Viral inhibitory potential of hyoscyamine in Japanese encephalitis virus-infected embryonated chicken eggs involving multiple signaling pathways.

Japanese encephalitis virus (JEV) is the leading cause of viral encephalitis worldwide. The emergence of new genotypes of the virus and a high rate of mutation make it necessary to develop alternative treatment strategies against this deadly pathogen. Although the antiviral properties of Atropa belladonna and some of its active components, such as atropine and scopolamine, have been studied, the effect of another important component, hyoscyamine, against JEV infection has not yet been investigated. In this study, we investigated the antiviral effect of hyoscyamine against JEV and its immunomodulatory activity in embryonated chicken eggs. Pretreatment with hyoscyamine sulphate resulted in a significant decrease in the viral load in both chorioallantoic membrane (CAM) and brain tissues at 48 and 96 hours postinfection. In silico studies showed stable binding and interaction between hyoscyamine and non-structural protein 5 (NS5), suggesting that this could be the basis of its antiviral effect. Embryonated eggs pretreated with hyoscyamine sulphate showed upregulation of Toll-like receptor 3 (TLR3), TLR7, TLR8, interleukin 4 (IL-4), and IL-10 as well as interferons and regulatory factors. Hyoscyamine sulphate was also found to cause significant downregulation of TLR4. The potential use of hyoscyamine for controlling JEV replication and its dissemination to the brain suggest that it may be a promising therapy option against JEV in the future.

Synthesis and biological evaluation of polyhydroxylated oxindole derivatives as potential antileishmanial agent.

The devastating appearance of numerous drug-unresponsive strains of Leishmania donovani and severe toxic side effects of conventional antileishmanial therapy necessitates the search for novel leads, to treat visceral leishmaniasis efficiently. The current study deals with the synthesis and biological evaluation of a unique C-5 functionalized oxindole based polyphenol to ascertain its activities against L. donovani infection, in vitro. The polyhydroxylated oxindole derivative (1) was generated by coupling styrene derivatives with 5-bromo bis-arylidene oxindole using Heck coupling reaction. The synthesized molecule 1 was tested for its antileishmanial activity using both promastigote and amastigote stages of L. donovani. Molecule 1 showed promising anti-promastigote and anti-amastigote activities with IC50 values 15 µM and 1 µM, respectively, with no cytotoxicity towards host splenocytes. The results revealed that this compound induced parasite death by promoting oxidative stress, thereby triggering apoptosis.

Oestrogen receptor-mediated liposomal drug delivery for treating melanoma.

Function of steroid hormone oestrogen that transactivates oestrogen receptor (ER) is expressed in multiple organs. Except for malignancies of gynaecological organs, ER remains largely unutilised as a target to treat cancers of ER-expressing brain, prostate, skin etc. We have previously developed oestrogen targeting cationic lipid molecule (ES-C10), which showed targeted killing of ER + breast and skin cancer cells. In this study, we explored the targeting ability of ES-C10 as a ligand as well as its additive killing effect (if any), when incorporated in two different liposomes (DCME and DCDE), carrying two anticancer molecules MCIS3 and Docetaxel™, respectively. DCME and DCDE exhibited higher cytotoxicity in ER + cancer cells than in ER - cancer or in non-cancer cells. Both liposomes induced ER-mediated cytotoxicity and caspase 3-induced apoptosis in ER + melanoma cells. Further, decreased levels of pAkt, and increased levels of PTEN and p53 were also observed. Both the targeted liposomes were least haemolytic. These selectively delivered drug-cargoes to tumour mass over other vital organs and induced better anti-tumour effect, which led to increased survivability than their respective controls. In conclusion, we demonstrated the development of two independent liposomal drug-delivery systems associated with an anticancer, oestrogen-structure based ligand for efficient, ER-mediated anti-melanoma effect.

Cationic folate-mediated liposomal delivery of bis-arylidene oxindole induces efficient melanoma tumor regression.

The folate receptor (FR) is a well-validated and common target for cancer due to its high over-expression in many different cancer cells. Herein, we developed a new FR-targeting ligand (FA8) by conjugating folic acid and a cationic lipid. Owing to its favorable structural property FA8 as a ligand could be accommodated at an unusually higher molar ratio for a ligand-targeted liposome. We then encapsulated a drug-like molecule, bis-arylidene oxindole (NME2), in the targeted liposome. The resulting formulation induced potent caspase-8 up-regulation even in FR-moderately expressing melanoma cells. The NME2-associated non-targeted liposome (i.e., without FA8) or pristine NME2 could not up-regulate caspase-8. Caspase-8, an important apoptotic protein involved in the extrinsic pathway of apoptosis-signalling and inhibition of acquired drug resistance, was induced in cancer cells due to the combination treatment of liposomally associated FA8 and NME2 through the activation and subsequent cleavage of RIP-1. Consistently, in a melanoma tumor model too wherein FR is moderately expressed, significant tumour regression was obtained with this liposomal combination of FA8 and NME2. In conclusion, we demonstrate the development of a new FR-targeting ligand molecule whose higher level of inclusion (>10 mol%) in the liposomal formulation altered the mode of anticancer action of the encapsulated drug, thereby indicating a new therapeutic possibility involving FR targeted cancer treatment.

