Preparation, Characterization And Targeting Anti-cancer Rearch Of Rapamycin Loaded Carboxymethylchitosan-coated Magnetic Nanoparticles

In recent years, with the in-depth study of the molecular bio-technique and the molecular mechanism of malignant tumors, the molecular-targeted therapy is emerging as a research focus for the treatment of malignant tumor. Magnetic-targeted drug delivery system, which possesses the targeting and long circulating properties, is comprised by the skeleton materials, superparamagnetic materials and the anti-cancer drug. When the magnetic nanocomposites were injected into body through blood vessels, they can locate in the targeted site under the control of an external magnetic field, and release drug slowly at the targeted site to kill tumor cells, resulting the reduction of side effects caused by systemic distribution of drug. In this study, we chose Fe3O4as the magnetic core, carboxymethylchitosan as the carrier and Rapa as the targeted drug to prepare Rapa loaded carboxymethylchitosan-coated magnetic nanoparticles(Fe3O4/CMCS-Rapa MNPs)with a new approach. Experimental results showed that the Fe3O4/CMCS-rapa drug delivery system possessed a high drug loading content, exhibited good drug release and magnetic targeting properties. And then its in vitro anti-hepatoma effect was detected.Firstly, the Fe3O4/CMCS-Rapa MNPs were prepared separately by emulsion cross-linking method and solvent evaporation method with CMCS as the basic skeleton, Fe3O4nanoparticles as magnetic materials and Rapa as the anti-tumor drug. The prescription procedure of Fe3O4/CMCS-Rapa MNPs was optimized with the help of the Box-Behnken experimental design based on second-order response surface methodology(RSM). The optimized prescription procedure was as following:the water/oil ratio was13.97:1; the Fe3O4/CMCS:Rapa ratio was8.42:1; the stirring rate was831rpm; the optimized drug loading content and encapsulation efficiency were6.32%±3.36%and62.9%±2.30%, respectively.Secondly, Fe3O4/CMCS-Rapa MNPs were characterized and analysized by FTIR, XRD, SEM, TEM, XPS, TG/DSC, VSM and the in vitro release experiment, and the possible reaction mechanism of Fe3O4/CMCS-Rapa MNPs was also dicussed. The in vitro drug release kinetics of Fe3O4/CMCS-Rapa MNPs was studied by the dialysis diffusion method, which showed that Fe3O4/CMCS-Rapa MNPs hold an obvious slow release characteristic. The in vitro release curve matched well with the Higuchi and Retiger-Pepper equation model, revealing the Fe3O4/CMCS-Rapa MNPs’dissolution rate is controlled by the drug diffusion rate.Finally, the magnetic targeting experiment, the Prussian blue staining experiment, the MTT assay, CCK-8assay, the Hoechst staining and the AnnexinV/FITC-PI double staining experiments were carried out to study the in vitro anti-tumor activity of Fe3O4/CMCS-Rapa MNPs from both qualitative and quantitative aspects by choosing HepG2cells as the tumor cell and LO2cells as the comparison. Results showed that the Fe3O4/CMCS-Rapa MNPs could inhibit the proliferation of HepG2cells in vitro, which was time and concentration dependent. The Fe3O4/CMCS-Rapa MNPs could locate in the targeted site under the control of an external magnetic field, release drug slowly at the targeted site which was able to reduce local drug concentration and avoid the damage to normal cells, and improve the effective utilization of Rapa by prolonging the circulation time.