Biochemistry And Microbial Sciences - Research Publications

Permanent URI for this collectionhttps://kr.cup.edu.in/handle/32116/27

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Subject: search.filters.subject.Bulbine frutescens

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    Bulbine frutescens phytochemicals as novel ABC-transporter inhibitor: A molecular docking and molecular dynamics simulation study
    (OAE Publishing Inc., 2021-01-08T00:00:00) Kushwaha, Prem Prakash; Maurya, Santosh Kumar; Singh, Amit; Prajapati, Kumari Sunita; Singh, Atul Kumar; Shuaib, Mohd; Kumar, Shashank
    Aim: The present in silico study aimed to evaluate the ATP-binding cassette (ABC) transporter inhibition potential of Bulbine frutescens (B. frutescens) phytochemicals. Methods: Several previous studies and databases were used to retrieve the ligands and target protein structure. The molecular docking study was performed using the Auto Dock Tools, and the GROMACS package was applied to accomplish molecular dynamics simulation. Results: Utilizing the molecular docking and simulation approach, ?25 phytochemicals were screened against the ABC transporter protein. Docking score analysis revealed that B. frutescens phytochemical 4?-Demethylknipholone 2?-?-D-glucopyranoside exhibited strong binding on the ABC transporter protein with a minimum binding score -9.8 kcal/mol in comparison to the standard ABC transporter inhibitor diltiazem (-6.86 kcal/mol). Furthermore, molecular dynamics simulation for 4?-Demethylknipholone 2?-?-D-glucopyranoside showed an acceptable root mean square deviation, radius of gyration, root mean square fluctuation, and hydrogen bond, in addition to other lead compounds. Conclusion: The in-silico study demonstrated that B. frutescens phytochemical 4?-Demethylknipholone 2?-?-D-glucopyranoside possesses anti-drug resistance properties and requires further testing in preclinical settings. � 2021 The Author(s).
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    In silico identification of natural anticancer product and their efficacy in breast cancer cells and cancer stem like cells
    (Central University of Punjab, 2020) Kushwaha, Prem Prakash; Kumar, Shashank
    Breast cancer is the most commonly diagnosed lethal cancer in women worldwide. Notch signaling pathway is directly linked to breast cancer recurrence and aggressiveness. Natural remedies are becoming a prime choice to overcome against cancer due to lesser side effect and cost-effectiveness. Literature survey and in silico study identified Bulbine frutescens (Asphodelaceae), Kurarinone (KU) and 3-O-(E)-p- coumaroylbetulinic acid (CB) as lead plant product/phytochemicals. Methanolic and hexane extract of B. frutescens (BME and BHE respectively), KU and CB were studied for their anticancer activity and notch signaling pathway inhibitory potential in breast cancer cells. Moreover, KU and CB were also studied for their effect in mammosphere. Literature-based identification of methanol soluble phytochemicals of B. frutescens and in silico docking study revealed Bulbineloneside D as a potent notch signaling inhibitor (ϒ-secretase). In silico docking potential of KU and CB were equal to standard gamma secretase inhibitor DAPT (-8.74 kcal/mol). KU-gamma secretase complex showed lower RMSD value, marginal fluctuation in Radius of gyration (Rg), more number of inter hydrogen bonding, and stable secondary structure of the protein which indicates KU as candidate gamma secretase inhibitor (GSI). B. frutescens extracts (IC50 4.8– 28.4 μg/ml), Kurarinone (IC50 0.43-3.42 µM) and CB (IC50 0.99-5.88 µM) significantly decreased cell viability in MDA-MB-231 and T47D cells in time dependent manner. B. frutescens, KU and CB induced cell cycle arrest at G1 phase in MDA-MB-231 and T47D cells. RT-PCR analysis of cell cycle (cyclin D1, CDK4, and p21) and apoptosis modulating genes (caspase 3, Bcl2 and survivin) revealed upexpression of p21, and caspase 3, and down expression of cyclin D1, CDK4, Bcl2 and survivin genes in test extract/phytochemicals treated breast cancer cells. Western Blot analysis showed reduced expression of cyclin D1 and increased procaspase 3 protein expression in extract/phytochemicals treated breast cancer cells in time dependent manner. Fluorescence spectrophotometry and confocal microscopy showed extract/phytochemicals induced nuclear morphology and mitochondrial integrity