A Comprehensive Computational Analysis for the Binding Modes of Hepatitis C Virus NS5A Inhibitors: The Question of Symmetry.

Direct-acting antivirals (DAAs) form the current standard of care (SOC) against hepatitis C virus (HCV). These drugs selectively target the viral proteins, offering a unique mechanism to avoid toxicity, to increase their efficacy, and to evolve from decades of interferon- and ribavirin-based therapy. Among the promising HCV targets for DAAs is the NS5A protein, and daclatasvir (DCV) forms a first-in-class compound that selectively targets this protein. Despite the exceptional potency of DCV (∼picomolar IC50) and although several DCV derivatives have been approved for human use or are close to approval, the exact mode of action of these drugs is still incomplete. This is simply due to the vast complexity of cocrystallizing DCV with NS5A in the absence of two amphipathic helices that are required for DCV binding. In this context, computational modeling provides a unique alternative to solve this problem. Here, we build upon our recent discovery of a completely symmetrical interaction between DCV and NS5A and investigate the mode of binding of six other structures similar to DCV. The selected compounds include both symmetric and asymmetric molecules. In addition, we show that our model correlates very well with mutations that can confer resistance to DCV. The current study enhances our understanding of the mode of action of this class of HCV inhibitors and helps in defining the origin of resistance to these drugs.

Bis-arylidene oxindole-betulinic Acid conjugate: a fluorescent cancer cell detector with potent anticancer activity.

Molecules offering simultaneous detection and killing of cancer cells are advantageous. Hybrid of cancer cell-selective, ROS generator betulinic acid and bis-arylidene oxindole with amino propyl-linker is developed. With intrinsic fluorescence, the molecule exhibited cancer cell-specific residence. Further, it generated ROS, triggered apoptosis, and exhibited potent cytotoxicity in cancer cells selectively. We demonstrate the first example and use of isatins as betulinic acid conjugate for selective detection of cancer and subsequent killing of cancer cells via apoptosis.

Synthesis and biological evaluation of ferrocenylquinoline as a potential antileishmanial agent.

The emergence of resistance against antileishmanial drugs in current use necessitates the search for new classes of antileishmanial compounds. Herein we report the design, synthesis, and evaluation of a novel ferrocenylquinoline for activity against Leishmania donovani. 7-Chloro-N-[2-(1H-5-ferrocenyl-1,2,3-triazol-1-yl)ethyl]quinolin-4-amine (1) was generated by coupling an iron(II) ethynylferrocene species with 4-(2-ethylazido)amino-7-chloroquinoline using click chemistry. The synthesized compound 1 was tested for its antileishmanial activity using both promastigote and amastigote stages of L. donovani. Compound 1 showed promising anti-promastigote activity, with an IC50 value of 15.26 µM and no cytotoxicity toward host splenocytes. From the battery of tests conducted in this study, it appears that this compound induces parasite death by promoting oxidative stress and depolarizing the mitochondrial membrane potential, thereby triggering apoptosis. These results suggest that ferrocenylquinoline 1 is a suitable lead for the development of new antileishmanial drugs.

Bis-arylidene oxindoles as anti-breast-cancer agents acting via the estrogen receptor.

We report a new family of bis-arylidene oxindole derivatives that show highly selective estrogen receptor (ER)-mediated anticancer activity at low-nanomolar concentrations in ER-positive (ER+) breast cancer cells. In terms of cell growth inhibition, IC50 values for these compounds in ER+ breast cancer cells are two to three orders of magnitude lower than in ER-negative (ER-) breast cancer cells and non-cancer cells. In comparison with known bis-arylidene drugs, these compounds are at least three orders of magnitude more toxic than tamoxifen and 1.5-4-fold more toxic than 4-hydroxytamoxifen in ER+ MCF-7 cancer cells. These oxindoles inhibit ER transactivation, and their anticancer activities are inhibited in ER-depleted MCF-7 cells. Some of these nonsteroidal molecules also exhibit essential properties of selective ER down-regulation. From the development of two series of bis-arylidene oxindole-based compounds, we report a new series of anticancer agents for estrogen-responsive breast cancer.