以聚乙烯亚胺-β-环糊精为骨架偶联短肽CY11的转基因载体  

Peptide CY11 conjugated polyethylenimine-β-cyclodextrin for gene delivery

作　　者：虎义平[1] 姜启英[1] 陈丹[1] 赵丹军[1] 陆晓[1] 余海[2] 王青青[2] 周峻[1] 胡秀荣[3] 汤谷平[1] 

机构地区：[1]浙江大学化学生物和药物研究所,浙江杭州310028 [2]浙江大学医学院免疫学研究所,浙江杭州310058 [3]浙江大学分析测试中心,浙江杭州310028

出　　处：《浙江大学学报（医学版）》2009年第1期24-30,共7页Journal of Zhejiang University(Medical Sciences)

基　　金：国家重点基础研究(973)资助项目(2004CB518802);国家高科技研究发展计划(863计划)项目(2007AA03Z355);国家自然科学基金项目(30571068);浙江省自然科学基金重点项目(Z207572);浙江省环境工程重中之重学科开放基金(20050215)

摘　　要：目的:以聚乙烯亚胺-β-环糊精(PEI-β-CyD)作为载体骨架,用靶向于肿瘤表面成纤维细胞生长因子受体(FGFR)的短肽CY11(CGMQLPLATWY)偶联于载体材料上,研究该载体材料的基因转染效率。方法:用琥珀酰亚胺-3-(2-嘧啶二硫)丙酸酯[N-succinimidy-3-(2-pyridyldithio)propionate,SPDP]将CY11偶联到PEI-β-CyD上,合成CY11-PEI-β-CyD。核磁共振氢谱(1H-NMR)、热重分析(TGA)对载体材料进行化学表征,凝胶电泳阻滞实验研究载体材料对质粒DNA的浓缩能力,MTT法测定载体材料的细胞毒性,在COS-7、Hela和B16细胞株上进行体外转染实验。结果:成功合成材料CY11-PEI-β-CyD,在N/P为4时能够有效地浓缩质粒DNA。在B16细胞上,材料CY11-PEI-β-CyD达到160μg/ml时,细胞存活率仍高于80%。在CY11-PEI-β-CyD/DNA复合物在N/P为20∶1时,转染效率是对照组PEI-β-CyD的17倍。结论:CY11-PEI-β-CyD是具有低毒性、高转染且有FGFR靶向性的新型非病毒转基因载体。Objective： To develop a novel non viral gene delivery vector CY11-PEI-β-CyD and to test its gene transfection efficiency. Methods： CY11 （CGMQLPLATWY） was conjugated to polyethylenimine-β-cyclodextrin to form CY11 PEI-β-CyD with a cross-linker [-N-succinimidy-3 （2-pyridyldithio） propionate,SPDP]. 1H-NMR and TGA were used to confirm the structure of vector. The DNA condensing ability of CY11- PEI-β-CyD was investigated by gel retardation assay. Cytotoxicity of CYll PEI-β-CyD was determined by MTT assay and transfection efficiency was investigated in COS-7,Hela and B16 cells. Results： CY11 was conjugated onto PEI-β-CyD successfully, confirmed bylH NMR and TGA. The novel vector effectively condensed DNA at N/P ratio of 4. It showed low cytotoxicity up to the concentration was 160μg/ml. The transfection efficiency was 17-fold higher than that of PEI 25 kDa at N/P ratio of 20. Conclusion： The novel vector CY11 -PEI-β-CyD with low eytotoxic and high transfection efficiency may be used as a potential carrier for gene delivery.

关 键 词：聚乙烯亚胺 β-环糊精类 遗传载体 转染 质粒 肽类 受体 成纤维细胞生长因子 

分 类 号：R73-36[医药卫生—肿瘤]