disruption, and increased reactive oxygen species production in MDA-MB-231 and T47D cells at IC50 and sub IC50 concentration. Flow cytometric apoptosis analysis of extract/phytochemicals treated MDA-MB-231 cells showed significant increase in early apoptotic population in comparison to non-treated cells at IC50 and sub IC50 (half of the IC50) concentration. Dual-Luciferase Reporter assay confirmed notch promoter inhibitory activity of B. frutescens, Kurarinone and CB in HEK293 transfected cells at IC50 concentration. Moreover, RT-PCR analysis showed down regulation of notch responsive genes (Hes1 and Hey1) at transcription levels in extract/phytochemical treated breast cancer cells in time dependent manner. Western Blot analysis showed reduced notch responsive protein (Hes1, Hey1 and E-cadherin) expression in extract/phytochemical treated breast cancer cells. KU and CB treatment decreased the mammosphere formation ability in MCF-7 cells at IC50 concentration by lowering the notch signaling target proteins (Hes1, Hey1, and E-cadherin) and proteins involved in cancer cell self-renewal (c-Myc, SOX-2, CD44). In conclusion, extract/phytochemicals have cell cycle arrest, ROS production, apoptosis induction, and mitochondria membrane potential disruption efficacy in breast cancer cells. KU and CB have the ability to downregulate the notch signaling pathway in breast cancer and cancer stem like cells.
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    Bulbine frutescens phytochemical inhibits notch signaling pathway and induces apoptosis in triple negative and luminal breast cancer cells
    (Elsevier Inc., 2019) Kushwaha P.P.; Vardhan P.S.; Kapewangolo P.; Shuaib M.; Prajapati S.K.; Singh A.K.; Kumar S.
    Breast cancer (BCa) is the most commonly diagnosed lethal cancer in women worldwide. Notch signaling pathway is directly linked to BCa recurrence and aggressiveness. Natural remedies are becoming a prime choice to overcome against cancer due to lesser side effect and cost-effectiveness. Bulbine frutescens (Asphodelaceae), a traditional medicinal plant in South Africa possess bioactive flavonoids and terpenoids. Polar (methanol) and non-polar (hexane) B. frutescens plant extracts were prepared. GC–MS analysis revealed the differential presence of secondary metabolites in both methanolic and hexane extracts. We hereby first time evaluated the anticancer potential of B. frutescens methanolic and hexane extract in triple-negative and luminal BCa cells. B. frutescens extracts significantly decreased cell viability (IC50 4.8–28.4 μg/ml) and induced cell cycle arrest at G1 phase in MDA-MB-231 and T47D cells as confirmed by spectrophotometry and flow cytometry technique. RT-PCR analysis of cell cycle (cyclin D1, CDK4, and p21) and apoptosis modulating genes (caspase 3, Bcl2 and survivin) revealed upexpression of p21, and caspase 3, and down expression of cyclin D1, CDK4, Bcl2 and survivin genes in extract-treated BCa cells. Fluorescence spectrophotometry and confocal microscopy showed B. frutescens induced nuclear morphology and mitochondrial integrity disruption, and increased reactive oxygen species production in MDA-MB-231 and T47D cells. Flow cytometric apoptosis analysis of B. frutescens extracts treated MDA-MB-231 cells showed ≈13% increase in early apoptotic population in comparison to non-treated cells. Dual-Luciferase Reporter assay confirmed notch promoter inhibitory activity of B. frutescens extracts. Moreover, RTPCR analysis showed down regulation of notch responsive genes (Hes1 and Hey1) at transcription levels in extract-treated BCa cells. Western Blot analysis showed increased procaspase 3 protein expression in extract-treated BCa cells. In all the assays methanolic extract showed better anti-cancer properties. Literature-based identification of methanol soluble phytochemicals in B. frutescens and in silico docking study revealed Bulbineloneside D as a potent ϒ-secretase enzyme inhibitor. In comparison to standard notch inhibitor, lead phytochemical showed two additional hydrophobic interactions with Ala80 and Leu81 amino acids. In conclusion, B. frutescens phytochemicals have cell cycle arrest, ROS production, apoptosis induction, and mitochondria membrane potential disruption efficacy in breast cancer cells. B. frutescens phytochemicals have the ability to downregulate the notch signaling pathway in triple-negative and luminal breast cancer cells.