Drug design and identification of potent leads against Mycobacterium tuberculosis thymidine monophosphate kinase

Antiviral chemotherapy often relies on nucleoside analogues, which, once phophorylated by intracellular kinases, target viral polymerases impeding DNA synthesis. In contrast, nucleoside analogues are much less explored as antibacterial drugs. Thymidine monophosphate kinase from Mycobacterium tuberculosis (TMPKmt), which is essential to DNA replication, was selected as a promising target for the design of new inhibitors. This review describes stepwise modifications of the TMPKmt substrate, guided by the feedback of enzyme assays and crystallographic analysis to afford potent enzyme inhibitors some of which also exhibited antitubercular activity. More importantly, several of the reported thymidine analogues provided a deeper understanding of the structure and catalytic mechanism of this intriguing enzyme

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

Identification of Novel Compounds with Anti-Mycobacterial Activity Using in Silico Screening and Pharmacophore Modeling Targeting Mycobacterium Thymidine Monophosphate Kinase

The increasing prevalence of drug-resistant tuberculosis (TB), which is resistant to effective multiple antibiotic, remains a major public health menace. The Mycobacterium tuberculosis (M. tuberculosis) thymidine monophosphate kinase (mtTMPK), which is an essential enzyme for the maintenance of the thymidine triphosphate pools, is considered a potential enzymatic target for the development of effective antibiotics against TB. In this study, I attempted to identify novel compounds with anti-mycobacterial activity specifically targeting mtTMPK. I performed in silico structure-based drug screening (SBDS) with a large-scale virtual compound library (6,192,932 compounds) and phramacophore-based in silico screening with a compound library of 461,383 compounds. I then evaluated the inhibitory effects of candidate compounds on model mycobacteria strains. As a result, I found that compounds K10, KTP3, KTPS1, and KTPS2, completely inhibited the growth of Mycobacterium vanbaalenii (M. vanbaalenii) and/or Mycobacterium smegmatis (M. smegmatis). In addition, I experimentally demonstrated that two compounds (KTPS1 and KTPS2) directly inhibitedmtTMPK catalytic activity to some extent. Moreover, the most potent chemical compounds, KTPS1, did not exhibit any significant toxic effects on the growth of model intestinal bacteria (Escheichila coli: E. coli) and several mammalian cells. The structural and experimental information regarding these chemical compounds is likely useful for the development of novel antibiotics for the treatment of TB.九州工業大学博士学位論文 学位記番号:情工博甲第298号 学位授与年月日:平成27年3月25日1. Introduction|2. Results|3. Discussion|4. Conclusion|5. Materials and methods九州工業大学平成26年

Anticancer and antibiotic leads from marine organis

During the past three decades, the development of marine natural products as drug leads has become a promising avenue for research. As our efforts towards the discovery of anticancer and antibiotic drug leads from marine organisms, modifications of anticancer drug candidate kahalalide F, isolations of new peptides from the mollusk, Elysia rufescens and anticancer drug leads from the NCI repository, as well as chemical regulation of antibiotic production from marine Pseudomonas aeruginosa were investigated. Kahalalide F (KF) is a potent anticancer lead isolated from the herbivorous marine mollusk E. rufescens and its algal diet Bryopsis pennata. Our semisynthesis approach was aimed to improve the efficacy or prolong the half-life resulted in 15 KF analogues. These analogues included eight elongation products with nonpolar amino acid residues, four analogues with modified amino group at the Orn residue, one dehydration product of KF, and two new cyclization products of kahalalide G. All the analogues are being evaluated by Wayne State University for solid tumor cytotoxicity. In the course of the isolation of KF as starting material for its analogues, nine new and 10 known peptides were isolated from E. rufescens. However, only five structures were determined and the remaining four structures could not be assigned due to limited sample amounts (less than 1 mg). Future work will focus on the structure assignment using FTMS amino acid sequence analysis or 700 MHz NMR. During the screening of anticancer drug leads from the NCI repository, 27 extracts from marine organisms were tested, and 12 known compounds including tetracyclic aromatic alkaloids, diterpenoids and pyrroloaminopropylimidazole alkaloids were purified. Bioassay-guided isolation from marine P. aeruginosa collected in the Gulf of Mexico afforded 15 known antibiotics including two phenazines, six 2-alkyl-4-quinolones, and seven rhamnolipids. 2-Nonyl-4(1H)-quinolone and 2-(1-nonenyl)-4(1H)-quinolone displayed potent antimalarial activity, which was first reported here. Based on the metabolic profile of antibiotics from P. aeruginosa, chemical regulation and its impact on the yield of these metabolites were investigated. Treatment of P. aeruginosa with sceptrin and co-culturing with another Pseudomonas sp. increased antibiotic production significantly. This could be attributed to the activation of antibiotic biosynthetic gene expression under stress conditions

