Marshleen Yadav

Antifilarial Activity of 1,3-Diarylpropen-1-One: Effect on Glutathione-S-Transferase, a Phase II Detoxification Enzyme

Chalcone derivatives were evaluated for their antifilarial activity on Setaria cervi using glutathione-S-transferase (GST) as a drug target. The compounds 1-(4-benzotriazol-1-yl-phenyl)-3-(4-methoxyphenyl)prop-2-en-1-one (5), and 3-(4-methoxyphenyl)-1-(4-pyrrolidin-1-yl-phenyl) prop-2-en-1-one (7) showed a significant suppression (P < 0.01) in GST activity of adult female parasite extract at 3 microM concentration in vitro. However, GST activity was detected along with depletion in GSH level. Except Compounds 1 and 2, all exhibited a significant effect on the motility and viability of adult parasites. Compounds 3-(4-chlorophenyl)-1-(4-piperidin-1-yl-phenyl)prop-2-en-1-one (3), 1-(4-benzotriazol-1-yl-phenyl)-3-(4-methoxyphenyl)prop-2-en-1-one (5), and 3-(4-methoxyphenyl)-1-(4-pyrrolidin-1-yl-phenyl) prop-2-en-1-one (7) exhibited major irreversible effects on viability and resulted in parasite death and also inhibited the GST activity by 84-100% in vitro. We report for the first time the antifilarial activity of chalcones on GST of adult parasites. This study also strengthens our previous findings where GST is reported as a potential drug target for antifilarials.

Filarial glutathione-S-transferase: A potential vaccine candidate against lymphatic filariasis

Present report enumerates the vaccine potential of a glutathione-S-transferase purified from Setaria cervi against lymphatic filariasis. In jirds (Meriones unguiculatus) vaccination trial, a very significant 82.75% (p<0.005) reduction in adult parasite burden was observed in ScGST immunized group after 90 days post Brugia malayi L3 challenge. An inverse correlation between the antibody level and worm burden was found in ScGST immunized group (Persons correlation r=0.943, p<0.05). No recoveries of worms were obtained in heart and lungs of vaccinated group. The Antibodies reactive to ScGST appeared within four weeks of first dose and were able to neutralize the GST activity up to 86%. In an earlier study we have shown vaccine potential of ScGST against B. malayi by ADCC. Evaluation of cytokine profile in T-cells isolated from BALB/c mice immunized with ScGST were also showed predominance of Th2 response which further maintained the humoral immunity generated by ScGST administration in mice. The overall observations prompted us to envisage ScGST as a potential vaccine candidate against lymphatic filariasis.

Structural modeling and simulation studies of Brugia malayi glutathione-S-transferase with compounds exhibiting antifilarial activity: implications in drug targeting and designing.

Since glutathione-S-transferase (GST) mediated xenobiotic detoxification is a crucial mechanism in nematodes survival, we aimed to conduct an in silico analysis of filarial GST in order to predict the possible interactions for antifilarials. Present report depicts the homology modeling approach applied in the construction of molecular structure of Brugia malayi GST (BmGST) followed by its docking simulation with available antifilarials such as diethylcarbamazine, albendazole, Butylated Hydroxyanisole (BHA) and substituted chalcones. A very low root mean square deviation (0.82A) from template structure and stereochemical quality of constructed BmGST model proposed it as a significant framework for further analysis. In docking studies antifilarials and chalcones exhibited demarcation in their binding affinity and modes. Amongst all the compounds studied, albendazole and methyl-substituted chalcone showed the lowest binding energy and occupied binding pocket near to substrate binding site of GST. The side chain of these compounds interplayed as a potential interaction site which targeted mainly hydrophilic residues of the BmGST. The structural information and binding site mapping of BmGST for different antifilarials obtained from this study could aid in screening and designing new antifilarials or selective inhibitors for chemotherapy against filariasis.

A Review of Organic Farming for Sustainable Agriculture in Northern India

In the post independence period, the most important challenge in India has been to produce enough food for the growing population. Hence, high-yielding varieties are being used with infusion of irrigation water, fertilizers, or pesticides. This combination of high-yielding production technology has helped the country develop a food surplus as well as contributing to concerns of soil health, environmental pollution, pesticide toxicity, and sustainability of agricultural production. Scientists and policy planners are, therefore, reassessing agricultural practices which relied more on biological inputs rather than heavy usage of chemical fertilizers and pesticides. Organic farming can provide quality food without adversely affecting the soil’s health and the environment; however, a concern is whether large-scale organic farming will produce enough food for India’s large population. Certified organic products including all varieties of food products including basmati rice, pulses, honey, tea, spices, coffee, oilseeds, fruits, cereals, herbal medicines, and their value-added products are produced in India. Non edible organic products include cotton, garments, cosmetics, functional food products, body care products, and similar products. The production of these organic crops and products is reviewed with regard to sustainable agriculture in northern India.

