Assessment of mutagenic, genotoxic, and cytotoxic potential of water samples of Harike wetland: a Ramsar site in India using different ex vivo biological systems.

Harike is a wetland of international importance under the Ramsar Convention. The present study entails the investigation of mutagenic, genotoxic and cytotoxic effect of surface water samples collected from five different areas of the Harike wetland by using the histidine reversion point mutation assay in Salmonella typhimurium (TA98 and TA100) strain with or without S9, bioluminescence mutagenicity assay using Vibrio harveyi (A16) strain, plasmid-nicking assay using pBR322 and 3(4,5-dimethyl-thiazol-2-yl)2,5-diphenyl-tetrazolium bromide assay as well as confocal imaging studies using Chinese hamster ovarian cell line, respectively. It was observed that although, the water sample of all the areas of wetland demonstrated mutagenic, genotoxic as well as cytotoxic activity, the effect was quite significant with the water samples from River Satluj and Khatan area (i.e. reservoir mainly contains Satluj water). The metal analysis of water samples was also conducted with atomic absorption spectrophotometer. The mutagenicity, genotoxicity and cytotoxicity of water samples emerged to be correlated with metal concentration. The source of toxic components seems to be associated with various industrial effluents and agricultural run-off. The results of the present study carry great importance in documenting the water quality monitoring data of the wetland.

Potent and selective inhibitors of PI3Kδ: obtaining isoform selectivity from the affinity pocket and tryptophan shelf.

A potent inhibitor of PI3Kδ that is ≥ 200 fold selective for the remaining three Class I PI3K isoforms and additional kinases is described. The hypothesis for selectivity is illustrated through structure activity relationships and crystal structures of compounds bound to a K802T mutant of PI3Kγ. Pharmacokinetic data in rats and mice support the use of 3 as a useful tool compound to use for in vivo studies.

Oxidative stress--implications, source and its prevention.

Oxidative stress has been a major predicament of present day living. It has been the product of imbalance between the processes involved in free radical generation and their neutralization by enzymatic and non-enzymatic defence mechanisms. The oxidative stress has been contributed by numerous factors including heavy metals, organic compound-rich industrial effluents, air pollutants and changing lifestyle pattern focussing mainly on alcohol consumption, dietary habits, sun exposure, nuclear emissions, etc. The most common outcome of oxidative stress is the increased damage of lipid, DNA and proteins that resulted in the development of different pathologies. Among these pathologies, cancer is the most devastating and linked to multiple mutations arising due to oxidative DNA and protein damage that ultimately affect the integrity of the genome. The chemopreventive agents particularly nutraceuticals are found to be effective in reducing cancer incidences as these components have immense antioxidative, antimutagenic and antiproliferative potentials and are an important part of our dietary components. These secondary metabolites, due to their unique chemical structure, facilitate cell-to-cell communication, repair DNA damage by the downregulation of transcription factors and inhibit the activity of protein kinases and cytochrome P450-dependent mixed function oxidases. These phytochemicals, therefore, are most appropriate in combating oxidative stress-related disorders due to their tendency to exert better protective effect without having any distinct side effect.

Discovery of novel PI3-kinase δ specific inhibitors for the treatment of rheumatoid arthritis: taming CYP3A4 time-dependent inhibition.

PI3Kδ is a lipid kinase and a member of a larger family of enzymes, PI3K class IA(α, ß, δ) and IB (γ), which catalyze the phosphorylation of PIP2 to PIP3. PI3Kδ is mainly expressed in leukocytes, where it plays a critical, nonredundant role in B cell receptor mediated signaling and provides an attractive opportunity to treat diseases where B cell activity is essential, e.g., rheumatoid arthritis. We report the discovery of novel, potent, and selective PI3Kδ inhibitors and describe a structural hypothesis for isoform (α, ß, γ) selectivity gained from interactions in the affinity pocket. The critical component of our initial pharmacophore for isoform selectivity was strongly associated with CYP3A4 time-dependent inhibition (TDI). We describe a variety of strategies and methods for monitoring and attenuating TDI. Ultimately, a structure-based design approach was employed to identify a suitable structural replacement for further optimization.

Unsupervised guided docking of covalently bound ligands.

An approach for docking covalently bound ligands in protein enzymes or receptors was implemented in MacDOCK, a similarity-driven docking program based on DOCK 4.0. This approach was tested with a small number of covalent ligand-protein structures, using both native and non-native protein structures. In all cases, MacDOCK was able to generate orientations consistent with the known covalent binding mode of these complexes, with a performance similar to that of other docking programs. This method was also applied to search for known covalent thrombin inhibitors in a medium-sized molecular database (ca. 11,000 compounds). Detection of functional groups suitable for covalent docking was carried out automatically. A significant enrichment in known active molecules in the first 5% of the database was obtained, showing that MacDOCK can be used efficiently for the virtual screening of covalently bound ligands.

Exploring the active site of human factor Xa protein by NMR screening of small molecule probes.

A collection of small molecules (MW < 350 Da) was screened for binding to human factor Xa using saturation transfer difference NMR spectroscopy to detect binding. The NMR screening experiments identified four hits. Binding isotherms constructed from NMR linewidth data showed that the binding affinities of the hits were all in the 30-210 microM range. Competition binding experiments showed that three of the ligands were displaced by a known microM inhibitor of factor Xa. The success of the method for identifying new ligands and the relevance of this information to the design of new factor Xa inhibitors are discussed.

Oxyaniliniums as acetylcholinesterase inhibitors for the reversal of neuromuscular block.

A series of oxyanilinium-based AChE inhibitors have been synthesised and tested for the reversal of vecuronium-induced neuromuscular block. Several compounds, for example 2-hydroxy- and 2-methoxy-N,N-dimethyl-N-allylanilinium bromide (3 and 6) showed comparable reversal potencies to edrophonium and clean in vivo cardiovascular profiles.