Immune-Mediated Drug Induced Liver Injury: A Multidisciplinary Approach

This thesis presents an approach to expose relationships between immune mediated drug induced liver injury (IMDILI) and the three-dimensional structural features of toxic drug molecules and their metabolites. The series of analyses test the hypothesis that drugs which produce similar patterns of toxicity interact with targets within common toxicological pathways and that activation of the underlying mechanisms depends on structural similarity among toxic molecules. Spontaneous adverse drug reaction (ADR) reports were used to identify cases of IMDILI. Network map tools were used to compare the known and predicted protein interactions with each of the probe drugs to explore the interactions that are common between the drugs. The IMDILI probe set was then used to develop a pharmacophore model which became the starting point for identifying potential toxicity targets for IMDILI. Pharmacophore screening results demonstrated similarities between the probe IMDILI set of drugs and Toll-Like Receptor 7 (TLR7) agonists, suggesting TLR7 as a potential toxicity target. This thesis highlights the potential for multidisciplinary approaches in the study of complex diseases. Such approaches are particularly helpful for rare diseases where little knowledge is available, and may provide key insights into mechanisms of toxicity that cannot be gleaned from a single disciplinary study

Immune-Mediated Drug Induced Liver Injury: A Multidisciplinary Approach

This thesis presents an approach to expose relationships between immune mediated drug induced liver injury (IMDILI) and the three-dimensional structural features of toxic drug molecules and their metabolites. The series of analyses test the hypothesis that drugs which produce similar patterns of toxicity interact with targets within common toxicological pathways and that activation of the underlying mechanisms depends on structural similarity among toxic molecules. Spontaneous adverse drug reaction (ADR) reports were used to identify cases of IMDILI. Network map tools were used to compare the known and predicted protein interactions with each of the probe drugs to explore the interactions that are common between the drugs. The IMDILI probe set was then used to develop a pharmacophore model which became the starting point for identifying potential toxicity targets for IMDILI. Pharmacophore screening results demonstrated similarities between the probe IMDILI set of drugs and Toll-Like Receptor 7 (TLR7) agonists, suggesting TLR7 as a potential toxicity target. This thesis highlights the potential for multidisciplinary approaches in the study of complex diseases. Such approaches are particularly helpful for rare diseases where little knowledge is available, and may provide key insights into mechanisms of toxicity that cannot be gleaned from a single disciplinary study

Computer-aided approaches in drug design: the exigent way forward: dynamic perspectives into the mechanistic activities of small molecule inhibitors toward antiviral, antitubercular and anticancer therapeutic interventions.

