HIGHLY EFFICIENT DNA DELIVERY MEDIATED BY PH-SENSITIVE IMMUNOLIPOSOMES
HIGHLY EFFICIENT DNA DELIVERY MEDIATED BY PH-SENSITIVE IMMUNOLIPOSOMES
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DOI：
10.1021/bi00450a039
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发表时间：
1989-11-28
期刊：
影响因子：
2.9
通讯作者：
HUANG, L
中科院分区：
文献类型：
--
作者：
WANG, CY;HUANG, L
我们先前已经表明，pH敏感性免疫脂质体可以介导质粒DNA向小鼠模型中生长的肿瘤细胞的靶特异性递送[Wang，C.是的，ANG。黄湖，澳-地等人（1987）Proc. Acad. Sci. U.S.A.84，7851-7855]。在靶细胞转化频率方面的递送效率现在已经被表征为在组织培养系统中的短期和长期基因表达。单纯疱疹病毒胸苷激酶（TK）基因作为报告基因。它被置于大鼠磷酸烯醇丙酮酸羧激酶基因启动子的控制下，该基因含有cAMP调节元件。因此，外源基因在靶细胞-小鼠Ltk-细胞中的表达可以通过cAMP药物调节。使用去污剂透析法将质粒DNA包封在脂质体中。基因递送的效率相对于脂质体相关DNA的时间过程和剂量进行了优化。脂质体中抗体的存在是保证DNA最大限度释放的必要条件。递送也依赖于脂质体的脂质组成。pH敏感性脂质组合物的效率比相应的pH不敏感性组合物高8倍。靶细胞的转化效率也依赖于基因表达的调节;与二丁酰-cAMP和茶碱孵育的细胞显示出比没有药物孵育的细胞高得多的转化频率水平。当优化所有脂质体和孵育参数时，Ltk-细胞显示出47%的短期转化效率和2%的长期转化效率。这些效率水平显著高于用常规磷酸钙沉淀法获得的效率水平，所述常规磷酸钙沉淀法作为对比平行进行。
We have previously shown that pH-sensitive immunoliposomes can mediate a target-specific delivery of plasmid DNA to tumor cells grown in a mouse model [Wang, C.-Y., .ANG. Huang, L. (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 7851-7855]. The efficiency of delivery in terms of the target cell transformation frequency has now been characterized for both short- and long-term gene expression in a tissue culture system. Herpes simplex virus thymidine kinase (TK) gene was used as a reporter gene. It was placed under the control of the promoter for the rat phosphoenolpyruvate carboxykinase gene, which contains a cAMP regulatory element. Therefore, the expression of the exogenous gene in the target cell, mouse Ltk- cells, can be regulated by cAMP drugs. The plasmid DNA was encapsulated in liposomes using a detergent dialysis method. The efficiency of gene delivery was optimized with respect to the time course and dose of liposome-associated DNA. The existence of antibody of the liposomes was essential for the maximal level of DNA delivery. Delivery was also dependent on the lipid composition of the liposome. The pH-sensitive lipid composition gave 8-fold higher efficiency than the corresponding pH-insensitive composition. The transformation efficiency of the target cell also depended on the regulation of gene expression; cells incubated with dibutyryl-cAMP and theophylline showed a much higher level of transformation frequency than cells incubated without the drugs. When all liposome and incubation parameters are optimized, the Ltk- cells showed a 47% efficiency for the short-term transformation, and 2% for the long-term transformation. These levels of efficiency are considerably higher than those obtained with the conventional calcium phosphate precipitation method which was performed in parallel as a comparison.