Potential Antituberculosis Drugs Research Articles

Deletion of the Mycobacterium tuberculosis cyp138 gene leads to changes in membrane-related lipid composition and antibiotic susceptibility.

Mycobacterium tuberculosis (Mtb), the main cause of tuberculosis (TB), has brought a great burden to the world's public health. With the widespread use of Mtb drug-resistant strains, the pressure on anti-TB treatment is increasing. Anti-TB drugs with novel structures and targets are urgently needed. Previous studies have revealed a series of CYPs with important roles in the survival and metabolism of Mtb. However, there is little research on the structure and function of CYP138. In our study, to discover the function and targetability of CYP138, a cyp138-knockout strain was built, and the function of CYP138 was speculated by the comparison between cyp138-knockout and wild-type strains through growth curves, growth status under different carbon sources, infection curves, SEM, MIC tests, quantitative proteomics, and lipidomics. The knockout of cyp138 was proven to affect the Mtb's macrophage infection, antibiotics susceptibility, and the levels of fatty acid metabolism, membrane-related proteins, and lipids such as triacylglycerol. We proposed that CYP138 plays an important role in the synthesis and decomposition of lipids related to the cell membrane structure as a new potential anti-tuberculosis drug target.

Application of Enantioselective Sulfur Ylide Epoxidation to a Short Asymmetric Synthesis of Bedaquiline, a Potent Anti-Tuberculosis Drug.

A highly selective asymmetric synthesis of a potent anti-TB drug (-)-bedaquiline is accomplished using sulfur ylide asymmetric epoxidation, employing (+)-isothiocineole as an inexpensive and readily available chiral sulfide. Excellent enantioselectivity (er 96:4) and diastereoselectivity (dr 90:10) were obtained for the construction of the key diaryl epoxide, which was subsequently subjected to a highly regioselective ring opening (96:4). The synthesis was completed in nine steps starting from commercially available aldehyde in 8% overall yield.

Simultaneous determination of NTB-3119, a novel anti-tuberculosis agent, and its major metabolites in mouse plasma by LC-MS/MS and its application in preclinical pharmacokinetics study

NTB-3119, a novel benzothiopyranone derivative, has been developed as a potential anti-tuberculosis(TB) drug with strong activity. In this study, three major metabolites of NTB-3119 were firstly identified in vitro and in vivo. A sensitive and specific liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was established for the quantitative analysis of NTB-3119 and its major metabolites NTB-3190, NTB-3202 and NTB-3204 in mouse plasma. The plasma samples were processed by protein precipitation with organic solvent. NTB-3119, NTB-3190, NTB-3202, NTB-3204 and NTB-4A (Internal Standard, IS) were separated by a Zorbax-SB C18 column (100 mm × 2.1 mm, 3.5 µm) with a gradient mobile phase of acetonitrile/water at a flow rate of 0.25 mL/min. The analytes were detected by electrospray positive ion mode in Parallel Reaction Monitoring (PRM) mode on a high resolution mass spectrum (HRMS, Thermo Q Executive). The monitored transitions were m/z 456.15632 → 360.06137 for NTB-3119, m/z 426.18214 → 246.01891 for NTB-3190, m/z 472.15124 → 360.06143 for NTB-3202, m/z 442.17706 → 246.01903 for NTB-3204 and m/z 337.13691 → 163.02081 for NTB-4A, respectively. Good linearity was conducted in the range of 5–2000 ng/mL for NTB-3119, NTB-3202 and NTB-3204 as well as 2.5–1000 ng/mL for NTB-3190. The inter- and intra-batch precision (RSD%) were both lower than 13.3 %, with the accuracy ranged from 88.0 % to 108.1 %. The analytes were proved to be stable during all samples storage, preparation and analytic procedures. The validated method was successfully applied to study the pharmacokinetics and bioavailability of NTB-3119 after oral treatment in Balb/c mouse.