Doctoral Degree. University of KwaZulu-Natal, Durban.The crucial role of CADD in the drug design process is now indisputable and has proven over the years that it can accelerate the discovery potential drug candidates while reducing the associated cost. Using knowledge and information about biological target or knowledge about a ligand with proven bioactivity, CADD, and its techniques can influence various drug discovery pipeline stages. The ability CADD approaches to elucidate drug-target interactions at the atomistic level allows for investigations of the mechanism of drugs' actions, revealing atomistic insights that influence drug design and improvement. CADD approaches also seek to augment traditional in vitro and in vivo experimental techniques and not replace them since CADD approaches can also allow modeling complex biological processes that hitherto seemed impossible to explore using experimental methods. According to the World Health Organization (WHO), featuring prominently in the top ten causes of death are cancer, lower respiratory tract infection, tuberculosis (TB), and viral infections such as HIV/AIDS. Collectively, these diseases are of global health concerns, considering a large number of associated deaths yearly. Over the years, several therapeutic interventions have been employed to treat, manage, or cure these diseases, including chemotherapy, surgery, and radiotherapy. Of these options, small molecule inhibitors have constituted an integral component in chemotherapy, thereby undoubtedly playing an essential role in patient management. Although significant success has been achieved using existing therapeutic approaches, the emergence of drug resistance and the challenges of associated adverse side effects has prompted the need for the drug design processes against these diseases to remain innovative, including combining existing drugs and establishing improved therapeutic options that could overcome resistance while maintaining minimal side effects to patients. Therefore, an exploration of drug target interactions towards unraveling mechanisms of actions as performed in the reports in this thesis are relevant since the molecular mechanism provided could form the basis for the design and identification of new therapeutic agents, improvement of the therapeutic activity of existing drugs, and also aid in the development of novel therapeutic strategies against these diseases of global health concern. Therefore the studies in this thesis employed CADD approaches to investigates molecular mechanisms of actions of novel therapeutic strategies directed towards some crucial therapeutics implicated in viral infections, tuberculosis, and cancer. Therapeutic targets studied included; SARS-CoV-2 RNA dependent RNA polymerase (SARS-CoV-2 RdRp), Human Rhinovirus B14 (HRV-B14) and human N-myristoyltransferases in viral infections, Dihydrofolate reductase (DHFR) and Flavin-dependent thymidylate synthase (FDTS) in TB, human variants of TCRCD1d, and Protein Tyrosine Phosphatase Receptor Zeta (PTPRZ) in cancer. The studies in this thesis is divided into three domains and begins with a thorough review of the concept of druggability and drug-likeness since the crux of the subsequent reports revolved around therapeutic targets and their inhibitions by small molecule inhibitors. This review highlights the principles of druggability and drug-likeness while detailing the recent advancements in drug discovery. The review concludes by presenting the different computational, highlighting their reliability for predictive analysis. In the first domain of the research, we sought to unravel the inhibitory mechanism of some small molecule inhibitors against some therapeutic targets in viral infections by explicitly focusing on the therapeutic targets; SARS-CoV-2 RdRp, HRV-B14, and N-myristoyltransferase. Therapeutic targeting of SARS-CoV-2 RdRp has been extensively explored as a viable approach in the treatment of COVID-19. By examining the binding mechanism of Remdesivir, which hitherto was unclear, this study sought to unravel the structural and conformational implications on SARS-CoV-2 RdRp and subsequently identify crucial pharmacophoric moieties of Remdesivir required for its inhibitory potency. Computational analysis showed that the modulatory activity of Remdesivir is characterized by an extensive array of high-affinity and consistent molecular interactions with specific active site residues that anchor Remdemsivir within the binding pocket for efficient binding. Results also showed that Remdesivir binding induces minimal individual amino acid perturbations, subtly interferes with deviations of C-α atoms, and restricts the systematic transition of SARS-CoV-2 RdRp from the “buried” hydrophobic region to the “surface exposed” hydrophilic region. Based on observed high-affinity interactions with SARS-CoV-2 RdRp, a pharmacophore model was generated, which showcased the crucial functional moieties of Remdesivir. The pharmacophore was subsequently employed for virtual screening to identify potential inhibitors of SARS-CoV-2 RdRp. The structural insights and the optimized pharmacophoric model provided would augment the design of improved analogs of Remdesivir that could expand treatment options for COVID-19. The next study sought to explore the therapeutic targeting of human rhinoviruses (HRV) amidst challenges associated with the existence of a wide variety of HRV serotypes. By employing advanced computational techniques, the molecular mechanism of inhibition of a novel benzothiophene derivative that reportedly binds HRV-B14 was investigated. An analysis of the residue-residue interaction profile revealed of HRV upon the benzothiophene derivative binding revealed a distortion of the hitherto compacted and extensively networked HRV structure. This was evidenced by the fewer inter-residue hydrogen bonds, reduced van der Waals interactions, and increased residue flexibility. However, a decrease in the north-south wall's flexibility around the canyon region also suggested that the benzothiophene derivative's binding impedes the “breathing motion” of HRV-B14; hence its inhibition. The next study in the first domain of the research investigated the structural and molecular mechanisms of action associated with the dual inhibitory activity of IMP-1088. This novel compound reportedly inhibits human N-myristoyltransferase subtypes 1 and 2 towards common cold therapy. This is because it has emerged that the pharmacological inhibition of Nmyristoyltransferase is an efficient non-cytotoxic strategy to completely thwart the replication process of rhinovirus toward common cold treatment. Using augmentative computational and nanosecond-based analyses, findings of the study revealed that the steady and consistent interactions of IMP-1088 with specific residues; Tyr296, Phe190, Tyr420, Leu453, Gln496, Val181, Leu474, Glu182, and Asn246, shared within the binding pockets of both HNMT subtypes, in addition to peculiar structural changes account for its dual inhibitory potency. Findings thus unveiled atomistic and structural perspectives that could form the basis for designing novel dualacting inhibitors of N-myristoyltransferase towards common cold therapy. In the second domain of the research, the mechanism of action of some small molecule inhibitors against DHFR, FDTS, and Mtb ATP synthase in treating