Anti-proliferative, apoptosis inducing, and antioxidant potential of Callistemon lanceolatus bark extracts: an in vitro and in silico study.

The present study reports anticancer and antioxidant activities of Callistemon lanceolatus bark extracts. Anticancer activity was studied against MDA-MB-231 cells. Antioxidant assessment of the chloroform and methanol extracts showed considerable free radical scavenging, metal ion chelating, and reducing power potential. Chloroform extract exhibited potent inhibition of cancer cell proliferation in MTT assay (IC50 9.6 µg/ml) and promoted programmed cell death. Reactive oxygen species (ROS) generation, mitochondria membrane potential (MMP) disruption ability, and nuclear morphology changes were studied using H2-DCFDA, JC-1, and Hoechst dyes, respectively, using confocal microscopy. Apoptotic cells exhibited fragmented nuclei, increased ROS generation, and altered MMP in dose- and time-dependent manner. Chloroform extract upregulated the BAX-1 and CASP3 mRNA expression coupled with downregulation of BCL-2 gene. Further, in silico docking of phytochemicals present in C. lanceolatus with anti-apoptotic Bcl-2 protein endorsed apoptosis by its inhibition and thus corroborated the experimental findings. Obatoclax, a known inhibitor of Bcl-2 was used as a reference compounds.

Identification of Natural Inhibitors Against SARS-CoV-2 Drugable Targets Using Molecular Docking, Molecular Dynamics Simulation, and MM-PBSA Approach.

The present study explores the SARS-CoV-2 drugable target inhibition efficacy of phytochemicals from Indian medicinal plants using molecular docking, molecular dynamics (MD) simulation, and MM-PBSA analysis. A total of 130 phytochemicals were screened against SARS-CoV-2 Spike (S)-protein, RNA-dependent RNA polymerase (RdRp), and Main protease (Mpro). Result of molecular docking showed that Isoquercetin potentially binds with the active site/protein binding site of the Spike, RdRP, and Mpro targets with a docking score of -8.22, -6.86, and -9.73 kcal/mole, respectively. Further, MS 3, 7-Hydroxyaloin B, 10-Hydroxyaloin A, showed -9.57, -7.07, -8.57 kcal/mole docking score against Spike, RdRP, and Mpro targets respectively. The MD simulation was performed to study the favorable confirmation and energetically stable complex formation ability of Isoquercetin and 10-Hydroxyaloin A phytochemicals in Mpro-unbound/ligand bound/standard inhibitor bound system. The parameters such as RMSD, RMSF, Rg, SASA, Hydrogen-bond formation, energy landscape, principal component analysis showed that the lead phytochemicals form stable and energetically stabilized complex with the target protein. Further, MM-PBSA analysis was performed to compare the Gibbs free energy of the Mpro-ligand bound and standard inhibitor bound complexes. The analysis revealed that the His-41, Cys145, Met49, and Leu27 amino acid residues were majorly responsible for the lower free energy of the complex. Drug likeness and physiochemical properties of the test compounds showed satisfactory results. Taken together, the study concludes that that the Isoquercetin and 10-Hydroxyaloin A phytochemical possess significant efficacy to bind SARS-Cov-2 Mpro active site. The study necessitates further in vitro and in vivo experimental validation of these lead phytochemicals to assess their anti-SARS-CoV-2 potential.

Phytochemicals present in Indian ginseng possess potential to inhibit SARS-CoV-2 virulence: A molecular docking and MD simulation study.

Coronaviruses are deadly and contagious pathogens that affects people in different ways. Researchers have increased their efforts in the development of antiviral agents against coronavirus targeting Mpro protein (main protease) as an effective drug target. The present study explores the inhibitory potential of characteristic and non-characteristic Withania somnifera (Indian ginseng) phytochemicals (n ≈ 100) against SARS-Cov-2 Mpro protein. Molecular docking studies revealed that certain W. somnifera compounds exhibit superior binding potential (-6.16 to -12.27 kcal/mol) compared to the standard inhibitors (-2.55 to -6.16 kcal/mol) including nelfinavir and lopinavir. The non-characteristic compounds (quercetin-3-rutinoside-7-glucoside, rutin and isochlorogenic acid B) exhibited higher inhibitory potential in comparison to characteristic W. somnifera compounds withanolide and withanone. Molecular dynamics (MD) simulation studies of the complex for 100 ns confirm favorable and stable binding of the lead molecule. The MMPBSA calculation of the last 10 ns of the protein-ligand complex trajectory exhibited stable binding of quercetin-3-rutinoside-7-glucoside at the active site of SARS-Cov-2 Mpro. Taken together, the study demonstrates that the non-characteristic compounds present in W. somnifera possess enhanced potential to bind SARS-Cov-2 Mpro active site. We further recommend in vitro and in vivo experimentation to validate the anti-SARS-CoV-2 potential of these lead molecules.