Increased Susceptibility of Mycobacterium tuberculosis to Ethionamide by Expressing PPs-Induced Rv0560c

Tuberculosis, an infectious disease, is one of the leading causes of death worldwide. Drug-resistant tuberculosis exacerbates its threat. Despite long-term and costly treatment with second-line drugs, treatment failure rates and mortality remain high. Therefore, new strategies for developing new drugs and improving the efficiency of existing drug treatments are urgently needed. Our research team reported that PPs, a new class of potential anti-tuberculosis drug candidates, can inhibit the growth of drug-resistant Mycobacterium tuberculosis. Here, we report a synergistic effect of PPs with ethionamide (ETH), one of the second-line drugs, as a result of further research on PPs. While investigating gene expression changes based on microarray and 2DE (two-dimensional gel electrophoresis), it was found that PPs induced the greatest overexpression of Rv0560c in M. tuberculosis. Based on this result, a protein microarray using Rv0560c protein was performed, and it was confirmed that Rv0560c had the highest interaction with EthR, a repressor for EthA involved in activating ETH. Accordingly, a synergistic experiment was conducted under the hypothesis of increased susceptibility of ETH to M. tuberculosis by PPs. As a result, in the presence of 0.5× MIC PPs, ETH showed a growth inhibitory effect on drug-sensitive and -resistant M. tuberculosis even at a much lower concentration of about 10-fold than the original MIC of ETH. It is also suggested that the effect was due to the interaction between PPs and Rv2887, the repressor of Rv0560c. This effect was also confirmed in a mouse model of pulmonary tuberculosis, confirming the potential of PPs as a booster to enhance the susceptibility of M. tuberculosis to ETH in treating drug-resistant tuberculosis. However, more in-depth mechanistic studies and extensive animal and clinical trials are needed in the future.

In silico design of novel quinazoline-based compounds as potential Mycobacterium tuberculosis PknB inhibitors through 2D and 3D-QSAR, molecular dynamics simulations combined with pharmacokinetic predictions

Serine/threonine protein kinase B (PknB) is essential to Mycobacterium tuberculosis (M. tuberculosis) cell division and metabolism and a potential anti-tuberculosis drug target. Here we apply Hologram Quantitative Structure Activity Relationship (HQSAR) and three-dimensional QSAR (Comparative Molecular Similarity Indices Analysis (CoMSIA)) methods to investigate structural requirements for PknB inhibition by a series of previously described quinazoline derivatives. PknB binding of quinazolines was evaluated by molecular dynamics (MD) simulations of the catalytic domain and binding energies calculated by Molecular Mechanics/Poisson Boltzmann Surface Area (MM-PBSA) and Molecular Mechanics/Generalized Born Surface Area (MM-GBSA) methods. Evaluation of a training set against experimental data showed both HQSAR and CoMSIA models to reliably predict quinazoline binding to PknB, and identified the quinazoline core and overall hydrophobicity as the major contributors to affinity. Calculated binding energies also agreed with experiment, and MD simulations identified hydrogen bonds to Glu93 and Val95, and hydrophobic interactions with Gly18, Phe19, Gly20, Val25, Thr99 and Met155, as crucial to PknB binding. Based on these results, additional quinazolines were designed and evaluated in silico, with HQSAR and CoMSIA models identifying sixteen compounds, with predicted PknB binding superior to the template, whose activity spectra and physicochemical, pharmacokinetic, and anti-M. tuberculosis properties were assessed. Compound, D060, bearing additional ortho- and meta-methyl groups on its R2 substituent, was superior to template regarding PknB inhibition and % caseum fraction unbound, and equivalent in other aspects, although predictions identified hepatotoxicity as a likely issue with the quinazoline series. These data provide a structural basis for rational design of quinazoline derivatives with more potent PknB inhibitory activity as candidate anti-tuberculosis agents.

