Dougan Hayes, Weitz Jeffrey I, Stafford Alan R, Gillespie Kris D, Klement Petr, Hobbs John B, Lyster Donald M
TRIUMF, Vancouver, BC, V6T 2A3, Canada.
Nucl Med Biol. 2003 Jan;30(1):61-72. doi: 10.1016/s0969-8051(02)00378-5.
Two DNA aptamers directed against two separate exosites on human alpha-thrombin were evaluated for thrombus-imaging potential. Aptamer ODN 1 is directed to the thrombin substrate binding site (exosite 1). Our finding that ODN 1 competes with fibrin for binding to exosite 1 on thrombin suggests that ODN 1 will not be useful for thrombus imaging. Aptamer ODN 2 is directed against the thrombin heparin binding site (exosite 2). ODN 2 bound to model thrombi that were formed either by clotting purified fibrinogen with thrombin, or by recalcifying citrated plasma. As the thrombin content of thrombi was increased the rate of ODN 2 uptake into preformed thrombi increased, whereas the rate of release of ODN 2 out of preformed thrombi decreased. This in vitro data suggested that ODN 2 might be useful for thrombus imaging because it can bind to exosite 2 on fibrin-bound thrombin. However, in a rabbit jugular vein model using thrombus supplemented with human thrombin, ODN 2 uptake was equal to the ovalbumin control, and did not reflect thrombin content. While the in vitro results with ODN 2 were consistent with thrombus imaging, the rapid clearance of ODN 2 from circulation, combined with slow mass transfer in the clot, seem to work against in vivo thrombin-dependent imaging or washout analysis.
评估了两种针对人α-凝血酶上两个不同外位点的DNA适配体的血栓成像潜力。适配体ODN 1靶向凝血酶底物结合位点（外位点1）。我们发现ODN 1与纤维蛋白竞争结合凝血酶上的外位点1，这表明ODN 1对血栓成像无用。适配体ODN 2靶向凝血酶肝素结合位点（外位点2）。ODN 2能与通过用凝血酶使纯化的纤维蛋白原凝血或使枸橼酸盐血浆再钙化形成的模型血栓结合。随着血栓中凝血酶含量的增加，ODN 2进入预先形成血栓的摄取速率增加，而ODN 2从预先形成血栓中的释放速率降低。这些体外数据表明ODN 2可能对血栓成像有用，因为它可以结合纤维蛋白结合的凝血酶上的外位点2。然而，在使用补充有人凝血酶的血栓的兔颈静脉模型中，ODN 2的摄取量与卵清蛋白对照相等，且未反映凝血酶含量。虽然ODN 2的体外结果与血栓成像一致，但ODN 2从循环中的快速清除，加上在凝块中的缓慢质量传递，似乎不利于体内凝血酶依赖性成像或洗脱分析。
