Nomura T, Yasuda K, Yamada T, Okamoto S, Mahato R I, Watanabe Y, Takakura Y, Hashida M
Department of Drug Delivery Research, Graduate School of Pharmaceutical Sciences, Kyoto University, Japan.
Gene Ther. 1999 Jan;6(1):121-9. doi: 10.1038/sj.gt.3300792.
Gene expression was assessed in three types of mouse solid tumors after direct injection of naked plasmid DNA encoding the luciferase gene (pCMV-Luc) and its DC-chol liposome complexes. Intratumoral injection of 5 or 100 micrograms naked pCMV-Luc into subcutaneously inoculated mouse colon tumor (CT-26), fibrosarcoma (MCA-15) and bladder carcinoma (MBT-2) resulted in significant gene expression. A DC-chol liposome formulation (5 micrograms pCMV-Luc complexed with 25 micrograms DC-chol liposome) showed lower level of gene expression in the tumor models. Based on the results using the reporter gene, we examined the antitumor effect after direct interferon-gamma (IFN-gamma) gene transfer into CT-26 tumors. A significant IFN-gamma production and growth inhibition were obtained following direct intratumoral injection of IFN-gamma gene with naked plasmid DNA (pCMV-Mu gamma). Interestingly, pCMV-Mu gamma/DC-chol liposome complexes exhibited more pronounced growth inhibitory effect despite lower IFN-gamma production. Induction of CT-26 specific antitumor immunity by IFN-gamma gene transfer was confirmed by rejection of a CT-26 tumor challenge in the mice showing complete regression of CT-26 tumors after both treatments. Further analysis demonstrated that a significant cDNA-independent induction of IFN-beta and TNF-alpha occurred following injection with the liposome complexes, suggesting a nonspecific suppressive effect on CT-26 tumor growth by these cytokines. Thus, the present study has demonstrated that tumor tissue might be a promising target for direct IFN-gamma gene transfer with plasmid-based nonviral vectors. It is also suggested that immunomodulatory effects by various cytokines could be involved in antitumor effects after direct intratumoral injection of plasmid DNA formulations.
在直接注射编码荧光素酶基因的裸质粒DNA（pCMV-Luc）及其DC-胆固醇脂质体复合物后，对三种类型的小鼠实体瘤中的基因表达进行了评估。向皮下接种的小鼠结肠癌（CT-26）、纤维肉瘤（MCA-15）和膀胱癌（MBT-2）瘤内注射5或100微克裸pCMV-Luc，可导致显著的基因表达。一种DC-胆固醇脂质体制剂（5微克pCMV-Luc与25微克DC-胆固醇脂质体复合）在肿瘤模型中显示出较低水平的基因表达。基于使用报告基因的结果，我们检测了将干扰素-γ（IFN-γ）基因直接导入CT-26肿瘤后的抗肿瘤效果。在瘤内直接注射含裸质粒DNA（pCMV-Muγ）的IFN-γ基因后，获得了显著的IFN-γ产生和生长抑制。有趣的是，尽管IFN-γ产生较低，但pCMV-Muγ/DC-胆固醇脂质体复合物表现出更明显的生长抑制作用。在两种治疗后CT-26肿瘤完全消退的小鼠中，通过对CT-26肿瘤攻击的排斥反应证实了IFN-γ基因转移诱导的CT-26特异性抗肿瘤免疫。进一步分析表明，注射脂质体复合物后发生了显著的不依赖于cDNA的IFN-β和TNF-α诱导，提示这些细胞因子对CT-26肿瘤生长有非特异性抑制作用。因此，本研究表明肿瘤组织可能是基于质粒的非病毒载体直接进行IFN-γ基因转移的一个有前景的靶点。还提示在瘤内直接注射质粒DNA制剂后，各种细胞因子的免疫调节作用可能参与抗肿瘤作用。