Antimicrobial activity of IDD-B40 against drug-resistant Mycobacterium tuberculosis

The emergence of multi-drug resistant (MDR) and extensively drug-resistant (XDR) Mycobacterium tuberculosis creates the urgency for new anti-tuberculosis drugs to improve the efficiency of current tuberculosis treatment. In the search for a new potential tuberculosis drug, we synthesized an isoindole based chemical library and screened a potential candidate with significant anti-tuberculosis activity. The compound named 2-hydroxy-4-(4-nitro-1,3-dioxoisoindolin-2-yl) benzoic acid (IDD-B40) showed strong activity against all the tested drug-susceptible and drug-resistant strains of M. tuberculosis, with the 50% minimum inhibitory concentrations (MIC50) of 0.39 μg/ml both in culture broth and inside Raw 264.7 cells. Also, IDD-B40, in combination with rifampicin, exhibited a direct synergistic effect against both XDR and H37Rv M. tuberculosis. Besides, IDD-B40 showed a better post-antibiotic effect (PAE) than did some first-line drugs and showed no significant cytotoxicity to any cell line tested, with a selectivity index of ≥ 128. Although IDD-B40 showed a result similar to isoniazid in the preliminary mycolic acid inhibition assay, it did not exhibit any effect against other mycolic acid-producing nontuberculous mycobacterial strains (NTM), and different non-mycobacterial pathogenic strains, so further studies are required to confirm the mode of action of IDD-B40. Considering its results against M. tuberculosis, IDD-B40 is a potential anti-tuberculosis drug candidate. However, further studies are required to evaluate its potential in vivo effect and therapeutic potential.

Quantitative structure–activity relationship (QSAR) and molecular docking of xanthone derivatives as anti-tuberculosis agents

Quantitative structure–activity relationship (QSAR) and molecular docking approach were carried out to design novel anti-tuberculosis agents based on xanthone derivatives. QSAR designed new compounds were calculated by Austin Model 1 (AM1) methods and analysis of multi-linear regression (MLR). The result showed that the best model as follows: Log IC50 = 3.113 + 11.627 qC1 + 15.955 qC4 + 11.702 qC9, this result has appropriate some statistical parameters (PRESS = 2.11, r2 = 0.730, SEE = 0. 3545, R = 0.6827, FCal/FTab = 4.68), and being used to design a potential anti-tuberculosis drugs with substituted amide, sulfoxide, and carboxylate group xanthone scaffold by a number of their inhibitory concentration (IC50). The mechanism action of sulfonamide substituted on the xanthone scaffold as anti-tuberculosis was carried out using molecular docking. Docking inhibition studies were carried out on MTB C171Q receptor (4C6X.pdb) as KasA inhibitors using by the discovery studio. Based on the binding interaction showed, the sulfonamide substituted xanthone has potential being the anti-tuberculosis drugs by KasA inhibitor for target drug activity.

Aureolic Acid Group of Agents as Potential Antituberculosis Drugs.

Mycobacterium tuberculosis is one of the most dangerous pathogens. Bacterial resistance to antituberculosis drugs grows each year, but searching for new drugs is a long process. Testing for available drugs to find active against mycobacteria may be a good alternative. In this work, antibiotics of the aureolic acid group were tested on a model organism Mycobacterium smegmatis. We presumed that antibiotics of this group may be potential G4 ligands. However, this was not confirmed in our analyses. We determined the antimicrobial activity of these drugs and revealed morphological changes in the cell structure upon treatment. Transcriptomic analysis documented increased expression of MSMEG_3743/soj and MSMEG_4228/ftsW, involved in cell division. Therefore, drugs may affect cell division, possibly disrupting the function of the Z-ring and the formation of a septum. Additionally, a decrease in the transcription level of several indispensable genes, such as nitrate reductase subunits (MSMEG_5137/narI and MSMEG_5139/narX) and MSMEG_3205/hisD was shown. We concluded that the mechanism of action of aureolic acid and its related compounds may be similar to that bedaquiline and disturb the NAD+/NADH balance in the cell. All of this allowed us to conclude that aureolic acid derivatives can be considered as potential antituberculosis drugs.

