THROMBOSIS AND HEMOSTASIS Platelet-delivered ADAMTS 13 inhibits arterial thrombosis and prevents thrombotic thrombocytopenic purpura in murine models

ADAMTS13, a plasma metalloprotease that cleaves von Willebrand factor (VWF), is produced primarily in hepatic stellate cells as well as in other cells, including endothelial cells, megakaryocytes, and platelets. Plasma ADAMTS13 concentration ranges from 0.5 to 1.5 mg/L. Deficiency of plasma ADAMTS13 activity resulting from hereditary or acquired etiologies results in an accumulation of ultralarge (UL) VWF on endothelial cell surface and in plasma, leading to the formation of disseminated thrombi in small arterioles and capillaries. This is the characteristic feature of TTP. Patients with TTP often present with severe thrombocytopenia and microangiopathic hemolytic anemiawith various degrees of end organ damage. TTP is seen in neonates, young children, and adults. The annual incidence rate is 3 to 10 per million populations. Hereditary TTP is caused by mutations in the ADAMTS13 gene, but acquired TTP is caused by autoantibodies against ADAMTS13. Without treatment, the mortality rate is ;85% to 90%. Plasma infusion suffices in treating hereditary TTP, but plasma exchange is often required to achieve remission for acquired TTP with inhibitors. Plasma exchange removes immunoglobulin G (IgG) autoantibodies against ADAMTS13, ULVWF multimers, and perhaps inflammatory cytokines while replenishing ADAMTS13 enzyme. In patients with high-titer inhibitors, the infused ADAMTS13 from plasma is often not sufficient to override autoantibodies and correct the underlying ADAMTS13 deficiency. In this case, a prolonged course of plasma exchange plus adjunctive immunosuppressive therapies, including cyclophosphamide, cyclosporine, or rituximab, may be required. Severe ADAMTS13 deficiency and persistence of autoantibodies correlate with increased mortality rate and/or relapses. Our groups and others have shown that antiADAMTS13 IgGs bind to multiple domains on ADAMTS13, but their inhibition appears to be mediated most commonly through the binding to the spacer domain. Conservative modifications in the antibody-binding epitopes have generated 2 ADAMTS13 variants that are more resistant to inhibition by autoantibodies, which may be developed as a novel therapeutic for acquired TTP resulting from inhibitors. Here, we characterize transgenic mice that express human ADAMTS13 in platelets and report its efficacy in murine models of arterial thrombosis and TTP. A similar strategy has been used to express factor VIII to treat hemophilia A with inhibitors, with limited success. Our results demonstrate that platelet-delivered rADAMTS13 is efficacious for inhibiting arterial thrombosis after vascular injury and prevents the onset of Shigatoxin-2 (Stx-2)or recombinant murine VWF (rmVWF)-induced TTP in the absence or the presence of inhibitors. The findings provide a proof of concept that platelet-derived ADAMTS13 may be explored as a novel