Abstract Number: OC 22.2
Meeting: ISTH 2022 Congress
Theme: Platelet Disorders, von Willebrand Disease and Thrombotic Microangiopathies » von Willebrand Factor Biology
Background: Capture of platelets by von Willebrand factor (VWF) is an essential process carefully controlled by mechanosensitive elements within VWF. We recently demonstrated that, during activation, VWF undergoes a local conformational change in the autoinhibitory module (AIM) to expose the A1 domain for platelet binding. We further demonstrated that caplacizumab binds to the N-terminal AIM (NAIM), inhibits AIM unfolding, and prevents platelet binding to VWF. The C-terminal AIM (CAIM) is used by ristocetin to activate VWF.
Aims: We sought to discover nanobody binders to the CAIM and test if they could modulate VWF activity.
Methods: Using an immunized nanobody library displayed on yeast, we used flow sorting to selectively enrich nanobodies that would bind specifically to the CAIM and deplete those that would bind to either the NAIM or A1. A parallel plate flow chamber was utilized to evaluate VWF-dependent platelet adhesion under shear.
Results: Two monoclonal nanobodies, Nd4 and Nd6, were identified to bind human AIM-A1 protein, although with different affinities. Neither showed binding towards a VWF fragment lacking the CAIM, and neither could immunoblot VWF, thus localizing their epitope to a conformationally sensitive region that includes residues 1459-1472. Interestingly, Nd4 and Nd6 incompletely inhibited platelet adhesion in whole blood on the collagen surface at high shear rates, preserving VWF function at higher shear rates to a better degree than m(monomeric)Caplacizumab. In comparison, ARC1172, a well-known inhibitor of VWF, completely inhibited adhesion at all shear rates.
Conclusion(s): Inhibition of VWF activity can be achieved through binding to residues in the CAIM, including those that ristocetin uses to activate VWF. Inhibition of certain parts of either the AIM or A1 can lead to differential effects on VWF activation and platelet capture under force.
Figure depicting yeast display and isolation of nanobodies Nd4 and Nd6, their affinity towards VWF constructs, and inhibition of platelet adhesion.
To cite this abstract in AMA style:
Arce N, Li R. Inhibition of von Willebrand factor through stabilization of the ristocetin-binding site [abstract]. https://abstracts.isth.org/abstract/inhibition-of-von-willebrand-factor-through-stabilization-of-the-ristocetin-binding-site/. Accessed March 21, 2024.« Back to ISTH 2022 Congress
ISTH Congress Abstracts - https://abstracts.isth.org/abstract/inhibition-of-von-willebrand-factor-through-stabilization-of-the-ristocetin-binding-site/