转铁蛋白受体介导的自杀基因转移系统的建立及其体外抗瘤效应

目的 构建转铁蛋白受体介导的自杀基因系统并观察其对肝癌细胞的体外杀伤效应。方法 SPDP法制备抗转铁蛋白受体与多聚赖氯酸(PLL)的复合物并用分子筛层析纯化。根据DNA阻断试验结果，pEBAF／tk重组质粒与Ab-PLL按1：6混合使二者结合形成PEBAF／tk—Ab PLL复合物。将此复合物转入人肝癌细胞株SMMC7721、HepG2和肺癌细胞株A549，并以脂质体转移为对照。加入不同浓度的更昔洛韦(GCV)以观察细胞的自杀效应。结果 加入GCV后HepG2／tk的增殖受抑制最明显。100mg／L和1mg／L时抑制率分别为60．5％和24．3％。SMMC7721／tk受抑制较低，而A549的增殖不受抑制。结论 本基因治疗体系具仃很好的靶向性和杀肿瘤效果。

The construction of transferrin receptor-mediated HSV-TK gene transfer system and its effect on human hepatocellular carcinoma cells in vitro

Objective To construct the localization system involving anti-TfR monoclonal antibody (McAb) and AFP promoters and assess its effect on human hepatoma cell lines. Methods The conjugate of anti-TfR McAb and polylysine (PLL) was made by SPDP and purified by molecular screen chromatography. DNA blocking test determined that the ratio of one pEBAF/tk to six Ab-PLL was the most suitable to couple them. The pEBAF/tk recombinant plasmid bearing HSV-TK gene was coupled to Ab-PLL by noncovalent bond. The pEBAF/tk was transferred into human hepatoma cell line HepG2, SMMC7721 and pulmonary cancer cell line A549 by receptor-mediated gene delivery (Ab-PLL-DNA) and liposome procedure. The growth inhibitory rates of HepG2, SMMC7721 and A549 cells were measured by MTT assay. Results The inhibitory rates of HepG2/tk in 100 mg/L and 1 mg/L of GCV were 60.5% and 24.3%, respectively. The inhibitory rate of GCV to SMMC7721 was 23.2% in 3 days. The pulmonary cancer cell A549, A549/tk(Ab) and A549 /tk (lipo) could not be inhibited by the addition of GCV. Conclusion The localization system employed in this paper has high specificity, effectiveness and safety for gene therapy. It would be a promising strategy for gene therapy.