Shunsuke Kageyama

Anti‐thrombotic effects and bleeding risk of AJvW‐2, a monoclonal antibody against human von Willebrand factor

1 A murine anti‐human vWF monoclonal antibody, AJvW‐2, was developed that inhibited the interaction between platelet glycoprotein Ib (GPIb) and von Willebrand factor (vWF) during the ristocetin‐ (IC50=0.7±0.1 μg ml−1) and botrocetin‐ (IC50=1.8±0.3 μg ml−1) induced aggregation of human platelets. 2 AJvW‐2 inhibited the high shear stress (10.8 N m−2) induced aggregation of human platelets dose‐dependently with an IC50=2.4±0.3 μg ml−1, but had no effect on low shear stress induced platelet aggregation (1.2 N m−2) up to 100 μg ml−1. 3 AJvW‐2 also inhibited the high shear stress (5.0 N m−2) induced adhesion of human platelets to collagen I with the same efficacy (IC50=2.4±0.3 μg ml−1), but had no effect at low shear conditions (1.5 N m−2). 4 AJvW‐2 inhibited the botrocetin‐induced aggregation of platelets from guinea‐pig, rat, rabbit, dog and pig at the same concentration range as human platelets; it likewise also inhibited the high shear stress induced aggregation and adhesion to collagen I of guinea‐pig platelets. 5 AJvW‐2 prevented arterial thrombus formation in guinea‐pigs at a dose of 100 μg kg−1 without prolonging the template bleeding time, whereas the GPIIb/IIIa antagonist lamifiban mediated inhibition of thrombosis at 1000 μg kg−1 was accompanied by a significant prolongation of the bleeding time. 6 These results suggest that AJvW‐2 is a potent inhibitor of the GPIb‐vWF interaction and a potential novel antithrombotic agent with lower bleeding risk than GPIIb/IIIa antagonists.

Anti-Human vWF Monoclonal Antibody, AJvW-2 Fab, Inhibits Repetitive Coronary Artery Thrombosis without Bleeding Time Prolongation in Dogs

The antithrombotic and antihaemostatic effects of the monoclonal antibody against human vWF (AJvW-2 Fab) were investigated in comparison with those of the monoclonal antibody against platelet GPIIb/IIIa (abciximab) in dogs. The ex vivo platelet aggregation and template bleeding time were measured before, 5, 90, 210 min and 24 h after injection of either AJvW-2 Fab or abciximab in anesthetized beagle dogs. Plasma concentration, vWF occupancy and plasma vWF antigen level were also measured by ELISA. In addition, the antithrombotic effect was evaluated in a canine model of repetitive coronary thrombosis (Folts model). AJvW-2 Fab significantly inhibited the ex vivo botrocetin-induced platelet aggregation at 0.18 mg/kg (53% plasma vWF occupancy) and also inhibited cyclic flow reductions (CFRs) at 0.06 mg/kg (31% occupancy). A significant prolongation of the bleeding time was observed at 1.8 mg/kg (95% occupancy), which was 30 times as high as the antithrombotic effective dose. Whereas, abciximab significantly inhibited both the ex vivo ADP-induced platelet aggregation and CFRs at 0.8 mg/kg, which was the minimally effective dose, also resulting in a significant prolongation of the bleeding time. These results suggest that blockade of the GPIb-vWF axis with AJvW-2 Fab leads to the inhibition of thrombus formation in the stenosed coronary arteries without less bleeding time prolongation than the GPIIb/IIIa blockade with abciximab.

Effect of a humanized monoclonal antibody to von Willebrand factor in a canine model of coronary arterial thrombosis

The purpose of this study was to investigate the antithrombotic effect and bleeding time prolongation of AJW200, a humanized monoclonal antibody to von Willebrand factor (vWF), in a canine model of coronary arterial thrombosis. AJW200 significantly inhibited cyclic flow reductions, as well as botrocetin-induced platelet aggregation, at 0.1 mg/kg. A significant prolongation of bleeding time was observed at 0.3-1 mg/kg. Approximately 50% occupancy of vWF (approximately 0.7 microg/ml AJW200 in plasma) and 80-100% occupancy (approximately 20 microg/ml AJW200 in plasma) were needed for the antithrombotic effect and the extensive prolongation of bleeding time, respectively. On the contrary, the minimal effective dose of abciximab (0.8 mg/kg) was associated with a significant prolongation of bleeding time. These results suggest that the pharmacological blockade of platelet glycoprotein (GP) Ib-vWF interaction with AJW200 results in a safer antithrombotic profile than platelet GPIIb/IIIa blockade with abciximab in dogs.

Anti-Human von Willebrand Factor Monoclonal Antibody AJvW-2 Prevents Thrombus Deposition and Neointima Formation After Balloon Injury in Guinea Pigs

Immediately after angioplasty, platelet adhesion to the injured arterial wall and subsequent release of various mitogens may contribute to neointima formation. The purpose of this study was to evaluate the inhibitory effect of AJvW-2, a monoclonal antibody against human von Willebrand factor (vWF), on neointima formation in a guinea pig model. The carotid artery was injured with a balloon catheter, and AJvW-2 was administered by a single bolus injection. AJvW-2 dose-dependently prevented neointima formation 14 days after injury. Significant inhibition was observed at 1.8 mg/kg, at which dose significant inhibition of platelet aggregation was achieved for 2 days. By elastic-Masson staining, organized thrombi were observed in the neointimal lesion on day 14. The thrombus area was significantly correlated with neointimal thickness. Furthermore, thrombus deposition, immunostained for vWF and fibrin(ogen), was observed on the media immediately after balloon injury. AJvW-2 significantly reduced the deposition of both adhesive proteins and reduced the incidence of organized thrombus formation, which might affect subsequent neointima formation. However, the proliferation of cultured smooth muscle cells was not affected by AJvW-2. These results suggest that AJvW-2 prevents neointima formation by inhibition of initial platelet-mediated thrombus formation rather than by direct inhibition of smooth muscle cell proliferation.