可控电穿孔原位消融及原位致敏DC综合抗肿瘤的实验研究

During the ablation treatment of tumor, occasionally, metastases may vanish spontaneously, the essence of its immunological reaction may be similar to the principle of cytotoxic response induced by tumor cell lysate vaccine. According to the above mentioned hypothesis, we propose an anti-tumor project combined controllable electroporation ablation in situ with fusion vaccine formed by in- situ sensitized dendritic cells. The technology of controllable electroporation is adopted, we realize reversible and irreversible regulation of cell membrane electroporation, just by adjusting electric pulse parameters. The following is the procedure of combined treatment: firstly, we ablate the tumor using irreversible electroporation, then DC cells in vitro are injected into the tumor and reversible electroporation will used to enhance the sensitization of DC. Through the combined treatment, the permeability of DC cell membrane will be boosted, so the fusion effect of DC cell with the former necrosis lysate can be enhanced, thus forming tumor lysate-sensitized DC vaccine as expected. According to the project design, we build the rat models with osteosarcoma and lung metastasis, and then we observe the changes of immunology, biology and the survival time before and after the combined treatment. Meanwhile, treatment process and technical parameters can be optimized. In this project, electroporation technology is a link combining the two methods to relieve the risk of in-situ tumor invasion effectively under the imaging guidance. The tumor lysate-sensitized DC vaccine, which breaks the mechanism of immune tolerance and escape during tumor treatment, is expected to provide a new strategy for clinic.

在临床实践中偶发肿瘤原发灶消融导致转移灶意外消退的现象可能和消融所致肿瘤溶胞产物所诱导的免疫细胞毒反应有关。本研究拟以可控电穿孔技术为纽带,通过调控电穿孔参数，希望依次完成以下步骤:首先以不可逆电穿孔电场条件原位灭活肿瘤细胞，继以可逆电穿孔电场条件增加原位注射体外诱导的树突状细胞的通透性，利用前一步不可逆电穿孔形成的大量溶胞产物进行强化致敏。本研究的重点在于原位灭活原发肿瘤的同时,促成树突状细胞与肿瘤坏死溶胞产物的结合,强化原位肿瘤溶胞抗原致敏树突状细胞,希望达到局部治疗和全身免疫治疗的双重目的。研究拟观察大鼠骨肉瘤及其肺转移模型在这种双效治疗前后的生物免疫学变化，建立起消融和免疫治疗相结合的可行方案。

在临床实践中偶发肿瘤原发灶消融导致转移灶意外消退的现象可能和消融所致肿瘤溶胞产物所诱导的免疫细胞毒反应有关。本研究拟以可控电穿孔技术为纽带, 通过调控电穿孔参数， 希望依次完成以下步骤: 首先以不可逆电穿孔电场条件原位灭活肿瘤细胞，继以可逆电穿孔电场条件增加原位注射体外诱导的树突状细胞的通透性，利用前一步不可逆电穿孔形成的大量溶胞产物进行强化致敏。不可逆电穿孔消融技术近年来已经被中美等国批准上市，不可逆电穿孔消融技术已经被广泛肝、脑，乳腺、肺、胰腺以及肾等各部位肿瘤。得益于这一利好消息。在人体中直接验证不可逆消融所致肿瘤溶胞产物激发的机体特异性或非特异性的免疫反应将为后续实验提供直接的临床指导。因此，我们测试了10例肿瘤患者术前术后的机体免疫反应。令人鼓舞的是，术后第3天，较术前并无明显的CD4+ T细胞，CD8+ T细胞，IL-2，IL-10，TNF-β明显增加，p值均小于0.05。然而，第10天却较术前无明显改善。这有利的佐证了我们的假设即不可逆电穿孔原位消融恶性肿瘤可诱发机体免疫。目前，本项目已完成部分实验，取得了部分有影响和特色的成果。项目按照计划展开工作，执行的结果集中反应在已发表的3篇SCI收录论文、举办的一次高质量学术交流会议和培养了的两名较为优秀的硕士研究生中。