Evaluation of AAV FVIII variants in vivo. (A) Mean hFVIII:Ag levels ± SD in murine plasma derived from male wild-type C57Bl/6 mice (N = 6) at 6 weeks after tail vein injection of 2 × 1012 vg/kg of rAAV8-HLP-codop-hFVIII variants. (B) Sodium dodecyl sulfonate polyacrylamide gel electrophoresis/western blotting of murine plasma following gene transfer with rAAV-HLP-codop-BDD-hFVIII (BDD) and rAAV8-HLP-codop-hFVIII-V3 (V3) using a polyclonal rhesus anti-hFVIII antibody showing the heavy (∼90 kDa) and light (∼80 kDa) chains as well as a 170-kDa nonprocessed, primary translation product. Negative control (−ve) is naive mouse plasma and positive control consists of recombinant BDD hFVIII (ReFacto, first lane) diluted in murine plasma containing protease inhibitors. (C) Relationship between rAAV8-HLP-codop-hFVIII-V3 dose and hFVIII:Ag levels (mean ± SD) in murine plasma and (D) transgene copy number (mean ± SD) at 12 weeks following gene transfer.
