Tie2介导的基因导入系统转导p53基因对肺癌的抑制作用

Inhibition of lung cancer by transfer of p53 gene using Tie2-mediated gene delivery system

目的:探讨Tie2介导的基因导入系统体内靶向转导p53基因治疗肺癌的效果。方法:应用双功能交联剂SMCC将Tie2配体寡肽GA3与PEI连接构建靶向Tie2的基因载体。体外试验将GA3-PEI/luciferase导入Tie2表达阳性的SPC-A1肺癌细胞与Tie2表达阴性的SMMC7721肝癌细胞中,24 h后检测luciferase活性。体内导入试验将GA3-PEI/β-gal复合物注射SPC-A1细胞裸鼠种植瘤周围皮下,同时设立PEI/β-gal组为对照;24 h后牺牲裸鼠,检测心、肝、脾、肺、肾和瘤体的β-gal表达。另外将另一批四周龄SPC-A1肺癌种植瘤裸鼠随机分为5组:(1)NS;(2)GA3;(3)p53;(4)PEI/p53;(5)GA3-PEI/p53;每组6只;皮下注射GA3-PEI/p53基因复合物,隔2天1次,并测量肿瘤长短径,计算瘤体积及抑瘤率。结果:GA3-PEI/luciferase组的luciferase活性在SPC-A1细胞中明显高于SMMC7721细胞(P<0.05)。在种植瘤中可见较多蓝染细胞,心、肝、脾、肾组织中几乎不见篮染,肺支气管黏膜除外。与对照组比较,GA3-PEI/p53治疗组对肿瘤生长有明显抑制作用(P<0.05),其抑瘤率为61.29%。结论:GA3基因导入系统靶向转导p53基因,显著抑制肿瘤生长。

Objective： To explore the therapeutic efficacy of transduction ofp53 gene with Tie2-mediated gene delivery system in the treatment of lung cancer and provide experimental basis for future clinical application. Methods： GA3, a small peptide against Tie 2, was conjugated with PEI by bifunctional crosslinking agent SMCC to construct Tie2-targeted gene vector. Then pCMV-luciferase eDNA was combined with GA3-PEI to form an electrostatic complex. GA3-PEI/lueiferase eDNA complex was transduced into Tie2- positive SPC-A1 lung cancer cells and Tie2-negative SMMC7721 hepatic cancer cells in vitro. The activity of luciferase was measured 24 h later. The GA3-PEI/β-gal complex was injected into peri-tumorous tissues of the transplanted SPC-Al tumors in nude mice in vivo. Meanwhile, PEI/β-gal gene was injected as control. The nude mice were decapitated 24 h later. The heart, liver, spleen, lung, kidney and tumor tissues were excised and stained with X-gal. Human lung cancer SPC-Al cells were inoculated subcutaneously into the 4- week-old female athymie mice （BALB/e）. The mice were randomly divided into 5 groups （ NS, GA3, p53, PEI/p53, and GA3-PEI/ p53） with 6 mice in each group. GA3-PEI/p53 1 μg was subcutaneously injected into peri-tumorous tissues once every two days. The tumor volume was calculated according to formula （V = πab2/6）. The tumor-inhibiting ratio was calculated regarding NS group as control. Results： The activity of luciferase of GA3-PEI/luciferase was significantly higher in SPC-Al cells than that in SMMC7721 cells （P 〈 0.05 ）. A lot of blue-stained cells were observed in transplanted tumor tissues but not in heart, liver, spleen, and kidney tissues except lung bronchial mueosa. Compared with NS group, GA3-PEI/p53 significantly inhibited the growth of tumor by 61.29%. Conclusion： Transfer of p53 gene byGA3 gene delivery system remarkably inhibits tumor growth.