一种新型基因传输载体-磷酸钙纳米颗粒用于肿瘤基因治疗的研究

Calcium phosphate nanoparticles as a novel nonviral vector in cancer gene therapy

目的研究新型非病毒载体磷酸钙纳米颗粒的生物学特性,评估其在肿瘤基因治疗中应用的前景。方法化学方法制备,氯化钙修饰纳米颗粒,用琼脂糖凝胶电泳分析磷酸钙纳米颗粒与DNA的结合效率及对DNA的保护作用,用绿色荧光蛋白基因作报告基因,将磷酸钙纳米颗粒为基因载体转染鼻咽癌(CNE-2)细胞和在动物体内转染肿瘤细胞;以及将磷酸钙纳米与自杀基因yCDglyTK结合,并在体外对鼻咽癌进行基因治疗。结果电镜观察证实磷酸钙纳米颗粒进入细胞内,磷酸钙纳米颗粒与DNA结合后,能对DNA起保护作用;磷酸钙纳米颗粒作为基因转染的载体,将绿色荧光蛋白基因导入CNE-2细胞,用荧光显微镜观察到高水平的绿色荧光蛋白表达;体内转染结果显示,纳米介导的基因在肿瘤组织中有很好的表达;以磷酸钙纳米为载体介导自杀基因yCDglyTK在体外治疗鼻咽癌的实验中显示,在加入5-FC后大量的CNE-2细胞出现死亡。结论磷酸钙纳米颗粒具有优良的生物学特性,是有应用前景的基因转染和基因治疗载体之一。

[Objective] To study the biological characteristics of calcium phosphate nanoparticles and evaluate the promising in cancer gene therapy. [Methods] Calcium chloride modified nanoparticles prepared by means of chemistry. Analyse the combination of calcium phosphate nanoparticles and DNA, the protection to DNA as well by gelose gelatin electrophoresis. When the green fluorescence protein gene was regarded as report gene, gene vector of calcium phosphate nanoparticles transfected CNE-2 cell in vitro and vivo. Combine the calcium phosphate nanoparticles and suicide gene yCDglyTK for Nasopharyngeal Carcinoma （NPC） therapy in vitro. [Results] Calcium phosphate nanoparticles went into cell and could protect DNA after its combination with DNA by electro-microscopy. Regard calcium phosphate nanoparticles as vector of gene transfection, transduct green fluorescence protein gene into CNE-2 cell, the green fluorescence protein high expression can be seen by fluorescence micresopy. The results of transfection in vivo showed that gene introduced by nanoparticles had a good expression in tumor tissue. The results of transducting suicide gene yCDglyTK by calcium phosphate nanoparticles for Nasopharyngeal Carcinoma （NPC） therapy in vitro indicated that large numbers of CNE-2 cells were dead after the addition of 5-FC. [Conclusion]Calcium phosphate nanoparticles has great biological characteristics and it is one of promising vectors of gene transfection and gene therapy.