uPAR靶向性MRI探针的制备及其与乳腺癌细胞靶向结合的研究  

The Preparation of uPAR-Targeted MRI Probe and its Targetability to Breast Cancer Cells

作　　者：杨扬 孟洁[1] 温涛 陈博[2] 刘飞 顾宁[2] 许海燕[1] 于薇[4] 刘健[1] Yang Yang;Meng Jie;Wen Tao;Chen Bo;Liu Fei;Gu Ning;Xu Haiyan;Yu Wei;Liu Jian(Institute of Basic Medical Sciences Chinese Academy of Medical Sciences,School of Basic Medicine Peking Union Medical College,Beijing 100005,China;Jiangsu Key Laboratory for Biomaterials and Devices,School of Biological Science and Medical Engineering,Southeast University,Nanjing 210096,China;Research Institute of Chia Tai Tianqing Pharmaceutical Group Limited by Share Ltd,Nanjing 210018,China;Department of Radiology,Beijing An Zhen Hospital,Capital Medical University,Beijing 100029,China)

机构地区：[1]中国医学科学院基础医学研究所,北京协和医学院基础学院,北京100005 [2]东南大学生物科学与医学工程学院,江苏省生物材料与器件重点实验室,南京210096 [3]正大天晴药业集团股份有限公司,南京210018 [4]首都医科大学附属北京安贞医院,北京100029

出　　处：《中国生物医学工程学报》2018年第4期481-488,共8页Chinese Journal of Biomedical Engineering

基　　金：国家自然科学基金(81771969,81071196); 中国医学科学院医学与健康科技创新工程项目(2016-I2M-3-004)

摘　　要：乳腺癌是女性最常患的恶性肿瘤之一,实现对乳腺癌原发性及转移性微小病灶的早期检测对于提高患者生存率具有重要的意义。研究表明,尿激酶受体(uPAR)在乳腺癌肿瘤细胞及肿瘤相关细胞中高表达而在正常组织细胞中低表达。通过将次氮基三乙酸(NTA)连接到超顺磁性氧化铁纳米颗粒(SPIO)表面,获得一种通用的磁共振成像(MRI)模块(SPIO-NTA)。进而将uPAR天然配体尿激酶(uPA)的氨基末端片段(ATF)通过组氨酸标签(His tags)偶联到SPIO-NTA上,获得一种具有uPAR靶向性的MRI探针(SPIO-ATF)。其γ-Fe_2O_3核心的直径小于10 nm,平均水合直径约为77 nm,Zeta电位约为-13 m V。蛋白凝胶电泳及考马斯亮蓝染色结果证明ATF被成功导入到SPIO上。细胞实验结果表明,SPIO-ATF与乳腺癌细胞4T1的结合量与细胞uPAR的表达量正相关,表明SPIO-ATF对4T1细胞具有良好的特异性结合能力,而这种结合是通过ATF与uPAR的相互作用介导的。另外,4T1细胞对SPIO-ATF的摄取具有浓度依赖性。在实验条件下,SPIO与SPIO-ATF都没有表现出明显的细胞毒性。通过研究构建一种可用于乳腺癌靶向诊断的新型MRI探针,从而为乳腺癌的早期诊断提供一种新策略。Breast cancer is one of the most common malignancies for women. Early detection of primary and metastatic lesions of breast cancer is a key point to improve the survival rate of patients. Studies have shown that urokinase-type plasminogen activator receptor（ uPAR） is highly expressed in breast cancer cells and tumorassociated cells,while lowly expressed in normal tissue cells. In this study,by conjugating amino-nitrilotriacetic acid（ NTA） to the surface of superparamagnetic iron oxide nanoparticles（ SPIO）,we obtained an universal magnetic resonance imaging（ MRI） module（ SPIO-NTA）. Then, amino-terminal fragment（ ATF） ofurokinase-type plasminogen activator（ uPA）,the native ligand of uPAR,was introduced to SPIO-NTA via histidine tags（ His tags） to obtain an uPAR-targeted MRI agent（ SPIO-ATF）. The diameter of the γ-Fe_2O_3 core was less than 10 nm. The hydrodynamic diameter of SPIO-ATF was about 77 nm and Zeta potential was about-13 mV. The results of protein gel electrophoresis and Coomassie brilliant blue staining indicated the successful conjugation of ATF to SPIO. In vitro experiments showed a positive correlation between the amount of SPIO-ATF bound with 4 T1 cells and the expression level of uPAR,indicating that SPIO-ATF can specifically bind to 4 T1 cells via the interaction between ATF and uPAR. In addition,the cellular uptake of SPIO-ATF was increased with the increase of incubating concentration of SPIO-ATF. Under experimental conditions,neither SPIO nor SPIO-ATF showed obvious cytotoxicity. In conclusion,we developed a novel MRI probe for the target detection of breast cancer,providing a new strategy for the early diagnosis of breast cancer.

关 键 词：乳腺癌 早期诊断 磁共振成像 超顺磁性氧化铁纳米颗粒 尿激酶受体 

分 类 号：R318[医药卫生—生物医学工程]