在前项国家自然基金中，我们完成了胶乳微球式人工抗原提呈细胞在体内靶向扩增肿瘤抗原特异性T细胞的观察性研究。本项目拟接着进行机制性探讨，并全面改造实验系统：用可生物降解的聚乳酸-羟基乙酸（PLGA）微粒做载体，表面负载MHC/peptide、共刺激分子、粘附分子和抗吞噬分子，内部包裹细胞因子、趋化因子和抑制性信号通路封闭剂，制备新型多功能人工抗原提呈细胞，在黑色素瘤肺转移鼠和皮下瘤鼠体内，通过表面提呈和旁分泌缓释方式募集和粘附T细胞，提供第一、二、三信号靶向活化和扩增肿瘤抗原特异性T细胞，同时封闭抑制性信号通路，扩增记忆T细胞，降低抑制性细胞浸润，并防止单核巨噬细胞对微粒的吞噬。利用活体量子点荧光成像和磁共振示踪、激光共聚焦、电镜、免疫组化、流式分析及细胞功能性检测等手段，研究该系统的更有效治疗方案、体内作用机制、时空运行、毒副作用及对整体免疫功能的影响，促使这一新免疫疗法早日走近临床研究。

Priming and amplifying antigen-specific T lymphocytes in vivo is one of the fundamental strategies for the tumor immunotherapy. Based on our previous works about latex bead-based artificial antigen-presenting cells, in this project, a novel artificial antigen-presenting cell system (aAPCs) will be developed by co-coupling MHC/peptide complexes, anti-CD28, anti-4-1BB, anti-CD2 and CD47 molecules onto the biodegradable poly (lactic-co-glycolic acid) (PLGA) microparticles and encapsulating the mixture of IL-2, IL-15, CCL21, blockade of both CTLA-4 and PD-1 inside. With the presentation of surface-bound ligands and paracrine delivery of soluble factors, the aAPCs are expected to possess multifunctional activities in vivo, such as recruiting antitumor effectors, targeting, priming and expanding tumor antigen-specific T cells by presenting antigen (signal 1) and costimulation (signal 2), releasing cytokines (signal 3) and providing adhesion molecules. Simultaneously, the aAPCs will be able to promote the expansion of specific T cells as memory T cells, block inhibitory pathways to enhance T cell proliferation and reduce the frequencies of suppressor cells. In addition, surface-bound CD47, as a marker of self, is expected to inhibit phagocytosis of PLGA particles by monocytes and macrophages to prolong their circulation time. In this study, the novel aAPCs will be administrated into B6 mice for the active immunotherapy of B16 melanoma pulmonary metastases and subcutaneous tumor. Antigen-specific T cell responses and tumor growth will be monitored. The optimal therapy regimens, precise mechanisms by which aAPCs achieve their therapeutic effects, spatial and temporal location，fate after injection, side effects and organ toxicity as well as effects on intact immune function will be elucidated. A series of modern techniques will be utilized, such as small animal optical imaging and tracking assay with double-fluorescence quantum dots, nuclear magnetic resonance, laser scanning confocal assay, electron microscopy, tissue sections and immunohistochemical staining, multicolor immunophenotyping by flow cytometry, and functional analysis for immune cells. This approach will provide initially the mechanistic insights in animal tests and pave the path to clinical trials for the novel active immunotherapy of cancers or persistent infections.