用宫颈癌细胞Hela表面高表达G250抗原的单克隆抗体G250修饰非病毒基因载体,获得肿瘤靶向基因载体.通过注射G250杂交瘤细胞于小鼠腹腔,制备富含G250mAb的腹水,用正辛酸-硫酸铵沉淀法和Protein A Agarose分离纯化,获得高纯度的G250mAb.通过二硫键将PEI与G250mAb偶联,得到修饰的基因载体G250mAb-PEI,研究其转基因靶向性.结果表明,G250mAb-PEI对Hela细胞的基因转染具有显著的靶向性,对Hela细胞的转基因效率是肝癌细胞HepG2(G250阴性)的2倍;而对正常血管平滑肌细胞(SMC)的基因转染效率比Hela低近20倍,G250mAb修饰与否对SMC没有靶向性;对3T3细胞的毒性显著低于未修饰的PEI,表明G250mAb-PEI是一种高效、低毒和具有靶向性的基因载体.
A tumor-targeting gene vector G250mAb-PEI-PEG has been prepared by modification of polyethylenimine (PEI) with polyethyleneglycol (PEG) and G250, a monoclonal antibody against the G250 antigen on tumor cell surface. The transfection efficiency was as high as 70% in G250 positive HeLa cells, whereas the transfection efficiency was relatively low (30%) in normal NIH3T3 cells. A plasmid encoding the short hairpin RNA (shRNA) specific for nucleostemin gene (NS) was efficiently transfected into the HeLa cells with this nonviral gene vector. RNA interference down-regulated the expression of NS gene in HeLa cells, inhibited cells proliferation and induced apoptosis. However, the growth and activity of the NIH3T3 cells were not affected under the same treatment. These results indicate that the reported nonviral gene vector, G250mAb-PEI-PEG, can target and efficiently deliver genes into HeLa cells, and has the potential for the cervical cancer treatment.
