基于肿瘤氧供与高渗透策略构建多功能红细胞及其联合光热/光动力与免疫疗法治疗三阴性乳腺癌

基于近红外光敏剂的光热/光动力疗法联合PD-1/PD-L1阻断治疗具有抗三阴性乳腺癌（TNBC）协同作用，但肿瘤乏氧与光敏剂的低组织渗透性限制了协同效应的充分发挥。本项目拟对红细胞进行iRGD靶向修饰，以pH-敏感化学键为桥梁连接PD-1/PD-L1小肽抑制剂（PPI）与红细胞膜，通过偶联TAT制备纳米光敏剂并利用TAT的穿膜功能将其导入红细胞内，构建多功能红细胞。利用红细胞的长循环特性与iRGD的肿瘤血管靶向性，多功能红细胞能有效蓄积于肿瘤部位，随后在瞬时高能聚焦超声下发生破碎，释放纳米光敏剂、PPI囊泡与血红蛋白。TAT介导纳米光敏剂深入肿瘤内部，在激光照射下发挥光热/光动力疗效；血红蛋白改善肿瘤乏氧，增强光动力效应；iRGD兼具肿瘤穿透性能，促进PPI囊泡进入弱酸性肿瘤微环境，释放PPI阻断PD-1/PD-L1信号通路，激活抗肿瘤免疫；各种效应相互呼应，协同抑制TNBC的生长与转移。

Near-infrared photosensitizer-mediated photothermal (PPT)/photodynamic therapy (PDT) combined with PD-1/PD-L1 immune checkpoint blockade exhibits synergistic effects against triple-negative breast cancer (TNBC). However, tumor hypoxia and poor tissue-penetration ability of photosensitizers have dramatically limited their synergistic efficacy. In this project, a therapeutic system based on multifunctional red blood cells (MFRBCs) is designed for TNBC treatment through combining PTT/PDT and immunotherapy. MFRBCs are prepared using the following method. Red blood cells (RBCs) are surface-modified with iRGD peptide and then PD-1/PD-L1 peptide inhibitor (PPI) is covalently linked to the cell membranes through a highly acid-labile benzoic-imine bond. Photosensitizer is conjugated with TAT peptide and used to prepare nanophotosensitizer by self-assembly method. Afterwards, nanophotosensitizer is imported into RBCs prepared above by using the strong cell-penetrating ability of TAT, thus obtain MFRBCs. Several mechanisms are involved in the synergistic effects of MFRBCs against TNBC. Due to the long-circulation characteristics of RBCs and the active targeting ability of iRGD for tumor neovasculature, MFRBCs can be effectively accumulated in the tumor. Next, MFRBCs will disrupt under the instantaneous high intensity focused ultrasound and thus release nanophotosensitizer, PPI vesicles and hemoglobin. Nanophotosensitizer can penetrate deep into the tumor via the TAT mediation to exert PTT/PDT therapeutic effects under the laser irradiation. iRGD also possesses strong tumor penetration ability and can mediate PPI vesicles to enter the tumor interior. Afterwards, PPI can be released from these vesicles by responding to the acidic pH of tumor microenvironment to block the PD-1/PD-L1 signal path, and thus activate the host antitumor immunity. Because of the strong oxygen-carrying and delivering capacity, hemoglobin can overcome the tumor hypoxia, which will further benefit for enhancing the PDT efficiency and improving the immune activation. All these effects will echo each other and thus efficiently suppress the growth and metastasis of TNBC. In this project, we will evaluate the synergistic anti-TNBC effects of MFRBCs both in vitro and in vivo and also preliminarily illuminate their relevant functional mechanisms.