Cancer gene therapy by the in vivo transfer of cytokine-genes in to the artery that leads to tumors with fusogenic liposomes.

我们报道了当肿瘤坏死因子α在体内通过基因转移在导致肿瘤的动脉中产生时，它诱导了强烈的抗肿瘤免疫反应。我们使用了在右足垫携带肉瘤-18O肿瘤的小鼠模型，并将包裹人肿瘤坏死因子-α基因的融合脂质体注射到右股动脉。在这种情况下，只在通向肿瘤的动脉和肿瘤中检测到人肿瘤坏死因子-α。当将肿瘤坏死因子-α基因转移到右侧股动脉时，肿瘤生长显著消退，11只小鼠中有4只完全治愈。当将肿瘤坏死因子-α基因导入左股动脉或肿瘤或给予荧光素酶基因时，没有观察到肿瘤消退。注射抗肿瘤坏死因子-α、抗CD_4、抗CD_8单抗和CD8+T细胞可抑制肿瘤消退。这些结果表明，肿瘤坏死因子-α表达L…。在更局部的情况下，导致肿瘤的动脉通过加强抗肿瘤免疫反应有效地抑制肿瘤的生长。此外，开发基因转移到中枢神经系统(CNS)的方法对于基因治疗的基础研究和临床研究都是重要的。阳离子脂质体(CL)因其安全性和易用性而成为一种极具吸引力的载体。然而，到目前为止，只有很低的成功率报告。我们成功地通过CL和基于T7RNA聚合酶和T7RNA聚合酶的细胞质基因表达系统以及带有T7启动子序列的荧光素酶基因在体内获得了高效率的转基因。该系统的效率比使用CL和荧光素酶基因与Rous肉瘤病毒启动子pRSVL的标准系统高2个数量级。此外，使用LLCMK2细胞进行的体外实验表明，基因在细胞质中迅速表达(在6h内)。相反，pRSVL需要24-48小时才能诱导荧光素酶的表达。较少

We report that tumor necrosis factor (TNF) alpha induced a strong antitumor immune reaction when it was produced in arteries leading to tumors by gene transfer in vivo. We used a mouse model carrying a sarcoma-18O tumor in the right footpad and injected the fusogenic liposomes encapsulating the human TNF-alpha gene into the right femoral artery. Under this condition, human TNF-alpha was detected only in the artery leading to the tumor and in the tumor. There was a significant regression in tumor growth when the TNF-alpha gene was delivered into the right femoral artery, with 4 of 11 mice completely cured. No regression was observed when the TNF-alpha gene was delivered into the left femoral artery or into the tumor or when the luciferase gene was administered. Tumor regression was inhibited by the injection of anti-TNF-alpha, anti-CD4, or anti-CD 8 monoclonal antibody, and CD8+ T cells accumulated in the tumors of TNF-alpha-treated mice. These results suggest that TNF-alpha expressed l … More ocally in the arteries leading to tumors efficiently suppresses tumor growth through reinforcement of an antitumor immune reaction. Additionally, Development of methodologies for gene transfer into the central nervous system (CNS) is important for fundamental research as well as clinical studies for gene therapy. Cationic liposomes (CL) are attractive vectors because of their safety and ease of use. However, to date only low rates of success have been reported. We succeeded in obtaining high transfection efficiencies into the newborn mouse brain in vivo by CL and a cytoplasmic gene expression system based on T7 RNA polymerase and T7 RNA polymerase- and the luciferase-gene with the T7 promoter sequence. This system showed an efficiency rate 2 orders of magnitude higher than the standard system, which used CL and luciferase genes with a Rous sarcoma virus promoter, pRSVL.In addition, in vitro experiments using LLCMK2 cells showed that cytoplasmic gene expression occurred rapidly (within 6 h) after transfection. In contrast, pRSVL required 24-48 h for induction of luciferase expression. Less