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

引言和说明：细胞毒性化疗对癌症的抗肿瘤活性很小。免疫疗法对黑色素瘤相对有效。在本研究中，我们研究了使用电穿孔和瘤内注射博莱霉素的瘤内递送白细胞介素（IL）12基因对免疫敏感的黑色素瘤细胞系B 16的联合治疗的有效性。方法：为了治疗小鼠中已建立的免疫敏感性黑色素瘤，通过电穿孔进行博来霉素和/或IL 12表达EB病毒质粒载体的肿瘤内转移。尽管单独的博来霉素或单独的IL 12基因显著抑制了皮下肿瘤，但是联合治疗显著改善了治疗结果。接受博来霉素和IL 12基因的8只小鼠中有3只（37.5%）表现出预先建立的肿瘤的完全缓解，并拒绝随后用肿瘤细胞再激发。我们还研究了皮下肿瘤块的电化学基因治疗是否诱导了通过静脉接种黑色素瘤细胞建立的肺转移的抑制。尽管单独给予IL 12基因或博莱霉素加IL 12基因的组中转移灶的数量显著减少，但只有联合治疗才显著延长了荷瘤动物的平均生存期。在接受化学基因治疗的小鼠中，自然杀伤（NK）细胞和细胞毒性T淋巴细胞的细胞溶解活性显著增强，而单独的IL 12基因治疗部分地提高了NK细胞的细胞毒性。目前的研究表明，电穿孔介导的IL 12基因和博来霉素的递送协同增强了先天性和适应性抗黑色素瘤免疫应答，导致治疗的肿瘤以及旁观者转移性病变的显着抑制。

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.