[摘 要] 目的：构建负载STING激动剂DMXAA的锰卟啉金属有机框架纳米颗粒（DPM），探讨其对三阴性乳腺癌（TNBC）细 胞4T1及其小鼠移植瘤的治疗效果。方法：通过物理吸附法制备 DPM 纳米颗粒，利用透射电镜、扫描电镜及纳米粒度电位仪表 征其形貌与理化性质。常规培养4T1细胞，细胞实验分为对照组、超声辐照组（US组）、DPM治疗组（DPM组）和DPM治疗联合超 声辐照组（DPM + US组），用CCK-8法检测细胞活性，免疫荧光法检测高迁移率族蛋白B1（HMGB1）和钙网蛋白（CRT）的表达， WB法检测STING通路相关蛋白的表达。构建4T1细胞移植瘤小鼠模型，分为四组，处理同细胞实验，测量肿瘤体积，免疫荧光法 检测移植瘤组织中Ki-67、HMGB1、CRT和缺氧诱导因子-1ɑ（HIF-1ɑ）蛋白的表达，TUNEL法检测细胞凋亡，流式细胞术检测免疫 细胞活化情况，对主要器官进行H-E染色，以评估纳米材料的体内安全性。结果：DPM呈梭形，平均粒径（268 ± 3.302）nm，电位 （33.1 ± 0.87）mV。细胞实验中 ，DPM 联合超声辐照可明显抑制 4T1 细胞的增殖（P < 0.001），提高 4T1细胞中ROS水平 （P < 0.001），诱导4T1细胞CRT表达上调（P < 0.001），HMGB1从细胞核中移至细胞质，激活STING信号通路[p-STING、p-TBK1、 p-IRF3蛋白表达均显著增加（均P < 0.001）]。体内实验中，DPM联合超声辐照可显著抑制4T1细胞移植瘤生长（P < 0.001）并促 进免疫细胞表型转化（P < 0.001），抑制移植瘤组织中Ki-67、HIF-1α蛋白表达（均P < 0.01），谷胱甘肽（GSH）产生（P < 0.01），促进 CRT、HMGB1蛋白表达、ROS产生（P < 0.001），对主要器官结构无明显影响。结论： DPM联合超声辐照可通过激活STING通路 显著抑制4T1细胞及其移植瘤的生长，诱导抗肿瘤免疫应答，且对主要器官无明显毒性。

[Abstract] Objective: To construct manganese porphyrin metal-organic framework nanoparticles (DPM) loaded with the STING agonist DMXAA, and to investigate their therapeutic effect against triple-negative breast cancer (TNBC) cells (4T1) and 4T1 cellmouse xenografts. Methods: DPM nanoparticles were prepared by physical adsorption. Their morphology and physicochemical properties were characterized using transmission electron microscopy (TEM), scanning electron microscopy (SEM), and a nanoparticle size analyzer. 4T1 cells were cultured and divided into the following experimental groups: Control, Ultrasound Irradiation (US), DPM Treatment (DPM), and DPM Treatment combined with Ultrasound Irradiation (DPM + US). Cell viability was assessed using the CCK-8 assay. The expression of high mobility group box 1 (HMGB1) and calreticulin (CRT) was detected using immunofluorescence. Expression of STING pathway-related proteins was analyzed using WB. A 4T1 cell xenograft model was constructed and randomly divided into four groups. Following the treatments as those in the cell experiments, tumor volume was measured, and immunofluorescence was used to detect the expression of Ki-67, HMGB1, CRT, and hypoxia inducible factor-1ɑ (HIF-1α) in the transplanted tumor tissues. Additionally, TUNEL assay was used to detect cell apoptosis, flow cytometry was applied to assess immune cell activation, and H-E staining was used to evaluate the safety of the nanoparticles in major organs. Results: DPM exhibited a spindle shape with an average particle size of (268 ± 3.302) nm and a zeta potential of (33.1 ± 0.87) mV. In cell experiments, DPM combined with ultrasound irradiation significantly inhibited 4T1 cell growth (P < 0.001), elevated ROS levels (P < 0.001), induced upregulation of CRT expression (P < 0.001), and caused the translocation of HMGB1 from the nucleus to the cytoplasm. In addition, the STING signaling pathway was activated, as evidenced by significantly increased expression of p-STING, p-TBK1, and p-IRF3 proteins (all P < 0.001). In vivo, DPM combined with ultrasound irradiation significantly inhibited the growth of 4T1 cell xenograft (P < 0.001), promoted immune cell phenotypic transformation (P < 0.001), suppressed Ki-67 and HIF-1α expression in xenograft tissues, and reduced GSH production (P < 0.01). It also promoted CRT and HMGB1 protein expression and ROS production (P < 0.001). No significant effects on major organ structures were observed. Conclusion: DPM combined with ultrasound irradiation can significantly inhibit the growth of 4T1 cells and the xenografts through activation of the STING pathway, induce antitumor immune responses, and show no obvious toxicity to major organs.

[基金项目] 国家自然科学基金（81901759，82271999）；武汉大学人民医院交叉创新人才项目（JCRCFZ-2022-004）