Priyanka Suryadevara

Thiazole–aminopiperidine hybrid analogues: Design and synthesis of novel Mycobacterium tuberculosis GyrB inhibitors

A series of ethyl-4-(4-((substituted benzyl)amino)piperidin-1-yl)-2-(phenyl/pyridyl)thiazole-5-carboxylates was designed by molecular hybridization and synthesized from aryl thioamides in five steps. The compounds were evaluated for their in vitro Mycobacterium smegmatis (MS) GyrB ATPase assay, Mycobacterium tuberculosis (MTB) DNA gyrase super coiling assay, antituberculosis activity and cytotoxicity. Among the twenty four compounds studied, ethyl-4-(4-((4-fluorobenzyl)amino)piperidin-1-yl)-2-phenylthiazole-5-carboxylate (14) was found to be the promising compound which showed activity against all test with MS GyrB IC50 of 24.0 ± 2.1 μM, 79% inhibition of MTB DNA gyrase at 50 μM, MTB MIC of 28.44 μM, and not cytotoxic at 50 μM.

Development of novel N-linked aminopiperidine-based mycobacterial DNA gyrase B inhibitors: scaffold hopping from known antibacterial leads.

DNA gyrase of Mycobacterium tuberculosis (MTB) is a type II topoisomerase that ensures the regulation of DNA topology and has been genetically demonstrated to be a bactericidal drug target. We present the discovery and optimisation of a novel series of mycobacterial DNA gyrase inhibitors with a high degree of specificity towards the mycobacterial ATPase domain. Compound 5-fluoro-1-(2-(4-(4-(trifluoromethyl)benzylamino)piperidin-1-yl)ethyl)indoline-2,3-dione (17) emerged as the most potent lead, exhibiting inhibition of MTB DNA gyrase supercoiling assay with an IC50 (50% inhibitory concentration) of 3.6 ± 0.16 μM, a Mycobacterium smegmatis GyrB IC50 of 10.6 ± 0.6 μM, and MTB minimum inhibitory concentrations of 6.95 μM and 10 μM against drug-sensitive (MTB H37Rv) and extensively drug-resistant strains, respectively. Furthermore, the compounds did not show any signs of cardiotoxicity in zebrafish ether-à-go-go-related gene (zERG), and hence constitute a major breakthrough among the otherwise cardiotoxic N-linked aminopiperidine analogues.

4-Aminoquinoline derivatives as novel Mycobacterium tuberculosis GyrB inhibitors: Structural optimization, synthesis and biological evaluation.

Mycobacterial DNA gyrase B subunit has been identified to be one of the potentially underexploited drug targets in the field of antitubercular drug discovery. In the present study, we employed structural optimization of the reported GyrB inhibitor resulting in synthesis of a series of 46 novel quinoline derivatives. The compounds were evaluated for their in vitro Mycobacterium smegmatis GyrB inhibitory ability and Mycobacterium tuberculosis DNA supercoiling inhibitory activity. The antitubercular activity of these compounds was tested over Mtb H37Rv strain and their safety profile was checked against mouse macrophage RAW 264.7 cell line. Among all, three compounds (23, 28, and 53) emerged to be active displaying IC₅₀ values below 1 μM against Msm GyrB and were found to be non-cytotoxic at 50 μM concentration. Compound 53 was identified to be potent GyrB inhibitor with 0.86 ± 0.16 μM and an MIC (minimum inhibitory concentration) of 3.3 μM. The binding affinity of this compound towards GyrB protein was analysed by differential scanning fluorimetry which resulted in a positive shift of 3.3 °C in melting temperature (Tm) when compared to the native protein thereby reacertaining the stabilization effect of the compound over protein.

Exploring the gyrase ATPase domain for tailoring newer anti-tubercular drugs: hit to lead optimization of a novel class of thiazole inhibitors.

