Next generation factor VIIa with enhanced half-life

Background: Recombinant coagulation factor VIIa (rFVIIa) is used to control bleeding episodes in hemophilia patients with inhibitors. However, its therapeutic efficacy is hampered by an extremely short in vivo plasma half-life. Although genetic fusion of rFVIIa to wild-type human albumin prolongs its half-life, it is still very short. As the neonatal Fc receptor (FcRn) is a a key regulator of albumin homeostasis, engineered albumin with improved FcRn binding properties may extends the half-life beyond that of wild-type albumin. Aims: To develop the next-generation rFVIIa with superior plasma half-life by taking advantage of a novel engineered human albumin variant with tailored FcRn binding. Methods: Wild-type and the engineered (QMP) rFVIIa albumin fusions were expressed in HEK293E cells, purified and characterized in vitro through PT-based and thrombin generation assays, surface plasmon resonance and ELISA, followed by studies in state-of-the art mouse models. Results: The designed rFVIIa-QMP fusion efficiently restored coagulation in FVII-depleted plasma and, most importantly, showed a by-passing activity similar to that of commercial rFVIIa in plasma from hemophilia A patients with high-titer inhibitors. In vitro, rFVIIa-QMP bound human FcRn much more strongly compared to the wild-type fusion. After injection in hemophilia B mice (expressing the mouse FcRn), the by-passing activity of rFVIIa-QMP in plasma was still detectable after 48-73 hours, whereas the activity in plasma from mice given rFVIIa was undetectable at 3-6 hours. Strikingly, in human FcRn transgenic mice, rFVIIa-QMP showed a half-life of 2.9 days, compared to only 0.8 days for the wild-type fusion. Conclusion: Fusion of engineered albumin to rFVIIa preserved the by-passing activity both in vitro and in vivo, and extended the plasma half-life by impressively 4-fold compared with the wild-type fusion. Thus, the novel engineered albumin should be an attractive carrier for half-life extension of other coagulation proteins