近红外聚集诱导发光和活性氧参与的高效基因载体的构建

It is an important method to reveal the mechanism of action by investigating the cellular distribution of nucleic acid/polymer. However, fluorophore-labeled DNA will probably induce aggregation-caused quenching, reduce the transfection efficacy and the cell viability. In this project, a NIR fluorophore with distinct aggregation-induced-emissive (AIE) effects was introduced into reduction-responsive cationic polymers to probe intracellular trafficking of polymer/DNA complex and elucidate the transfection routes after being complexed plasmid DNA through electrostatic interactions. Using NIR fluorophores as fluorescence labeling materials could minimize the background interference from tissue auto-fluorescence. While their AIE property is postulated to circumvent photo bleaching and aggregation-induced quenching. A high quantum yield will help the AIE molecular probe the cellular distribution efficiently with minimized cytotoxicity. AIE fluorophore is assumed to not only promote the colloidal stability of formulated complexes due to its hydrophobicity, but also produce reactive oxygen species (ROS), consequently, synergistically resulting in improved the transfection efficiency. The tryptophan was then incorporated into the above polymers, which is supposed to improve their solubility of the polymers and binding ability to DNA and membranes. Finally, a pH-responsive functional group with spontaneous negative-to-positive charge conversion property was decorated on the polymer for facilitated internalization into the target cancer cells while resisting serum adsorption. The structure of AIE molecular and the composition of the polymer will be optimized to obtain traceable gene vector with good transfection efficacy and biocompatibility.

研究核酸/高分子在细胞内的分布是揭示其作用机制的一种重要方法。然而，通常荧光标记DNA会发生聚集诱导猝灭，影响转染效率，引起细胞毒性。本课题拟合成在近红外(NIR)具有聚集诱导发光(AIE)特性的阳离子高分子，与质粒DNA通过正、负电荷作用发生复合，并研究其示踪性能，阐明基因复合体的转染途径。NIR发光有望降低生物组织自发荧光对荧光信号的干扰；AIE有望避免荧光的聚集猝灭；而高量子产率的AIE分子有望在其安全浓度有效示踪。AIE分子呈现疏水性，有望提高基因复合体的稳定性，AIE荧光团还可以产生活性氧(ROS)，两者均有望提高转染效率。为提高AIE高分子与DNA和细胞膜的结合能力，提高其水溶性，使用色氨酸对高分子进行修饰，进一步提高转染效率。最后，引入具有pH响应性的可电荷翻转基团，实现靶向癌细胞和抗血清吸附的功能。优化AIE分子的结构和高分子组成，获得具有示踪作用的高效、低毒基因载体。

研究核酸/高分子在细胞内的分布是揭示其作用机制的一种重要方法。然而，通常荧光标记DNA会发生聚集诱导猝灭，影响转染效率，引起细胞毒性。以近红外发光分子示踪降低了生物组织自发荧光对荧光信号的干扰；AIE避免了荧光的聚集猝灭；而高量子产率的AIE分子实现了其安全浓度有效示踪。AIE分子呈现疏水性，能够提高基因复合体的稳定性；而AIE荧光团还可以产生活性氧(ROS)，两者均有效提高了转染效率。另外，在含有AIE的基因载体中引入ROS响应的二硫键/二硒键、肿瘤微酸性响应的聚2-（5,5-二甲基-1,3-二恶烷-2-基氧基）丙烯酸乙酯（PDM）、超分子单元、天然蛋白质组分，有效促进了内涵体逃逸，进一步提高了其生物相容性和转染效率。与基因转染“黄金标准”PEI25k相比较，细胞存活率提高了1.5倍，转染效率提高了6倍。上述具有示踪功能的高效、低毒的基因递送系统的制备和性能研究，为进一步开发利用奠定了实验和理论基础。