FICI) and fractional bactericidal concentration index (FBCI) determinations, using the Loewe Additivity Model. RESULTS: In planktonic cultures, CFZ demonstrated synergistic growth inhibitory activity in combination with RMP and INH individually and collectively. With respect to bactericidal activity, CFZ exhibited synergistic activity only in a two-drug combination with RMP. However, in biofilm-forming cultures, all CFZ-containing anti-TB drug combinations exhibited synergistic inhibitory and bactericidal effects, particularly in combination with RIF and INH. CONCLUSION: Clofazimine exhibited synergistic effects in combination with primary anti-TB drugs against both planktonic and biofilm-forming cultures, showing potential benefit in augmenting treatment outcome when used during standard TB chemotherapy.

Dual-Acting Small-Molecule Inhibitors Targeting Mycobacterial DNA Replication.

Mycobacterium tuberculosis (Mtb) is a pathogenic bacterium and a causative agent of tuberculosis (TB), a disease that kills more than 1.5 million people worldwide annually. One of the main reasons for this high mortality rate is the evolution of new Mtb strains that are resistant to available antibiotics. Therefore, new therapeutics for TB are in constant demand. Here, we report the development of small-molecule inhibitors that target two DNA replication enzymes of Mtb, namely DnaG primase and DNA gyrase (Gyr), which share a conserved TOPRIM fold near the inhibitors' binding site. The molecules were developed on the basis of previously reported inhibitors for T7 DNA primase that bind near the TOPRIM fold. To improve the physicochemical properties of the molecules as well as their inhibitory effect on primase and gyrase, 49 novel compounds have been synthesized as potential drug candidates in three stages of optimization. The last stage of chemical optimization yielded two novel inhibitors for both Mtb DnaG and Gyr that also showed inhibitory activity toward the fast-growing non-pathogenic model Mycobacterium smegmatis (Msmg).

Discovery and evaluation of new compounds targeting ribosomal protein S1 in antibiotic-resistant Mycobacterium Tuberculosis.

The emergence of antibiotic-resistant Mycobacterium Tuberculosis (Mtb) infections compels new treatment strategies, of which targeting trans-translation is promising. During the trans-translation process, the ribosomal protein S1 (RpsA) plays a key role, and the Ala438 mutant is related to pyrazinamide (PZA) resistance, which shows its effects after being hydrolysed to pyrazinoic acid (POA). In this study, based on the structure of the RpsA C-terminal domain (RpsA-CTD) and POA complex, new compounds were designed. After being synthesized, the compounds were tested in vitro with saturation transfer difference (STD), fluorescence quenching titration (FQT) and chemical shift perturbation (CSP) experiments. Finally, six of the 17 new compounds have high affinity for both RpsA-CTD and its Ala438 deletion mutant. The active compounds provide new choices for targeting trans-translation in Mtb, and the analysis of the structure-activity relationships will be helpful for further structural modifications based on derivatives of 2-((hypoxanthine-2-yl)thio)acetic acid and 2-((5-hydroxylflavone-7-yl)oxy)acetamide.

Association of adverse drug reaction to anti-tuberculosis medication with quality of life in patients in a tertiary referral hospital

Abstract INTRODUCTION: Adverse drug reactions can develop when using anti-tuberculosis medication, and the effects of the drugs can also significantly hinder the treatment of patients. METHODS: A cross-sectional survey was conducted in 73 patients using two standardized questionnaires and the World Health Organization Quality of Life-Bref. RESULTS: All patients reported the presence of adverse drug reactions, 71.6% of which are minor and 28.3% both major and minor. The global quality of life analysis showed that patients with tuberculosis have a good average (67.3%). CONCLUSIONS: There is an association between quality of life and adverse drug reaction, educational level, and vulnerability.

Synthesis and evaluation of functionalized benzoboroxoles as potential anti-tuberculosis agents.

Several derivatives of aminobenzoboroxole have been prepared starting from 2-boronobenzaldehyde. All of these derivatives have been evaluated for their anti-mycobacterial activity on Mycobacterium smegmatis and cytotoxicity on breast cancer cell line MCF7. Based on these studies, all the tested molecules have been found to be generally non-toxic and benzoboroxoles with unsubstituted (primary) amines have been found to exhibit good anti-mycobacterial activity. Some of the key compounds have been evaluated for their anti-tubercular activity on Mycobacterium tuberculosis H37Rv using 7H9 and GAST media. 7-Bromo-6-aminobenzoboroxole 4 has been identified as the lead candidate compound for further development.

Rational design of InhA inhibitors in the class of diphenyl ether derivatives as potential anti-tubercular agents using molecular dynamics simulations.

A series of diphenyl ether derivatives were developed and showed promising potency for inhibiting InhA, an essential enoyl acyl carrier protein reductase involved in mycolic acid biosynthesis, leading to the lysis of Mycobacterium tuberculosis. To understand the structural basis of diphenyl ether derivatives for designing more potent inhibitors, molecular dynamics (MD) simulations were performed. Based on the obtained results, the dynamic behaviour in terms of flexibility, binding free energy, binding energy decomposition, conformation, and the inhibitor-enzyme interaction of diphenyl ether inhibitors were elucidated. Phe149, Tyr158, Met161, Met199, Val203 and NAD+ are the key residues for binding of diphenyl ether inhibitors in the InhA binding pocket. Our results could provide the structural concept to design new diphenyl ether inhibitors with better enzyme inhibitory activity against M. tuberculosis InhA. The present work facilitates the design of new and potentially more effective anti-tuberculosis agents.

Anti-Tuberculosis Agents Induced Pseudomembranous Colitis Treated with Maintaining Anti-Tuberculosis Drugs / 대한소화기내시경학회지

Tuberculosis is one of the main infectious health problems in Korea, and a combination of antibiotics is required to treat this illness. The combination therapy with rifampicin, isoniazid, ethambutol and pyrazinamide has many adverse reactions and there have been several case reports about pseudomembranous colitis (PMC) after anti- tuberculosis treatment. Rifampicin is regarded as a main cause of anti-tuberculosis induced PMC because of its bacteriocidal effect, and interruption of the offending drug, such as rifampicin, is usually necessary to treat the PMC. However, in patents with uncompensated tuberculosis, the discontinuance of anti-tuberculosis medication accentuates the disease severity, and continuance of the anti-tuberculosis medication is necessary to overcome the tuberculosis. We report here on a case in which the anti- tuberculosis agents induced PMC in 32 year old female who was diagnosed with active pulmonary tuberculosis. She was treated with maintenance of the anti-tuberculosis medication and also the addition of both oral metronidazole and probiotics.