The Potential of Natural Diterpenes Against Tuberculosis: An Updated Review.

Every year, 10 million people are affected by tuberculosis (TB). Despite being a preventable and curable disease, 1.5 million people die from TB each year, making it the world's top infectious disease. Many of the frontline antibiotics cause painful and disagreeable side effects. To mitigate the side effects from the use of chemically synthesized or clinical anti-tubercular drugs, there are many research studies focussed on natural products as a source of potential anti-tuberculosis drugs. Among different phytoconstituents, several classes of diterpenoids exert significant antimicrobial effects. This review explores diterpenoids as potential anti-tubercular drugs from natural sources. A total of 204 diterpenoids isolated from medicinal plants and marine species are discussed that inhibit the growth of Mycobacterium tuberculosis. The literature from 1994-2018 is reviewed, and 158 diterpenoids from medicinal plants, as well as 40 diterpenoids from marines, are alluded to have antituberculosis properties. The antitubercular activities discussed in the review indicate that the type of diterpenoids, the Mtb strains, substituents attached to diterpenoids and their position in the diterpenoids general skeleton can change the compounds antimycobacterial inhibitory effects.

A review on Hydroxamic Acids: Widespectrum Chemotherapeutic Agents

Recent developments in drug discovery have highlighted the ability of hydroxamic acids to form complexes with various metal ions, in particular iron, zinc, magnesium and calcium, and this imparts them with a number of unique biological and pharmacological properties. This review provides information on the most significant developments of the hydroxamate compounds in the medicinal area with a focus on Suberoylanilide hydroxamic acid (SAHA) and its derivatives with antibacterial, antifungal, antiviral, antimalarial, antitubercular and anticancer effect and their possible molecular mechanisms. One of the latest favorable developments has been on developing SAHA and its hybrids as potent anti-tuberculosis drugs. Moreover, the combination of the hydroxamic acids with other pharmacophore groups in one molecule can lead to the creation of a huge number of new compounds with promising properties as a multifunctional drug. Also, within the last decade, active research has focused to create these compounds into a class of new antiviral drugs designed to treat influenza, hepatitis C, and other viral infections. Thus, both linear hydroxamic acids and cyclic acids having the uniqueness of the set of bioactive properties, have been compiled here and could be looked forward as the basis for the development of effective drugs and forpilot scale productions through nitrile and amides via chemical and biological transformations using amidase and acyltransferase enzymes. Such synthesis of medicinally significant compounds from amide containing waste derived feedstocks using microbial agents is also a key factor for stimulating bioeconomy.

OPC-167832, a Novel Carbostyril Derivative with Potent Antituberculosis Activity as a DprE1 Inhibitor.

There is an urgent need for new, potent antituberculosis (anti-TB) drugs with novel mechanisms of action that can be included in new regimens to shorten the treatment period for TB. After screening a library of carbostyrils, we optimized 3,4-dihydrocarbostyril derivatives and identified OPC-167832 as having potent antituberculosis activity. The MICs of the compound for Mycobacterium tuberculosis ranged from 0.00024 to 0.002 μg/ml. It had bactericidal activity against both growing and intracellular bacilli, and the frequency of spontaneous resistance for M. tuberculosis H37Rv was less than 1.91 × 10-7 It did not show antagonistic effects with other anti-TB agents in an in vitro checkerboard assay. Whole-genome and targeted sequencing of isolates resistant to OPC-167832 identified decaprenylphosphoryl-β-d-ribose 2'-oxidase (DprE1), an essential enzyme for cell wall biosynthesis, as the target of the compound, and further studies demonstrated inhibition of DprE1 enzymatic activity by OPC-167832. In a mouse model of chronic TB, OPC-167832 showed potent bactericidal activities starting at a dose of 0.625 mg/kg of body weight. Further, it exhibited significant combination effects in 2-drug combinations with delamanid, bedaquiline, or levofloxacin. Finally, 3- or 4-drug regimens comprised of delamanid and OPC-167832 as the core along with bedaquiline, moxifloxacin, or linezolid showed efficacy in reducing the bacterial burden and preventing relapse superior to that of the standard treatment regimen. In summary, these results suggest that OPC-167832 is a novel and potent anti-TB agent, and regimens containing OPC-167832 and new or repurposed anti-TB drugs may have the potential to shorten the duration of treatment for TB.