tuberculosis is extensively investigated and reportedly subsequently. To begin with, the dual therapeutic targeting of crucial enzymes in the folate biosynthetic pathway was explored towards developing novel treatment methods for TB. Therefore, the study investigated the molecular mechanisms and structural dynamics associated with dual inhibitory activity of PAS-M against both DHFR and FDTS, which hitherto was unclear. MD simulations revealed that PAS-M binding towards DHFR and FDTS is characterized by a recurrence of strong conventional hydrogen bond interactions between a peculiar site residue the 2-aminov decahydropteridin-4-ol group of PAS-M. Structural dynamics of the bound complexes of both enzymes revealed that, upon binding, PAS-M is anchored at the entrance of hydrophobic pockets by a strong hydrogen bond interaction while the rest of the structure gains access to deeper hydrophobic residues to engage in favorable interactions. Further analysis of atomistic changes of both enzymes showed increased C-α atom deviations and an increase C-α atoms radius of gyration consistent with structural disorientations. These conformational changes possibly interfered with the enzymes' biological functions and hence their inhibition as experimentally reported. Additionally, in this domain, the therapeutic targeting of the ATP machinery of Mtb by Bedaquiline (BDQ) was explored towards unravelling the structures and atomistic perspectives that account for the ability of BDQ to selectively inhibits mycobacterial F1Fo-ATP synthase via its rotor c-ring. BDQ is shown to form strong interaction with Glu65B and Asp32B and, consequently, block these residues' role in proton binding and ion. BDQ binding was also revealed to impede the rotatory motion of the rotor c-ring by inducing a compact conformation on the ring with its bulky structure. Complementary binding of two molecules of BDQ to the rotor c-ring, proving that increasing the number of BDQ molecule enhances inhibitory potency. The last study in this research domain investigated the impact of triple mutations (L59V, E61D, and I66M) on the binding of BDQ to Mtb F1F0 ATP-synthase. The study showed that the mutations significantly impacted the binding affinity of BDQ, evidenced by a decrease in the estimated binding free energy (ΔG). Likewise, the structural integrity and conformational architecture of F1F0 ATP-synthase was distorted due to the mutation, which could have interfered with the binding of BDQ. The third domain of the research in this thesis investigated some small molecule inhibitors' inhibitory mechanism against some therapeutic targets in cancer, specifically PTPRZ and hTCRvi CD1d. Studies in the third domain of the research in the thesis began with the investigation of the investigation of the inhibitory mechanism of NAZ2329, an allosteric inhibitor of PTPRZ, by specifical investigating its binding effect on the atomic flexibility of the WPD-loop. Having been established as crucial determinant of the catalytic activity of PTPRZ an implicated protein in glioblastoma cells, its successfully therapeutic modulation could present a viable treatment option in glioblastoma. Structural insights from an MD simulation revealed that NAZ2329 binding induces an open conformation of the WPD-loop which subsequently prevents the participation of the catalytic aspartate of PTPRZ from participating in catalysis hence inhibiting the activity of PTPRZ. A pharmacophore was also created based of high energy contributing residues which highlighted essential moieties of NAZ2329 and could be used in screening compound libraries for potential inhibitors of PTPRZ. A second study in this domain sought to explore how structural modification could improve a therapeutic agent's potency from an atomistic perspective. This study was based on an earlier report in which the incorporation of a hydrocinnamoyl ester on C6’’ and C4-OH truncation of the sphingoid base of KRN7000 generated a novel compound AH10-7 high therapeutic potency and selectivity in human TCR-CD1d and subsequently results in the activation of invariant natural killer T cells (iNKT). The hydrocinnamoyl ester moiety was shown to engage in high-affinity interactions, possibly accounting for the selectivity and higher potency of AH10-7. Molecular and structural perspectives provided could aid in the design of novel α-GalCer derivatives for cancer immunotherapeutics. Chapter 3 provides theoretical insights into the various molecular modeling tools and techniques employed to investigate the various conformational changes, structural conformations, and the associated mechanism of inhibitions of the studied inhibitors towards viral, tuberculosis, and cancer therapy. Chapter 4 provided sufficient details on druggability and drug-likeness principles and their recent advancements in the drug discovery field. The study also presents the different computational tools and their reliability of predictive analysis in the drug discovery domain. It thus provides a comprehensive guide for computational-oriented drug discovery research. Chapter 5 provides an understanding of the binding mechanism of Remdesivir, providing structural and conformational implications on SARS-CoV-2 RdRp upon its binding and identifying its crucial pharmacophoric moieties. Chapter 6 explains the mechanism of inhibition of a novel benzothiophene derivative, revealing its distortion of the native extensively networked and compact residue profile. Chapter 7 unravels molecular and structural bases behind this dual inhibitory potential of the novel inhibitor IMP-1088 toward common cold therapy using augmentative computational and cheminformatics methods. The study also highlights the pharmacological propensities of IMP- 1088. Chapter 8 unravels the molecular mechanisms and structural dynamics of the dual inhibitory activity of PAS-M towards DHFR and FDTS. Chapter 9 reports the structural dynamics and atomistic perspectives that account for the reported ability of BDQ to halt the ion shuttling ability of mycobacterial c-ring. Chapter 10 presents the structural dynamics and conformational changes that occur on Mtb F1F0 ATP-synthase binding as a result of the triple mutations using molecular dynamics simulations, free energy binding, and residue interaction network (RIN) analyses. Chapter 11 explored the impact of NAZ2329, a recently identified allosteric inhibitor of Protein Tyrosine Phosphatase Receptor Zeta (PTPRZ), on the atomic flexibility of the WPD-loop, an essential loop in the inhibition of PTPRZ. The study also presents the drug-likeness of NAZ2329 using in silico techniques and its general inhibitory mechanism. Chapter 12 provides atomistic insights into the structural dynamics and selective mechanisms of AH10-7 for human TCR-CD1d towards activating iNKT cells. The studies in this thesis collectively present a thorough and comprehensive in silico perspective that characterizes the pharmacological inhibition of some known therapeutic targets in viral infections, tuberculosis, and cancer. The augmentative integration of computational methods to provide structural insights could help design highly selective inhibitors of these therapeutic targets. Therefore, the findings presented are fundamental to the design and development of next generation lead compounds with improved therapeutic activities and minimal toxicities

