新型肿瘤疫苗Eps8-CRT-chi9558抗乳腺癌转移的作用及机制研究

Tumor metastasis is the leading cause of refractory or relapsed breast cancer. Epidermal growth factor receptor pathway substrate No.8 (Eps8) facilitates metastasis of breast cancer and represents an attractive target for preventing and treating metastasis of breast cancer. In our previous studies, we have successfully constructed Eps8-CRT-chi9558 vaccine and it preliminarily demonstrated satisfactory anti-tumor effect on murine breast cancer model. However, further elucidation of the mechanism is urgently needed. In this study, we plan to target at Eps8 and establish transplanted and spontaneous murine breast cancer models. Moreover, in vivo optical imaging animal experiments will be performed to elucidate the anti-growth-and-metastasis effect of Eps8-CRT-chi9558 vaccine against breast cancer. Furthermore, we will examine humoral and celluar immune responses induced by the combined vaccine in anti-tumor immunity, and will evaluate gene and protein expression in anti-metastasis-related signaling pathways influenced by the combined vaccine by gene chip and Westernblot in vitro. Taken together, our study tries to unveil the anti-metastasis effect and mechanism of the novel Eps8-targeted vaccine against breast cancer. These findings will estabilish an experimental basis for preventing and treating metastasis of breast cancer, and thus will provide a new method and strategy for radical treatment of breast cancer.

乳腺癌转移导致乳腺癌难治和复发。表皮生长因子受体通路底物8(Eps8)促进乳腺癌转移，是预防乳腺癌转移和根治乳腺癌的新靶点。我们已在前期研究中成功构建了Eps8-CRT-chi9558疫苗，并在小鼠荷4T1乳腺癌模型中初步显示出良好的抗肿瘤免疫效应，但其抗乳腺癌转移的作用及机制亟待进一步阐明。因此，本课题设计以Eps8为靶点切入，复制小鼠移植乳腺癌和自发乳腺癌模型，通过荧光活体成像等体内动物实验明确Eps8-CRT-chi9558抗肿瘤生长及转移的效果；体外免疫学实验分析复合疫苗对抗肿瘤体液免疫和细胞免疫的影响；基因芯片和Westernblot评价复合疫苗对肿瘤转移相关信号通路中基因和蛋白表达的作用，可望阐明Eps8这一新靶点疫苗在防治乳腺癌转移中的作用及其机制，为该疫苗用于临床预防和治疗乳腺癌转移提供实验依据，为根治乳腺癌提供新方法和策略。

乳腺癌是危害女性健康的首要恶性肿瘤，传统的手术及放疗、化疗不能有效阻止乳腺癌复发和转移。免疫疗法可为乳腺癌的复发和转移提供新的治疗策略。目前，大部分乳腺癌疫苗以ErbB 家族（如 HER/neu）、 MUCI-1、 Survivin 等基因为靶点，虽已进入临床研究阶段，但治疗效果有限。表皮生长因子受体通路底物8(Epidermal Growth Factor Receptor pathway substrate No.8, Eps8)能促进乳腺癌转移，是预防乳腺癌转移和根治乳腺癌的新靶点。针对乳腺癌疫苗的上述缺陷，课题组设计了以Eps8为靶点，采用钙网蛋白(Calreticulin, CRT)为免疫佐剂及新一代调节性体内延迟减毒的沙门氏菌χ9558为载体的新型口服疫苗，旨在获得最优化抗乳腺癌疫苗，通过便捷的免疫途径诱导机体产生抗肿瘤免疫，有效防治乳腺癌转移和复发。.课题组成功构建高免疫原性Eps8-CRT-χ9558复合疫苗，该疫苗在小鼠4T1乳腺癌显示良好抗肿瘤作用及在Lewis肺癌模型显示预防肺癌进展的效果。在抗肿瘤作用的免疫机制研究中显示，Eps8-CRT-χ9558复合疫苗可以通过提高IFN-γ的分泌，提高CD8+T细胞比例，以及下调CD4+Foxp3+调节性T细胞比例从而提高对肿瘤杀伤作用。在分子机制方面，与对照组相比Eps8-CRT-χ9558复合疫苗免疫的荷瘤小鼠肿瘤组织中，EGFR、Eps8、p-ERK、pAKT、p-GSK3β及p-c-Raf显著低表达，提示复合疫苗经口服途径进入机体后，被巨噬细胞吞噬并递呈抗原，机体产生 Eps8特异性细胞及体液免疫反应，靶向性作用于 Eps8 高表达的肿瘤细胞，通过抑制EGFR下游分子活化如ERK、AKT等，从而抑制肿瘤的生长与转移。.课题组构建Eps8-CRT-x9558复合疫苗是以钙网蛋白为免疫佐剂、采用新一代调节性体内延迟减毒的沙门氏菌x9558为载体的新型口服疫苗，具有Eps8靶向性，高免疫原性、抗肿瘤效果显著的特点，可应用于所有Eps8高表达的肿瘤，此外该疫苗为口服疫苗，在临床应用上更安全与简便，这也利于后期复合疫苗的临床转化及产业化。.课题相关研究成果共发表SCI论文9篇；申请专利4项，授权1项。.培养优秀研究生共7位，其中4位获得博士学位，3位获得硕士学位。