Biological properties of hybrid plasminogen activators.
A number of hybrid plasminogen activator genes were constructed from the t-PA and u-PA cDNAs and expressed using a bovine papilloma virus vector and mouse C-127 cells. Hybrid A was constructed by replacing the finger (F) and EGF domains of t-PA with the EGF and Ku domains of u-PA, while hybrids B and C had an extra Ku inserted before or after the double kringle (K1-K2) region of t-PA respectively. While all the hybrids showed comparable enzymatic activities towards a small substrate (S-2288), they had different activities in binding to fibrin clots as well in the fibrin-dependent plasminogen activation, the order of activities being: t-PA greater than or equal to hybrid B greater than hybrid C greater than hybrid A. Carbohydrate analysis showed that while hybrid C, like rt-PA, had at least one high-mannose type sugar chain (probably at residue 117 in K1), the other hybrids had only complex-type carbohydrates suggesting that domain interaction in t-PA might influence glycan processing. Pharmacokinetic studies in dog showed that hybrid B had a significantly longer plasma half-life than rt-PA. Thrombolytic efficacies of hybrid B and rt-PA were compared in dog model using an artificially induced coronary thrombus. Complete thrombolysis was achieved with 18 mg and 50 mg dosages for hybrid B and rt-PA respectively. These data show the superior pharmacokinetic and thrombolytic properties of hybrid B compared to rt-PA.
Hung
P P
PP
Wyeth-Ayerst Research, Philadelphia, PA 19101.
Wilhelm
J
J
Kalyan
N K
NK
Cheng
S M
SM
James
H L
HL
Nachowiak
D
D
Weinheimer
C J
CJ
Sobel
B E
BE
Bergmann
S R
SR
Lee
S G
SG
D000818:Animals; D002460:Cell Line; D004792:Enzyme Precursors / Q000378:metabolism; D005822:Genetic Vectors; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D051379:Mice; D010958:Plasminogen / Q000378:metabolism; D010960:Plasminogen Activators / Q000235:genetics / Q000378:metabolism*; D055503:Protein Multimerization; D011994:Recombinant Proteins / Q000378:metabolism; D010959:Tissue Plasminogen Activator / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D014568:Urokinase-Type Plasminogen Activator / Q000235:genetics / Q000378:metabolism*
D004792:Enzyme Precursors; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D010958:Plasminogen; D010960:Plasminogen Activators; D010959:Tissue Plasminogen Activator; D014568:Urokinase-Type Plasminogen Activator