In silico search for novel bacterial inhibitors targeting RNA polymerase switch region

The arise of antibiotic-resistant bacterial strains in an alarming rate has increased the interest in the discovery of novel antibiotics. The rifamycins are a valuable class of antibiotics that target bacterial Ribonucleic Acid polymerase (RNAP) and are considered the first-line treatment for tuberculosis. Consequently, bacterial strains resistant to rifamycin constitute a public health threat. RNAP switch region is an attractive target for the development of new antibacterial agents as it lies away from the rifamycin binding region and thus the compounds that target the switch region would not show cross-resistance with rifamycins. In this work, we developed a virtual screening pipeline to identify new bacterial RNAP inhibitors that target the enzyme switch region. The screening pipeline involved docking of the designated libraries using the Maestro Glide docking tool, and the compounds with the best docking scores were submitted for binding free energy calculations using the molecular mechanics-generalised born surface area (MM-GBSA)-based method. Moreover, a quantitative structure-activity relationship (QSAR) model was developed, and it was applied to predict the biological activity of the compounds with the most favourable calculated binding free energies. Based on the results of docking, MM-GBSA binding free energies and the activities predicted by the QSAR model, the most promising compounds were chosen to be evaluated by molecular dynamics (MD) simulations. The results of the MD simulation of each docked candidate compound in the RNAP binding site were compared with the MD simulations carried out with the apo protein and with a reference co-crystallized ligand in the RNAP binding site. The candidate compounds showing comparable binding to the RNAP site to the reference ligand were selected for further biological testing