Grupo de Química Coloidal y Supramolecular, Departamento de Química Física I, Facultad de Ciencias Químicas, Universidad Complutense de Madrid, 28040 Madrid, Spain.
Department of Molecular Medicine, "Sapienza" University of Rome, Viale Regina Elena 291, 00161 Rome, Italy.
Biochim Biophys Acta Gen Subj. 2017 Jul;1861(7):1737-1749. doi: 10.1016/j.bbagen.2017.03.010. Epub 2017 Mar 16.
The self-assembling processes underlining the capabilities of facially differentiated ("Janus") polycationic amphiphilic cyclodextrins (paCDs) as non-viral gene nanocarriers have been investigated by a pluridisciplinary approach. Three representative Janus paCDs bearing a common tetradecahexanoyl multitail domain at the secondary face and differing in the topology of the cluster of amino groups at the primary side were selected for this study. All of them compact pEGFP-C3 plasmid DNA and promote transfection in HeLa and MCF-7 cells, both in absence and in presence of human serum. The electrochemical and structural characteristics of the paCD-pDNA complexes (CDplexes) have been studied by using zeta potential, DLS, SAXS, and cryo-TEM. paCDs and pDNA, when assembled in CDplexes, render effective charges that are lower than the nominal ones. The CDplexes show a self-assembling pattern corresponding to multilamellar lyotropic liquid crystal phases, characterized by a lamellar stacking of bilayers of the CD-based vectors with anionic pDNA sandwiched among them. When exposed to human serum, either in the absence or in the presence of pDNA, the surface of the cationic CD-based vector becomes coated by a protein corona (PC) whose composition has been analyzed by nanoLC-MS/MS. Some of the CDplexes herein studied showed moderate-to-high transfection levels in HeLa and MCF-7 cancer cells combined with moderate-to-high cell viabilities, as determined by FACS and MTT reduction assays. The ensemble of data provides a detail picture of the paCD-pDNA-PC association processes and a rational base to exploit the protein corona for targeted gene delivery on future in vivo applications.
通过多学科方法研究了具有面差异化（“Janus”）聚阳离子两亲性环糊精（paCD）作为非病毒基因纳米载体的自组装过程。选择了三种具有共同十四烷酰多尾域的代表性 Janus paCD 作为研究对象，它们在主面上的氨基簇拓扑结构上有所不同。所有这些都可以压缩 pEGFP-C3 质粒 DNA，并在 HeLa 和 MCF-7 细胞中促进转染，无论是在没有还是存在人血清的情况下。使用 zeta 电位、DLS、SAXS 和 cryo-TEM 研究了 paCD-pDNA 复合物（CDplex）的电化学和结构特性。paCD 和 pDNA 在组装成 CDplex 时，呈现出的有效电荷低于标称电荷。CDplex 表现出自组装模式，对应于溶致液晶相的多层结构，其特征是基于 CD 的载体的双层以层状堆叠，阴离子 pDNA 夹在它们之间。当暴露于人血清中时，无论是在没有 pDNA 的情况下还是存在 pDNA 的情况下，阳离子基于 CD 的载体的表面都会被蛋白质冠（PC）覆盖，通过纳米 LC-MS/MS 分析了其组成。本文研究的一些 CDplex 表现出中等至高的转染水平在 HeLa 和 MCF-7 癌细胞中，与中等至高的细胞活力相结合，通过 FACS 和 MTT 还原测定确定。这些数据提供了 paCD-pDNA-PC 缔合过程的详细图片，并为利用蛋白质冠进行未来体内应用的靶向基因传递提供了合理的基础。
