Defining the Optimal FVIII Transgene for Placental Cell-Based Gene Therapy to Treat Hemophilia A

Abstract The delivery of FVIII through gene- and/or cellular platforms has emerged as a promising hemophilia A treatment. Herein, we investigated the suitability of human placental cells (PLC) as delivery vehicles for FVIII, and determined an optimal FVIII transgene to produce/secrete therapeutic FVIII levels from these cells. Using 3 PLC cell banks we demonstrated PLC constitutively secreted low levels of FVIII, suggesting their suitability as a transgenic FVIII production platform. Furthermore, PLC significantly increased FVIII secretion after transduction with a lentiviral vector (LV) encoding a myeloid codon-optimized bioengineered FVIII containing high expression elements from porcine FVIII. Importantly, transduced-PLC didn’t upregulate cellular stress or innate immunity molecules, demonstrating that after transduction and FVIII production/secretion, PLC retained low immunogenicity and cell stress. When LV encoding 5 different bioengineered FVIII-transgenes were compared for transduction efficiency, FVIII production, and secretion, data showed that PLC transduced with LV encoding hybrid human/porcine FVIII transgenes secreted substantially higher levels of FVIII than LV encoding B domain-deleted human FVIII. In addition, data showed that in PLC, myeloid codon optimization is needed to increase FVIII secretion to therapeutic levels. These studies have identified an optimal combination of FVIII transgene and cell source to achieve clinically meaningful levels of secreted FVIII.