Labeling of adenovirus particles with PARACEST agents
Labeling of adenovirus particles with PARACEST agents
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DOI：
10.1021/bc7002605
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发表时间：
2008-03-01
影响因子：
4.7
通讯作者：
Sherry, A. Dean
中科院分区：
文献类型：
--
作者：
Vasalatiy, Olga;Gerard, Robert D.;Sherry, A. Dean
用两种不同的双功能配体标记重组腺病毒5型颗粒（AdCMVLuc），这些配体能够与顺磁性镧系离子形成稳定的配合物。每个腺病毒颗粒的共价配体数量在630到1960之间变化，这取决于双功能配体（NHS酯与异硫氰化物）的化学反应活性、过量配体的添加量和反应时间。每个标记的腺病毒衍生物的生物活性，通过病毒感染细胞和表达荧光素酶的能力来测量，被证明高度依赖于共价附着配体的数量。这表明，某些氨基，可能在腺病毒纤维蛋白的表面，细胞结合已知发生，是至关重要的病毒附着和感染。将Lu-177（3+）添加到化学修饰的病毒中，与对照病毒相比，显示出大量的Lu-177（3+）与病毒颗粒的非特异性结合，而添加过量的DTPA无法将其隔离。因此，有必要实施预先标记策略，将预先形成的镧系配体螯合物偶联到腺病毒颗粒上。利用预形成的Tm3+-L2，大量具有化学交换饱和转移（CEST）性质的螯合物附着在AdCMVLuc的表面残基上，而不会与病毒颗粒上其他地方的金属离子发生非特异性结合。这些缀合物作为PARACEST显像剂的潜力使用非共振华尔兹序列进行CEST激活测试。与对照组相比，观察到散装水信号强度降低了12%。这表明用paracest型显像剂标记的病毒颗粒可以潜在地作为分子成像的靶向剂。
Recombinant adenovirus type 5 particles (AdCMVLuc) were labeled with two different bifunctional ligands capable of forming stable complexes with paramagnetic lanthanide ions. The number of covalently attached ligands varied between 630 and 1960 per adenovirus particle depending upon the chemical reactivity of the bifunctional ligand (NHS ester versus isothiocyanide), the amount of excess ligand added, and the reaction time. The bioactivity of each labeled adenovirus derivative, as measured by the ability of the virus to infect cells and express luciferase, was shown to be highly dependent upon the number of covalently attached ligands. This indicates that certain amino groups, likely on the surface of the adenovirus fiber protein where cell binding is known to occur, are critical for viral attachment and infection. Addition of Lu-177(3+) to chemically modified versus control viruses demonstrated a significant amount of nonspecific binding of Lu-177(3+) to the virus particles that could not be sequestered by addition of excess DTPA. Thus, it became necessary to implement a prelabeling strategy for conjugation of preformed lanthanide ligand chelates to adenovirus particles. Using preformed Tm3+-L2, a large number of chelates having chemical exchange saturation transfer (CEST) properties were attached to the surface residues of AdCMVLuc without nonspecific binding of metal ions elsewhere on the virus particle. The potential of such conjugates to act as PARACEST imaging agents was tested using an on-resonance WALTZ sequence for CEST activation. A 12% decrease in bulk water signal intensity was observed relative to controls. This demonstrates that viral particles labeled with PARACEST-type imaging agents can potentially serve as targeted agents for molecular imaging.