3D QSAR Pharmacophore Modeling, in Silico Screening, and Density Functional Theory (DFT) Approaches for Identification of Human Chymase Inhibitors

Human chymase is a very important target for the treatment of cardiovascular diseases. Using a series of theoretical methods like pharmacophore modeling, database screening, molecular docking and Density Functional Theory (DFT) calculations, an investigation for identification of novel chymase inhibitors, and to specify the key factors crucial for the binding and interaction between chymase and inhibitors is performed. A highly correlating (r = 0.942) pharmacophore model (Hypo1) with two hydrogen bond acceptors, and three hydrophobic aromatic features is generated. After successfully validating “Hypo1”, it is further applied in database screening. Hit compounds are subjected to various drug-like filtrations and molecular docking studies. Finally, three structurally diverse compounds with high GOLD fitness scores and interactions with key active site amino acids are identified as potent chymase hits. Moreover, DFT study is performed which confirms very clear trends between electronic properties and inhibitory activity (IC50) data thus successfully validating “Hypo1” by DFT method. Therefore, this research exertion can be helpful in the development of new potent hits for chymase. In addition, the combinational use of docking, orbital energies and molecular electrostatic potential analysis is also demonstrated as a good endeavor to gain an insight into the interaction between chymase and inhibitors

Mapk-activated protein kinase 2 contributes to Clostridium difficile-associated inflammation

Clostridium difficile infection (CDI) results in toxin-induced epithelial injury and marked intestinal inflammation. Fecal markers of intestinal inflammation correlate with CDI disease severity, but regulation of the inflammatory response is poorly understood. Previous studies demonstrated that C. difficile toxin TcdA activates p38 kinase in tissue culture cells and mouse ilium, resulting in interleukin-8 (IL-8) release. Here, we investigated the role of phosphorylated mitogen-activated protein kinase (MAPK)-activated protein kinase (MK2 kinase, pMK2), a key mediator of p38-dependent inflammation, in CDI. Exposure of cultured intestinal epithelial cells to the C. difficile toxins TcdA and TcdB resulted in p38-dependent MK2 activation. Toxin-induced IL-8 and GROα release required MK2 activity. We found that p38 and MK2 are activated in response to other actin-disrupting agents, suggesting that toxin-induced cytoskeleton disruption is the trigger for kinase-dependent cytokine response. Phosphorylated MK2 was detected in the intestines of C. difficile-infected hamsters and mice, demonstrating for the first time that the pathway is activated in infected animals. Furthermore, we found that elevated pMK2 correlated with the presence of toxigenic C. difficile among 100 patient stool samples submitted for C. difficile testing. In conclusion, we find that MK2 kinase is activated by TcdA and TcdB and regulates the expression of proinflammatory cytokines. Activation of p38-MK2 in infected animals and humans suggests that this pathway is a key driver of intestinal inflammation in patients with CDI

Development and elucidation of mechanism of action of new leads to attenuate neuroinflammation in stroke model.

In vitro anti-inflammatory activity screening in LPS intoxicated Raw 264.7 murine macrophage cell line showed tribulusterine/perlolyrine had its activity by down regulating inflammatory mediators such as iNOS, COX2, TNFα and IL1β • Screening of aqueous extract of Tribulus terrestris has shown neuroprotective activity by regulating the GSK-3β pathway protein level and elevating β-catenin and S9-phospho GSK-3β protein level. It also stabilizes GSK-3 β / β-catenin pathway, maintains GSK-3β in inactive state, prevents dephosphorylation thereby reduces β-catenin degradation and mediates its translocation resulting in neuroprotection. • Docking studies revealed tribulusterine / perlolyrine can be a possible prototype from natural source Tribulus terrestris. • Tribulusterine / perlolyrine protected oxygen glucose deprived reperfusion injured differentiated SHSY5Y cells which is revealed by cell viability studies. • The molecular pathway through which tribulusterine/perlolyrine exhibits it neuroprotection is by down regulating both caspase dependent and independent pathway by inhibiting stress kinasess. • Five novel leads have been identified through pharmacophore based structure search which can be explored further for their neuroprotective activity. The study can be concluded that tribulusterine/perlolyrine exhibits it neuroprotection by down regulating both caspase dependent and independent pathway by inhibiting stress kinases. Further it possesses anti inflammatory activity so it can be efficacious in ischemia condition because stroke pathology involves neuro inflammation as a major cascade for disease prognosis. IMPACT OF STUDY: Treatment of neurodegenerative disease such as stroke is a major challenge for the physicians. Neuroprotective effects of voltage dependent calcium channel blockers, NMDA antagonists and radical scavengers were found to be effective in experimental models of cerebral ischemia. The outcome of these drugs as therapeutic regimen is still beyond reach. Medicinal plants have been historically known a rich source for successful lead identification, and always signify an important pool for the discovery of new pharmacological hits as on date. Tribulusterine/perlolyrine, a possible prototype exhibited it neuroprotection is by down regulating both caspase dependent and independent pathway by inhibiting stress kinasess. Five novel leads have been identified from natural source which can be potential candidates as novel anti- inflammatory and neuroprotective agents. In revelation of the increasing cost of development of new drugs, alternative approaches like development from herbal extracts could be beneficial