FIGURE 5.

Removal of 2118 mannose ending glycans does not modulate FVIII immunogenicity in FVIII-deficient mice, neither in double VWF/FVIII-deficient mice. (A) Half-life of wild type and N2118Q mutated FVIII. FVIII^HSQ and FVIII^2118Q were administered to FVIII-deficient mice and the residual FVIII:Ag was measured at different time points (n = 6 mice per time point) using a sandwich ELISA. The data is plotted as percentage of initial FVIII activity (measured 5 min after administration) versus time (mean ± SEM). Representative of two independent experiments. Inset. The slow and fast phases of FVIII clearance were determined by fitting the data to a two phase decay curve. The immunogenicity of FVIII^2118Q was evaluated in FVIII-deficient mice: (B,C), seven mice for both FVIII^HSQ and FVIII^2118Q group and VWF/FVIII double-deficient mice; (D,E) 8 and 6 mice for FVIII^HSQ and FVIII^2118Q group, respectively. One week after the fourth injection, blood samples were collected and anti-FVIII IgG titers (B,D) quantified using a purified mouse monoclonal anti-FVIII IgG (Mab6) as a standard. Results are expressed in arbitrary units (AU). Inhibitory titers toward FVIII (C,E) were measured by chromogenic assay. Statistical analysis was performed using the non-parametric two-tailed Mann-Whitney U test. ns, not significant.