OC 77.4 - Pharmacological targeting of GPV in a novel humanized mouse model to modulate thrombus formation in vivo

Background: Platelet glycoprotein (GP) V is part of the GPIb-V-IX complex, which mediates the initial platelet recruitment to sites of vascular injury. Despite its abundant expression, the physiological function of GPV has remained elusive. We have recently shown that GPV spatio-temporally controls fibrin formation and thereby provides a platelet-based mechanism that locally limits excessive fibrin formation and thrombus growth (Beck et al., ID: 1201886). Thus, pharmacological modulation of GPV might provide a therapeutical strategy to treat both thrombotic and hemorrhagic disease states.

Aims: To generate a model system to pharmacologically modulate human GPV in vitro and in vivo.

Methods: The extracellular domain of mouse GPV was replaced by the human sequence and the resulting hGP5KI mice, as well as their platelets, were characterized. Monoclonal antibodies (mAbs) against human GPV (LUM/B, LUM/C, LUM3-5) were generated and characterized.

Results: hGP5KI mice displayed normal expression levels of hGPV and all other tested platelet membrane glycoproteins. Also, their activation and aggregation responses to different agonists were unaltered compared to controls. Thrombin-mediated cleavage of hGPV and protease-activated receptor (PAR)-dependent activation was indistinguishable from control platelets, and no hyperreactivity to thrombin, as seen with Gp5-/- platelets, was observed. Screening of newly generated anti-hGPV mAbs identified LUM/C to block the collagen-binding site on hGPV and LUM/B to interfere with its thrombin-induced cleavage, while all other mAbs had no effect on platelet function. In vivo, none of the LUM antibodies affected platelet counts in hGP5KI mice. However, LUM/B significantly accelerated fibrin formation and thrombosis in vivo, confirming the proposed role of GPV in these processes.

Conclusion(s): Together, these results demonstrate that hGP5KI mice are a valuable model system for studying the impact of GPV on thrombus formation in a translational approach. Furthermore, the LUM/B antibody very consistently modulates fibrin and thrombus formation by interfering with thrombin-dependent cleavage of hGPV.