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Title: Discovery of novel mechanisms and molecular targets for the inhibition of activated thrombin activatable fibrinolysis inhibitor
Authors: Hillmayer, Kerstin
Vancraenenbroeck, Renée
De Maeyer, Marc
Compernolle, Griet
Declerck, Paul
Gils, Ann # ×
Issue Date: Nov-2008
Series Title: Journal of Thrombosis and Haemostasis vol:6 issue:11 pages:1892-1899
Abstract: Background: Thrombin activatable fibrinolysis inhibitor (TAFI) is an important regulator of fibrinolysis and an attractive target to develop profibrinolytic drugs. Objective: To analyze the (inhibitory) properties of five monoclonal antibodies (MA) directed towards rat TAFI (i.e. MA-RT13B2, MA-RT30D8, MA-RT36A3F5, MA-RT36B2 and MA-RT82F12). Methods and results: Direct interference of the MA with rat TAFIa activity was assayed using a chromogenic activity assay. This revealed a reduction of 79+/-1%, 54+/-4% and 19+/-2% activity in the presence of a 16-fold molar excess of MA-RT13B2, MA-RT36A3F5 and MA-RT82F12, respectively whereas MA-RT30D8 and MA-RT36B2 had no direct inhibitory effect. Additionally, MA-RT13B2 and MA-RT36A3F5 reduced rat TAFIa half-life with 56+/-2% and 61+/-3%. T-PA mediated in vitro clot lysis was determined using rat plasma. Compared to PTCI, MA-RT13B2, MA-RT30D8, MA-RT36A3F5 and MA-RT82F12 reduced clot lysis times with 86+/-14%, 100+/-5%, 100+/-10% and 100+/-11%, respectively. During epitope mapping, Arg(227) and Ser(251) were identified as major residues interacting with MA-RT13B2. Arg(188) and His(192) contribute to the interaction with MA-RT36A3F5. Arg(227,) Ser(249), Ser(251) and Tyr(260) are involved in the binding of MA-RT30D8 and MA-RT82F12 with rat TAFI(a). Following mechanisms of inhibition have been deduced: MA-RT13B2 and MA-RT36A3F5 have a destabilizing effect on rat TAFIa whereas MA-RT30D8 and MA-RT82F12 partially block the access to the active site of TAFIa or interact with the binding of TAFIa to the blood clot. Conclusions: The described inhibitory MA towards rat TAFIa will facilitate TAFI research in murine models. Additionally, we reveal novel molecular targets for the direct inhibition of TAFIa through different mechanisms.
URI: 
ISSN: 1538-7933
Publication status: published
KU Leuven publication type: IT
Appears in Collections:Biochemistry, Molecular and Structural Biology Section
Laboratory for Pharmaceutical Biology (-)
× corresponding author
# (joint) last author

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