CTLA-4单抗增强mRNA纳米疫苗抗小鼠三阴型乳腺癌免疫应答的机制研究

Triple negative breast cancer (TNBC) that constitutes 10%-20% of all breast cancers is associated with aggressive histology, high metastases and poor prognosis. The treatment of patients with triple negative breast cancer remains a great clinical challenge. The therapeutic induction of systemic anti-tumor immunity is a promising approach. The immunotherapy is a hotspot for triple negative breast cancer treatment research. Our previous research has demonstrated that tumor growth of triple negative breast cancer in mice immunized with anti-CTLA-4 monoclonal antibody (anti-CTLA-4 mAb) and mRNA-loaded nanoparticle vaccine is significantly slower than mice immunized with either vaccine or anti-CTLA-4 monoclonal antibody and the combination group induces a significant increase in tumor-infiltrating CD8+ T cells compared to the vaccine and monoclonal antibody group. In this project, we would still focus on the combination treatment based on previous work. It would be proved that therapeutic effect of mRNA-loaded nanoparticle vaccine on triple negative breast cancer is associated with an increased anti-tumor cytotoxic T-lymphocyte activity by the additional immunization with anti-CTLA-4 monoclonal antibody. Because tumor microenvironment could be improved after combination treatment, the expression of immunosuppressive molecules and the infiltration of inhibitory immune cells into tumor microenvironment would be investigated. These studies would help us to understand enhancement mechanism of therapeutic efficacy of nanoparticle-based vaccine against triple negative breast cancer by the combination treatment with anti-CTLA-4 monoclonal antibody and discover new therapeutic target, which may provide fundamental basis and experimental evidence for clinical therapy against triple negative breast cancer.

三阴型乳腺癌约占乳腺癌病例的10-20%，具有侵袭性强、转移率高和预后差等特征，已成为临床治疗的难点。免疫疗法表现出巨大潜力，逐渐成为该病治疗研究的热点。申请人前期研究证实免疫检查点分子CTLA-4 单抗能增强mRNA纳米疫苗抗小鼠三阴型乳腺癌肿瘤的生长，联合免疫组肿瘤浸润性CD8+ T细胞比例显著高于疫苗免疫组和单抗免疫组。本项目旨在前期基础上借助三阴型乳腺癌小鼠模型，在细胞和分子水平上深入研究CTLA-4单抗增强mRNA纳米疫苗诱导的细胞毒性T淋巴细胞抗肿瘤免疫反应，以及联合治疗后肿瘤微环境的改变，特别是抑制性免疫细胞浸润和免疫抑制分子表达的变化，探讨CTLA-4 单抗增强mRNA纳米疫苗抗小鼠三阴型乳腺癌免疫应答的机制，获得新的治疗靶点，为该病的临床治疗提供理论基础和实验依据。

乳腺癌是女性最常见的癌症之一，而三阴型乳腺癌（Triple-negative breast cancer, TNBC）是最具侵袭性的乳腺癌分型之一。由于缺乏治疗靶点，给TNBC的临床治疗带来了巨大的挑战。免疫疗法正成为一种有潜力的治疗TNBC的方法。申请人前期研究证实免疫检查点分子CTLA-4 单抗能增强MUC1 mRNA纳米疫苗抗小鼠三阴型乳腺癌肿瘤的生长，联合免疫组肿瘤浸润性CD8+ T细胞比例显著高于疫苗免疫组和单抗免疫组。本项目开展了CTLA-4 单抗增强mRNA纳米疫苗抗小鼠三阴型乳腺癌免疫应答的机制研究。研究结果表明，MUC1 mRNA纳米疫苗和CTLA-4单抗联合治疗可诱导更强的抗肿瘤CTL反应，导致肿瘤微环境（Tumor microenvironment, TME）中骨髓来源的抑制性细胞（MDSC）、Treg细胞、肿瘤相关成纤维细胞（TAF）和肿瘤脉管系统显著减少，白细胞介素-6、肿瘤坏死因子-α和转化生长因子-β表达下调，IFN-γ和白细胞介素-12显著上调，CD8+ T细胞数量增加，并且比mRNA纳米疫苗或CTLA-4单抗更有助于肿瘤细胞凋亡。此外，联合治疗可显著下调信号转导和转录激活因子3（STAT3）信号通路。综合转录组测序和蛋白质组分析结果也证实联合治疗使促进细胞增殖、抑制凋亡的PI3K-Akt信号通路发生了显著性变化，比单独治疗更有助于肿瘤细胞的凋亡，从而有利于启动获得性免疫应答。因此，我们认为CTLA-4单抗与MUC1 mRNA纳米疫苗的联合通过降低免疫抑制性TME、抑制促肿瘤STAT3信号通路和促进肿瘤细胞凋亡来增加CD8+ T细胞浸润肿瘤和提高对肿瘤细胞的杀伤效应，为临床治疗三阴型乳腺癌提供了理论基础和实验依据。