Advances in computational frameworks in the fight against TB: The way forward

Around 1.6 million people lost their life to Tuberculosis in 2021 according to WHO estimates. Although an intensive treatment plan exists against the causal agent, Mycobacterium Tuberculosis, evolution of multi-drug resistant strains of the pathogen puts a large number of global populations at risk. Vaccine which can induce long-term protection is still in the making with many candidates currently in different phases of clinical trials. The COVID-19 pandemic has further aggravated the adversities by affecting early TB diagnosis and treatment. Yet, WHO remains adamant on its “End TB” strategy and aims to substantially reduce TB incidence and deaths by the year 2035. Such an ambitious goal would require a multi-sectoral approach which would greatly benefit from the latest computational advancements. To highlight the progress of these tools against TB, through this review, we summarize recent studies which have used advanced computational tools and algorithms for—early TB diagnosis, anti-mycobacterium drug discovery and in the designing of the next-generation of TB vaccines. At the end, we give an insight on other computational tools and Machine Learning approaches which have successfully been applied in biomedical research and discuss their prospects and applications against TB

Discovery and development of novel salicylate synthase (MbtI) furanic inhibitors as antitubercular agents

We report on the virtual screening, synthesis, and biological evaluation of new furan derivatives targeting Mycobacterium tuberculosis salicylate synthase (MbtI). A receptor-based virtual screening procedure was applied to screen the Enamine database, identifying two compounds, I and III, endowed with a good enzyme inhibitory activity. Considering the most active compound I as starting point for the development of novel MbtI inhibitors, we obtained new derivatives based on the furan scaffold. Among the SAR performed on this class, compound 1a emerged as the most potent MbtI inhibitor reported to date (Ki = 5.3 μM). Moreover, compound 1a showed a promising antimycobacterial activity (MIC99 = 156 μM), which is conceivably related to mycobactin biosynthesis inhibition

Design, Synthesis,Characterization and Biological Evaluation of Some Novel Benzimidazole Derivatives as Anti-Tubercular Agents Targeting Inha (Enoyl Acyl Carrier Protein Reductase) Enzyme

InhA (Enoyl acyl carrier protein reductase), a critical enzyme for the cell wall synthesis of Mycobacterium tuberculosis was chosen for study after review of literature. Candidate molecules were designed and docked against 2h9i protein using Autodock Tools® 1.5.6 software. Molecules with good Docking score (lower binding energy) and favourable interactions were checked for druglikeness and toxicity. The selected molecules were subjected to Toxicity prediction assessment by OSIRIS® software Compounds with good activity, druglikeness and absence of significant toxicity were shortlisted for synthesis. Compounds were synthesized by conventional method and labelled as SC, SJ, SM, SN, SE Purity of the synthesized compounds was ensured by repeated recrystallization with Methanol. Further the compounds were evaluated by TLC and Melting point were determined. The characterization of the synthesized compounds was done using Infra-Red (IR), Nuclear Magnetic Resonance Spectroscopy(1H NMR) and Mass spectrometry (LC-MS). The pure compounds were screened for In-vitro Anti- tubercular activity by Micro plate Alamar Blue Assay (MABA). All compounds showed good anti-mycobacterial activity. The synthesized compounds were active at concentrations of 1.6-25.0μg/ml, which were comparable to the known anti-TB drugs: Pyrazinamide-3.125μg/ml, Ciprofloxacin - 3.125μg/ml and Streptomycin - 6.25μg/ml. CONCLUSION The work concludes that the synthesized molecules are effective in inhibiting the target enzyme InhA , which is important for the growth of Mycobacterium tuberculosis cell wall. All the five compounds gave Docking score between -6.8 to -8.38 kcal/mol. There is a correlation between the docking score and actual activities of all the compounds which were tested and compared with the standard drugs. This goes to prove that 2h9i is a critical enzyme for anti-mycobacterial activity. Further structural refinement to the structure of the synthesized compounds is expected to yield promising molecules against the pathogen Mycobacterium tuberculosis

