Trentin Diana, Hubbell Jeffrey, Hall Heike
Institute for Biomedical Engineering and Department of Materials, ETH and University of Zurich, Moussonstrasse 18 CH-8044 Zurich, Switzerland.
J Control Release. 2005 Jan 20;102(1):263-75. doi: 10.1016/j.jconrel.2004.09.029.
Non-viral DNA delivery systems show important advantages vs. viral systems that are usually associated with an immunological response and safety risks. In this study, disulfide cross-linked peptide-DNA condensates were investigated for local gene delivery. Two different 21 amino acid peptides were designed to have a DNA binding sequence in combination with a transglutaminase substrate site or a nuclear localization site. The peptides were used in different ratios to each other to form stable cross-linked DNA-peptide condensates with a mean diameter of 164 nm and a size distribution from 43 to 204 nm. Such aggregates showed similar stability compared to condensates formed between DNA and high molecular weight poly-L-lysine (PLL). Peptide-DNA condensates were covalently immobilized into fibrin matrices by the activity of factor XIII and were used for gene delivery in vitro. After internalization, reduction of the cross-linked peptide-DNA condensates yielded increased transfection efficiencies into different cell types cultured in 2D sandwich assays, and comparable values for HUVECs cultured in a 3D fibrin matrix, as compared to PLL-DNA condensates. Cell viability 24 h after transfection remained above 95%. The target was to develop a transfection system based on small peptides that can be covalently cross-linked into fibrin-matrices where DNA-release takes place upon cellular degradation of the matrix. This approach provides an interesting tool in non-viral gene delivery.
与通常与免疫反应和安全风险相关的病毒系统相比，非病毒DNA递送系统具有重要优势。在本研究中，研究了二硫键交联的肽-DNA缩合物用于局部基因递送。设计了两种不同的21个氨基酸的肽，使其具有与转谷氨酰胺酶底物位点或核定位位点结合的DNA结合序列。将这些肽以不同比例相互使用，以形成平均直径为164nm、尺寸分布为43至204nm的稳定交联的DNA-肽缩合物。与DNA和高分子量聚-L-赖氨酸（PLL）之间形成的缩合物相比，这种聚集体显示出相似的稳定性。肽-DNA缩合物通过因子XIII的活性共价固定在纤维蛋白基质中，并用于体外基因递送。内化后，与PLL-DNA缩合物相比，交联的肽-DNA缩合物的还原在二维夹心试验中培养的不同细胞类型中产生了更高的转染效率，对于在三维纤维蛋白基质中培养的人脐静脉内皮细胞（HUVEC）也有相当的值。转染后24小时细胞活力保持在95%以上。目标是开发一种基于小肽的转染系统，该系统可以共价交联到纤维蛋白基质中，在基质细胞降解时发生DNA释放。这种方法为非病毒基因递送提供了一个有趣的工具。
