Efficient adenoviral vector transduction and expression of functional human factor VIII in cultured primary human hepatocytes

Hemophilia A is a severe bleeding disorder caused by a deficiency in blood coagulation factor VIII (FVIII). Adenoviral vectors containing a potent hum an FVIII expression cassette encoding a truncated FVIII cDNA were developed that mediated sustained FVIII expression in normal and haemophiliac mice a nd complete phenotypic correction of the bleeding disorder in haemophiliac mice and dogs (Connelly and Kaleko, Haemophilia, 1998; 4: 380-8). Here, we evaluated two E1/E2a/E3-deleted adenoviral vectors encoding human FVIII, on e containing the full-length cDNA and the second containing a truncated cDN A lacking the B-domain. Viral vectors encoding the human full-length FVIII cDNA have not been described previously. Hepatocyte transduction was effici ent and dose dependent, ranging from 50% to 100%. High levels of functional FVIII were secreted from transduced cells at amounts up to 6000 mU(-1) 10( 6)cells(-1) 60 h. B-domain deleted FVIII was expressed at levels at least 8 -fold higher than the full-length FVIII protein, whereas FVIII RNA levels w ere similar with both vectors. These data provide the first demonstration o f FVIII adenoviral vector function in primary human cells and verify the po tential clinical utility of adenoviral vectors for the treatment of haemoph ilia A.