文献：Dimeric Her2-specifc afbody mediated cisplatin-loaded nanoparticles for tumor enhanced chemo-radiotherapy

文献链接：https://jnanobiotechnology.biomedcentral.com/articles/10.1186/s12951-021-00885-6

作者：Haijun Wang, Dianlong Jia, Dandan Yuan, Xiaolei Yin , Fengjiao Yuan , Feifei Wang , Wenna Shi , Hui Li , LiMin Zhu, Qing Fan

相关产品：ZHer2 with 6-FAM

原文摘要：

Background: Solid tumor hypoxic conditions prevent the generation of reactive oxygen species (ROS) and the formation of DNA double-strand breaks (DSBs) induced by ionizing radiation, which ultimately contributes to radio therapy (RT) resistance. Recently, there have been signifcant technical advances in nanomedicine to reduce hypoxia by facilitating in situ O2 production, which in turn serves as a “radiosensitizer” to increase the sensitivity of tumor cells to ionizing radiation. However, of-target damage to the tumor-surrounding healthy tissue by high-energy radiation is often unavoidable, and tumor cells that are further away from the focal point of ionizing radiation may avoid damage. Therefore, there is an urgent need to develop an intelligent targeted nanoplatform to enable precise enhanced RTinduced DNA damage and combined therapy.

Results: Human epidermal growth factor receptor 2 (Her2)-specifc dimeric afbody (ZHer2) mediated cisplatinloaded mesoporous polydopamine/MnO2/polydopamine nanoparticles (Pt@mPDA/MnO2/PDA-ZHer2 NPs) for MRI and enhanced chemo-radiotherapy of Her2-positive ovarian tumors is reported. These NPs are biodegradable under a simulated tumor microenvironment, resulting in accelerated cisplatin release, as well as localized production of O2. ZHer2, produced using the E. coli expression system, endowed NPs with Her2-dependent binding ability in Her2positive SKOV-3 cells. An in vivo MRI revealed obvious T1 contrast enhancement at the tumor site. Moreover, these NPs achieved efcient tumor homing and penetration via the efcient internalization and penetrability of ZHer2. These NPs exhibited excellent inhibition of tumor growth with X-ray irradiation. An immunofuorescence assay showed that these NPs signifcantly reduced the expression of HIF-1α and improved ROS levels, resulting in radiosensitization.

Conclusions: The nanocarriers described in the present study integrated Her2 targeting, diagnosis and RT sensitiza tion into a single platform, thus providing a novel approach for translational tumor theranostics.

ZHer2是一种抗HER2亲和体分子，具有与HER2受体高度特异性和亲和力的特性。其化学结构中，C端含有一个带有巯基的半胱氨酸，N端和C端可能分别含有和/或不含有6个氨基酸的HIS标签，这使得ZHer2能够与目标受体紧密结合。人表皮生长因子受体2（Her2）特异性二聚体（ZHer2）介导的顺铂负载介孔多孔聚多巴胺/MnO2/聚多巴胺纳米颗粒（Pt@mPDA/MnO2/PDA-ZHer2 NPs）用于MRI和Her2 tumor的增强效果。

图为：Pt@mPDA/MnO2/PDA-ZHer2 NPs的制备和体内mri引导下增强Chemotherapy -放疗的实验工作

ZHer2在Pt@mPDA/mnO2/PDAZHer2NPs制备和表征中的应用：

由于聚合可能提高游离的稳定性和亲和力，制备了一种新的与Her2结合的二聚体，Te ZHer2 afbody用大肠杆菌表达系统表达，并用Ni-NTA免疫层析纯化。与诱导前从细胞中分离出的蛋白相比，IPTG诱导了一个蛋白。细胞破坏后，SDS-PAGE分析可溶性中的总蛋白组分显示，重组蛋白主要以可溶性蛋白表达。与Ni-NTA树脂结合后，通过部分中重组蛋白的数量减少，表明高的纯化性。Te纯化的ZHer2蛋白在SDS-PAGE凝胶上显示为单条蛋白带，根据电泳带的扫描密度测定法计算纯度。从培养的大肠杆菌细胞中获得蛋白质。其目标体外的二聚体形式的性质通常高于单体形式。因此，在c端添加一个半胱氨酸残基后，ZHer2游离体二聚。与预期的一样，ZHer2在自然条件下的ZHer2的分子量大约是还原条件下的ZHer2的两倍，表明ZHer2在自然条件下形成二硫键和二聚体。用荧光树脂剂6-羧基荧光树脂（6-FAM）标记ZHer2后，观察到分子量略有增加。该标签允许视觉评估FAMZHer2和her2阳性细胞之间的结合能力。最后，ZHer2 afbody通过Pichaer加法/Schif与Pt@ mPDA/MnO2/PDA NPs结合在弱碱性条件下，将氨基与邻苯二酚的氧化醌形式结合的碱反应。ZHer2游离偶联后，NPs的平均水动力尺寸增加。两种NPs均表现出较高的胶体稳定性，并可在PBS中分散，且在一天内不聚集。mPDA/ MnO2/PDA-ZHer2 NPs的Te元素作图像显示C、N、O、S、Mn和Pt的均匀分布，进一步限制了顺铂的成功加载和ZHer2的偶联。

图为：直接注射Pt-mPDA/mno2/PDANPs前后SKOV-3荷瘤小鼠的T1-MR图像

结果：her2靶向顺铂负载mPDA/MnO2/PDA NPs用于缺氧tumor的Mr成像。Tese NPs在模拟tumor微环境下可生物降解，通过触发过氧化氢的分解，可以加速顺铂的释放和o2的产生。细胞摄取实验表明，ZHer2赋予NPs与Her2结合的能力，实现NPs向Her2阳性SKOV-3细胞的内化。