Preparation and characterization of the farnesiferol C-loaded solid lipid nanoparticles decorated with folic acid-bound chitosan and evaluation of its in vitro anti-cancer and anti-angiogenic activities

Abstract

Farnesiferol C (FC) has long been used as a cancer prevention agent. In the present study, FC was loaded into solid lipid nanoparticles (SLN) coated with folic acid (FA) bound chitosan (CS), FC-SLN-CS-FA, by homogenization followed by ultrasonication. Then its toxicity effect on cancer cells compared to normal cells was investigated by the MTT method. In the following, the pro-apoptotic effects of synthesized nanoparticles were evaluated by flow cytometry and acridine orange, propidium iodide, and DAPI staining, and the expression of genes involved in apoptosis was evaluated. The anti-oxidant effects of formulations have been evaluated using DPPH and ABTS assay. To investigate the anti-angiogenic effects of nanoparticles, the chicken chorioallantoic membrane (CAM) assay was used and the expression of vascular endothelial growth factor (VEGF) and VEGF receptor (VEGFR) genes was used by real-time quantitative PCR (qPCR). The formed nanoparticles with a size of 150 nm and a dispersion index of 0.30 had a surface charge of +17 mV. These nanoparticles exhibited significant growth and proliferation inhibitory effects in PANC, AGS, HT-29, MCF7, and A549 human cancer cells from the lowest concentration (15.6 µg/mL, *** p < 0.001). The flow cytometry analysis shows 10.1%, 48.8%, and 74.4% subG1 arrest at the concentrations of 6, 36, and 66 µg/mL of the FC-SLN-CS-FA, respectively. The formulation caused apoptotic cell death by 3-fold overregulating BAX at concentrations of 36 and 66 µg/mL, and 2- and 2.5-fold overexpression of caspase-9 at concentrations of 36 and 66 µg/mL respectively. The treatment with FC-SLN-CS-FA resulted in decreasing BCL2 and NF-κB gene expression. The ABTS and DPPH assays showed the anti-oxidant properties of formulation in a concentration-dependent manner. FC-SLN-CS-FA significantly inhibited angiogenesis in CAM by reducing the length and number of blood vessels. Considering the pro-apoptotic and anti-angiogenic effects of FC-SLN-CS-FA, it can be said that these nanoparticles can effectively suppress the carcinogenesis of multipotent cancer stem cells. Therefore, they have the potential to be studied as an efficient anticancer agent.