超分子纳米探针的构建及其标记肝素酶单抗对肿瘤转移MR分子成像

MR molecular imaging is of great value in early diagnosis of tumor metastasis. our studies and other reports have found that the common problems are absorption of non-specific uptake by plasma proteins and reticular-endothelial system(RES)confronted in USPIO nanoparticles in vivo applications. It leads to reduce the efficiency in nanoparticle based diagnostics and target. 100nm nanoparticles stabilized colloidal nanocapsule can escape from the RES for longer circulation times and passive targeting designed to accumulate in tumor. Heparanase is associate with tumor invasion and metastasis, and highly expressed in most malignant tumors, especially in metastases. This study plans to make tumor heparanase antigen as target and to use nanocapsules magnetic particles labeled heparanase monoclonal antibody. Nanoparticle surface modification technology, the interface self-assembly techniques and covalent cross-linking technology will be used in order to reduce non-specific uptake by plasm protein and RES, as well as in order to prolong the circulation times. Besides researching construction method of heparanase nanoparticles stabilized colloidal nanocapsule, in vivo kinetics and biodistribution characteristics and heparin antibodies mediated characteristics, this study will use MR molecular imaging with SWI combined and explore the targeted binding mechanism of the heparanase nanoparticles stabilized colloidal nanocapsule in order for the early diagnosis of malignant tumor metastasis

MR分子影像在肿瘤转移早期诊断具有重要价值，文献及我们既往的研究表明，USPIO作为常用的载体存在网状内皮系统和血浆蛋白的非特异吞噬，影响MR分子显像的靶向性和特异性。100 nm左右内的胶体纳米胶囊既可以有效躲避网状内皮系统的捕获从而获得更长的体内循环时间，又具有被动靶向沉积的特点。肝素酶与肿瘤转移密切相关，肝素酶在大多数恶性肿瘤中都有表达，尤其是在转移瘤中强表达。本研究拟以肿瘤肝素酶抗原为靶点，采用胶体纳米磁微粒标记肝素酶单克隆抗体，运用纳米颗粒的表面修饰技术、界面自组装技术和共价交联技术，降低胶体纳米胶囊的非特异性吞噬，延长其循环时间，研究肝素酶胶体纳米胶囊的构建方法和体内动力学、生物学分布特点和肝素酶抗体介导的结合特性，利用磁敏感成像（SWI）进行MR分子成像，探讨胶体纳米肝素酶胶囊的靶向结合机制，以期为恶性肿瘤早期转移的诊断。

成功获得5株抗肝素酶单抗，经过多重细胞筛选方案后获得特异性较好细胞株2株进入活体实验，细胞株专利申请准备中；采用微乳液法，在表面活性剂NP-5与环己烷的体系中加入15nm左右的四氧化三铁超顺磁纳米颗粒，在30%氨水的体系中引入TEOS以形成二氧化硅包裹的四氧化三铁超顺磁颗粒，随后对纳米颗粒进行氨基改性，采用共价作用引入近红外功能团和聚乙二醇功能团。该方法包括纳米二氧化硅/四氧化三铁壳核颗粒的制备， 磁性二氧化硅微球表面近红外功能团、胺基化以及聚乙二醇化修饰。成功制备纳米二氧化硅/四氧化三铁壳核颗粒并完成表面生物功能修饰，提交专利申请一项；所修饰的功能团能够稳定的与多种小分子化学物质，蛋白质，短肽，抗体等结合，并能被三种以上成像手段监测。以该方法制备出的磁性纳米材料可根据不同的大小分子化学物质，蛋白质，短肽，抗体和放射性元素等结合用于多模态成像，实现对疾病的诊断治疗与监测。