Non-hemostatic effects of recombinant FVIII and emicizumab on endothelial cell function in hemophilia A

Hemophilia A (HA) is a rare x-linked bleeding disorder caused by the absence or dysfunction of coagulation factor VIII (FVIII). Beyond its role in coagulation, FVIII contributes to endothelial cell biology, but the relevance of these extra-coagulative effects remains unclear. Treatments include prophylaxis with FVIII concentrates or non-factor therapies such as emicizumab (Hemlibra). Recombinant FVIII (rFVIII) products differ in molecular design, including the host cell line used for production and modifications introduced to extend half-life, which may influence their biological activity. We assessed the impact of rFVIII products with molecular modifications to extend half-life (Elocta [Fc-fusion], Adynovate and Jivi [PEGylated], Altuviiio [Fc-fusion, vWF D'D3 domain, and XTEN polypeptides]) and those without modifications (Nuwiq, Advate), as well as emicizumab (a bispecific antibody), on blood outgrowth endothelial cells (BOECs) derived from HA patients, whose cellular functions were previously shown to be defective. All rFVIII products significantly improved HA-BOECs proliferation, migration, tubulogenesis, and permeability except Elocta, which consistently exhibited reduced activity and reduced binding to BOECs compared with Nuwiq. In a mouse model of severe HA, both Nuwiq and Altuviiio restored vascular permeability, with Nuwiq showing the strongest effect whereas Elocta provided partial correction. Similarly, Nuwiq promoted greater in vivo angiogenesis than Elocta, while Altuviiio had an intermediate effect. Emicizumab had no measurable effect either in vitro or in vivo. These data underscore the emerging importance of FVIII in HA ECs biology and vascular homeostasis, with potential long-term clinical implications for biomarker development and the optimization of HA treatment beyond conventional bleeding control.