主客体组装的金刚烷甲酸-阿霉素/聚阳离子材料及体外抗肿瘤活性的研究

Synthesis of a supermolecular nanoparticle γ-hy-PC/Ada-Dox and its antitumor activity

目的:合成以金刚烷甲酸阿霉素(Ada-Dox)为客体,聚乙烯亚胺PEI600-γ-羟丙基环糊精(γ-hy-PC)为主体的超分子纳米材料,观察其理化特性和转基因功能。方法:通过主客体相互作用将Ada-Dox组装到γ-hy-PC材料上,制成γ-hy-PC/Ada-Dox的载药纳米粒子。用1H-NMR、NOESY、UV-Vis、XRD、TGA对其结构进行表征,粒径和电势的测定和凝胶电泳阻滞实验观察其浓缩质粒DNA的能力,在人肝癌细胞BEL-7402和SMMC-7721细胞上对载体材料的细胞毒性进行评价,并进行细胞迁移和细胞形态学鉴定实验,在HEK293细胞上进行体外转染实验以及在BEL-7402细胞上进行细胞吞噬实验。结果:1H-NMR、NOESY、UV-Vis、XRD、TGA证实成功地合成了γ-hy-PC/Ada-Dox复合物;UV-Vis测得其载药量分别是0.5%和5.5%;载药量0.5%和5.5%的γ-hy-PC/Ada-Dox分别在N/P为3和4时可以浓缩质粒DNA;MTT实验结果显示材料的毒性均低于PEI 25 kDa;细胞迁移与H/E染色实验表明合成的材料都有一定的抗肿瘤活性;体外转染实验显示,γ-hy-PC具有较高的转染效率,且随着载药量的增加,转染效率逐渐降低;细胞吞噬实验结果表明γ-hy-PC/Ada-Dox可同时携带药物和FAM-siRNA进入细胞。结论:成功地合成了γ-hy-PC/Ada-Dox复合物,并表现出一定基因转染效率和抗肿瘤活性。

Objective： To synthesize a（2-Hydroxypropyl）-γ-cyclodextrin-polyethylenimine/adamantane-conjugated doxorubicin（γ-hy-PC/Ada-Dox）based supramolecular nanoparticle with host-guest interaction and to identify its physicochemical characterizations and antitumor effect.Methods： A novel non-viral gene delivery vector γ-hy-PC/Ada-Dox was synthesized based on host-guest interaction.1H-NMR,NOESY,UV-Vis,XRD and TGA were used to confirm the structure of the vector.The DNA condensing ability of complexes was investigated by particle size,zeta potential and gel retardation assay.Cytotoxicity of complexes was determined by MTT assay in BEL-7402 and SMMC-7721 cells.Cell wound healing assay was performed in HEK293 and BEL-7404 cells.The transfection efficiency was investigated in HEK293 cells.H/E staining and cell uptake assay was performed in BEL-7402 cells.Results： The structure of γ-hy-PC/Ada-Dox was characterized by 1H-NMR,NOESY,UV-Vis,XRD,TGA.The drug loading was 0.5% and 5.5%.Gel retardation assay showed that γ-hy-PC was able to completely condense DNA at N/P ratio of 2;0.5% and 5.5% γ-hy-PC/Ada-Dox was able to completely condense DNA at N/P ratio of 3 and 4,respectively.The cytotoxicity of polymers was lower than that of PEI25KDa.The transfection efficiency of γ-hy-PC was higher than that of γ-hy-PC/Ada-Dox at N/P ratio of 30 in HEK293 cells;and the transfection efficiency was decreasing when Ada-Dox loading was increasing.Cell uptake assay showed that γ-hy-PC/Ada-Dox was able to carry drug and FAM-siRNA into cells.Conclusion： The novel vector γ-hy-PC/Ada-Dox has been developed successfully,which has certain transfection efficiency and antitumor activity.