Labeling of human clots in vitro with an active-site mutant of t-PA.
Prompt detection of acute thrombosis and its response to treatment with thrombolytic agents generally require angiography. Scintigraphic approaches with labeled antibodies to or components of the coagulation and fibrinolytic systems have been disappointing because of prolonged circulating half-lives of tracers and relatively slow or limited binding to thrombi. Accordingly, we developed and characterized a thrombolytically inactive, active-site mutant (Ser-478----Thr) of tissue-type plasminogen activator (t-PA) designed to detect thrombi in vivo. Binding of iodine-125-(125I) labeled Ser----Thr t-PA to thrombi in vitro was time- and concentration-dependent, and specific judging from inhibition by pre-incubation with anti-t-PA IgG. Clearance of 125I-labeled mutant t-PA in rabbits was rapid and biexponential (alpha t1/2 = 1.9 +/- 0.4 min, beta t1/2 = 39.8 +/- 11.2 min). Thus, the amidolytically inactive mutant of t-PA designed binds rapidly and specifically to human thrombi in vitro and is cleared rapidly from the circulation in vivo--properties rendering it attractive as a potentially useful clot imaging agent.
Fry
E T
ET
Cardiovascular Division, Washington University School of Medicine, St. Louis, Missouri 63110.
Mack
D L
DL
Monge
J C
JC
Billadello
J J
JJ
Sobel
B E
BE
D006801:Humans; D066298:In Vitro Techniques; D007457:Iodine Radioisotopes; D007553:Isotope Labeling; D009154:Mutation; D011877:Radionuclide Imaging; D012694:Serine; D013912:Threonine; D013927:Thrombosis / Q000000981:diagnostic imaging*; D010959:Tissue Plasminogen Activator / Q000235:genetics*
D007457:Iodine Radioisotopes; D013912:Threonine; D012694:Serine; D010959:Tissue Plasminogen Activator