Nanobodies against factor XI apple 3 domain inhibit binding of factor IX and reveal a novel binding site for high molecular weight kininogen. (20th July 2022)

• Record Type: Journal Article
• Authors: Bar Barroeta, Awital; Marquart, J. Arnoud; Bakhtiari, Kamran; Meijer, Alexander B.; Urbanus, Rolf T.; Meijers, Joost C. M.
• Is Part Of: Journal of thrombosis and haemostasis. Volume 20:Number 11(2022)
• Page Start: 2538
• Page End: 2549
• Publication Date: 2022-07-20
• Subjects: anticoagulants -- factor XI -- high molecular weight kininogen -- hydrogen–deuterium exchange mass spectrometrynanobody; Thrombosis -- Periodicals; Hemostasis -- Periodicals; Blood coagulation disorders -- Periodicals; 616.1
• DOI: 10.1111/jth.15815
• Languages: English
• ISSNs: 1538-7933

Abstract

Abstract: Background: Factor XI (FXI) is a promising target for novel anticoagulants because it shows a strong relation to thromboembolic diseases, while fulfilling a mostly supportive role in hemostasis. Anticoagulants targeting FXI could therefore reduce the risk for thrombosis, without increasing the chance of bleeding side effects. Objectives: To generate nanobodies that can interfere with FXIa mediated activation of factor IX (FIX). Methods: Nanobodies were selected for binding to the apple 3 domain of FXI and their effects on FXI and coagulation were measured in purified protein systems as well as in plasma‐based coagulation assays. Additionally, the binding epitope of selected nanobodies was assessed by hydrogen–deuterium exchange mass spectrometry. Results: We have identified five nanobodies that inhibit FIX activation by FXI by competing with the FIX binding site on FXI. Interestingly, a sixth nanobody was found to target a different binding epitope in the apple 3 domain, resulting in competition with the FXI–high molecular weight kininogen (HK) interaction. Conclusions: We have characterized a nanobody targeting the FXI apple 3 domain that elucidates the binding orientation of HK on FXI. Moreover, we have produced five nanobodies that can inhibit the FXI–FIX interaction.