Departments of ‡Biomedical Sciences, §Chemistry, and ∥Physics, Missouri State University , Springfield, Missouri 65897, United States.
Biomacromolecules. 2013 Nov 11;14(11):4108-15. doi: 10.1021/bm4012425. Epub 2013 Oct 28.
Dendrimer chemistries have virtually exploded in recent years with increasing interest in this class of polymers as gene delivery vehicles. An effective nucleic acid delivery vehicle must efficiently bind its cargo and form physically stable complexes. Most importantly, the nucleic acid must be protected in biological fluids and tissues, as RNA is extremely susceptible to nuclease degradation. Here, we characterized the association of nucleic acids with generation 4 PEGylated poly(amidoamine) dendrimer (mPEG-PAMAM-G4). We investigated the formation, size, and stability over time of the nanoplexes at various N/P ratios by gel shift and dynamic light scatter spectroscopy (DLS). Further characterization of the mPEG-PAMAM-G4/nucleic acid association was provided by atomic force microscopy (AFM) and by circular dichroism (CD). Importantly, mPEG-PAMAM-G4 complexation protected RNA from treatment with RNase A, degradation in serum, and various tissue homogenates. mPEG-PAMAM-G4 complexation also significantly enhanced the functional delivery of RNA in a novel engineered human melanoma cell line with splice-switching oligonucleotides (SSOs) targeting a recombinant luciferase transcript. mPEG-PAMAM-G4 triconjugates formed between gold nanoparticle (GNP) and particularly manganese oxide (MnO) nanorods, poly IC, an anticancer RNA, showed enhanced cancer-killing activity by an MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) cell viability assay.
近年来，树枝状聚合物化学领域取得了飞速发展，人们对作为基因传递载体的这类聚合物的兴趣日益浓厚。有效的核酸传递载体必须能够有效地结合其货物并形成物理稳定的复合物。最重要的是，核酸必须在生物流体和组织中得到保护，因为 RNA 极易被核酸酶降解。在这里，我们对核酸与第四代聚乙二醇化聚（酰胺-胺）树枝状大分子（mPEG-PAMAM-G4）的结合进行了表征。我们通过凝胶电泳迁移率和动态光散射光谱（DLS）研究了不同 N/P 比下纳米复合物的形成、大小和随时间的稳定性。原子力显微镜（AFM）和圆二色性（CD）进一步提供了 mPEG-PAMAM-G4 与核酸结合的特性。重要的是，mPEG-PAMAM-G4 复合物能够保护 RNA 免受 RNase A 处理、血清降解以及各种组织匀浆的影响。mPEG-PAMAM-G4 复合物还显著增强了新型工程化人黑素瘤细胞系中 RNA 的功能传递，该细胞系中含有针对重组荧光素酶转录本的剪接转换寡核苷酸（SSO）。金纳米颗粒（GNP）和特别的氧化锰（MnO）纳米棒、多聚肌苷酸（poly IC）之间形成的 mPEG-PAMAM-G4 三连接体，与一种抗癌 RNA 结合后，通过 MTT（3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四唑溴盐）细胞活力测定法显示出增强的抗癌活性。
