Toxicological Profiling of Potential Shikimate Kinase Inhibitors Against Mycobacterium tuberculosis.

Over the last decade, Mycobacterium tuberculosis has mutated into a putative 'superbug', as treatments against it have failed due to increasing antimicrobial resistance. As a result, the rising incidence of multidrug-resistant tuberculosis (MDR-TB) is posing a significant public health threat, thus, the need to develop effective drugs for MDR-TB has become an urgent priority. To identify new drug candidates for the treatment of MDR-TB, the present study was based on mycobacterial shikimate kinase (MtSK) as the pharmacological target. One hundred potential MtSK inhibitors were identified from literature and database searches to identify compounds that were designed to specifically function as MtSK antagonists. The ADME properties of these compounds were evaluated by using the SwissADME web tool. ProTox-II software was also used to investigate any potential endocrine disrupting effects, mediated through their interaction with oestrogenic and/or androgenic receptors. This study also aimed to predict LD50 values of potential drug candidates that would be active against the standard H37Rv strain of M. tuberculosis, by using the ProTox-II in silico tool. The molecules for which no structural hazard alerts were identified with these software tools were further subjected to molecular docking analyses and molecular dynamic simulations to estimate their ability to interact with the MtSK enzyme. Preliminary results from SwissADME indicated that 30 molecules were drug-like, due to their physicochemical and pharmacokinetic properties. However, subsequent analysis with ToxTree and ProTox-II indicated that only three of these 30 drug-like molecules were suitable for taking forward into further in vitro experiments. This study, which is based on the use of commonly used open-source in silico tools, identified new MtSK ligands for potential use in the development of new drugs for the therapeutic management of tuberculosis. An initial prediction of their safety profile was also generated.

Design, synthesis and biological evaluation of substituted flavones and aurones as potential anti-influenza agents.

We designed a series of substituted flavones and aurones as non-competitive H1N1 neuraminidase (NA) inhibitors and anti-influenza agents. The molecular docking studies showed that the designed flavones and aurones occupied 150-cavity and 430-cavity of H1N1-NA. We then synthesized these compounds and evaluated these for cytotoxicity, reduction in H1N1 virus yield, H1N1-NA inhibition and kinetics of inhibition. The virus yield reduction assay and H1N1-NA inhibition assay demonstrated that the compound 1f (4-methoxyflavone) had the lowest EC50 of 9.36 nM and IC50 of 8.74 µM respectively. Moreover, kinetic studies illustrated that compounds 1f and 2f had non-competitive inhibition mechanism.

Structure-aided drug development of potential neuraminidase inhibitors against pandemic H1N1 exploring alternate binding mechanism.

The rate of mutability of pathogenic H1N1 influenza virus is a threat. The emergence of drug resistance to the current competitive inhibitors of neuraminidase, such as oseltamivir and zanamivir, attributes to a need for an alternative approach. The design and synthesis of new analogues with alternate approach are particularly important to identify the potential neuraminidase inhibitors which may not only have better anti-influenza activity but also can withstand challenge of resistance. Five series of scaffolds, namely aurones (1a-1e), pyrimidine analogues (2a-2b), cinnamic acid analogues (3a-3k), chalcones (4a-4h) and cinnamic acid linkages (5a-5c), were designed based on virtual screening against pandemic H1N1 virus. Molecular modelling studies revealed that the designed analogues occupied 430-loop cavity of neuraminidase. Docking of sialic acid in the active site preoccupied with the docked analogues, i.e. in 430-loop cavity, resulted in displacement of sialic acid from its native pose in the catalytic cavity. The favourable analogues were synthesized and evaluated for the cytotoxicity and cytopathic effect inhibition by pandemic H1N1 virus. All the designed analogues resulting in displacement of sialic acid suggested alternate binding mechanism. Overall results indicated that aurones can be measured best among all as potential neuraminidase inhibitor against pandemic H1N1 virus.

A Computational Model for Docking of Noncompetitive Neuraminidase Inhibitors and Probing their Binding Interactions with Neuraminidase of Influenza Virus H5N1.