Gyrase ATPase domain, the pharmaceutical underexploited segment of DNA gyrase, the sole Type II topoisomerase present in Mycobacterium tuberculosis represents an attractive target for anti-tubercular drug discovery. Here we report, the development of a novel series of MTB DNA gyraseB inhibitor identified through a medium throughput screening (MTS) of BITS in-house chemical library (3000 compounds). The MTS hit was further remodeled by chemical synthesis to identify the most potent analogue 27 exhibiting an in vitro gyrB inhibitory IC50 of 0.15 μM. The series also demonstrated well correlating gyrase super coiling activity and in vitro anti-mycobacterial potency against MTB H37Rv strain. Furthermore the compounds displayed good safety profile in their subsequent cytotoxicity and hERG toxicity evaluations, to be worked out from a pharmaceutical point of view as potential anti-tubercular agents.

Structure-guided design and development of novel benzimidazole class of compounds targeting DNA gyraseB enzyme of Staphylococcus aureus

The gyraseB subunit of Staphylococcus aureus DNA gyrase is a well-established and validated target though less explored for the development of novel antimicrobial agents. Starting from the available structural information in PDB (3TTZ), we identified a novel series of benzimidazole used as inhibitors of DNA gyraseB with low micromolar inhibitory activity by employing structure-based drug design strategy. Subsequently, this chemical class of DNA gyrase inhibitors was extensively investigated biologically through in vitro assays, biofilm inhibition assays, cytotoxicity, and in vivo studies. The binding affinity of the most potent inhibitor 10 was further ascertained biophysically through differential scanning fluorimetry. Further, the most potent analogues did not show any signs of cardiotoxicity in Zebra fish ether-a-go-go-related gene (zERG), a major breakthrough among the previously reported cardiotoxic gyraseB inhibitors.

Depletion of M. tuberculosis GlmU from Infected Murine Lungs Effects the Clearance of the Pathogen.

M. tuberculosis N-acetyl-glucosamine-1-phosphate uridyltransferase (GlmUMtb) is a bi-functional enzyme engaged in the synthesis of two metabolic intermediates N-acetylglucosamine-1-phosphate (GlcNAc-1-P) and UDP-GlcNAc, catalyzed by the C- and N-terminal domains respectively. UDP-GlcNAc is a key metabolite essential for the synthesis of peptidoglycan, disaccharide linker, arabinogalactan and mycothiols. While glmU Mtb was predicted to be an essential gene, till date the role of GlmUMtb in modulating the in vitro growth of Mtb or its role in survival of pathogen ex vivo / in vivo have not been deciphered. Here we present the results of a comprehensive study dissecting the role of GlmUMtb in arbitrating the survival of the pathogen both in vitro and in vivo. We find that absence of GlmUMtb leads to extensive perturbation of bacterial morphology and substantial reduction in cell wall thickness under normoxic as well as hypoxic conditions. Complementation studies show that the acetyl- and uridyl- transferase activities of GlmUMtb are independently essential for bacterial survival in vitro, and GlmUMtb is also found to be essential for mycobacterial survival in THP-1 cells as well as in guinea pigs. Depletion of GlmUMtb from infected murine lungs, four weeks post infection, led to significant reduction in the bacillary load. The administration of Oxa33, a novel oxazolidine derivative that specifically inhibits GlmUMtb, to infected mice resulted in significant decrease in the bacillary load. Thus our study establishes GlmUMtb as a strong candidate for intervention measures against established tuberculosis infections.

Discovery and Structure Optimization of a Series of Isatin Derivatives as Mycobacterium tuberculosis Chorismate Mutase Inhibitors

In this study, the crystal structure of the Mycobacterium tuberculosis (MTB) enzyme chorismate mutase (CM) bound to transition state analogue (PDB: 2FP2) was used as a framework for virtual screening of the BITS‐Pilani in‐house database (2500 compounds) to identify new scaffold. We identified isatin as novel small molecule MTB CM inhibitors; further twenty‐four isatin derivatives were synthesized and evaluated in vitro for their ability to inhibit MTB CM, and activity against M. tuberculosis as steps towards the derivation of structure–activity relationships (SAR) and lead optimization. Compound 3‐(4‐nitrobenzylidene)indolin‐2‐one, 24 emerged as the most promising lead with an IC50 of 1.01 ± 0.22 μm for purified CM and MIC of 23.5 μm for M. tuberculosis, with little or no cytotoxicity.