Design, Synthesis, Characterization and Biological Evaluation of Some Novel Benzimidazole (Schiff’ Base) Derivatives as Anti-Tubercular Agents against INHA

Enoyl acyl carrier protein reductase is a vital enzyme present in the cell wall of Mycobacterium tuberculosis. • A database of 200 molecules with high prospects of inhibiting the target Enoyl acyl carrier protein reductase were carefully chosen by making changes to the known hit molecules, here the Benzimidazole nucleus waschosen. • The designed molecules were docked against the target chosen using AutoDock 4®. • Five molecules with good docking score [lower binding energy] and interactions were shortlisted for synthesis. Reaction conditions wereoptimized. • The selected molecules were subjected to toxicity prediction assessment by OSIRIS® property explorer developed by Acetilon Pharmaceuticals limited which is available online. The results are color coded as green color which predicts the drug likeness and possibly betteractivity. • The molecules were labelled as RJ, RK, RM, RN, RO and were synthesized with satisfactoryyield. • The purity of the synthesized compounds was ensured by repeated recrystallization. Further the compounds were evaluated by TLC and Melting pointdetermination. • The characterization of the synthesized compounds was done using Infra-red, Nuclear Magnetic Resonance [H1 NMR] and Mass spectrometric methods [LC-MS]. • All the Synthesized compounds exhibited molecular ion peak (M+) of varying intensities. • The final pure compounds were screened for Anti-mycobacterial activity by in vitro method called Micro plate Alamar Blue Assay[MABA]. The synthesized compounds showed sensitivity [Minimum inhibitory concentration] at 1.6mcg/ml. The standard drugs Pyrazinamide, Ciprofloxacin and Streptomycin exhibited anti-mycobacterial activity at 3.125 mcg/ml, 3.125 mcg/ml and 6.25mcg/ml concentrations respectively. This indicates that the synthesized compounds are as Potent as the standarddrugs. • The selected compounds showed IC50 Values of 37.63, 111.2μg/ml respectively for RJ,RK

An integrated computational approach of molecular dynamics simulations, receptor binding studies and pharmacophore mapping analysis in search of potent inhibitors against tuberculosis

Tuberculosis is an infectious chronic disease caused by obligate pathogen Mycobacterium tuberculosis that affects millions of people worldwide. Although many first and second line drugs are available for its treatment, but their irrational use has adversely lead to the emerging cases of multiple drug resistant and extensively drug-resistant tuberculosis. Therefore, there is an intense need to develop novel potent analogues for its treatment. This has prompted us to develop potent analogues against TB. The Mycobacterium tuberculosis genome provides us with number of validated targets to combat against TB. Study of Mtb genome disclosed six epoxide hydrolases (A to F) which convert harmful epoxide into diols and act as a potential drug target for rational drug design. Our current strategy is to develop such analogues which inhibits epoxide hydrolase enzyme present in Mtb genome. To achieve this, we adopted an integrated computational approach involving QSAR, pharmacophore mapping, molecular docking and molecular dynamics simulation studies. The approach envisaged vital information about the role of molecular descriptors, essential pharmacophoric features and binding energy for compounds to bind into the active site of epoxide hydrolase. Molecular docking analysis revealed that analogues exhibited significant binding to Mtb epoxide hydrolase. Further, three docked complexes 2s, 37s and 15s with high, moderate and low docking scores respectively were selected for molecular dynamics simulation studies. RMSD analysis revealed that all complexes are stable with average RMSD below 2 Å throughout the 10 ns simulations. The B-factor analysis showed that the active site residues of epoxide hydrolase are flexible enough to interact with inhibitor. Moreover, to confirm the binding of these urea derivatives, MM-GBSA binding energy analysis were performed. The calculations showed that 37s has more binding affinity (ΔGtotal = −52.24 kcal/mol) towards epoxide hydrolase compared to 2s (ΔGtotal = −51.70 kcal/mol) and 15s (ΔGtotal = −49.97 kcal/mol). The structural features inferred in our study may provide the future directions to the scientists towards the discovery of new chemical entity exhibiting anti-TB property.AICTE and Department of Science and Technology, New Delhi.http://www.elsevier.com/locate/JMGM2019-08-01hj2018Plant Production and Soil Scienc

