Key Laboratory of Biomedical Polymers, Ministry of Education, Department of Chemistry, Wuhan University, Wuhan 430072, PR China.
Biomaterials. 2013 May;34(15):3858-69. doi: 10.1016/j.biomaterials.2013.01.101. Epub 2013 Feb 26.
Reduction-sensitive micelles were prepared from monomethoxy-poly(ethylene glycol)-S-S-hexadecyl (mPEG-S-S-C16), an amphiphilic poly(ethylene glycol) derivative containing a disulfide bond. The micelles were then used for the intracellular delivery of the anticancer drug doxorubicin (DOX) into tumor cells, and the cellular uptake mechanisms of the micelles were determined. To serve as a control, monomethoxy-poly(ethylene glycol)-C-C-hexadecyl (mPEG-C-C-C16) with an analogous structure but without a disulfide bond was also prepared. The polymer could self-assemble into micelles in an aqueous solution and be loaded with high-content DOX. In vitro release studies revealed that DOX-loaded mPEG-S-S-C16 micelles released DOX faster than DOX-loaded mPEG-C-C-C16 micelles in the presence of dithiothreitol (DTT), but showed similar release rates in the absence of DTT. MTT assay demonstrated significantly enhanced cytotoxicity of DOX-loaded mPEG-S-S-C16 micelles against the human cervical cancer cells (HeLa) compared with DOX-loaded mPEG-C-C-C16 micelles, but there was no significant difference in the cytotoxicity between the two DOX-loaded micelles against the african green monkey SV40-transformed kidney fibroblast cells (COS-7). Confocal laser scanning microscopy observation and flow cytometry analyses indicated that DOX-loaded mPEG-S-S-C16 micelles were efficiently internalized into HeLa cells, released DOX into the cytoplasm, and entered the nuclei. By contrast, in the case of DOX-loaded mPEG-C-C-C16 micelles, little DOX was found in the nuclei. Endocytosis inhibition results proved that both mPEG-S-S-C16 and mPEG-C-C-C16 micelles entered the HeLa cells mainly through the clathrin-mediated endocytosis pathway, and caveolae-mediated endocytosis was involved to a small extent. These results indicated that the different behaviors of cell uptake between reduction-sensitive and -insensitive micelles may occur after the micelles were internalized into the cells, but not during endocytosis, and the potential of this reduction-sensitive polymer for the effective intracellular delivery of anticancer drugs.
采用含有二硫键的双亲性聚乙二醇衍生物单甲氧基聚乙二醇-S-S-十六烷基（mPEG-S-S-C16）制备了还原敏感胶束。然后将胶束用于将抗癌药物阿霉素（DOX）递送到肿瘤细胞内，并确定了胶束的细胞摄取机制。作为对照，还制备了具有类似结构但不含二硫键的单甲氧基聚乙二醇-C-C-十六烷基（mPEG-C-C-C16）。聚合物可以在水溶液中自组装成胶束，并负载高含量的 DOX。体外释放研究表明，在存在二硫苏糖醇（DTT）的情况下，载 DOX 的 mPEG-S-S-C16 胶束比载 DOX 的 mPEG-C-C-C16 胶束更快地释放 DOX，但在不存在 DTT 的情况下，释放速率相似。MTT 测定表明，与载 DOX 的 mPEG-C-C-C16 胶束相比，载 DOX 的 mPEG-S-S-C16 胶束对人宫颈癌（HeLa）细胞的细胞毒性显著增强，但两种载 DOX 胶束对非洲绿猴 SV40 转化的肾成纤维细胞（COS-7）的细胞毒性没有显著差异。共聚焦激光扫描显微镜观察和流式细胞术分析表明，载 DOX 的 mPEG-S-S-C16 胶束有效地被 HeLa 细胞内化，将 DOX 释放到细胞质中，并进入细胞核。相比之下，在载 DOX 的 mPEG-C-C-C16 胶束的情况下，很少有 DOX 存在于细胞核中。内吞作用抑制结果证明，mPEG-S-S-C16 和 mPEG-C-C-C16 胶束均主要通过网格蛋白介导的内吞作用途径进入 HeLa 细胞，并且小程度涉及小窝介导的内吞作用。这些结果表明，还原敏感和不敏感胶束之间的细胞摄取行为的不同可能发生在胶束被内化到细胞内之后，而不是在胞吞作用期间，并且这种还原敏感聚合物具有有效递送到细胞内的潜力抗癌药物。
