Fe3O4@Au-C225复合靶向磁性纳米粒子介导胶质瘤综合治疗研究

Glioma is one of the malignant tumors which are serious threat to human health. So the search for new effective treatment methods and drugs and developing an integrated treatment or combination therapy have attracted the widespread concern of researchers. A lot of basic and clinical research have been carried out on the MNPs-mediated glioma hyperthermia. But how to give the MNPs active targeting, improve glioma cells MNPs distribution, still is hot. This project intends to build Fe3O4@Au-C225 composite targeted MNPs based on the introduction of targeting EGFR McAb-C225 and construction of Fe3O4@Au composite MNPs successfully, further to characterize it, to evaluate the biocompatibility. Then we evaluate the targeting effects of Fe3O4@Au-C225 composite MNPs to established U251 cell models in vitro and xenograft models on the nude mouses in vivo by MRI. Furthermore we apply the magnetic induction heating combined with near-infrared light on both models to realize "efficient targeted, joint damage", to verify and compare the treatment efficacy of integrated therapy and single-factor treatment. Meanwhile we detect the expression of EGFR and apoptosis related genes and the distribution of Fe3O4@Au MNPs in the tumor tissue to explore the mechanism of the integrated therapy on glioma mediated by Fe3O4@Au-C225 composite targeted MNPs.

脑胶质瘤是严重威胁人类健康的恶性肿瘤之一，寻求新的有效治疗方法和药物，开展综合治疗、联合治疗受到研究者的广泛关注。MNPs介导的胶质瘤热疗已进行了大量基础和临床研究，如何赋予MNPs的主动靶向性、提高胶质瘤细胞内MNPs分布，仍是目前研究热点。本课题拟在成功构建Fe3O4@Au复合MNPs的基础上，引入靶向EGFR的McAb-C225，进一步构建Fe3O4@Au-C225复合靶向MNPs，对其进行表征，评价其生物相容性；建立U251细胞模型和裸鼠异种移植瘤模型，通过MRI在体外和体内评价Fe3O4@Au-C225复合纳米制剂对胶质瘤细胞的靶向作用，体外磁感应联合近红外光加热，实现对胶质瘤细胞的"高效靶向、联合杀伤"，验证和比较综合治疗和单因素治疗的疗效；检测EGFR和凋亡相关基因表达，病理学检查瘤组织内Fe3O4@Au分布情况，探讨Fe3O4@Au-C225介导胶质瘤综合治疗的作用机制。

1. 项目背景：脑胶质瘤是严重威胁人类健康的恶性肿瘤之一，MNPs介导的胶质瘤热疗已进行了大量基础和临床研究，如何赋予MNPs的主动靶向性、提高胶质瘤细胞内MNPs分布，仍是目前研究热点。.2.研究内容：本项目在成功构建Fe3O4@Au复合MNPs的基础上，引入靶向EGFR的McAb-C225，进一步构建了Fe3O4@Au-C225复合靶向MNPs，进行表征，评价其生物相容性；建立U251细胞模型和裸鼠异种移植瘤模型，通过MRI在体外和体内评价Fe3O4@Au-C225复合纳米制剂对胶质瘤细胞的靶向作用，体外磁感应联合近红外光加热，验证和比较综合治疗和单因素治疗的疗效；检测凋亡相关基因表达，探讨Fe3O4@Au-C225介导胶质瘤综合治疗的作用机制。.3.重要结果和关键数据：①Fe3O4@Au-C225制备和表征：其平均直径为46nm；在pH 7.4中性环境中，其zeta电位值为11.1±1.8 mV；表面C225活性保持90%以上；最大光吸收峰在630nm；其磁滞回线接近为重合“S”型曲线； ②Fe3O4@Au-C225生物相容性研究：其体外细胞毒性被列为1级；小鼠LD50确定为8.39g/kg，其95％CI为6.58-10.72g/kg。③Fe3O4@Au-C225体外综合治疗胶质瘤疗效评价：流式细胞分析发现，Fe3O4@Au-C225复合MNPs介导的MFH联合NIR热疗组凋亡率最高。RT-PCR和Western blot检测表现为Caspase9和凋亡执行分子Caspase 3的蛋白表达明显升高。④Fe3O4@Au-C225体内靶向评价：活体MRI结果显示，注射后肿瘤组织不同时间点的T2WI、T2*WI强度及信号变化率均有明显下降，具有统计学差异。⑤Fe3O4@Au-C225体内综合治疗胶质瘤疗效评价：Fe3O4@Au-C225靶向MNPs联合热疗组与对照组相比其肿瘤质量和体积均有显著减小（p<0.05）。.4.科学意义： Fe3O4@Au-C225复合靶向MNPs对比于Fe3O4@Au复合MNPs，具有更好的靶向性，该材料注射入人胶质母细胞瘤细胞系U251荷瘤鼠模型后，可在MRI进行示踪，然后进行联合或序贯体外磁感应定向加热和近红外光加热，实现“高效靶向、联合杀伤”作用，为脑胶质瘤联合治疗和影像诊断一体化开辟一条的新途径。