CRT促进DC疫苗抗肿瘤特异性免疫应答研究

Tumor cells are capable of evading the immunosurveillance and elimination of the immune system by various means, this is the very reason that why immunotherapy of anti-tumor is hardly successful. To reverse this situation, it is necessary to promote the specific identification of antigen by the immune system to enhance immune response. Recent studies showed that after treated with anthracycline-based chemical drugs, tumor cells entered into the apoptosis process followed by the CRT ecto. Thus, exposure of CRT to the immune cells could induce the clearance of apoptotic cells by the immune system. Our previous works demonstrated that the high level expression CRT in tumor cells B16 mediates the tumor antigen E2 ecto, increasing the specificity of tumor cell identification and enhancing specific anti-tumor immune response. In present study, DC was sensitized by CRT-MUC1, which was delivered positively and efficiently by CPP, a cellular delivery system. A series of in vivo/vitro experiments were performed to explore the potential mechanism by which CRT enhanced primed-DC to present antigen and induced anti-tumor specific immune response. It will provide a new pattern for the research of DC vaccine in the future.

肿瘤细胞具备多种免疫逃逸机制，如何克服这些机制是目前肿瘤免疫疗法所面临的最大挑战。因此，增强机体免疫系统特异性识别肿瘤细胞的能力，对于肿瘤的根治至关重要。最新研究发现，蒽环类化学药物诱导的肿瘤细胞凋亡，伴随钙网蛋白（Calreticulin，CRT）膜外侧向移动定位（Ecto），诱使吞噬细胞主动清除凋亡的肿瘤细胞。我们前期数据表明，肿瘤细胞B16内表达的外源性CRT介导了肿瘤抗原E2的定位改变，显著提高了DC（Dendritic cells）对肿瘤细胞的识别并产生特异性抗肿瘤免疫应答，且具有浓度依赖性。本项目拟在此基础上，利用细胞穿膜肽（Cell penetrating peptide，CPP）可主动高效向胞内递送CRT-MUC1致敏DC的特点，通过体内外细胞与动物学实验，探讨CRT提高致敏DC的抗原呈递能力，诱导机体抗肿瘤特异性免疫应答的可能机制，为抗肿瘤DC疫苗的研制提供一种新思路。

钙网蛋白（calreticulin，CRT）作为一个参与细胞识别的关键性分子，可外翻到（前）凋亡细胞表面向树突状细胞（dendritic cells，DCs）或其它抗原递呈细胞（antigen presenting cells，APCs）发出“吞噬我”的信号，进而诱导吞噬作用。肿瘤相关抗原（tumor associate antigen，TAA）可被DCs 进一步呈递给T 细胞，从而诱导特异性抗肿瘤免疫应答。基于前期基础，我们认为CRT 是一种理想的抗肿瘤免疫“分子佐剂”，可介导TAA 移动定位于细胞膜表面，诱导肿瘤抗原特异性免疫反应。在本研究中，我们首先证实CRT 可携带乳腺癌抗原MUC1 外翻到乳腺癌细胞4T1 表面，诱导时间依赖性细胞凋亡。进一步实验证实，当腺病毒包装的MUC1-CRT 致敏原代DC 后，可有效诱导T 细胞增殖及细胞因子的分泌。表明CRT 可作为MUC1 的“免疫佐剂”诱导高效的基于DC 的抗肿瘤免疫应答反应。此外，通过hpp10-3DRBD 携带SOCS1 siRNA 入胞，可有效沉默DC 中的负调控因子SOCS1，高效解除其对DC 抗原呈递能力的抑制作用，进一步提高CRT 促DC 抗肿瘤免疫应答的能力。