放射诱导启动子介导双自杀基因治疗舌鳞癌的实验研究

Radiation-inducible promoters mediated CDglyTK gene in the treatment of Xenograft of human tongue squamous cell carcinoma in naked mice

目的探讨放射诱导启动子介导双融合自杀基因CDglyTK靶向治疗人舌鳞癌裸鼠移植瘤的疗效。方法构建人舌鳞癌裸鼠移植瘤模型,以脂质体为载体介导携放射诱导启动子调控双融合自杀基因的真核表达质粒pcDNA3.1(+)/E-CDglyTK瘤内转染,辅以3Gy放疗,腹腔注射5-氟胞嘧啶和丙氧鸟苷,描绘肿瘤生长曲线,RT-PCR检测CDglyTK的mRNA水平,免疫组化检测细胞核增殖抗原的表达,原位末端标记法检测肿瘤细胞凋亡。结果放射诱导启动子介导的CDglyTK对肿瘤生长有明显抑制作用,RT-PCR可检测到转染后肿瘤细胞CDglyTK的mRNA表达,诱导放疗增强了其表达水平,电泳条带灰度值从0.213±0.023上调至0.279±0.038(P<0.01);诱导放疗显著提高疗效,凋亡指数从21.52%上升为32.23%(P<0.01),增殖指数由28.36%下降至20.07%(P<0.01)。结论放射诱导启动子可作为基因治疗分子开关调节CDglyTK基因在人舌鳞癌裸鼠移植瘤中靶向表达,低剂量放射性照射可显著提高其疗效。

Objective To observe the therapeutic effect of CDglyTK gene mediated by radiation-inducible promoters in the treatment of Xenograft of human tongue squamous carcinoma in nude mice. Methods The model of transplanted Tca8113 cell line in nude mice was established. The plasmids pcDNA （＋）3.1/E-CDglyTK were transfected into tumors by lipofectamine. 24 h later, the tumors were exposed to 3 Gy irradiation, and then 5-fluorocytosine （5-FC） and ganeiclovir（GCV） were injected into nude mice at intervals. Animals were examed at regular intervals for tumor volume. CDglyTK mRNA was assayed by RT-PCR. Apoptosis and proliferating cell nuclear antigen were detected respectively by in situ end-labeling and immunohistioehemical method. Results Compared with control groups, the tumor was suppressed obviously by CDglyTK gene therapy combined with 3 Gy induction radiation. The expression of CDglyTK geue could be detected by RT-PCR in the transfeeted tumor, and up-regnlation of CDglyTK expression was found in tumor exposed to radiation （P 〈 0.01 ）. There was significant difference between the apoptosis index or proliferation index in tumor without irradiation and that in tumor with irradiation （P 〈 0.01 ）. Conclusions The radiation-inducible promoter can be served as a molecular switch to improve the expression of CDglyTK gene in transplanted human tongue squamous carcinoma in nude mice, and low dose induction radiation can significantly improve the therapeutic efficiency.