Research of FTC-CD133 Nanoparticles Inhibiting the Drug Resistance of Liver Cancer Stem Cell

Abstract

Objective: To prepare the nanoparticles packing FTC, a specific inhibitor of multidrug transporter BCRP/ABCG2, and connecting CD133 antibody, and to evaluate its features and therapeutic effect.

Methods: Nanoparticles packing FTC and with PLGA as carrier were prepared using multiple emulsion-solvent evaporation method, and carbodiimide reaction was undertaken to connect CD133 antibody on the surface of the nanoparticles, forming the nanoparticles of FTC-PLGA-NP-C; the particle size distribution, FTC encapsulation efficiency, drug loading capacity and the antibody connection efficiency of nanoparticles were analyzed; CD+133 SP cells were sorted by CD+133 magnetic activated cell sorting (MACS), MTT method was used to measure the multiplication capacity of CD+133 SP cells, and western blotting method was employed to detect the expression of ABCG2 in SP cells; the combination efficiency of FTC-PLGA-NP-C and CD+133 SP was observed under fluorescence microscope; CD+133 SP cells were added into nanoparticles of FTC-PLGA-NP-C of different concentrations, and western blotting was used to detect the expression of ABCG2; with PBS of the same volume taken as control, FTC-PLGA-NP-C nanoparticles were added into CD+133 SP cells with adriamycin added at the same time, and the concentration of adriamycin in the cells was determined by HPLC at different time points of incubation. Tumor was transplanted to the armpit of nude mice, and tail intravenous injection of FTC-PLGA-NP-C or 0.9% sodium chloride solution was undertaken after the formation of tumor; 48 hours later, tail intravenous injection of adriamycin was conducted, tumor diameter was measured, and high efficiency liquid chromatography was employed to measure the concentration of adriamycin in the tumor tissue of mude mice.

Results: FTC-PLGA-NP-C was in sphere shape, with a size distribution of 60-120 nm, a drug loading capacity of 11.27% and an encapsulation efficiency of 37.18%, and had a better combining capacity with CD+133 SP cells; western blotting showed that ABCG2 expression existed in CD+133 SP cells and decreased with the increase of the dosage of FTC-PLGA-NP-C nanoparticles with no ABCG2 expression detected when the dosage of FTC-PLGA-NP-C was 4 mg/ml. At 24 hours of incubation, the concentration of adriamycin began to decrease quickly to below 1 μg/ml in the CD+133 SP cells in PBS, while the concentration of adriamycin in the CD+133 SP cells in FTC-PLGA-NP-C remained at 2-3 μg/ml. The diameter of the mice tumor of normal saline group was (1.845±0.076) cm, higher than (1.238±0.072)cm of FTC-PLGA-NP-C group (t=5.802, P＜0.001); the concentration of adriamycin in the tumor of normal saline group was (2.005±0.154)μg/ml, lower than (1.238±0.072)cm of FTC-PLGA-NP-C group (t=6.088, P＜0.001).

Conclusion: Prepared drug-loading nanoparticle FTC-PLGA-NP-C could effectively target liver cancer stem cells, inhibit the expression of drug resistance protein ABCG2, reduce the drug resistance of tumor cells to conventional drugs and improve the therapeutical effect of chemotherapeutic drug.