Neuroprotective and Neurorescue Mode of Action of Bacopa monnieri (L.) Wettst in 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine-Induced Parkinson's Disease: An In Silico and In Vivo Study.

Ethnopharmacological Relevance: Parkinson's disease (PD) is characterized by progressive death of dopaminergic neurons. The presently used medicines only tackle the symptoms of PD, but none makes a dent on the processes that underpin the disease's development. Herbal medicines have attracted considerable attention in recent years. Bacopa monnieri (L.) Wettst (Brahmi) has been used in Indian Ayurvedic medicine to enhance memory and intelligence. Herein, we assessed the neuroprotective role of Bacopa monnieri (L.) Wettst on Parkinson's disease. Aim of the Study: Bacopa monnieri (L.) Wettst, a medicinal herb, is widely used as a brain tonic. We investigated the neuroprotective and neurorescue properties of Bacopa monnieri (L.) Wettst extract (BME) in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced mice model of PD. Materials and Methods: The mice model of MPTP-induced PD is used in the study. In the neuroprotective (BME + MPTP) and neurorescue (MPTP + BME) experiments, the animals were administered 40 mg/kg body weight BME orally before and after MPTP administration, respectively. Effect of BME treatment was evaluated by accessing neurobehavioral parameters and levels of dopamine, glutathione, lipid peroxide, and nitrites. An in silico study was performed using AutoDock Tools 1.5.6 (ADT). Results: A significant recovery in behavioral parameters, dopamine level, glutathione level, lipid peroxides, and nitrite level was observed in BME-treated mice. Treatment with BME before or after MPTP administration has a protective effect on dopaminergic neurons, as evidenced by a significant decrease in GFAP immunostaining and expression of inducible nitric oxide synthase (iNOS) in the substantia nigra region; however, the degree of improvement was more prominent in mice receiving BME treatment before MPTP administration. Moreover, the in silico study revealed that the constituents of BM, including bacosides, bacopasides, and bacosaponins, can inactivate the enzyme monoamine oxidase B, thus preventing the breakdown of MPTP to MPP+. Conclusion: Our results showed that BME exerts both neuroprotective and neurorescue effects against MPTP-induced degeneration of the nigrostriatal dopaminergic neurons. Moreover, BME may slow down the disease progression and delay the onset of neurodegeneration in PD.

Induction of apoptosis in breast cancer cells by naphthylisoquinoline alkaloids.

Breast cancer is one of the most common types of cancer in the world and a major cause of mortality. Present therapeutic strategies against breast cancer have severe drawbacks such as allergies, damage to healthy tissues, reoccurrence of cancer, and emergence of drug resistance. Naphthylisoquinoline alkaloids are a group of structurally unique natural products produced by tropical lianas belonging to the plant families Dioncophyllaceae and Ancistrocladaceae indigenous to Asia and Africa. These secondary metabolites have been reported to show anti-infective activity, but they also act against leukemic and pancreatic cancer cells. In the present study we have tested the potential of eleven mono- and dimeric naphthylisoquinoline compounds against two breast cancer cell lines, MCF-7 and MDA-MB-231. Three out of the compounds (agents 1, 4, and 11) showed significant activities against both tested cancer cell lines. Further mechanistic investigations revealed that all of the three substances induce apoptotic cell death via its intrinsic pathway by causing deformation of the nuclear membrane, disruption of the mitochondrial membrane potential (MMP), and elevated reactive oxygen species (ROS) production in both cell lines. Flow cytometric analysis using Annexin V - FITC/PI double staining showed an increased number of apoptotic cells in both, the early and the late phases.

Bulbine frutescens phytochemical inhibits notch signaling pathway and induces apoptosis in triple negative and luminal breast cancer cells.

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.