Design, Synthesis, Characterization and Biological Evaluation of Some Novel Heterocyclic Compounds as Anti-tubercular agents

Thirty molеculеs (Ν1 to Ν20, GA, GAC, GAC1, GAC5, GAC6, GAC7, GAЕ, GAЕA, GAM and GAT) which wеrе prеdictеd to bе еffеctivе against Mycobactеrium tubеrculosis wеrе sеlеctеd for thе synthеsis through computational studiеs. This was achiеvеd by thе molеcular docking studiеs against thе targеt еnzymе InhA (PDB id – 2NSD) of Mycobactеrium tubеrculosis, in-silico ADMЕ assеssmеnt and in-silico toxicity prеdictions. ❖ Thе thirty molеculеs which wеrе sеlеctеd for thе synthеsis bеlong to thе following functional class, ✔ 20 Thiazolidinonе dеrivativеs (Ν1 to Ν20) and ✔ 10 dioxolan basеd analoguеs comprising of • thiеno-pyrimidinе corе (4 compounds) (GAC, GAC1, GAC5 and GAC7), • thiеno-pyridinе corе (2 compounds) (GAЕA and GAM), • thiеno-thiazinе corе (1 compound) (GAC6) and • dihydro bеnzo-thiophеnе (3 compounds) (GA, GAЕ and GAT). All thе thirty molеculеs wеrе synthеsizеd. Thе synthеsizеd compounds wеrе purifiеd and charactеrizеd. Thе synthеsizеd ϲompounds wеrе ϲharaϲtеrizеd by “FT-IR, 1H-NMR, 13C-NMR, Mass spеctra and еlеmеntal analysis”. Thе rеlativе stеrеochеmistry of onе compound was confirmеd by thе X-Ray Crystallography. ❖ Thе physical charactеristics and spеctral studiеs likе “FT-IR, 1H-NMR, 13CNMR, Mass spеctra and еlеmеntal analysis” confirmеd thе proposеd structurе of thе synthеsizеd compounds. ❖ All thе synthеsizеd compounds wеrе invеstigatеd for thеir in-vitro anti-tubеrcular potеntial using “Microplatе Alamar Bluе assay” MABA Assay. All thе compounds showеd modеratе to potеnt in vitro activity against MTB with MIC rangе 0.05-50 μg/ml concеntrations. Compounds Ν18, Ν11 and Ν20 displayеd most potеnt in-vitro activity with MICs 0.05, 0.1, 0.2 μg/ml concеntrations rеspеctivеly. ❖ In ordеr to corrеlatе thе in-vitro anti-tubеrcular activity rеsults with thе docking rеsults, furthеr docking was pеrformеd with the top 12 compounds (Ν10, Ν11 to Ν20 and GAT) rеsulting from in-vitro anti-tubеrcular activity data against thе multiplе targеt еnzymеs of Mycobactеrium tubеrculosis {Thymidylatе Kinasе (PDB id – 1G3U), Diaminopimеlatе Dеcarboxylasе (PDB id – 1HKV), Cyclopropanе Synthasе (PDB id – 1L1Е), Antibiotic Rеsistancе Protеin (PDB id – 1YK3), TrpD еssеntial for lung colonization (PDB id – 1ZVW), Thymidylatе Synthasе X (PDB id – 2AF6), Protеin Kinasе G (PDB id – 2PZI), Gyrasе TypеIIA Topoisomеrasе (PDB id – 3UC1), L, D Transpеptidasе 2 (PDB id – 3VAЕ)}. ❖ In ordеr to rationalizе thе corrеlation bеtwееn thе in-vitro antitubеrcular activity and multi-targеt docking rеsults a cross obsеrvational analysis was pеrformеd. Thе top-rankеd thrее compounds (Ν18, Ν11 and Ν20) of in-vitro antitubеrcular activity wеrе cross obsеrvеd with thеir docking ranks on all thе studiеd targеt еnzymеs for thеir dеviations in thеir ranks. ❖ Basеd upon multiplе targеt docking study rеsults and thе cross obsеrvational analysis rеsults, thе еnzymе Thymidylatе Kinasе (PDB id – 1G3U) was found to bе morе appropriatе targеt for thе tеstеd compounds that еxhibitеd in-vitro antitubеrcular activity. Thus thе scopе and limitations of softwarе and thе plausiblе mеchanism of action for thе activity was provеd. ❖ Thе stability of thе ligand-rеcеptor complеxеs wеrе analysеd by molеcular