C. Gopi Mohan

Inhibition of acetylcholinesterase by coumarins: The case of coumarin 106

In this contribution, from a coumarin library consisting of 29 compounds including natural and synthetic derivatives, an active acetylcholinesterase (AChE) inhibitor (coumarin 106) was found. This circumstance leaded us to continue with the pharmacological characterization of coumarin 106. The first study with the coumarin library was performed using a 96-microtiter well plate assay based on Ellmans reaction. Coumarins were assayed at 5 and 30 microM, and coumarin 106 was found the most active inhibitor at both concentrations. The follow-up analysis using kinetic studies demonstrated that coumarin 106 displays mixed-type AChE inhibition with a pIC(50)=4.97+/-0.09 and K(i)=2.36+/-0.17 microM. The ability of this molecule to interact with AChE was further confirmed through computational studies, in which a primary binding was proved to occur at the active gorge site, while a secondary binding was demonstrated at the peripheral anionic site. Also, coumarin 106 was shown to inhibit butyrylcholinesterase (BChE) with slightly lower potency (pIC(50)=4.56+/-0.06), and found to be non-toxic in Caco-2 cells. The combination of these findings makes coumarin 106 an attractive molecule for further investigation. This is the first report where AChE inhibitory activity has been associated with coumarin 106, and proof has been given of its convenience as a lead molecule.

Antibiotic resistance in Pseudomonas aeruginosa and alternative therapeutic options

Pseudomonas aeruginosa is a leading cause of nosocomial infections and is responsible for ∼10% of all hospital-acquired infections worldwide. It continues to pose a therapeutic challenge because of the high rate of morbidity and mortality associated with it and the possibility of development of drug resistance during therapy. Standard antibiotic regimes against P. aeruginosa are increasingly becoming ineffective due to the rise in drug resistance. With the scope for developing new antibiotics being limited, alternative treatment options are gaining more and more attention. A number of recent studies reported complementary and alternative treatment options to combat P. aeruginosa infections. Quorum sensing inhibitors, phages, probiotics, anti-microbial peptides, vaccine antigens and antimicrobial nanoparticles have the potential to act against drug resistant strains. Unfortunately, most studies considering alternative treatment options are still confined in the pre-clinical stages, although some of these findings have tremendous potential to be turned into valuable therapeutics. This review is intended to raise awareness of several novel approaches that can be considered further for combating drug resistant P. aeruginosa infections.

Computer-assisted methods in chemical toxicity prediction.

In Silico predictive ADME/Tox screening of compounds is one of the hottest areas in drug discovery. To provide predictions of compound drug-like characteristics early in modern drug-discovery decision making, computational technologies have been widely accepted to develop rapid high throughput in silico ADMET analysis. It is widely perceived that the early screening of chemical entities can significantly reduce the expensive costs associated with late stage failures of drugs due to poor ADME/Tox properties. Drug toxic effects are broadly defined to include toxicity, mutagenicity, carcinogenicity, teratogenicity, neurotoxicity and immunotoxicity. Toxicity prediction techniques and structure-activity relationships relies on the accurate estimation and representation of physico-chemical and toxicological properties. This review highlights some of the freely and commercially available softwares for toxicity predictions. The information content can be utilized as a guide for the scientists involved in the drug discovery arena.

Antiobesity and lipid lowering effects of Glycyrrhiza chalcones: experimental and computational studies.

Twelve flavonoids (1-12), isolated from Glycyrrhiza glabra roots were evaluated for their pancreatic lipase (PL) inhibitory activity in vitro. The structures of the isolated compounds were elucidated by spectroscopic methods. Amongst all the compounds 7, 8, 10 and 11 showed strong inhibition against PL with IC(50) values of 7.3 μM, 35.5 μM, 14.9 μM and 37.6 μM, respectively. Molecular docking studies on the most active compound 7 revealed that it binds with the key amino acid residues of the PL active site. In silico absorption, distribution, metabolism and excretion (ADME) parameters were also computed on the active compounds to determine their preliminary pharmacokinetic properties. Further, investigations were carried out to determine the antiobesity and lipid lowering effects of 7 and 10 in high fat diet (HFD) fed male SD rats. In the rats supplemented with compound 7 the body weight increase was only 23.2±3.6 g as compared to 64.2±0.5 g in the HFD control group while in the rats treated with compound 10 showed 23.2±3.6 g weight gain only. Compound 7 decreased the levels of plasma total cholesterol (TC) to 84.6±1.4 mg/dl and plasma total triglycerides (TG) to 128.8±6.0 mg/dl. Compound 10 also lowered the plasma TC and TG levels considerably. The results indicate the potential of the chalcone scaffold as a source of PL inhibitors for preventing obesity.

