Preparation Of SiRNA Targeting HIF-1α Loaded TPGS-b-(PCL-ran-PGA) And Its Influence On The Growth Of Nasopharyngeal Carcinoma Cell

Objective To investigate the influence of small interference RNA(siRNA) targeting hypoxia inducible factor-1α(HIF-1α) loaded TPGS-b-(PCL-ran-PGA) nanoparticles(NPs) on the growth of nasopharyngeal carcinoma(NPC) cell in vitro and in vivo. Methods(1) The siRNA loaded NPs were prepared using a double emulsion method. The particle size, particle morphology, encapsulation efficiency and the in vitro release profile of siRNA loaded NPs were investigated.(2) The cellular uptake of the siRNA loaded NPs and the siRNA distribution in the cells were observed using a confocal laser scanning microscope. The expression level of HIF-1α in CNE-2 cells were detected by Real-time PCR and Western blot. The cell viability was determined using MTT assay.(3) Xenograft tumor models were developed by inoculation of CNE-2 in nude mice, and the tumors were injected with the naked siRNA targeting HIF-1α, blank TPGS-b-(PCL-ran-PGA) NPs, scrambled siRNA loaded TPGS-b-(PCL-ran-PGA) NPs or siRNA targeting HIF-1α loaded TPGS-b-(PCL-ran-PGA) NPs. The tumor volume, weight and inhibition ratio were measured and calculated. The expression level of HIF-1α in the xenograft tumors were detected by Real-time PCR and Western blot. Histopathological changes were observed by inverted phase contrast microscope. Results(1) The average diameter and encapsulation efficiency of the siRNA loaded NPs were 275.72±2.94 nm and 70.20±1.91%, respectively. The siRNA loaded NPs were spherical in shape and the gradual release pattern of siRNA loaded nanoparticles was observed from in-vitro release experiment.(2) The fluorescent of FAM- siRNA loaded NPs could be internalized into the cells and the siRNA were located in the cytoplasm and around the nuclei. The expression of HIF-1α m RNA and protein were inhibited after siRNA loaded NPs transfection. The siRNA loaded NPs had significant cytotoxic effect on cell survival and the cytotoxicity of the CNE-2 cells was significantly enhanced with dose increasing.(3) In comparison of naked siRNA targeting HIF-1α treatment group, the inhibitory ratio of blank TPGS-b-(PCL-ran-PGA) NPs, scrambled siRNA loaded TPGS-b-(PCL-ran-PGA) NPs and siRNA targeting HIF-1α loaded TPGS-b-(PCL-ran-PGA) NPs treatment group were 23.53%、28.63% and 52.94%, respectively. The expression level of HIF-1α were significant decreased in siRNA loaded NPs treatment group compared with the other three groups. Tumor cells necrosis and apoptosis were markedly increased in siRNA loaded NPs treatment group. Conclusions The siRNA targeting HIF-1α loaded TPGS-b-(PCL-ran-PGA) NPs could be transfected into cells, significantly decreased the expression level of HIF-1α and inhibited the growth of NPC and promoted tumor cells necrosis and apoptosis. TPGS-b-(PCL-ran-PGA) NPs could function as an efficient carrier for siRNA targeting HIF-1α delivery and may have a bright prospect in the research field of gene therapy.