雷帕霉素通过Foxo3调节DCs耐受性抑制移植后肿瘤生长的机制研究

De novo malignant tumour will be the first dead reason in long term survival in the transplantation patients. It is very narrow interspace of the immuno-equilibrium in the transplanted recipients and very fragility for the internal environment. It is the bottleneck of the research and clinical due to the intense contradictory between the transplantation tolerance and the anti-tumor reaction. In our prophase works we have found that the quantity of DCs were decreased and the MDSCs were accumulation in patients' peripheral blood and the section of tumor tissues. The quantity of DCs have been elevation highly after tumorectomy. It suggested that the host immune system and DCs were inhibited by the cancer. Although it have been evidenced the huge accumulation of MDSCs infiltrating in transplanted kidney and tumor in there DCs were induced into tolerangenic DCs and the detail mechanism is still not clearly. Recent it has been reported that DCs expressed the nuclear transfactor in the more kind of tumors and triggered tumor generation. DCs expressed Forkhead Box class O transcription factor(Foxo3) highly in tumors maybe is a key for regulating tolerangenic DCs, and it controlled by phosphatidylinositol-3 kinase/Akt/mTOR pathway. Blocked Foxo3 transduction could inversed inhibited condition of DCs in tumor. We will set up a mouse modal for load transitional cell carcinoma of Bladder after aorta transplantation. In this modal it will be clarificated that MDSCs induced tolerangenic DCs with expressing Foxo3, and determined that Foxo3 is a master switch for regulating tolerance or immunity in dendritic cells. The first we will prove that immunity capability of DCs could be recovered and reduced expression of Foxo3 after blocked mTOR by Rapamycin. The second we will approach to promote anti-tumor immunity without influence on the transplanted tolerance in the same times. The third we will quantifying the data of MDSCs/DCs/Foxo3 not only animal modal but also from the clinical cases and analysis the dose-effect relationship with immune suppressive agents which is for to use to diagnosis rejection and assessment the patients' immune level and molecule target treatment of cancer in the future.

癌症是移植后长期存活的主要死因。移植后治疗窗狭窄，内环境脆弱，移植耐受和肿瘤免疫尖锐矛盾是移植后肿瘤研究和治疗的瓶颈。我们前期工作证实膀胱癌患者血及瘤床中树突细胞（DCs）数量减少；髓源性抑制细胞（MDSCs）堆积；肿瘤切除后DCs大幅上升；提示癌症明显抑制宿主免疫和DCs。有证据移植肾和肿瘤中MDSCs蓄积并诱导DCs耐受但机制不详。最近报道肿瘤中DCs高表达核转录因子Foxo3促进肿瘤发生；Foxo3高表达是致DCs耐受总关键并受PI3K/Ak调控；阻断其表达可调节DCs耐受状态。课题拟建小鼠主动脉移植后负载膀胱癌模型中阐明耐受性DCs高表达Foxo3为调控DCs免疫或耐受的总开关；证明雷帕霉素阻断mTOR降低DCs表达Foxo3可恢复DCs免疫力，探讨保证移植耐受同时提高抗肿瘤免疫；分析实验及临床数据与免疫抑制剂的量效关系，为今后诊断排斥和评估免疫及肿瘤分子靶向防治提供新思路。

要点：主要研究计划要点是通过建立移植后荷瘤模型并给予雷帕霉素阻断mTOR受体，观察是否抑制DCs等免疫抑制细胞在肿瘤中的浸润，降低肿瘤微环境中的免疫抑制状态，恢复自然免疫力,促进局部抗肿瘤能力的同时尽量减轻对移植免疫状态的影响，提高移植后宿主的抗肿瘤免疫反应。执行情况：基本按计划完成。调整：增加了建立FOXO3基因敲除小鼠，在此基础上移植并荷瘤，比较与野生型模型中肿瘤微环境的差异，分析FOXO3在移植后肿瘤中的影响和作用，是否可成为未来检测和预防移植后肿瘤的靶点之一。主要进展和结果：实验结果提示，雷帕霉素在体外不能抑制肿瘤的生长，可以提高移植模型小鼠体内脾脏中Treg数；雷帕霉素也不能抑制荷瘤模型中肿瘤的生长。Foxo3基敲小鼠脾脏中成熟DC比例小于野生型小鼠。Foxo3基因缺失后脾脏内DC数量减少。肿瘤微环境中，野生型和基敲组移植后，经雷帕霉素处理，两组脾脏中DC成熟比例无差异、但两组DC数量均减少。提示：雷帕霉素可能抑制脾脏内成熟DC数量，但抑制过程可能与Foxo3基因无关。Foxo3基因敲除鼠移植模后，脾脏中Treg比例未见增加。提示Foxo3基因与脾脏中DC成熟有关，影响雷帕霉素提高Treg数量。病理检测结果示肿瘤微环境中，抑制性因子和细胞数量增高与肿瘤恶性程度相关。学术交流：2014年中国免疫学会移植免疫分会年会大会发言：肾移植后新发恶性肿瘤微环境中免疫细胞的变化及意义。2015年移植肾疾病岭南论坛大会发言：肾移植后病人新发恶性肿瘤微环境中免疫细胞和因子的变化。发表文章9篇，其中统计源及核心期刊8篇，SCI一篇。增加基因敲除后工作内容较多，资料整理进行中，相关文章正在加紧总结。人才培养：培养硕士2人，作为指导教师培养博士1人。