Synthesis, biological evaluation and molecular docking studies of stellatin derivatives as cyclooxygenase (COX-1, COX-2) inhibitors and anti-inflammatory agents

Stellatin (4), isolated from Dysophylla stellata is a cyclooxygenase (COX) inhibitor. The present study reports the synthesis and biological evaluation of new stellatin derivatives for COX-1, COX-2 inhibitory and anti-inflammatory activities. Eight derivatives showed more pronounced COX-2 inhibition than stellatin and, 17 and 21 exhibited the highest COX-2 inhibition. They also exhibited the significant anti-inflammatory activity in TPA-induced mouse ear edema assay and their anti-inflammatory effects were more than that of stellatin and indomethacin at 0.5mg/ear. The derivatives were further evaluated for antioxidant activity wherein 16 and 17 showed potent free radical scavenging activity against DPPH and ABTS radicals. Molecular docking study revealed the binding orientations of stellatin and its derivatives into the active sites of COX-1 and COX-2 and thereby helps to design the potent inhibitors.

Exploration of Natural Compounds as Sources of New Bifunctional Scaffolds Targeting Cholinesterases and Beta Amyloid Aggregation: The Case of Chelerythrine

The presented project started by screening a library consisting of natural and natural based compounds for their acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory activity. Active compounds were chemically clustered into groups and further tested on the human cholinesterases isoforms. The aim of the presented study was to identify compounds that could be used as leads to target two key mechanisms associated with the ADs pathogenesis simultaneously: cholinergic depletion and beta amyloid (Aβ) aggregation. Berberin, palmatine and chelerythrine, chemically clustered in the so-called isoquinoline group, showed promising cholinesterase inhibitory activity and were therefore further investigated. Moreover, the compounds demonstrated moderate to good inhibition of Aβ aggregation as well as the ability to disaggregate already preformed Aβ aggregates in an experimental set-up using HFIP as promotor of Aβ aggregates. Analysis of the kinetic mechanism of the AChE inhibition revealed chelerythrine as a mixed inhibitor. Using molecular docking studies, it was further proven that chelerythrine binds on both the catalytic site and the peripheral anionic site (PAS) of the AChE. In view of this, we went on to investigate its effect on inhibiting Aβ aggregation stimulated by AChE. Chelerythrine showed inhibition of fibril formation in the same range as propidium iodide. This approach enabled for the first time to identify a cholinesterase inhibitor of natural origin-chelerythrine-acting on AChE and BChE with a dual ability to inhibit Aβ aggregation as well as to disaggregate preformed Aβ aggregates. This compound could be an excellent starting point paving the way to develop more successful anti-AD drugs.

Discovery of dual binding site acetylcholinesterase inhibitors identified by pharmacophore modeling and sequential virtual screening techniques

Dual binding site acetylcholinesterase (AChE) inhibitors are promising for the treatment of Alzheimers disease (AD). They alleviate the cognitive deficits and AD-modifying agents, by inhibiting the β-amyloid (Aβ) peptide aggregation, through binding to both the catalytic and peripheral anionic sites, the so called dual binding site of the AChE enzyme. In this Letter, chemical features based 3D-pharmacophore models were developed based on the eight potent and structurally diverse AChE inhibitors (I-VIII) obtained from high-throughput in vitro screening technique. The best 3D-pharmacophore model, Hypo1, consists of two hydrogen-bond acceptor lipid, one hydrophobe, and two hydrophobic aliphatic features obtained by Catalyst/HIPHOP algorithm adopted in Discovery studio program. Hypo1 was used as a 3D query in sequential virtual screening study to filter three small compound databases. Further, a total of nine compounds were selected and followed on in vitro analysis. Finally, we identified two leads--Specs1 (IC(50)=3.279 μM) and Spec2 (IC(50)=5.986 μM) dual binding site compounds from Specs database, having good AChE enzyme inhibitory activity.

