肿瘤细胞与线粒体双靶向Bcl-2蛋白抑制剂载体系统

The tumor chemtherapy strategy is the key problem for the design of anti-tumor drug delivery system. In this project, with the selection and characteristics of Bcl-2 protein inhibition of (-) gossypol, tumor cells targeting of hyaluronic acid, as well as the mitochondrial targeting of triphenylphosphines (TPP)cation, the dual-targeting drug delivery system to tumor cells and their mitochondrials is prepared as follows: (-) gossypol amino m-PEG nanoparticles are prepared by the encapsulation of (-) gossypol with amino m-PEG using self -assemble method, then triphenylphosphine bromonexanoic acid cations are conjugated to the amino group of the nanoparticles by N-ethyl-N'-3-(dimethylamino)propylcabodiimid（EDC ）and N-hydroxysuccinimide（NHS）, the dual-targeting drug delivery system will be obtained by encapsulate the shell of hyaluronic acid on the (-) gossypol m-PEG nanoparticles. Their surface and chemical structures can be determined by IR , NMR spectra and so on. Their targeting characteristics for tumor cells and their mitochondrials can be measured by the fluorescent labeling，fluorescence lifetime， mitochondrial membrane potential，FQ-PCR and circular dichroism(CD), respectively. Their antitumor effects will be detected by cell tests in vitro and aminal tests in vivo. This kind of Bcl-2 protein inhabitor delivery system with the chracteristics of tumor cells and their mitochondrials targeting can be both targeted combined to the tumor cells with hyaluronic acid and combined to their mitochondrial membrane with triphenylphosphine cation after they are came into the tumor cells, (-) gossypol can inhibit Bcl-2 proteins more effectively than common gossypol delivery system, thereby obtain optimal inhibition of Bcl-2 proteins and antitumor effects.

肿瘤化疗策略是抗肿瘤药物载体设计的关键。本项目以肿瘤细胞及其线粒体为目标靶点，综合应用左旋棉酚等对Bcl-2蛋白的抑制，透明质酸、三苯基膦阳离子化合物对肿瘤细胞及其线粒体的目标选择和靶向性。将左旋棉酚等用氨基m-PEG自组装包裹形成纳米粒子，通过交联剂EDC和N-羟基琥珀酰亚胺与三苯基膦溴代己酸阳离子的羧基结合形成线粒体靶向纳米粒子。再将PEG修饰的透明质酸包裹在外层制成具有肿瘤细胞及其线粒体双靶向载体系统。红外光谱、核磁共振等确定其化学和表面构成；荧光标记、荧光寿命、线粒体膜电位检测、FQ-PCR、圆二色谱等研究其肿瘤和线粒体靶向特性和作用机制；体内外实验研究其抗肿瘤效果。这种具有肿瘤细胞与线粒体双靶向性的Bcl-2蛋白抑制剂载体系统，可在透明质酸介导下靶向结合到肿瘤细胞上，进入细胞后在三苯基膦阳离子介导下靶向结合到线粒体膜上与Bcl-2蛋白作用，达到最佳Bcl-2蛋白抑制和抗肿瘤效果。

本项目以肿瘤细胞及其线粒体为目标靶点，综合应用棉酚及其衍生物对Bcl-2蛋白的抑制，透明质酸、三苯基膦阳离子化合物对肿瘤细胞及其线粒体的目标选择和靶向性。先将棉酚衍生物Apogossypolone (ApoG2) 用阳离子淀粉包裹成为纳米微粒，再通过交联剂EDC和N-羟基琥珀酰亚胺与三苯基膦溴代己酸阳离子的羧基结合形成线粒体靶向纳米粒子。再将PEG修饰的透明质酸包裹阿霉素形成纳米小球，最后利用静电作用将它们组装成为“桑葚’状结构的复合纳米载体，该载体具有肿瘤细胞及其线粒体双靶向载体系统。通过红外光谱、核磁共振等确定其化学和表面构成；荧光标记、荧光寿命、线粒体膜电位检测等研究其肿瘤和线粒体靶向特性和作用机制；体内外实验研究其抗肿瘤效果。这种具有肿瘤细胞与线粒体双靶向性的Bcl-2蛋白抑制剂载体系统，可在透明质酸介导下靶向结合到肿瘤细胞上，进入细胞后在三苯基膦阳离子介导下靶向结合到线粒体膜上与Bcl-2蛋白作用，达到最佳Bcl-2蛋白抑制和抗肿瘤效果。