Cocculus hirsutus-Derived Phytopharmaceutical Drug Has Potent Anti-dengue Activity

Dengue is a serious public health concern worldwide, with ∼3 billion people at risk of contracting dengue virus (DENV) infections, with some suffering severe consequences of disease and leading to death. Currently, there is no broad use vaccine or drug available for the prevention or treatment of dengue, which leaves only anti-mosquito strategies to combat the dengue menace. The present study is an extension of our earlier study aimed at determining the in vitro and in vivo protective effects of a plant-derived phytopharmaceutical drug for the treatment of dengue. In our previous report, we had identified a methanolic extract of aerial parts of Cissampelos pareira to exhibit in vitro and in vivo anti-dengue activity against all the four DENV serotypes. The dried aerial parts of C. pareira supplied by local vendors were often found to be mixed with aerial parts of another plant of the same Menispermaceae family, Cocculus hirsutus, which shares common homology with C. pareira. In the current study, we have found C. hirsutus to have more potent anti-dengue activity as compared with C. pareira. The stem part of C. hirsutus was found to be more potent (∼25 times) than the aerial part (stem and leaf) irrespective of the extraction solvent used, viz., denatured spirit, hydro-alcohol (50:50), and aqueous. Moreover, the anti-dengue activity of stem extract in all the solvents was comparable. Hence, an aqueous extract of the stem of C. hirsutus (AQCH) was selected due to greater regulatory compliance. Five chemical markers, viz., Sinococuline, 20-Hydroxyecdysone, Makisterone-A, Magnoflorine, and Coniferyl alcohol, were identified in fingerprinting analysis. In a test of primary dengue infection in the AG129 mice model, AQCH extract at 25 mg/kg body weight exhibited protection when administered four and three times a day. The AQCH was also protective in the secondary DENV-infected AG129 mice model at 25 mg/kg/dose when administered four and three times a day. Additionally, the AQCH extract reduced serum viremia and small intestinal pathologies, viz., viral load, pro-inflammatory cytokines, and vascular leakage. Based on these findings, we have undertaken the potential preclinical development of C. hirsutus-based phytopharmaceutical, which could be studied further for its clinical development for treating dengue.

Neuroprotective Potentials of Cocculus hirsutus Leaf Extract Against 6,7-Epoxytropine Tropate-Induced Memory Impairment in Rats

Cocculus hirsutus, a tropical South Asian creeper,traditionally used as a diuretic, laxative, cardiotonic, anti-microbial, antidiabetic, anti-inflammatory and spermatogenic. However, the neuroprotective role was less explored; therefore, this researchwas conducted to investigate neuroprotective potentials of Cocculus hirsutus leaf hydroalcoholic extract in 6,7-Epoxytropine tropate (Scopolamine) induced cognitive impairment and oxidative lipid peroxidation in the brain of wistar albino rat. Scopolamine (1 mg/kg body weight, i.p.) was given in rats for 14 days to induce transient cognitive impairment. Donepezil (2 mg/kg body weight, orally) has been used for this research as a positive control. Behavioral studies were done using Morris water maze and elevated plus maze and neurobiochemical parameters such as acetylcholinesterase activity, reduced glutathione levels and activity of catalase were assessed in rats brain homogenate. Cocculus hirsutus leaf hydroalcoholic extract(200 mg/kg and 400 mg/kg) exhibited an improvement in spatial, exteroceptive learning and memory. The extract showed significant decline in the activity of acetylcholinesterase, enhancement of reduced glutathione levels and catalase activity (p<0.001). All the outcomes were assessed by Bonferroni post hoc tests with ANOVA for multiple comparison studies. This study reveals that hydroalcoholic extract of Cocculus hirsutusleaf acts as neuroprotective against scopolamine induced behavioral and neurobiochemical changes.