Synthesis, biological evaluation and molecular modeling study of pyrazole derivatives as selective COX-2 inhibitors and anti-inflammatory agents.

A novel series of pyrazole derivatives were synthesized and evaluated in vivo for their anti-inflammatory activity in carrageenan-induced rat paw edema model. Among all compounds, 5a, and 5b showed comparable anti-inflammatory activity to Nimesulide, the standard drug taken for the studies. In silico (docking) studies were carried out to investigate the theoretical binding mode of the compounds to target the cyclooxygenase (COX-2) using Autodock 4.2.

The exploration of thienothiazines as selective butyrylcholinesterase inhibitors

The role of butyrylcholinesterase (BChE) in the progression of Alzheimers disease (AD) has recently become more crucial. In the AD brain, selective BChE inhibitors have been demonstrated to have a beneficial effect in vivo, probably by recovering cholinergic activity and/or by restoring AChE:BChE activity ratios to the levels observed in the healthy brain. Thienothiazines are compounds sharing some structural features with phenothiazines, which are known to be potent BChE inhibitors. Thus, in this contribution 45 thienothiazines were investigated for their BChE inhibitory activity. Six of them were proven to be potent and selective inhibitors of equine BChEs hydrolase activity. Structure-activity relationships were laid out, and a tentative pharmacophore model for BChE inhibitors of the thienothiazine type was proposed. The most active compound, 3f, displayed a mixed type of inhibition and was also active against the human BChE (huBChE) with an IC(50) huBChE of 0.51 ± 0.07 μM. Computational studies suggested that 3f likely binds to the catalytic site and nearby to the peripheral site of the huBChE in an extended form. In addition, the chemical space occupied by the active thienothiazines, as opposed to phenothiazines and other representative chemical classes of BChE inhibitors, was explored with the aid of ChemGPS-NP, and the relevant chemical space regions were identified. This study shows for the first time that thienothiazines represent a new group of BChE inhibitors that can be used as molecular probes for studying the role of BChE in the brain or for developing newer drug leads for AD therapy.

Homology modeling and atomic level binding study of Leishmania MAPK with inhibitors.

The current therapy for leishmaniasis is not sufficient and it has two severe drawbacks, host-toxicity and drug resistance. The substantial knowledge of parasite biology is not yet translating into novel drugs for leishmaniasis. Based on this observation, a 3D structural model of Leishmania mitogen-activated protein kinase (MAPK) homologue has been developed, for the first time, by homology modeling and molecular dynamics simulation techniques. The model provided clear insight in its structure features, i.e. ATP binding pocket, phosphorylation lip, and common docking site. Sequence-structure homology recognition identified Leishmania CRK3 (LCRK3) as a distant member of the MAPK superfamily. Multiple sequence alignment and 3D structure model provided the putative ATP binding pocket of Leishmania with respect to human ERK2 and LCRK3. This analysis was helpful in identifying the binding sites and molecular function of the Leishmania specific MAPK homologue. Molecular docking study was performed on this 3D structural model, using different classes of competitive ATP inhibitors of LCRK3, to check whether they exhibit affinity and could be identified as Leishmania MAPK specific inhibitors. It is well known that MAP kinases are extracellular signal regulated kinases ERK1 and ERK2, which are components of the Ras-MAPK signal transduction pathway which is complexed with HDAC4 protein, and their inhibition is of significant therapeutic interest in cancer biology. In order to understand the mechanism of action, docking of indirubin class of molecules to the active site of histone deacetylase 4 (HDAC4) protein is performed, and the binding affinity of the protein-ligand interaction was computed. The new structural insights obtained from this study are all consistent with the available experimental data, suggesting that the homology model of the Leishmania MAPK and its ligand interaction modes are reasonable. Further the comparative molecular electrostatic potential and cavity depth analysis of Leishmania MAPK and human ERK2 suggested several important differences in its ATP binding pocket. Such differences could be exploited in the future for designing Leishmania specific MAPK inhibitors.