Copper-64 radiolabelling of the C2A domain of synaptotagmin I using a functionalised bis(thiosemicarbazone): A pre- and post-labelling comparison.
Copper-64 radiolabelling of the C2A domain of synaptotagmin I using a functionalised bis(thiosemicarbazone): A pre- and post-labelling comparison.
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使用功能化双（缩氨基硫脲）对突触结合蛋白 I 的 C2A 结构域进行铜 64 放射性标记：标记前和标记后的比较。
DOI：
10.1016/j.jinorgbio.2013.07.017
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发表时间：
2013
影响因子：
3.9
通讯作者：
Hueting R
中科院分区：
文献类型：
--
作者：
Hueting R
细胞凋亡和坏死的失调是许多疾病的核心，细胞死亡的无创成像是疾病分期或监测治疗进展的重要临床目标。大鼠synaptotagmin I的C2A结构域与细胞死亡过程中暴露的磷脂酰丝氨酸（PS）结合，对其赖氨酸残基的修饰已被证明会破坏PS的结合。位点特异性标记的99mtc (CO)3-C2AcH和68ga - c2ac已分别用于单光子发射计算机断层扫描（SPECT）和正电子发射断层扫描（PET）成像。由于64cu的半衰期较长，我们希望设计一种64cu标记的对应物。由于C2A中的钙结合位点可能干扰铜的结合，我们寻找了一种高亲和力、快速标记的螯合剂。我们合成了一种马来酰亚胺功能化的双（硫代氨基脲），H2ATSE/AMal，用于巯基功能化C2Ac的位点特异性铜64放射性标记。当通过配体-蛋白偶联物孵育（后标记方法）进行放射性标记时，对结果64cuatse /AMal-C2Ac的分析显示，C2Ac能够与螯合剂竞争放射性铜。相比之下，预标记的64cuatse /AMal-C2Ac偶联物在血清中显示出良好的稳定性，并在红细胞结合试验中保持靶亲和力。结果表明，由于蛋白质固有的铜结合特性，当铜是期望的放射性同位素时，对synaptotagmin I的C2Ac结构域采用预标记方法是首选的。
Dysregulation of apoptosis and necrosis is central to many diseases and non-invasive imaging of cell death is an important clinical objective to stage disease or to monitor treatment progress. The C2A domain of rat synaptotagmin I binds to phosphatidylserine (PS) exposed during cell death and modification to its lysine residues has been shown to disrupt PS binding. Site-specifically labelled99mTc(CO)3-C2AcH and68Ga-C2Ac have previously been investigated for single photon emission computed tomography (SPECT) and positron emission tomography (PET) imaging, respectively. We wished to design a64Cu-labelled counterpart due to the longer half-life of64Cu. Since the calcium binding sites in C2A may interfere with copper binding we sought a high affinity, fast labelling chelator. We synthesised a maleimide functionalised bis(thiosemicarbazone), H2ATSE/AMal, for the site-specific copper-64 radiolabelling of thiol-functionalised C2Ac. When radiolabelling was performed by incubation of the ligand–protein conjugate (post-labelling approach), analysis of the resultant64CuATSE/AMal-C2Ac revealed that the C2Ac was able to compete for radiocopper with the chelator. In contrast, the pre-labelled64CuATSE/AMal-C2Ac conjugate revealed good stability in serum and maintained target affinity in a red blood cell binding assay. The results suggest that due to the intrinsic copper binding properties of the protein, a pre-labelling approach is preferred for the C2Ac domain of synaptotagmin I when copper is the desired radioisotope.
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