Nano Formulation of Plant Extracts in Treatment of Glioblastoma

Abstract

Glioblastoma is one of the most prevalent and deadly types of solid brain tumors with overall survival of about 15 months even after therapy. pRB pathway has been identified as one of the most important genetic pathways whose alteration contribute to the development of GBM. pRB signaling disruption is found in about 78 percent of GBMs, with the most frequently altered genes being, CDK2, CDK4 and CDK6. The currently available anti-GBM drugs are unable to treat the disease because of their inability to cross blood brain barrier, have high dose related toxicity and cause drug resistance. Thus to address the both stated issues i.e discovering new potential drugs as well as Nano based delivery vehicles, we designed the current study. In-silico analysis was performed at the first phase of the study. Extensive literature search was performed to generate a library of 691 plant extracts having anti-cancer, anti-inflammatory or anti-proliferative properties and these compounds were analyzed through different softwares to shortlist the most potent drug for GBM. MOE and chimera were used to evaluate the binding of ligands with the targeted proteins (CDK2, CDK4, and CDK6). Ligands with high binding energies were further screened through Lipinski rule and DFT analysis to determine the nuclear structure of the compounds and predict their drug-like properties. Silymarin was identified as the top compound targeting CDK4 after the screening. Stearic acid solid lipid nanoparticles (SLNPs) were designed to pass the drug through BBB. Physio-chemical characterizations of SLNPs were performed to assess their size, stability, entrapment efficiency, and release rate. MTT assay on u87, u251 and HEK-293 cell lines for the validation of in-silico results. Silymarin and Silymarin-SLNPs showed obvious decrease in cell viabilities after 72 hours on u87 and u251 cell line. These results were paralleled with Temozolamide (TMZ) with Abstract xii is the standard GBM drug. The cytotoxic effect was also shown by TMZ and TMZ-SLNPs on u251 and this effect was almost equal to the effect shown by Silymarin on u251 at each concentration, however on u87 cell line, TMZ and TMZ-SLNPs showed no effect. Silymarin on HEK-293 after 72hrs showed negligible effect whereas TMZ had some cytotoxicity at higher concentrations. Lastly, Cell migration assay was performed on u87 and U251 cell lines. Cells treated with Silymarin and Silymarin loaded SLNPs showed that they significantly suppressed the migration of U87 and u251 cells and the wound remained unfilled after 24hrs however, cells treated with TMZ and TMZ loaded SLNPs showed very limited hindrance in cell migratory activities of U87 and U251 cells. In conclusion, Silymarin and Silymarin encapsulated SLNPs proved successful in decreasing in vitro GBM cancer cells proliferation and their migration.