Ding Bi-Sen, Gottstein Claudia, Grunow Andrea, Kuo Alice, Ganguly Kumkum, Albelda Steven M, Cines Douglas B, Muzykantov Vladimir R
Department of Pharmacology, IFEM, 1 John Morgan Bldg, University of Pennsylvania, 3620 Hamilton Walk, Philadelphia, PA 19104-6068, USA.
Blood. 2005 Dec 15;106(13):4191-8. doi: 10.1182/blood-2005-05-2002. Epub 2005 Sep 6.
Means to prevent thrombus extension and local recurrence remain suboptimal, in part because of the limited effectiveness of existing thrombolytics. In theory, plasminogen activators could be used for this purpose if they could be anchored to the vascular lumen by targeting stably expressed, noninternalized determinants such as platelet-endothelial-cell adhesion molecule 1 (PECAM-1). We designed a recombinant molecule fusing low-molecular-weight single-chain prourokinase plasminogen activator (lmw-scuPA) with a single-chain variable fragment (scFv) of a PECAM-1 antibody to generate the prodrug scFv/lmw-scuPA. Cleavage by plasmin generated fibrinolytically active 2-chain lmw-uPA. This fusion protein (1) bound specifically to PECAM-1-expressing cells; (2) was rapidly cleared from blood after intravenous injection; (3) accumulated in the lungs of wild-type C57BL6/J, but not PECAM-1 null mice; and (4) lysed pulmonary emboli formed subsequently more effectively than lmw-scuPA, thereby providing support for the concept of thromboprophylaxis using recombinant scFv-fibrinolytic fusion proteins that target endothelium.
预防血栓扩展和局部复发的方法仍然不够理想，部分原因是现有溶栓药物的有效性有限。从理论上讲，如果纤溶酶原激活剂能够通过靶向稳定表达且不内化的决定簇（如血小板内皮细胞黏附分子1，PECAM-1）锚定在血管腔内，就可以用于此目的。我们设计了一种重组分子，将低分子量单链尿激酶原激活剂（lmw-scuPA）与PECAM-1抗体的单链可变片段（scFv）融合，以生成前药scFv/lmw-scuPA。纤溶酶切割产生具有纤溶活性的双链lmw-uPA。这种融合蛋白：（1）特异性结合表达PECAM-1的细胞；（2）静脉注射后能迅速从血液中清除；（3）在野生型C57BL6/J小鼠的肺中积累，但在PECAM-1基因敲除小鼠中不积累；（4）比lmw-scuPA更有效地溶解随后形成的肺栓塞，从而为使用靶向内皮细胞的重组scFv-纤溶融合蛋白进行血栓预防的概念提供了支持。
