Electrochemo-gene therapy of Cancer using Epstein Barr virus-plasmid vector ; Intratumoral delivery of Interleukin-12 gene and bleomycin induce therapeutic immunity and suppressed subcutaneous and metastatic melanomas in mice

前言和目的：细胞毒化疗对肿瘤的抗肿瘤活性很低。免疫疗法对黑色素瘤相对有效。本研究通过电穿孔瘤内转导IL-12基因和瘤内注射博莱霉素瘤内注射对免疫敏感型黑色素瘤细胞株B16进行联合治疗。方法：采用电穿孔的方法瘤内转移博莱霉素和/或表达IL-12的Epstein-Barr病毒-质粒载体，治疗小鼠免疫敏感型黑色素瘤。同时接受博莱霉素和IL12基因治疗的八只小鼠中，有三只(37.5%)显示预先建立的肿瘤完全缓解，并拒绝随后与肿瘤细胞的再次攻击。我们还检查了皮下肿瘤的电化学基因治疗是否通过静脉接种黑色素瘤细胞来抑制肺转移。虽然单独给予IL12基因或博莱霉素+IL12基因治疗组的转移灶明显减少，但只有联合治疗才能显著延长荷瘤动物的平均生存期。在接受化疗基因治疗的小鼠中，自然杀伤细胞(NK)和细胞毒性T淋巴细胞的杀伤活性显著增强，而IL12基因单独治疗则部分提高了NK细胞的杀伤活性。结论：本研究表明，电穿孔介导的IL12基因和博莱霉素协同诱导了天然和适应性的抗黑色素瘤免疫反应，从而显著抑制了治疗后的肿瘤和旁观者的转移。

INTRODUCTION AND OBJECTIVES : Cytotoxic chemotherapy has shown little antitumor activity against carcinomas. Immunotherapy is relatively effective against melanoma. In the present study, we investigated the effectiveness of the combination therapy with Intratumoral delivery of Interleukin (IL) 12 gene using electroporation and intratumoral injection of bleomycin against immuno sensitive melanoma cell line, B 16.METODS : To treat established immuno sensitive melanoma tumors in mice, intratumoral transfer of bleomycin and/or an IL12 expression Epstein-Barr Virus-plasmid vector was performed by means of electroporation.RESULTS : Although either bleomycin alone or the IL12 gene alone significantly suppressed the subcutaneous tumors, the combination therapy drastically improved the therapeutic outcome. Three of eight mice (37.5%) that received both bleomycin and the IL12 gene showed complete remission of the preestablished tumors and rejected subsequent rechallenge with the tumor cells. We also examined whether electrochemo-gene therapy for subcutaneous tumor mass induced suppression of pulmonary metastasis that had been established by intravenous inoculation of the melanoma cells. Although metastatic foci were significantly reduced in number in groups that were given IL12 gene alone or bleomycin plus IL12 gene, it was only the combination therapy that significantly prolonged the mean survival period of the tumor-bearing animals. Natural killer(NK) and cytotoxic T lymphocyte cytolytic activities were markedly enhanced in the mice that received the chemo-gene therapy, while IL12 gene therapy alone partially elevated the NK cytotoxicity.CONCLUSIONS : The present study suggests that the electroporation-mediated delivery of the IL12 gene and bleomycin synergistically elicits innate and adaptive anti-melanoma immune responses, resulting in marked suppression of the treated tumors as well as bystander metastatic lesions.