PB0338 - Novel therapeutic antibodies targeting the C-domains of Von Willebrand Factor drives anti-thrombotic efficacy with reduced bleeding risk

Background: Von Willebrand Factor (VWF) plays a critical role in haemostasis by capturing platelets to sites of injury under shear stress. Elevated VWF levels are both genetically and pharmacologically associated with a risk of thrombosis. Gain-of-function mutations have been identified in the C-domains of VWF that increase the risk of pathological thrombosis. We have previously demonstrated that disrupting the VWF cysteine-rich C-domain region by blocking free-thiols reduces platelet capture in a shear-dependent manner, suggesting that this region of VWF represents a novel potential pharmacological target for inhibiting shear-dependent thrombosis.

Aims: To develop shear-dependent antithrombotic therapeutic monoclonal antibody (mAb) targeting the C-domains of VWF.

Methods: mAbs were raised against a C1-C6 fragment of VWF and tested for functionality using in vitro flow assays and static binding assays and also in a murine model of thrombotic thrombocytopenic purpura (TTP).

Results: The mAbs bound specifically to epitopes in the C1-C6 domains but not to other VWF regions. Screening revealed shear-dependent high flow inhibition of VWF platelet capture for a subset of mAbs, including AP21. AP21 bound to VWF with high affinity (13.8pM) and ablated VWF- dependent platelet capture only at high shear rates. AP21 prevented the formation of rolling VWF-platelet aggregates and platelet capture in a microfluidic model designed to mimic an occluded vessel at the high shear site and reduced the formation of VWF-platelet strings on the surface of HUVECS. In an in vivo TTP model AP21 achieved complete reversal of the thrombocytopenic phenotype induced following challenge with ultra-large VWF, with correction of biochemical and histological markers of tissue damage. AP21 did not induce any bleeding in vivo.

Conclusion(s): These data further describe a pathological role of the VWF C-domains observed solely at high shear. These findings offer significant therapeutic potential for a novel, differentiated anti-thrombotic therapy with lower bleeding risk.