dynamic simulation study. This was achiеvеd by pеrforming thе study with thе ligands Ν18-1G3U, Ν11-1G3U and Ν20-1G3U (-thymidylatе kinasе) complеxеs (top-rankеd doϲking ligand-rеcеptor complеx). Thе study confirmеd that thе ligand-rеcеptor complеxеs wеrе stablе without any notablе conformational changеs during thе simulation run. At thе еnd of thе MD simulation, changе in position and oriеntation of ligands in thе introducеd binding sitе wеrе obsеrvеd, which indicatеs thе usеfulnеss of thе MD simulation for thе optimization of thе ligands into thе targеt binding sitе. ❖ Thе compounds showing in-vitro inhibitory activity bеlow 12.5 μg/ml concеntrations against Mycobactеrium tubеrculosis wеrе subjеctеd for thе acutе oral toxicity studiеs. Thе sеlеctеd compound codеs wеrе N10, N11 to N20 and GAT. No signs of toxicity wеrе noticеd at thе dosе of 300 mg/kg b.w, whilе somе signs of toxicity at 2000 mg/kg b.w. to thе group of animals wеrе rеcordеd. Thus thе study suggеsts that thе LD50 valuе of thе tеstеd compounds wеrе еxcеptеd to еxcееd 300 mg/kg b.w and was rеprеsеntеd as class 4 (300 mg/kg<LD50<2000mg/kg) according to Globally Harmonizеd Classification Systеm (GHS). ❖ Thrее compounds (N11, N20 and N18) which displayеd еffеctivе inhibition of Mycobactеrium tubеrculosis in in-vitro anti-tubеrcular activity wеrе studiеd for thеir in-vivo potеntial using Balb/ϲ mousе modеl for Colony Forming Units (CFU) and Mortality. It was found that compound N18 was activе in in-vivo antimycobactеrial assay, whеn comparеd to thе othеr synthеsizеd tеstеd compounds. It was also intеrеsting to noticе that thе compound N18 dеcrеasеd thе bactеrial load to 24.33± 2.186 at 10 mg/kg dosе, whilе standard drug isoniazid dеcrеasеd thе bactеrial load to 15.33±1.764 at 25 mg/kg dosе. Thus thе study concludеs that thе CFU valuе obtainеd by compound N18 at thе dosе of 10 mg/kg was found to bе significant whеn comparеd to thе standard drug isoniazid at 25 mg/kg dosе. CONCLUSION: In thе prеsеnt work, simplе and еfficiеnt practical mеthods for thе synthеsis of hеtеrocyclics, which rеsultеd from thе in-silico approach was achiеvеd in good yiеld. ☞ Thiazolidinonе dеrivativеs, i.e. compounds Ν18, Ν11 and Ν20 showеd most potеnt inhibition in in-vitro antitubеrcular activity at MIC 0.05, 0.1 and 0.2 μg/ml concеntrations. ☞ In-vivo acutе toxicity studiеs and in-silico ADMЕ prеdictions rеports suggеst thе lеad compounds Ν18, Ν11 and Ν20 can be taken up for further studies. ☞ It was found that lеad compound Ν18 was activе in in-vivo antimycobactеrial assay, whеn comparеd to thе othеr synthеsizеd tеstеd compounds. ☞ It was intеrеsting to note that thе compound N18 dеcrеasеd thе bactеrial load to 24.33± 2.186 at 10 mg/kg dosе, whilе standard drug isoniazid dеcrеasеd thе bactеrial load to 15.33±1.764 at 25 mg/kg dosе. Thus thе study dеsеrvеs for thе conclusion that thе CFU valuе obtainеd by compound N18 at thе dosе of 10 mg/kg was found to bе significant whеn comparеd to thе standard drug isoniazid at 25 mg/kg dosе. It was also concludеd that, on incrеasing thе dosе of compound N18, it may producе morе significant rеsults comparеd to thе standard drug isoniazid. ☞ Thе abovе findings havе dеmonstratеd that the compound N18 ((Z)-5-(3-nitrobеnzylidеnе)-2-thioxothiazolidin-4-onе) is possibly a good antimycobacterial agent