Validation of a universal and highly sensitive two-dimensional liquid chromatography–tandem mass spectrometry methodology for the quantification of pyrazinamide, ethambutol, protionamide, and clofazimine in different biological matrices

A novel and potent anti-tuberculosis drug combination pyrazinamide (PZA), ethambutol (EMB), protionamide (PTO), and clofazimine (CFZ) that rapidly kills Mycobacterium tuberculosis (Mtb) in the lungs has been identified using the artificial-intelligence-enabled parabolic response surface approach. A universal and highly sensitive two-dimensional liquid chromatography-tandem mass spectrometry (2D-LC-MS/MS) method for the simultaneous determination of PZA, EMB, PTO, and CFZ in various biological samples in different states (liquid samples: plasma, bile, and urine; solid samples: tissue and feces) using simple pretreatment was established and validated. For the first dimension of this column-switching arrangement, the automated purification and enrichment of the drugs were achieved on a Polar-RP column. The subsequent analytical separation was performed on an Agilent Zorbax SB-Aq column, and the total loop time was 7.5 min. The positive-ionization mode with multiple reaction monitoring was used for detection. The sensitivity was good with no carry-over detected, and the lower limit of quantification ranged from 100 to 500 pg/mL. This quantification method was fully validated and proved to be robust in accordance with US Food and Drug Administration guidelines. High recoveries (85.3–111.4%) and accuracies (92.1–109.3%), together with high precision values (0.5–13.8%), were verified in all matrices. All standard curves showed favorable linearities with r2 > 0.995. This validated method was applied to study plasma pharmacokinetics, tissue distribution, and excretion in Sprague–Dawley rats after oral administration of the drug combination.

Structure of the N-terminal domain of ClpC1 in complex with the antituberculosis natural product ecumicin reveals unique binding interactions.

The biological processes related to protein homeostasis in Mycobacterium tuberculosis, the etiologic agent of tuberculosis, have recently been established as critical pathways for therapeutic intervention. Proteins of particular interest are ClpC1 and the ClpC1-ClpP1-ClpP2 proteasome complex. The structure of the potent antituberculosis macrocyclic depsipeptide ecumicin complexed with the N-terminal domain of ClpC1 (ClpC1-NTD) is presented here. Crystals of the ClpC1-NTD-ecumicin complex were monoclinic (unit-cell parameters a = 80.0, b = 130.0, c = 112.0 Å, β = 90.07°; space group P21; 12 complexes per asymmetric unit) and diffracted to 2.5 Å resolution. The structure was solved by molecular replacement using the self-rotation function to resolve space-group ambiguities. The new structure of the ecumicin complex showed a unique 1:2 (target:ligand) stoichiometry exploiting the intramolecular dyad in the α-helical fold of the target N-terminal domain. The structure of the ecumicin complex unveiled extensive interactions in the uniquely extended N-terminus, a critical binding site for the known cyclopeptide complexes. This structure, in comparison with the previously reported rufomycin I complex, revealed unique features that could be relevant for understanding the mechanism of action of these potential antituberculosis drug leads. Comparison of the ecumicin complex and the ClpC1-NTD-L92S/L96P double-mutant structure with the available structures of rufomycin I and cyclomarin A complexes revealed a range of conformational changes available to this small N-terminal helical domain and the minor helical alterations involved in the antibiotic-resistance mechanism. The different modes of binding and structural alterations could be related to distinct modes of action.

