Block catiomer polyplexes with regulated densities of charge and disulfide cross-linking directed to enhance gene expression
Block catiomer polyplexes with regulated densities of charge and disulfide cross-linking directed to enhance gene expression
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DOI：
10.1021/ja0379666
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发表时间：
2004-03-03
影响因子：
15
通讯作者：
Kataoka, K
中科院分区：
文献类型：
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作者：
Miyata, K;Kakizawa, Y;Kataoka, K
通过控制主链聚阳离子的阳离子电荷和二硫键交联密度，开发了一种嵌段阳离子聚合物复合物，它在细胞外介质中显示出高稳定性，并能在细胞内区室中高效释放质粒DNA（pDNA）。使用两种巯基化试剂（N - 琥珀酰亚胺基 - 3 -（2 - 吡啶二硫代）丙酸酯（SPDP）或2 - 亚氨基硫醇（Traut试剂））中的任意一种对聚（乙二醇） - 聚（L - 赖氨酸）嵌段共聚物（PEG - PLL）进行巯基化，以研究电荷和二硫键交联密度对聚合物复合物性质的影响。通过SPDP引入巯基是通过形成酰胺键进行的，同时降低了PLL链段的阳离子电荷密度，而Traut试剂通过引入阳离子亚氨基促进巯基化，使电荷密度保持恒定。这些巯基化的PEG - PLL与pDNA复合形成二硫键交联的嵌段阳离子聚合物复合物，其尺寸约为100nm。两种巯基化方法在引入二硫键交联以防止聚合物复合物在细胞外氧化条件下通过反聚阴离子交换而解离方面同样有效。另一方面，只有用SPDP巯基化的聚合物复合物才能在模拟细胞内环境的还原条件下实现pDNA的高效释放，该体系通过二硫键交联补偿了电荷密度的降低。这种对氧化和还原环境的独特敏感性与这两种巯基化聚合物复合物（SPDP型和Traut试剂型）之间转染效率的显著差异密切相关：前者对293T细胞的转染效率比后者高约50倍。显然，巯基化聚合物复合物中阳离子电荷密度和二硫键交联密度之间的平衡在将包封的pDNA递送至细胞内区室的微环境并实现其控释以达到高转染效率方面起着至关重要的作用。
A block catiomer polyplex, showing a high stability in the extracellular medium and an efficient release of plasmid DNA (pDNA) in the intracellular compartment, was developed by controlling both the cationic charge and disulfide cross-linking densities of the backbone polycations. Poly(ethylene glycol)poly(L-lysine) block copolymer (PEG-PLL) was thiolated using either of two thiolation reagents, N-succinimidyl 3-(2-pyridyldithio)propionate (SPDP) or 2-iminothiolane (Traut's reagent), to investigate the effects of both the charge and disulfide cross-linking densities on the properties of the polyplexes. The introduction of thiol groups by SPDP proceeded through the formation of amide linkages to concomitantly decrease the cationic charge density of PLL segment, whereas Traut's reagent promoted the thiolation with the introduction of cationic imino groups to keep the charge density constant. These thiolated PEG-PLLs were complexed with pDNA to form the disulfide cross-linked block catiomer polyplexes, which had the size of approximately 100 nm. Both thiolation methods were similarly effective in introducing disulfide cross-links to prevent the polyplex from the dissociation through a counter polyanion exchange in the extracellular oxidative condition. On the other hand, the efficient release of pDNA responding to the reductive condition mimicking the intracellular environment was only achieved for the polyplex thiolated with SPDP, a system compensating for the decrease in the charge density with the disulfide cross-linking. This distinctive sensitivity toward oxidative and reductive environments was nicely correlated with the remarkable difference in the transfection efficiency between these two types of thiolated polyplexes (SPDP and Traut's reagent types): the former revealed approximately 50 times higher transfection efficiency toward 293T cells than the latter. Obviously, the balance between the densities of the cationic charge and disulfide cross-linking in the thiolated polyplex played a crucial role in the delivery and controlled release of entrapped pDNA into the microenvironment of intracellular compartment to achieve the high transfection efficiency.