新型GM-CSF锚定的膀胱癌肿瘤干细胞疫苗的治疗作用及相关机制研究

Bladder cancer (BCa) is the most common malignancy of the urinary system, with high recurrence and metastasis rate. In our previous study, We have prepared cytokines surface-modified tumor vaccines using our novel protein-anchor technology. The vaccines can efficiently induce mice to produce specific anti-tumor immune response and make the tumor shrank or even dissipated. However, We found that part of the tumor regression and then grow again after the vaccine treatment, We speculate that the reason may be the existence of CSCs. So we plan to prepare a novel cytokines surface-modified bladder cancer stem cell (BCa-CSCs) vaccine, which could induce the specific anti-CSCs immune response,then achieve the purpose of the elimination of CSCs. Therefore, this project intends to isolate the BCa-CSCs, and then prepares a novel bladder cancer stem cell vaccine by immobilizing GM-CSF on BCa-CSCs on the basis of our protein-anchor technology,and this kind of BCa-CSCs vaccine will be used to treat the bladder cancer in mice. Flow cytometry, immunohistochemistry and CTL assay are used to evaluate the antitumor efficiency of the BCa-CSCs vaccine,and then explore the mechanism. This study intends to demonstrate whether the novel GM-CSF surface-modified BCa-CSCs vaccine could activate specific anti- BCa-CSCs immune response. The findings of this project may provide some experimental basis for the application of this kind of vaccines in human bladder cancer treatment.

膀胱癌（BCa）是泌尿系统最常见的恶性肿瘤，浸润性BCa术后易复发、转移。在前期研究中，我们用细胞因子锚定修饰BCa制成肿瘤细胞疫苗，能使小鼠肿瘤明显缩小甚至完全消退，但发现仍有部分肿瘤缩小后再次生长。推测可能与BCa肿瘤干细胞（CSCs）未被清除有关。因此，我们设想若研制一种新型细胞因子锚定的膀胱癌CSCs疫苗，有可能诱导机体产生特异性免疫反应，杀伤CSCs，达到彻底清除CSCs的目的。由此，本项目拟分离筛选出BCa的CSCs，采用我们特有的锚定技术，将细胞因子GM-CSF锚定修饰于CSCs细胞表面，研制出一种新型BCa肿瘤干细胞疫苗。将此疫苗接种于BCa模型小鼠，采用组织病理、免疫组化、CTL检测、流式细胞检测等技术，检测其疗效并研究其作用机制。以验证该新型疫苗能激发接种小鼠产生对同种CSCs特异性免疫反应，能有效地清除CSCs。为用这种新型疫苗防治膀胱癌术后复发、转移奠定实验基础。

膀胱癌是最常见的泌尿系统恶性肿瘤，术后容易复发、转移的原因可能是肿瘤干细胞没有被清除或消灭。我们成功筛选分离并鉴定出膀胱癌干细胞，并在前期蛋白修饰细胞表面的技术基础上，将GM-CSF、IL-2等细胞因子锚定于膀胱癌干细胞表面而制备成了一种新型肿瘤细胞疫苗，通过预防性、治疗性、记忆性、特异性动物实验，发现疫苗可显著增强疫苗的免疫原性，有效地激活机体的抗肿瘤免疫反应，诱导机体产生肿瘤特异性免疫反应，抑制和杀伤肿瘤细胞；促进树突状细胞的成熟，显著地增加浸润在肿瘤组织中效应性CD4＋、CD8＋T淋巴细胞量，增强特异性T淋巴细胞的杀伤肿瘤细胞的能力，从而有效地直接杀伤肿瘤干细胞；诱导机体产生长期的保护性免疫反应和对抗同种肿瘤细胞的特异性记忆性抗肿瘤免疫反应，能保护小鼠抵抗肿瘤的再次攻击。该研究为采用这种新型肿瘤干细胞疫苗靶向治疗膀胱癌奠定了实验基础。