Vitamin B and Vitamin C Affect DNA Methylation and Amino Acid Metabolism in Mycobacterium bovis BCG.

Vitamins are essential nutrients and key cofactors of enzymes that regulate cellular metabolism, and also activate the immune system. Recent studies have shown that vitamin B1 (VB1) and vitamin C (Vc) can inhibit Mycobacterium tuberculosis growth, but the precise mechanism is still not well understood. In the present study, we have used RNA-sequencing (RNA-seq), liquid chromatography coupled to mass spectrometry (LC-MS) and single-molecule real-time (SMRT) sequencing to analyze the transcriptional, metabolic and methylation profiles of Mycobacterium bovis BCG when treated with VB1 and Vc. Our results show that, after vitamin treatment, variant metabolites were mainly clustered in pathways related to amino acid metabolism. Treatment with both vitamins significantly up-regulated the gene encoding cysteine synthase A. Additionally, only BCG that was treated with VC showed m4c modifications. Genes harboring this methylation were up-regulated, suggesting that m4c methylation can promote gene transcription to some extent. Overall, this study contributes to the understanding of the effects of VB1 and VC, and suggests that these vitamins constitute potential anti-tuberculosis drugs.

Molecular detection of Multi-Drug Resistant Mycobacterium species in a Tertiary Care centre

Tuberculosis (TB) - one of the leading causes of adult death in the Asia-Pacific Region, is an infectious disease caused by Mycobacterium tuberculosis (MTB). TB remains a major public health issue especially in developing nations due to the lack of adequate rapid diagnostic testing facilities. Drug resistance in tuberculosis was observed nearly fifty years ago. The risk is now due to the emergence of new strains which are most resistant to the potent anti-tuberculosis drugs. The resistance to drugs in Mycobacterium tuberculosis is conferred by mutations witⱨ the genes encoding drug targets or drug converting enzymes. In suspected extra pulmonary tuberculosis cases also, fast and accurate laboratory diagnosis is of primary importance, since the techniques which are followed from past years for detecting acid-fast bacilli have many limitations. This study may also help in standardizing the technique for rapid identification of MDR TB in extra pulmonary TB patients in this Tertiary care Hospital. The current study describes the importance and use of Polymerase Chain Reaction (PCR) for the detection of Multidrug-resistant Mycobacterium species. The target genes encoding resistance to Isoniazid and Rifampicin were detected by Polymerase Cain reaction and Agarose gel electrophoresis. Among the 359 samples, 2% were resistant to botⱨ Isoniazid and Rifampicin and tⱨe prevalence of drug resistance was found to be more in adult age groups.

WQ-3810: A new fluoroquinolone with a high potential against fluoroquinolone-resistant Mycobacterium tuberculosis

Fluoroquinolone (FQ) resistance in Mycobacterium tuberculosis (Mtb), caused by amino acid substitutions in DNA gyrase, has been increasingly reported worldwide. WQ-3810 is a newly developed FQ that is highly active against FQ-resistant pathogens; however, its activity against Mtb has not been evaluated. Herein we examined the efficacy of WQ-3810 against Mtb through the use of recombinant Mtb DNA gyrases. In addition, in vitro antimycobacterial activity of WQ-3810 was evaluated against recombinant Mtb var. bovis Bacille Calmette–Guérin strains in which gyrase-coding genes were replaced with Mtb variants containing resistance-conferring mutations. WQ-3810 showed a higher inhibitory activity than levofloxacin against most recombinant DNA gyrases with FQ-resistance mutations. Furthermore, WQ-3810 showed inhibition even against a DNA gyrase variant harboring a G88C mutation which is thought to confer the highest resistance against FQs in clinical Mtb isolates. In contrast, the FQ susceptibility test showed that WQ-3810 had relatively weak mycobactericidal activity compared with moxifloxacin. However, the combination of WQ-3810 and ethambutol showed the greatest degree of synergistic activity against recombinant strains. Since FQs and ethambutol have been used in multi-drug therapy for tuberculosis, WQ-3810 might represent a new, potent anti-tuberculosis drug that can be effective even against FQ-resistant Mtb strains.