Identification of potential Mycobacterium tuberculosis topoisomerase I inhibitors: A study against active, dormant and resistant tuberculosis

Mycobacterium tuberculosis (Mtb) topoisomerase I (Topo I), involved in the relaxation of negatively supercoiled DNA, plays an important role in the viability of pathogen Mtb. Being one of the most significant enzymes; it also takes part in crucial biological pathways such as transcription and replication of the pathogen. The present study aims at the development of Mtb Topo I 3D protein structure which in turn was employed for the virtual screening of compound libraries in a process of identification of a hit molecule. The identified hit, hydroxycamptothecin, was active at 6.25 μM which was further derivatized synthetically into fifteen novel analogues. Among these, four compounds (3b, 3g, 3h and 3l) emerged to be active displaying IC50 values ranging from 2.9 to 9.3 μM against Mtb Topo I and were non-cytotoxic at 25 μM. These four compounds also proved their efficacy when tested against active, dormant and resistant forms of Mtb. The most potent inhibitor 3b was screened for in vivo anti-mycobacterial activity using zebrafish model and was found to be more effective when compared to first line anti-tubercular drugs, isoniazid and rifampicin. The binding affinity of this compound towards Mtb Topo I was analyzed by differential scanning fluorimetry which resulted in a positive shift in melting temperature when compared to the native protein thereby proving its stabilization effect over protein.

Computational Sampling and Simulation Based Assessment of Novel Mycobacterium tuberculosis Glutamine Synthetase Inhibitors: Study Involving Structure Based Drug Design and Free Energy Perturbation.

The highly persistent nature of Mycobacterium tuberculosis can be attributed to its lipophilic cell wall which acts as a major barrier in the process of drug discovery against tuberculosis. Glutamine synthetase plays a major role in nitrogen metabolism and cell wall biosynthesis of pathogenic mycobacteria. The current review focuses on the structural and functional aspects of Mtb glutamine synthetase and an overview of its reported inhibitors till date. Also in the present study, we employed a computational structure based drug design protocol for identifying novel inhibitors against Mtb glutamine synthetase (MtbGS). A total of 12 hits were identified based on e-pharmacophore related search and virtual screening, which were further tested for their in vitro MtbGS inhibitory activity. Three compounds (compound 6, 1 and 12) were found with IC50 less than 5 µM, of which compound 6 being top active with IC50 of 2.124 µM. Differential scanning fluorimetry studies were employed so as to measure the thermal stability of the protein complexed with the most active compound. Also the protein complexes with top three active compounds were subjected for molecular dynamics simulations to study their binding pattern and stabilization effect. The solvation free energies were also determined for these compounds, undertaking free energy perturbation studies, which can be used further for lead optimization in the process of anti-tubercular drug discovery targeting Mtb glutamine synthetase.

Design and Biological Evaluation of Furan/Pyrrole/Thiophene‐2‐carboxamide Derivatives as Efficient DNA GyraseB Inhibitors of Staphylococcus aureus

DNA topoisomerases are well‐validated targets in micro‐organisms. DNA gyraseB is one of the most important enzymes among them as per their clinical importance. In earlier study, a novel lead 4‐((4‐(furan‐2‐carboxamido)phenyl)amino)‐4‐oxobutanoic acid was identified as inhibitor against DNA gyraseB with an IC50 of 12.88 ± 1.39 μm. Subsequently, analogues of this lead were developed and evaluated through in vitro assays and in vivo studies. Among the 24 analogues, compound 22 was found to be the top hit with an improved DNA gyraseB activity of 5.35 ± 0.61 μm, and the binding affinity of this compound was further ascertained biophysically through differential scanning fluorimetry. The most potent ligand did not show any signs of cardiotoxicity in zebra fish ether‐ago‐go‐related gene, ascertaining the safety profile of this series a breakthrough among the previously reported cardiotoxic gyraseB inhibitors.