ANTI-INFLAMMATORY PROPERTIES OF UNEXPLORED PLANTS – CUSCUTA REFLEXA AND COCCULUS HIRSUTUS – AN EXPERIMENTAL STUDY

Objective: The study aimed to document the rarely explored plants, namely, Cuscuta reflexa (CRA) and Cocculus hirsutus (CHP) used by the ethnic people of a specific region. The anti-inflammatory (AI) property of kwath of CRA (KCRA), kwath of CHP (KCHP), and their blend (kwath blend [KB]) was also assessed. Methods: The KCRA and KCHP were prepared following standard Ayurvedic procedures. The AI property was determined by carrageenan-induced paw edema at doses; 250, 125, and 62.5 mg/kg. The KB (500 mg/kg) was prepared using equal parts of KCRA and KCHP in view of the potential AI property as compared to the individual plants. Ibuprofen (100 mg/kg) was used as the standard AI drug standard drug (SD). Results: The carrageenan-induced paw inflammation was highest and doubled at 3 h. The oral administration of SD (100 mg/kg) produced a high reduction of edema (78.47%) at 3 h. Both KCRA and KCHP had reduced edema and were equally potent (EC50; 139.8 and 147.3 mg/kg, respectively) at the early phase. However, the efficacy of KCRA was greater than KCHP at the second phase of inflammation (EC50=313.6 and 2760 mg/kg, respectively). KCRA was efficacious and potent as an AI agent. Unlike SD, KB had effectively inhibited paw edema from the 6th h onward. The AI activity of KB was superior compared to individual plant groups. Conclusions: The study demonstrated that the traditional formulation – kwath of rarely explored plants, namely, CRA, CHP, and KB has AI property and can be explored to develop them as AI agents.

Cocculus hirsutus-derived phytopharmaceutical drug has potent anti-dengue activity

BackgroundDengue is a serious public health concern worldwide, with ~3 billion people at risk of contracting dengue virus (DENV) infections. Currently, no effective vaccine or drug is available for the prevention or treatment of dengue, which leaves only anti-mosquito strategies to combat this disease. The present study was initiated to determine the in-vitro and in vivo protective effects of a plant-derived phytopharmaceutical drug for the treatment of dengue.Methodology/Principal FindingsIn our previous report, we had identified methanolic extract of the aerial parts of Cissampelos pareira to exhibit in vitro and in vivo anti-dengue activity against all the four DENV serotypes. In the current study, we have identified another Indian medicinal plant, Cocculus hirsutus, which has a more potent anti-dengue activity than C. pareira. The activity has been evaluated through flow-cytometry-based virus inhibition assay. Interestingly, the stem of C. hirsutus was found to be more potent than the aerial part irrespective of the extraction solvent used viz., denatured spirit, hydro-alcohol (50:50) and water. Hence, the aqueous extract of stem of C. hirsutus (AQCH) was further advanced for investigations because of greater regulatory acceptance. The AQCH exhibited dose-dependent inhibition of release of DENV and its secretory antigen, NS1. Five chemical markers viz. Sinococuline, 20-Hydroxyecdysone, Makisterone-A, Magnoflorine and Coniferyl alcohol were identified as the major chemical ingredients of the AQCH extract. These chemicals were subsequently used for extract standardisation. Importantly, AQCH completely protected AG129 mice at 25 mg/kg/dose body weight when fed 4 times a day post-infection with a lethal dose of DENV-2 S221 strain. Because of its potential as an effective phytopharmaceutical drug against dengue, AQCH, has been formulated into tablets for further pre-clinical and clinical developments.Conclusions/SignificanceWe provide evidence of the pan anti-dengue potential of C. hirsutus-based phytopharmaceutical drug as determined through in vitro and in vivo experiments. We have also characterized five chemical entities in the drug substance, which provides means for standardization of drug substance and drug product. Based on these findings, a program to develop a safe and effective C. hirsutus-derived phytopharmaceutical drug for the treatment of dengue has been initiated.Author summaryThere is an urgent need to develop a safe and effective drug against dengue, which is a rapidly expanding mosquito-borne viral disease. Half of the world’s population has been estimated to be at risk of contracting this disease and the situation remains grim due to lack of an approved drug. We aimed to develop an ethnopharmacological drug against dengue by exploring traditional Indian medicinal science, Ayurveda. This led us to identify a creeper, Cocculus hirsutus, as a more potent anti-dengue plant than Cissampelos pareira, reported in our earlier published study. The stem part of C. hirsutus was found to be more efficacious in inhibiting the propagation of dengue viruses (DENVs) in cell culture than its aerial part. Hence, we chose to advance aqueous extract of stem of C. hirsutus (AQCH) for further studies. Importantly, AQCH also protected immune-compromised mice from lethal DENV infection, which is suggestive of its potential clinical relevance. We have identified five chemical marker compounds in AQCH to gauge the quality and consistency of extract preparation and its formulation into stable tablets. Based on the findings of this study, we have undertaken the development of a safe and effective C. hirsutus-derived phytopharmaceutical drug for the treatment of dengue.