Applications of Molecular Simulation in the Discovery of Antituberculosis Drugs: A Review.

After decades of efforts, tuberculosis has been well controlled in most places. The existing drugs are no longer sufficient for the treatment of drug-resistant Mycobacterium tuberculosis due to significant toxicity and selective pressure, especially for XDR-TB. In order to accelerate the development of high-efficiency, low-toxic antituberculosis drugs, it is particularly important to use Computer Aided Drug Design (CADD) for rational drug design. Here, we systematically reviewed the specific role of molecular simulation in the discovery of new antituberculosis drugs. The purpose of this review is to overview current applications of molecular simulation methods in the discovery of antituberculosis drugs. Furthermore, the unique advantages of molecular simulation was discussed in revealing the mechanism of drug resistance. The comprehensive use of different molecular simulation methods will help reveal the mechanism of drug resistance and improve the efficiency of rational drug design. With the help of molecular simulation methods such as QM/MM method, the mechanisms of biochemical reactions catalyzed by enzymes at atomic level in Mycobacterium tuberculosis has been deeply analyzed. QSAR and virtual screening both accelerate the development of highefficiency, low-toxic potential antituberculosis drugs. Improving the accuracy of existing algorithms and developing more efficient new methods for CADD will always be a hot topic in the future. It is of great value to utilize molecular dynamics simulation to investigate complex systems that cannot be studied in experiments, especially for drug resistance of Mycobacterium tuberculosis.

Insights into an alternative benzofuran binding mode and novel scaffolds of polyketide synthase 13 inhibitors.

Small molecules targeting biosynthesis of mycolic acids in the tuberculosis causing bacterium carry high potential for anti-tuberculosis drug discovery. Hitherto, benzofuran containing molecules were identified to target the thioesterase domain of polyketide synthase 13 (Pks13), one of the crucial constituents of this pathway. Among these, TAM16 was also reported to be highly potent in vivo. Here we performed a multi-stage virtual screening methodology recruiting both ligand- and structure-based screening tools for identification of novel Pks13 inhibitors. The large ZINC15 database comprising more than 1 billion ligands was reduced to 21,277 ligands with benzofuran rings and similarity to TAM16. This collection was screened by docking and the 20 top scoring ligands were further analyzed by molecular dynamics simulations. Molecular mechanics/generalized Born surface area (MM-GBSA) based binding free energy predictions confirmed five molecules in the ZINC15 database lead to remarkable increases in the binding free energy of TAM16. Essentially, the most potent ligand ZINC840169872, which carries a distinct scaffold with a cyclohexane group fused to the furan ring, produced a twofold change in the ligand efficiency of TAM16. Further, assessments of the structure-based tools on five different Pks13-TAM complex structures suggested a high-level agreement with the experiments, substantiating the validity of our methodology for screening Pks13 inhibitors. Overall, these in silico insights into a low energy benzofuran-based scaffold and an alternative binding mode for the benzofuran ring unravel viable strategies to generate potent anti-tuberculosis drugs, accentuating the applications of virtual screening approaches for exploring large compound libraries that cannot be easily accessed by experimentation. Graphical abstract In silico virtual screening of ZINC15 database for the identification of potential inhibitors targeting PKS13.

