Preparation and characterization of the myricetin-loaded solid lipid nanoparticles decorated with folic acid-bound chitosan and evaluation of its antitumor and anti-angiogenic activities in vitro and in vivo in mice bearing tumor models

Abstract

Myricetin is a flavonoid with anticancer properties. This study aimed to formulate myricetin in the form of solid lipid nanoparticles (SLN), decorated with chitosan (CS) and active-targeted with folic acid (FA). After characterization, the in vitro release, cytotoxicity, antioxidant, and ability of the formulation to induce apoptosis using flow cytometry, fluorescent microscopy, and real-time qPCR were examined. Then in vivo anti-angiogenesis on chick chorioallantoic membrane (CAM) and antitumor activities on mice bearing tumor models were investigated. The present study showed that the size of 310 nm and zeta potential of + 30 mV were acceptable for oral administration. The Michaelis–Menten model fitted the drug release pattern with lag during 144 h of the study. The cytotoxicity assay showed that myricetin-SLN-CS-FA significantly killed cancer cells at the concentrations of 6.25, 12.5, 25, 50 and 100 µg/mL (*p < 0.05, **p < 0.01, and ***p < 0.001). The highest level of apoptosis was shown at the concentration of 45 µg/ml in flow cytometry, and fluorescent studies. These results showed the anticancer properties of myricetin-SLN-CS-FA in a dose-dependent manner. The real-time results also indicated that the formulation exerted its cytotoxic effect by activating apoptosis genes. The DPPH, ABTS, and FRAP studies also demonstrated the significant antioxidant properties of the myricetin-SLN-CS-FA (*p < 0.05, **p < 0.01, and ***p < 0.001). The anti-angiogenic activities of the formulations depicted in the CAM assay significantly decrease the number and length of the vessels (*p < 0.05, **p < 0.01, and ***p < 0.001), and also affect VEGF and VEGFR, genes involved in angiogenesis (**p < 0.01, and ***p < 0.001). The antitumor studies indicated the statistically significant effects of myricetin-SLN-CS-FA on reducing tumor volume (*p < 0.05 and ***p < 0.001). The H&E staining of the liver and monitoring of the animal weights also indicated the safety of the formulation. The analysis of mRNA expression in liver and tumor demonstrated that myricetin-SLN-CS-FA exerts its antitumor activities by modulating the inflammatory and oxidative responses in the tissues.