Investigation into the mechanisms of LRP1-mediated factor VIII clearance



 DESCRIPTION (provided by applicant): Hemophilia A is a well-known inherited blood disorder that affects 1 in 5,000 males caused by a deficiency in factor VIII (fVIII), a critical blood coagulation protein. In addition to the high economic cost of Hemophilia A, this disease has a significant impact on overall health. Over time recurrent bleeds result in hemophilic arthropathy and permanent joint damage. Further, each time a hemophiliac experiences a bleeding episode, blood pressure deviates from normal and many severe hemophiliacs develop hypertension later in life. Current treatment for hemophiliacs is replacement therapy by routine injection of recombinant fVIII to arrest bleeding episodes. Replacement therapy involves lifetime injection of fVIII in response to a bleed or approximately three times a week to prevent bleeds from occurring. Although effective, current Hemophilia A treatments are difficult to maintain for many patients due to cost and frequency of injections, leading to the need for a better prophylactic treatment. In the circulation, fVIII binds tightly to a protein known as von Willebrand Factor (vWF), which protects it from rapid clearance by preventing fVIII from binding to hepatic receptors. Studies have shown that the hepatic clearance of fVIII is mediated by the low-density lipoprotein receptor-related protein (LRP1), with some contribution by the LDL receptor. Upon injury within the vasculature, fVIII is activated by thrombin, resulting in its dissociation from vF. Activated fVIII binds to enzymatically active factor IXa to trigger the final steps of the coagulaton cascade. While it is known how fVIII functions and what receptor clears it from circulation, the molecular details of its clearance are not well understood. It is particularly of interest to revea molecular mechanisms of fVIII catabolism as a more stable fVIII that can stay in circulation for extended time would improve efficiency of replacement therapy. The objectives for the current project are to define if activation of fVIII is required for efficient LRP1-mediated hepatic clearance and identify mechanisms to delay fVIII clearance. In this project, the mechanisms of fVIII clearance will be elucidated with a combination of in vitro techniques, such as binding assays, and in vivo clearance assays in mice. Successful completion of this project will provide the fVIII research community with invaluable information regarding how fVIII is cleared from the circulation. Further, fVIII clearance from circulation can be delayed by mapping bindings sites to LRP1 on fVIII to develop a recombinant fVIII that will have a longer half-life in circulation and/o soluble LRP1 fragments will be designed to block fVIII binding to LRP1. Either of these strategies may represent an improved strategy for hemophilia therapy.