Pharmacologic ascorbate as a pro-drug for hydrogen peroxide release to kill mycobacteria

Background and purposeTuberculosis is one of the most highly fatal diseases worldwide, and one-third of the world's population has been infected with Mycobacterium tuberculosis (M. tuberculosis). A previous study showed that M. tuberculosis was highly susceptible to being killed by ascorbate (i.e. vitamin C, VC), but the molecular mechanisms of the bactericidal activity of VC against M. tuberculosis are still not well understood. Experimental approachWe assayed the effects of VC as an anti-tuberculosis drug against mycobacteria (i.e. M. bovis BCG or M. tuberculosis H37Rv) in macrophages (i.e. RAW 264.7 cells). Relative global protein expression changes in 5 mM VC-treated and control samples of H37Rv were investigated by Tandem mass tag (TMT)-based quantitative proteomic analysis. qRT-PCR was also used to measure the differential expression of six intracellular stress response mycobacteria genes. Key resultsQuantitative proteomic analysis showed that 11 peptide components including rip3, fdxA, Rv2028c, mtp, LH57_00670, hspX, pfkB, Rv1824, Rv1813c, LH57_08410 and Rv2030c were up-regulated and 17 peptide components were down-regulated in 5 mM VC-treated H37Rv compared with the control samples. qRT-PCR also verified that VC could induce the expression of six genes (hsp, fdxD, furA, devR, hspX, and dnaB) in BCG and H37Rv. We also found that exosomes from RAW 264.7 cells treated with pharmacologic VC could kill M. bovis BCG in vitro. Conclusion and implicationsOur results demonstrated that the bactericidal activity of VC against mycobacteria, as a pro-drug for hydrogen peroxide formation (H2O2), was dependent on reactive oxygen species production and the activated oxidative stress pathway, which suggested that pharmaceutical VC and exosomes from macrophages treated with VC could be used as potential anti-tuberculosis drugs.

1-hydroxy-5, 7-dimethoxy-2 naphthalene-carboxaldehyde inhibitors as novel antimycobacterial agents targeting H-InMyoFib cells and targeting enzymes involved in fatty acid biosynthesis of bacilli

Background: Gastrointestinal tuberculosis (Gastrointestinal TB) is a rapidly common infectious disease in low income countries due to poor life style that poses diagnostic challenge, as the broad spectrum features of the disease which may lead to diagnostic interruptions and complications. Early diagnosis and initiation of antituberculous therapy without drug-resistance is essential to prevent morbidity and mortality. 1-hydroxy-5, 7-dimethoxy-2 naphthalene-carboxaldehyde (HDNC) is a bioactive compound derived from Aegle marmelos (Rutaceae, ‘Bael’). It is a naturally occurring flavonoid having anticancer and wound healing properties. Methods & Materials: The present study demonstrated that HDNC had ant-mycobacterial effects on tubercle bacilli ATCC25618, multi-drug and extensively drugresistant clinical isolates with minimum inhibitory concentrations of 147 and 312 μM, respectively. Bacilli mainly affect the GIT, causing a strong local inflammatory response that is critical to the pathogenesis of tuberculosis. Results: We investigated the effects of HDNC on interferon (IFN)-γ-stimulated human GIT fibroblast H-InMyoFib cells. HDNC suppressed the release of tumor necrosis factor (TNF)-α and interleukin (IL)-12. A nontoxic dose of HDNC reduced mRNA expression of TNF-α, IL-1β, IL-6, IL-12, and matrix metalloproteinase-1 in IFN-γ-stimulated cells. HDNC inhibited IFN-γ-mediated stimulation of extracellular signal regulated kinase and p38 mitogen-activated protein kinase and showed high affinity binding to these kinases (binding constants: 3.95 × 104 M−1 and 6.8 × 103 M−1, respectively).HDNC inhibit targeting enzymes involved in fatty acid biosynthesis, such as enoyl-ACP-reductase, b-ketoacyl-ACP reductase and b-hydroxyacyl-ACP dehydratase of bacilli. A mouse in vivo study of lipopolysaccharide-induced GIT inflammation revealed that a nontoxic dose of HDNC reduced the levels of IL-1β, IL-6, IL-12, and INF-γ in GIT tissue. Conclusion: Gastrointestinal TB is generally managed with medical therapy with antituberculous drugs These data provide the first evidence that HDNC could be developed as a potent antituberculosis drug and represents as a strong candidate for the b-hydroxyacyl-ACP dehydratase enzyme of Bacilli.