Development Of Anisamide-targeted PEGylated Gold Nanorods To Deliver Epirubicin For Tumor Chemo-photothermal Therapy

Prostate cancer is a kind of common malignant tumors of the male urinary system.The incidence of prostate cancer is increasing year by year.It is the sixth common malignant tumour in China.And it has gradually become the primary problem threatening the health of middle-aged and old men in China.Melanoma is one of the most lethal skin cancers.Because of its easy escape from immune surveillance,strong invasiveness and migration ability,it is easy to induce drug resistance,it is easy to metastasis and other factors,it brings great trouble to clinical treatment.In recent years,targeting drug delivery system has become a hot issue in the field of cancer therapy.Epirubicin is one of the important chemotherapeutic drugs for prostate cancer and melanoma,but it has high cardiotoxicity.And cardiac toxicity may be one of the important reasons for the increase of non-cancer mortality after chemotherapy.With the development of many drug delivery vectors,new drug delivery vectors are used to achieve targeting delivery of epirubicin which can achieve more advantages for target tumors,more clearance efficiency and less biological toxicity possibly.Nanomaterial-assisted photothermal therapy?PTT?has recently emerged as a potential therapeutic strategy for solid tumours.Due to the optical and electronic properties namely the surface plasma resonance?SPR?,gold nanoparticles?AuNRs?with specific structures,including gold nanorods?AuNRs?,gold nanoshells,and gold nanocages,have been developed to absorb light strongly in certain wavelengths and convert the light energy into heat,resulting in cancer cell death by apoptosis and/or necrosis.Among these nanostructures,AuNRs demonstrate unique physicochemical properties,better biocompatibility,larger specific surface area and easy functionalization,which can be used as excellent drug delivery carriers to achieve targeted delivery of drug molecules,In that they may strongly absorb light in wavelengths between 650 and 900 nm within the near-infrared?NIR?region.It is known that for the light wavelengths between 650 and 2,000 nm,the tissue absorption is weak,and therefore,tumours deeply within the body may be destroyed by PTT induced by AuNRs.However,the heterogeneous heat distribution inside tumours caused by the uneven distribution of AuNRs has curtailed the PTT efficacy.Therefore,novel therapeutic strategies are required to facilitate synergistic effects when combined with PTT.In this study,based on the determination of epirubicin by high performance liquid chromatography and ultraviolet-visible spectrophotometry,AuNRs with tunable longitudinal SPR(?_max=700,725,765 and 800 nm)were initially synthesised in the presence of hexadecyltrimethylammonium bromide?CTAB,it promotes the growth of gold seeds in one direction?to produce positively charged gold nanorods?Au-CTAB?.The Au-CTAB was subsequently coated by polyacrylic acid?PAA?to generate negatively charged Au-CTAB-PAA.To improve physiological stability and target cancer cells,Au-CTAB-PAA was further modified with PEG and PEGylated anisamide?AA,a ligand known to target the sigma receptor overexpressed on a variety of human cancers,such as breast cancer,melanoma,non-small cell lung carcinoma,and prostate cancer?to achieve Au-CTAB-PAA-PEG and Au-CTAB-PAA-PEG-AA.The Au-CTAB-PAA-PEG-AA was able to complex Epirubicin?EPI,it is positively charged at physiological pH?via the electrostatic interaction forming Au-CTAB-PAA-PEG.EPI and Au-CTAB-PAA-PEG-AA.EPI complex.Then the optimum preparation conditions of Au-CTAB-PAA-PEG.EPI and Au-CTAB-PAA-PEG-AA.EPI were explored by taking the drug encapsulation rate as the evaluation index,and the change of particle size and Zeta potential.The optimum preparation conditions were as follows:the ratio of drug to EPI:AuNRs?w:w?=1:5,the incubation temperature was 45?,and the incubation time was 12 h.At this time,the encapsulation rate of EPI was the highest,reaching about 90%.Furthermore,we selected Au₈₀₀-CTAB-PAA-PEG.EPI and Au₈₀₀-CTAB-PAA-PEG-AA.EPI as the research objects of photothermal therapy for tumors,considering the photothermal absorption characteristics of gold nanorods and the difference of encapsulation efficiency of gold nanorods with different longitudinal absorption peaks.The nano-delivery system was characterized by uniform particle size,uniform distribution and good stability.And the resultant complex also demonstrated pH-dependent drug release,the cumulative release rate of epirubicin was about 80%within 48 hours in the slightly acidic environment of tumors,which was significantly different from that in the neutral environment.Laser irradiation at a specific wavelength?800 nm,2.5W/cm²?can indeed increase the surface temperature of gold nanorods and improve the photothermal conversion efficiency,which have facilitated nucleus trafficking of EPI,the surface temperature of gold nanorods can indeed be increased by laser irradiation at a specific wavelength.and inducedanti-proliferative effects in human prostate cancer PC-3 cells and melanoma B16 cells.And the study also confirmed that both targeted and non-targeted agents Au₈₀₀-CTAB-PAA-PEG-AA.EPI exhibited better cytotoxicity and induced cell apoptosis more efficiently,but targeted agents Au₈₀₀-CTAB-PAA-PEG-AA.EPI exhibited better therapeutic effects with higher intracellular accumulation and more significant endosomal escape.Targeted and non-targeted gold nano-dosage forms can inhibit the proliferation of cancer cells.But the targeted preparation is better.Moreover,when the targeted preparation was stimulated by 808 nm,2.5 W/cm²,Tumor cytotoxicity of the target preparation was better than that of the free EPI group and the non-targeted agents group.In vivo experimental results showed that:when stimulated with desired laser irradiation?808nm,2.5 W/cm²?,the resultant complex of Au₈₀₀-CTAB-PAA-PEG-AA with EPI demonstrated significantly greater anti-tumour effects in a prostate tumour xenograft mouse model when compared to either of individual monotherapies,the growth of tumor cells was further delayed,and there was no biological toxicity and other adverse reactions.In summary,the preparation of anisamide-targeted PEGylated gold nanorods to deliver epirubicin provides a promising CPTT strategy for cancer therapy and demonstrating a promising strategy for clinical application of CPTT in cancer.