Native-like aggregates of factor VIII are immunogenic in von Willebrand factor deficient and hemophilia a mice

The administration of recombinant factor VIII (FVIII) is the first-line therapy for hemophilia A (HA), but 25%-35% of patients develop an inhibitory antibody response. In general, the presence of aggregates contributes to unwanted immunogenic responses against therapeutic proteins. FVIII has been shown to form both native-like and nonnative aggregates. Previously, we showed that nonnative aggregates of FVIII are less immunogenic than the native protein. Here, we investigated the effect of native-like aggregates of FVIII on immunogenicity in HA and von Willebrand factor knockout (vWF^-/-) mice. Mice immunized with native-like aggregates showed significantly higher inhibitory antibody titers than animals that received native FVIII. Following restimulation in vitro with native FVIII, the activation of CD4+ T-cells isolated from mice immunized with native-like aggregates is approximately fourfold higher than mice immunized with the native protein. Furthermore, this is associated with increases in the secretion of proinflammatory cytokines IL-6 and IL-17 in the native-like aggregate treatment group. The results indicate that the native-like aggregates of FVIII are more immunogenic than native FVIII for both the B-cell and the T-cell responses.