[摘 要] 目的：制备并表征负载吲哚菁绿（ICG）的纳米胶束（F127-ICG），利用其光热效应、抗原吸附能力及淋巴结（LN）靶向 优势，探索F127-ICG的抗肿瘤作用。方法：采用薄膜水化法制备F127-ICG，通过粒度电位仪测量F127-ICG的粒径及Zeta电位， 通过紫外可见分光光度计及荧光分光光度计检测F127-ICG的吸收光谱及荧光光谱。通过比较F127胶束与肿瘤细胞裂解液孵育 前后基本性质及蛋白含量变化，分析F127-ICG的抗原吸附作用。Calcein-AM/PI双染法检测F127-ICG对乳腺癌细胞4T1的光热 杀伤作用。在小鼠皮下注射染料标记的F127胶束构建淋巴引流模型，使用小动物活体成像检测F127胶束的LN靶向作用，并使 用离体器官成像检测F127胶束在小鼠腹股沟LN及腋窝LN中蓄积和渗透情况。构建小鼠背部双侧乳腺癌肿瘤模型，小鼠瘤内 注射F127-ICG进行光热治疗，观察对侧肿瘤生长趋势。结果：成功构建负载ICG的F127胶束，粒径为（19.41 ± 0.49）nm， Zeta电位为-（2.78 ± 0.36）mV。F127-ICG与肿瘤抗原共孵育后粒径、负Zeta电位以及蛋白含量均增大（P < 0.05）。Calcein-AM/PI 双染法结果显示，F127-ICG可以发挥光热效应杀伤4T1细胞。活体成像结果显示，F127胶束可靶向LN。动物体内实验中，与 PBS及F127-ICG组相比，F127-ICG + 激光组的对侧肿瘤体积更小（P < 0.05）。结论：F127-ICG通过光热消融原位肿瘤组织，同 时捕获释放的肿瘤抗原并迁移至局部LN，促进机体抗肿瘤免疫应答，抑制远处肿瘤生长，增强原位疫苗效应。

[Abstract] Objective: To prepare and characterize indocyanine green (ICG)-loaded F127 nanomicelles (F127-ICG) and explore their anti-tumor effects through their photothermal effect, antigen-capturing capability, and lymph node (LN)-targeting properties. Methods: F127-ICG was prepared using the thin-film hydration method. The particle size and Zeta potential were measured using a Malvern particle size analyzer. The absorption and fluorescence spectra were determined using ultraviolet-visible and fluorescence spectrophotometry, respectively. The antigen-capturing capability of F127-ICG was analyzed by comparing the basic properties and protein content changes of F127 nanomicelles before and after incubation with tumor cell lysates. The photothermal cytotoxicity of F127-ICG against breast cancer cells (4T1) was evaluated using a Calcein-AM/PI double-staining assay. Dye-labeled F127 nanomicelles were subcutaneously injected into mice to establish a lymphatic drainage model. The LN targeting ability of F127 nanomicelles was detected using in vivo small animal imaging, and the accumulation and penetration of F127 nanomicelles in the inguinal and axillary lymph nodes of mice were detected by ex vivo organ imaging. A bilateral breast cancer model was established on the back of the mouse, and F127-ICG was injected intratumorally for photothermal therapy (PTT). Meanwhile, the growth of the contralateral tumor was monitored. Results: The F127-ICG nanomicelles were successfully prepared, with a mean particle size of (19.41 ± 0.49) nm and an average Zeta potential of –(2.78 ± 0.36) mV. After co-incubation of F127-ICG with tumor antigens, the particle size, Zeta potential negativity, and protein content all increased (P < 0.05). The results of the Calcein-AM/PI double staining assay revealed that F127-ICG could effectively exert a photothermal effect to mediate the killing of 4T1 cells. In vivo imaging results revealed that F127 nanomicelles can effectively target LN. In animal experiments, the contralateral tumor volume in the F127-ICG + Laser group was smaller than that in the PBS and F127-ICG groups (P < 0.05). Conclusion: F127-ICG-mediated photothermal effects can effectively ablate primary tumors and capture released tumor-specific antigens and then migrate to local LN, which promotes anti tumor immune response, inhibits distant tumor growth, and enhances in situ vaccine efficacy.

[基金项目] 肿瘤微环境与免疫治疗湖北省重点实验室开放基金（No. 2023KZL024）