Identification of tyrosine kinase inhibitors from Panax bipinnatifidus and Panax pseudoginseng for RTK-HER2 and VEGFR2 receptors, by in silico approach.

Breast and stomach cancer is reported as a leading cause for human mortality across the world. The overexpression of receptor tyrosine kinase (RTK) proteins, namely the human epidermal growth factor receptor2 (HER2) and the vascular endothelial growth factor receptor2 (VEGFR2), is reported to be responsible for development and metastasis of breast and stomach cancer. Although several synthetic tyrosine kinase inhibitors (TKIs) as drug candidates targeting RTK-HER2 and VEGFR2 are currently available in the market, these are expensive with the reported side effects. This confers an opportunity for development of alternative novel tyrosine kinase inhibitors (TKIs) for RTK-HER2 and VEGFR2 receptors from the botanical sources. In the present study, we characterized 47 bioactive phytocompounds from the methanol extracts of the rhizomes of Asiatic traditional medicinal herbs-Panax bipinnatifidus and Panax pseudoginseng, of Indian Himalayan landraces using HPLC, GC-MS and high-sensitivity LC-MS tools. We performed molecular docking and molecular dynamics simulation analysis using Schrödinger suite 2020-3 to confirm the TKI phytocompounds showing the best binding affinity towards RTK-HER2 and VEGFR2 receptors. The results of molecular docking studies confirmed that the phytocompound (ligand) luteolin 7-O-glucoside (IHP15) showed the highest binding affinity towards receptor HER2 (PDB ID: 3PP0) with docking score and Glide g score (G-Score) of - 13.272, while chlorogenic acid (IHP12) showed the highest binding affinity towards receptor VEGFR2 (PDB ID: 4AGC) with docking score and Glide g score (G-Score) of - 10.673. Molecular dynamics (MD) simulation analysis carried out for 100 ns has confirmed strong binding interaction between the ligand and receptor complex [luteolin 7-O-glucoside (IHP15) and HER2 (PDB ID: 3PP0)] and is found to be stabilized within 40 to 100 ns of MD simulation, whereas ligand-receptor complex [chlorogenic acid (IPH12) and VEGFR2 (PDB ID: 4AGC)] also showed strong binding interaction and is found to be stabilized within 18-30 ns but slightly deviated during 100 ns of MD simulation. In silico ADME-Tox study using SwissADME revealed that the ligands luteolin 7-O-glucoside (IHP15) and chlorogenic acid (IHP12) have passed majority parameters of the common drug discovery rules. The present study has confirmed luteolin 7-O-glucoside (IHP15) and chlorogenic acid (IHP12) as potential tyrosine kinase inhibitors (TKIs) which were found to inhibit RTKs-HER2 and VEGFR2 receptor proteins, and thus paving the way for development of alternative potential TKIs (drug molecules) for treatment of HER2- and VEGFR2-positive breast and stomach cancer.

In-silico based identification of phytochemicals from Houttuynia cordata Thunb. as potential inhibitors for overexpressed HER2 and VEGFR2 cancer genes.

Human epidermal growth factor receptor2 (HER2) and Vascular endothelial growth factor receptor2 (VEGFR2) - a tyrosine kinase receptors play a key role in breast and stomach cancers. The overexpression of HER2 and VEGFR2 genes increases the number of HER2 and VEGFR2 in the cell which initiates breast and stomach cancer respectively. The phytochemicals from traditional medicinal herb Houttuynia cordata Thunb. are reported to possess anti-inflammatory and anti-cancer potential. However, isolation of phytochemicals from this herb is fraught with uncertainly and time-consuming. Here, a molecular docking approach provides probable binding affinities between the receptors and phytochemicals (ligands) which initiate the first step of anticancer drug discovery and development. In the present study, In-silico docking approaches were used to identify the top-hit phytochemicals from H. cordata as potential inhibitors for overexpressed HER2 (breast) and VEGFR2 (stomach) cancer genes. A total of 100 biologically active phytochemicals from H. cordata were screened and docked against the ligand-binding pocket of HER2 and VEGFR2 kinase domains. Docking results revealed only a few phytochemicals (molecules) which appropriately fit into the ligand-binding pocket with higher binding affinity than the natural ATP ligand. A competitive docking was used to ascertain the top-hit phytochemicals that bind perfectly to the ATP ligand-binding pocket. Among the top-hit phytochemicals docked from H. cordata, the ß-sitosterol and Quercetin showed highest binding affinity towards HER2 and VEGFR2 receptors using both hydrogen and hydrophobic interactions. This study confirmed ß-sitosterol and Quercetin as potential drug candidates against breast and stomach cancer.Communicated by Ramaswamy H. Sarma.