BACKGROUND: With cases of emergence of drug resistance to the current competitive inhibitors of neuraminidase (NA) such as oseltamivir and zanamavir, there is a present need for an alternative approach in the treatment of avian influenza. With this in view, some flavones and chalcones were designed based on quercetin, the most active naturally occurring noncompetitive inhibitor. OBJECTIVE: We attempt to understand the binding of quercetin to H5N1-NA, and synthetic analogs of quercetin namely flavones and its precursors the chalcones using computational tools. METHODS: Molecular docking was done using Libdock. Molecular dynamics (MD) simulations were performed using Amber14. We synthesized the two compounds; their structures were confirmed by infrared spectroscopy, 1H-NMR, and mass spectrometry. These molecules were then tested for H5N1-NA inhibition and kinetics of inhibition. RESULTS: Molecular docking studies yielded two compounds i.e., 4'-methoxyflavone and 2'-hydroxy-4-methoxychalcone, as promising leads which identified them as binders of the 150-cavity of NA. Furthermore, MD simulation studies revealed that quercetin and the two compounds bind and hold the 150 loop in its open conformation, which ultimately perturbs the binding of sialic acid in the catalytic site. Estimation of the free energy of binding by MM-PBSA portrays quercetin as more potent than chalcone and flavone. These molecules were then determined as non-competitive inhibitors from the Lineweaver-Burk plots rendered from the enzyme kinetic studies. CONCLUSION: We conclude that non-competitive type of inhibition, as shown in this study, can serve as an effective method to block NA and evade the currently seen drug resistance.

Drug susceptibility of influenza A/H3N2 strains co-circulating during 2009 influenza pandemic: first report from Mumbai.

OBJECTIVE: From its first instance in 1977, resistance to amantadine, a matrix (M2) inhibitor has been increasing among influenza A/H3N2, thus propelling the use of oseltamivir, a neuraminidase (NA) inhibitor as a next line drug. Information on drug susceptibility to amantadine and neuraminidase inhibitors for influenza A/H3N2 viruses in India is limited with no published data from Mumbai. This study aimed at examining the sensitivity to M2 and NA inhibitors of influenza A/H3N2 strains isolated from 2009 to 2011 in Mumbai. METHODS: Nasopharyngeal swabs positive for influenza A/H3N2 virus were inoculated on Madin-Darby canine kidney (MDCK) cell line for virus isolation. Molecular analysis of NA and M2 genes was used to detect known mutations contributing to resistance. Resistance to neuraminidase was assayed using a commercially available chemiluminescence based NA-Star assay kit. RESULTS: Genotypically, all isolates were observed to harbor mutations known to confer resistance to amantadine. However, no know mutations conferring resistance to NA inhibitors were detected. The mean IC50 value for oseltamivir was 0.25 nM. One strain with reduced susceptibility to the neuraminidase inhibitor (IC50=4.08 nM) was isolated from a patient who had received oseltamivir treatment. Phylogenetic analysis postulate the emergence of amantadine resistance in Mumbai may be due to genetic reassortment with the strains circulating in Asia and North America. CONCLUSIONS: Surveillance of drug susceptibility helped us to identify an isolate with reduced sensitivity to oseltamivir. Therefore, we infer that such surveillance would help in understanding possible trends underlying the emergence of resistant variants in humans.

Rational development of neuraminidase inhibitor as novel anti-flu drug.

The highly pathogenic influenza virus has caused many human fatalities and poses an increasing pandemic threat. Neuraminidase inhibitors such as oseltamivir and zanamivir have been widely used in the treatment and have gained remarkable success. Although, they are effective in prevention of influenza; the concern for drug resistance still remains a question. Recently, the availability of crystal structures of the enzyme gave a new trend to the structure based drug designing of neuraminidase inhibitors. The article reviews a detailed understanding of the structural features within neuraminidase enzyme which turnouts to be crucial for future drug development. In depth analysis for the newly proposed spots within the 150 and 430-loop regions in N1 makes it distinguishable among the subtypes. Further we have discussed the various computational studies carried out in optimizing the designing of neuraminidase inhibitors thereby providing new clues to modify the currently available drugs.

De novo design of 7-aminocoumarin derivatives as novel falcipain-3 inhibitors.

The availability of the crystal structure of falcipain-3, knowledge of the peptides carrying the 7-aminocoumarin moiety as falcipain-3 ligands/substrates, and a need for new antimalarial agents stimulated us to look at the possibility of finding some novel falcipain-3 inhibitors. In this paper, we report the effect of substitution at the 7-amino position of the coumarin nucleus on the inhibition of falcipain-3, which is a well-validated antimalarial target. The de novo drug design was assisted by QSAR studies that shed light on the binding patterns of known and the newly designed inhibitors, thus taking this discovery process to the next level.