摘要
目的:多肽MC10(MARAKEGGGC)介导的聚乙烯亚胺-β-环糊精聚合物载体作为新型靶向性非病毒基因载体的研究。方法:低分子量的聚乙烯亚胺(PEI)通过羰二咪唑(1,1'-carbonyl-diimidazole,CDI)偶联β-环糊精(β-CyD)合成聚乙烯亚胺-β-环糊精(PEI-β-CyD)作为载体骨架,用琥珀酰亚胺-3-(2-嘧啶二硫)丙酸酯[N-succinimidy-3-(2-pyridyldithio)propionate,SPDP]将MC10偶联到PEI-β-CyD上,制成新型转基因载体材料MC10-PEI-β-CyD。用核磁共振氢谱(1H-NMR)对载体的结构进行化学表征,凝胶电泳阻滞实验研究载体材料对质粒DNA的浓缩能力,透射电镜观察其浓缩质粒DNA后形成的微粒粒径。MTT法检测聚合物的毒性,用SKOV-3、A549和MCF-7细胞株进行体外转基因实验,确定MC10-PEI-β-CyD具有基因携带能力及靶向选择性。结果:成功地合成了MC10-PEI-β-CyD,且MC10-PEI-β-CyD在N/P为5时,能够有效浓缩质粒DNA,形成粒径约为(170±35)nm的微粒。结论:MC10-PEI-β-CyD具有低毒和高转基因效率,对Her-2受体的肿瘤细胞具有很好的靶向性。
Objective： To develop a novel non-viral gene delivery vector based on polyethylenimine and β-cyclodextrin targeting to Her-2 receptor （MC10-PEL-β-CyD）. Methods： The PEI -β-CyD was synthesized by low molecular weight polyethylenimine （PEI, Mw 600） cross-linked 13-cyelodextrin （β-CyD） via N, N-carbonyldiimidazole （CDI）. The chemical linker [-N-succinimidy-3-（2-pyridyldithio） propionate, SPDP] was used to bind peptide MC10 （MARAKEGGGC） to PEI-β-CyD to form the vector MC10-PEI-β-CyD. The 1H-NMR was used to confirm the structure of vector. The DNA condensing ability,and the particle size of MC10 -PEI-β-CyD/DNA complexes were demonstrated by gel retardation assay and electron microscope observation （TEM）. Cell viability was tested by MTT assay. The transfection efficiency was determined on cultured SKOV 3,A549 and MCF-7 cells. Results： MC10 was linked onto PEI-β-CyD successfully. The vector was able to condense DNA at N/P ratio of 5 and particle size was about （170 ±35） nm. The vector showed low cytotoxicity and high transfection efficiency in cultured SKOV 3,A549 and MCF-7 cells. Conclusion： A novel non-viral vector MC10-PEI-β-CyD with low cytotoxicity and high transfection efficiency has been successfully synthesized.
出处
《浙江大学学报（医学版）》
CAS
CSCD
北大核心
2009年第1期7-14,共8页
Journal of Zhejiang University(Medical Sciences)
基金
国家高技术研究发展计划(863)资助项目(2007AA03Z355)
国家重点基础研究规划(973)资助项目(2004CB518802)
国家自然基金项目(30571068)
浙江省自然科学基金重点项目(Z207572)
关键词
聚乙烯亚胺
β环糊精类
遗传载体
转染
质粒
肽类
肿瘤/治疗
Polyethyteneimine
beta-Cyclodextrins
Genetic vectors
Transfection
Plasmids
Peptides
Neoplasms/ther