Evaluation of antioxidant, anti-inflammatory and anticancer activities of diosgenin enriched Paris polyphylla rhizome extract of Indian Himalayan landraces.

ETHNOPHARMACOLOGICAL RELEVANCE: Traditional medicinal plants have gained attention as a potential therapeutic agent to combat cancer and inflammation. Diosgenin rich fresh extracts of Paris polyphylla rhizome from Indian Himalaya is traditionally used as wound healing, anti-bleeding, anti-inflammatory and anti-cancer agent by the folk healers. AIM OF THE STUDY: Present study was aimed to prepare two types of extracts from Paris polyphylla rhizome of Indian Himalayan landraces - 1. ethanolic extract of Paris polyphylla rhizome (EEPPR) and 2. Diosgenin enriched Paris polyphylla rhizome extract (DPPE), quantification of diosgenin content, and to evaluate their in vitro anti-oxidant, in vivo anti-inflammatory and in vitro cytotoxicity and anti-cancer activities of the DPPE. MATERIALS AND METHODS: Diosgenin content of EEPPR was quantified through GC-MS while diosgenin content of DPPE was quantified through HPTLC, and the diosgenin yield from EEPPR and DPPE were compared. In vitro antioxidant activities of DPPE were performed using DPPH, NOD, RP and SOD assay while in vivo anti-inflammatory activity of DPPE were evaluated in dextran induced hind paw edema in rats. In vitro cytotoxicity and anti-cancer activities of DPPE were evaluated in human breast cancer cell lines (MCF-7, MDA-MB-231), cervical cancer cell lines (HeLa) and Hep-2 cell lines. RESULTS: EEPPR obtained through cold extraction method using 70% ethanol showed maximum diosgenin content of 17.90% quantified through GC-MS while similar compounds pennogenin (3.29%), 7ß-Dehydrodiosgenin (1.90%), 7-Ketodiosgenin acetate (1.14%), and 7 ß-hydroxydiosgenin (0.55%) were detected in low concentration, and thus confirmed diosgenin as major and lead phytochemical. However, DPPE obtained through both cold and repeated hot extraction with the same solvent (70% ethanol) showed diosgenin content of 60.29% which is significantly higher (p < 0.001) than the diosgenin content in EEPPR. DPPE demonstrated significant in vitro antioxidant activities by dose-dependently quenched (p < 0.001) SOD free radicals by 76.66%, followed by DPPH (71.43%), NOD (67.35%), and RP (63.74%) at a max concentration of 2 µg/µl of ascorbic acid and test drugs with remarkable IC50 values (p < 0.01). Further, DPPE also showed potent anti-inflammatory activities by dose-dependently suppressed dextran induced paw edema in rats (p < 0.01) from 2 h to 4 h. DPPE suppressed the proliferation of MCF-7, MDA-MB-231, Hep-2 and HeLa cell lines. Maximum activity was observed in MCF-7 cells. The DPPE also induced apoptosis in MCF-7 cell lines as measured by AO/PI and DAPI staining, as well as DNA laddering, cell cycle analysis and phosphatidylserine externalization assay. The growth-inhibitory effect of DPPE on MCF-7 breast cancer cells was further confirmed from the colony-formation assay. DPPE upregulated expression of Bax and downregulated Bcl-2 and survivin mRNA transcripts. CONCLUSION: DPPE obtained through both cold and repeated hot extraction using ethanol showed significantly higher content of diosgenin than the diosgenin content detected in EEPPR. However, diosgenin yield of both the extracts (EEPPR & DPPE) clearly confirmed diosgenin as major and lead phytochemical of Paris polyphylla rhizome of Indian Himalayan landraces. Further, DPPE also demonstrated potent in vitro anti-oxidative and in vivo anti-inflammatory activities and showed in vitro cytotoxicity and significant anti-cancer (apoptosis) effects in MCF-7 breast cancer cells.