Engineering rFVIIa-loaded platelets; a novel approach to treating acute bleeding

Abstract

Haemorrhage remains a leading cause of mortality around the world, resulting from both trauma and surgery. Current treatments for acute haemorrhage include blood products such as fresh frozen plasma, as well as platelet transfusion and recombinant clotting factors such as rFVIIa. However, the use of recombinant clotting factors is limited due to cost and adverse side effects resulting from increased thrombosis, such as increased rate of myocardial infarction and cerebrovascular accidents. Platelets are small anucleate cells that exist in very large quantities in our blood and, along with cross-linked fibrin from the coagulation cascade, are involved in forming a haemostatic plug upon exposure to damaged endothelium in a wound. They are metabolically active and undergo a process of platelet activation when stimulated by pro-thrombotic agonists such as thrombin resulting from the coagulation cascade. This then triggers a process whereby the contents of their granules are released locally to facilitate coagulation. This thesis explores the possibility of loading these platelet granules with recombinant clotting factors, in this case rFVIIa which has already been shown in clinical trials to result in a significant decrease in mortality from acute haemorrhage. The targeted delivery of this drug is explored, both through endocytosis and genetic engineering of megakaryocytes differentiated in vitro from induced pluripotent stem cells (iPSCs). These are evaluated with in vitro assays to model the process of clot formation, as well as an in vivo model of haemostasis to compare their efficacy to conventional treatments. Overall, this serves as a proof of concept of the engineering of platelet granules as a novel drug delivery system.