GRP受体介导的、铃蟾肽-125碘标记的PEG化壳聚糖衣磁性纳米粒子制备及检测微小胰腺癌的MRI研究

Pancreatic cancer is the third most common cause of.gastrointestinal malignancies.Despite significant improvements in surgical.and non-surgical treatment modalities , the 5-year survival rate remains poor. Early detection of panreatic cancer is crucial for the improvement of clinical treatment outcomes .It is difficult to detect the micro pancreatic tumor correctly and to distinguised benign and malignent pancreatic lesions using conventional clinical imaging methods. Novel approaches for early detection will make the most significant impact on clinical management of this disease. .Previously, we have developed a colloidal dextran-stabilized iron oxide nanocompound (SPIO) as a magnetic label . In an effort to use this magnetic label to visualize pancreatic tumor by MRI in vivo, we synthesis a new nanoparticles ,mediated by gastrin releasing peptide (GRP)receptor which is high expression biomarker on pancreastic cancer cells. In current study，we try to conjugate bombesin －125 I to PEG－Chitosan－SPIO and analysis its chemical and physical characteristics. In vitro and vivo competitive binding experiments to assess the magnetically labeled retained affinity to the GRP receptors. Two animals models ,including a rat model of pancreatic intraepithelial carcinoma induced by DMBA and red fluorescent protein orthotopic transpalntation nude mice metastasis model of pancreatic cancer, were established ，respectively.we evaluated the target-specific agents potential use in early MR imaging of micro panreatic tumors.If successful，this new approach may be valuable for other tumors targeting enhancemenc.

胰腺癌是我国常见的消化道恶性肿瘤，临床治疗效果差。如何早期准确诊断是提高其临床疗效的关键。本课题拟制备出GRPR受体介导的、具有长循环特性、铃蟾肽－125I标记的 PEG-CS-SPIO磁性纳米粒子，测定其理化特性，并在7，12．二甲基苯并蒽(7，12－dimethyl－l，2-benzanthracene，DMBA)可以诱导形成大鼠的胰腺导管腺癌模型及裸鼠皮下荷人胰腺癌模型上进行研究，探讨靶向增强显示微小胰腺癌灶的可能。一旦成功，将对早期胰腺癌及胰腺癌与慢性胰腺炎的鉴别提供新的途径，具有十分重要的意义。对其它肿瘤如前列腺癌与前列腺增生的鉴别也有一定的借鉴作用。

背景：前列腺癌早期诊断对治疗效果和预后有着重要意义。胃泌素释放肽受体（GRPr）在前列腺癌细胞表面高表达，是分子成像的合适靶点。本研究尝试制备以GRPr为靶点，铃蟾肽BBN修饰、荧光素标记的可用于MR和光学成像的双模态纳米微粒Gd2O3-FI-PEG-BBN，研究其体内外主动靶向能力及双模态成像性能。.目的：1、探索铃蟾肽和荧光素双标记的Gd2O3纳米颗粒Gd2O3-FI-PEG-BBN的构建方法及理化性能。2、培养GRPr阳性的人前列腺癌细胞系PC-3，评价Gd2O3-FI-PEG-BBN NPs的细胞毒性和体外靶向前列腺癌细胞的有效性。3、建立前列腺癌荷瘤裸鼠模型，探讨Gd2O3-FI-PEG-BBN活体肿瘤靶向成像的可能性和体内成像的特点。.材料和方法：1、采用非多元醇途径制备Gd2O3 NPs，PEG化后表面修饰FI及BBN获得Gd2O3-FI-PEG-BBN；通过透射电镜、及激光粒度分析仪、傅里叶变换红外光谱分析仪、热重力分析、MR成像对其理化及磁共振性质进行表征。2、使用高表达GRPr的PC-3细胞为研究对象，进行MTT实验、体外荧光及MR双模态细胞摄取实验。3、建立前列腺癌荷瘤裸鼠模型，进行肿瘤靶向MRI成像、活体荧光成像及肿瘤组织的荧光免疫病理学检测。.结果：1、成功构建Gd2O3-FI-PEG-BBN双模态纳米颗粒，平均粒径约50-60nm，水合粒径约为90nm。Gd2O3-FI-PEG-BBN的弛豫率为4.23 M-1s-1。2、MTT实验示与Gd2O3-FI-PEG-BBN孵育的PC-3细胞存活率在80%以上。体外细胞摄取实验示与Gd2O3-FI-PEG-BBN共孵育PC-3细胞内见绿色荧光；流式细胞术检测FI荧光强度为15326，对照组强度256；体外MR细胞摄取实验Gd2O3-FI-PEG-BBN共孵育的实验组信噪比（SNR）与其他组差异均有统计学意义（P<0.05）。3、体外成像，2h肿瘤区荧光及MR信号均达到峰值。Gd2O3-FI-PEG-BBN组2h强化率为27.95±5.86%，Gd2O3-FI-PEG 组2h的强化率为4.02±2.24%，差异有统计学意义（P＜0.05）。.结论：构建的Gd2O3-FI-PEG-BBN双模态纳米颗粒，理化性质优良，对PC-3细胞具有较好的荧光和MRI成像能力。