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VhH anti-thrombomodulin clone 1 inhibits TAFI activation and enhances fibrinolysis in human whole blood under flow.
van Moorsel, Marc V A; Poolen, Geke C; Koekman, Cornelis A; Verhoef, Sandra; de Maat, Steven; Barendrecht, Arjan; van Kleef, Nadine D; Meijers, Joost C M; Schiffelers, Raymond M; Maas, Coen; Urbanus, Rolf T.
Afiliación
  • van Moorsel MVA; Central Diagnostic Laboratory, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands.
  • Poolen GC; Central Diagnostic Laboratory, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands.
  • Koekman CA; Center for Benign Haematology, Thrombosis and Haemostasis, Van Creveldkliniek, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands.
  • Verhoef S; Central Diagnostic Laboratory, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands.
  • de Maat S; Central Diagnostic Laboratory, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands.
  • Barendrecht A; Central Diagnostic Laboratory, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands.
  • van Kleef ND; Central Diagnostic Laboratory, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands.
  • Meijers JCM; Central Diagnostic Laboratory, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands.
  • Schiffelers RM; Department of Experimental Vascular Medicine, Amsterdam Cardiovascular Sciences, Amsterdam UMC, University of Amsterdam, Amsterdam, The Netherlands.
  • Maas C; Department of Molecular Hematology, Sanquin Research, Amsterdam, The Netherlands.
  • Urbanus RT; Central Diagnostic Laboratory, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands.
J Thromb Haemost ; 20(5): 1213-1222, 2022 05.
Article en En | MEDLINE | ID: mdl-35170225
BACKGROUND: Thrombomodulin on endothelial cells can form a complex with thrombin. This complex has both anticoagulant properties, by activating protein C, and clot-protective properties, by activating thrombin-activatable fibrinolysis inhibitor (TAFI). Activated TAFI (TAFIa) inhibits plasmin-mediated fibrinolysis. OBJECTIVES: TAFIa inhibition is considered a potential antithrombotic strategy. So far, this goal has been pursued by developing compounds that directly inhibit TAFIa. In contrast, we here describe variable domain of heavy-chain-only antibody (VhH) clone 1 that inhibits TAFI activation by targeting human thrombomodulin. METHODS: Two llamas (Lama Glama) were immunized, and phage display was used to select VhH anti-thrombomodulin (TM) clone 1. Affinity was determined with surface plasmon resonance and binding to native TM was confirmed with flow cytometry. Clone 1 was functionally assessed by competition, clot lysis, and thrombin generation assays. Last, the effect of clone 1 on tPA-mediated fibrinolysis in human whole blood was investigated in a microfluidic fibrinolysis model. RESULTS: VhH anti-TM clone 1 bound recombinant TM with a binding affinity of 1.7 ± 0.4 nM and showed binding to native TM. Clone 1 competed with thrombin for binding to TM and attenuated TAFI activation in clot lysis assays and protein C activation in thrombin generation experiments. In a microfluidic fibrinolysis model, inhibition of TM with clone 1 fully prevented TAFI activation. DISCUSSION: We have developed VhH anti-TM clone 1, which inhibits TAFI activation and enhances tPA-mediated fibrinolysis under flow. Different from agents that directly target TAFIa, our strategy should preserve direct TAFI activation via thrombin.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Carboxipeptidasa B2 Tipo de estudio: Prognostic_studies Límite: Humans Idioma: En Revista: J Thromb Haemost Asunto de la revista: HEMATOLOGIA Año: 2022 Tipo del documento: Article País de afiliación: Países Bajos Pais de publicación: Reino Unido
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Carboxipeptidasa B2 Tipo de estudio: Prognostic_studies Límite: Humans Idioma: En Revista: J Thromb Haemost Asunto de la revista: HEMATOLOGIA Año: 2022 Tipo del documento: Article País de afiliación: Países Bajos Pais de publicación: Reino Unido