Synergic Therapy Of Melanoma Using GNRs-MUA-PEI/siIDO2-FA Through Targeted Gene Silencing And Plasmonic Photothermia

Objectives:To construct the GNR-MUA-PEI-FA nanometer gene carrier, to detect the target of the nanometer gene carrier to melanoma and the efficiency of gene silencing, and to evaluate the therapeutic effect of photothermal therapy in mouse melanoma using GNR-MUA-PEI-FA Methods:1. The gold nanoparticles(GNRs) were synthesized according to E-Eealy's method. syThe GNRs were characterized by ultraviolet spectrum, infrared spectrum and the transmission electron microscopy(TEM).2. The cytotoxicity of GNRs was determined by MTT test.3. The loading capacity of siRNA by GNRs was determined by by the gel block experiment. The effects of transfecting siRNA by was determined by flowcytometry.4. In vitro phontothermal effects of GNR-MUA-PEI-FA was induced by near infrared laser(808 nm). The killing ability to tumor cells was evaluated by MTT test.5. The efficacy of gene silencing by GNRs-MUA-PEI-FA/siIDO2 was detected by qPCR and WB. The apoptosis of was determined by by Annexin V and PI staining then using flow cytometry to detect cell apoptosis rate.6. The dermal administration of GNR-MUA-PEI-FA/cy3-siGAPDH was achieved by mix with 50% glycerol, 10% DMSO and apply it to the skin of tumor-bearing mice. The efficacy of interdrmal distribution of GNRs-MUA- PEI-FA/ cy3-siGAPDH was detected in the frozen section of skin by fluorescence microscope..7. The therapeutic effects of GNR-MUA-PEI-FA/siIDO2 were evaluated in a murine melanoma model. Results:1. Ultraviolet spectrum absorption peak of the gold nanorods after modification occurs obvious redshift, The particle size, dispersion were observed using TEM, and after modification there were no obvious changes.2. The gold nanorods after modification possessed low cytotoxicity, good biocompatibility and biological safety.3. The gel block experiments show,when the weight ratio of the gold nanorods to siRNA was 8:1, gold nanorods combined all siRNAs; In order to improve the cell intake of siRNA, so we increase the weight ratio of gold nanorods to siRNA to 15:1.4. The B16-BL6 cells treated with GNR-MUA-PEI-FA/siIDO2 achieved the maximum killing effects under the condition of 2 w/cm2, 5 min of laser irradiation u5. Treatment with GNR-MUA-PEI-FA/siIDO2 showed therapeutic effective to melanoma. Conclusion:1. The gold nanorods gene carrier can deliver siRNA into melanoma cells effectively, and silence genes subsequently.2. Treatment of GNR-MUA-PEI-FA/siIDO2 is an alternative method photothermal therapy, which combines phototheral effect and gene silencing of IDO2, achieving a synergistic therapeutic effects for melanoma.