Effect of diethylcarbamazine, butylated hydroxy anisole and methyl substituted chalcone on filarial parasite Setaria cervi: Proteomic and biochemical approaches

For survival, parasite exerts several lines of defense of which drug neutralization is one of the major phenomena. Lack of phase I cytochrome P450 in some of the nematode render them depend on the phase II detoxification system involving GST as a major detoxifying enzymes. In present study, the antifilarial DEC, phenolic compound BHA and methyl chalcone have been evaluated for proteomic and biochemical studies in Setaria cervi. BHA and methyl chalcone showed cytotoxic effect leading to irreversible inhibition in motility and viability of parasites. These drugs showed marked alteration in proteomic profile of S. cervi at 100 μM concentration with 10.82, 8.52 and 6.75% downregulated (<0.5) and 7.64, 31.78 and 24.32% upregulated (>1.5) in DEC, BHA and methyl chalcone treatment respectively. Significant depletion in GSH level with increase in NO production was observed. Amongst these compounds, methyl chalcone demonstrated significant inhibitory effect (p<0.05) on GST, PGHS and PTP activity leading to loss of metabolic homeostasis and parasite death. The cytotoxic response and altered expression profile of major enzymes under drug exposure suggested the oxidative stress induced apoptosis as a major cause of parasite killing which was further supported by DNA fragmentation in BHA and methyl chalcone.

Identification of major antigenic peptide of filarial glutathione-S-transferase.

In our earlier report, a 26kDa Setaria cervi glutathione-S-transferase showed significant protection (82%) in jirds infected with L3 larvae of Brugia malayi. In the present study we have identified the major antigenic epitopes in ScGST. Carboxypeptidase B has been used to digest the ScGST in to smaller fragments. The digested products were separated as four protein bands on SDS-PAGE. The smallest fragment of 6kDa (P4) from ScGST was identified as major antigenic epitope because of its significant reactivity with jird anti ScGST sera and human filarial sera in immunoblotting. The MALDI-LC/MS sequencing of ScGST P4 peptide ((5)KLTYFSIRGRGLAEPIRL(20), (22)KVPDDQQFLDDLISR(36) and (47)VFHFGQGPHHGPPR(62)) suggested that this protein band has a fragment of 5-62 residues long that matched with the N-terminal end of filarial GST. The antigenicity plot of ScGST was compared with BmGST model and both exhibited three immunogenic peaks within the first 60 residues towards N-terminal. In BmGST the N-terminal region was also detected with N-glycosylation signal peptide NAS adding to its high immunogenic property. Further, P4 showed strong reactivity with IgG1 and IL-4 response in endemic normal sera suggested its role in Th2 response which in turn is correlated with antibody dependent cell mediated cytotoxicity. Thus taking these results into account we propose 5-62 residues long N-terminal peptide of GST as a potential target for further vaccination studies against filarial infection.

Evidence for the presence of prostaglandin H synthase like enzyme in female Setaria cervi and its inhibition by diethylcarbamazine

Experimental evidence has shown that Setaria cervi a bovine filarial parasite contains significant amount of prostaglandin H synthase like activity in the somatic extract of its different life stages. A protein with characteristics of prostaglandin H synthase was purified to homogeneity from female somatic extract using a combination of affinity and gel filtration chromatography. Molecular weight of purified enzyme was 70kDa as determined by SDS-PAGE. Purified enzyme showed high activity with arachidonic acid and TMPD substrates suggests the presence of both cyclooxygenase and peroxidase activity in enzyme. Fluorescence spectroscopy and hemin-associated peroxidase activity confirmed presence of heme in purified enzyme. The K(m) and V(max) values using arachidonic acid were determined to be 79+/-1.5microM and 0.165+/-0.2U/ml, respectively. Further, indomethacin and aspirin, specific inhibitors for PGHS, significantly inhibited the enzyme activity. Diethylcarbamazine, an antifilarial drug inhibited the microfilarial PGHS like activity as well as their motility. Here we are reporting for the first time PGHS like activity in filarial parasite and its inhibition with DEC which provide that this enzyme could be used as a drug target.