Naked plasmid transfer of hFVIII plasmids into hemophilia A mice with immunosuppressive agents. hFVIII levels and inhibitory activity were assessed in hemophilia A mice after treatment with pBS-HCRHPI-hFVIIIA beginning on day 1. No transient immunosuppression (A,B), CSA (C,D), RAP (E,F), and MMF (G,H). Fifty μg plasmid in 2 mL saline solution was injected into the tail vein of mice (n = 4) in 5 to 8 seconds. Immunosuppressive drugs were administered intraperitoneally for 14 days starting from the day of the plasmid injection. Mice were then bled at regular intervals. Circulating hFVIII activities in plasma were evaluated by a modified clotting assay (A,C,E,G) and confirmed by a COATEST assay. Inhibitory antibody titers were evaluated by Bethesda assay and are expressed as BU/mL (B,D,F,H). For transient immunosuppression with combined agents, CSA and MMF (I,J) and RAP and MMF (K,L) are shown. Combined immunosuppressive drugs were given at the same schedule as the respective single agent (Table 1). Each symbol represents an individual mouse's results from both assays.