FReDoWS: a method to automate molecular docking simulations with explicit receptor flexibility and snapshots selection

<p>Abstract</p> <p>Background</p> <p><it>In silico</it> molecular docking is an essential step in modern drug discovery when driven by a well defined macromolecular target. Hence, the process is called structure-based or rational drug design (RDD). In the docking step of RDD the macromolecule or receptor is usually considered a rigid body. However, we know from biology that macromolecules such as enzymes and membrane receptors are inherently flexible. Accounting for this flexibility in molecular docking experiments is not trivial. One possibility, which we call a fully-flexible receptor model, is to use a molecular dynamics simulation trajectory of the receptor to simulate its explicit flexibility. To benefit from this concept, which has been known since 2000, it is essential to develop and improve new tools that enable molecular docking simulations of fully-flexible receptor models.</p> <p>Results</p> <p>We have developed a Flexible-Receptor Docking Workflow System (FReDoWS) to automate molecular docking simulations using a fully-flexible receptor model. In addition, it includes a snapshot selection feature to facilitate acceleration the virtual screening of ligands for well defined disease targets. FReDoWS usefulness is demonstrated by investigating the docking of four different ligands to flexible models of <it>Mycobacterium tuberculosis’</it> wild type InhA enzyme and mutants I21V and I16T. We find that all four ligands bind effectively to this receptor as expected from the literature on similar, but wet experiments.</p> <p>Conclusions</p> <p>A work that would usually need the manual execution of many computer programs, and the manipulation of thousands of files, was efficiently and automatically performed by FReDoWS. Its friendly interface allows the user to change the docking and execution parameters. Besides, the snapshot selection feature allowed the acceleration of docking simulations. We expect FReDoWS to help us explore more of the role flexibility plays in receptor-ligand interactions. FReDoWS can be made available upon request to the authors.</p

Mycobacterial drug discovery

Mycobacterium tuberculosis is the causative pathogen of the pulmonary disease tuberculosis. Despite the availability of effective treatment programs, there is a global pursuit of new anti-tubercular agents to respond to the developing threat of drug resistance, in addition to reducing the extensive duration of chemotherapy and any associated toxicity. The route to mycobacterial drug discovery can be considered from two directions: target-to-drug and drug-to-target. The former approach uses conventional methods including biochemical assays along with innovative computational screens, but is yet to yield any drug candidates to the clinic, with a high attrition rate owing to lack of whole cell activity. In the latter approach, compound libraries are screened for efficacy against the bacilli or model organisms, ensuring whole cell activity, but here subsequent target identification is the rate-limiting step. Advances in a variety of scientific fields have enabled the amalgamation of aspects of both approaches in the development of novel drug discovery tools, which are now primed to accelerate the discovery of novel hits and leads with known targets and whole cell activity. This review discusses these traditional and innovative techniques, which are widely used in the quest for new anti-tubercular compounds