Development of a novel hemophilia A mouse model to study emicizumab and factor VIII inhibitor risk

The introduction of emicizumab, a bispecific antibody that mimics factor VIII (FVIII) function, has raised critical questions regarding the potential for FVIII inhibitor development when patients receiving emicizumab prophylaxis are administered FVIII during high-risk clinical scenarios. To investigate this concern, we developed a novel human hemophilia A (HemoAhfIX/hfX) mouse model by adeno-associated virus delivery of human F9 and F10 genes as episomal copies, which expressed sufficient levels of human FIX and FX (158.0 ± 57.9 international units [IU]/dL and 372.8 ± 170.7 IU/dL, respectively, at 12 weeks). Emicizumab treatment in HemoAhfIX/hfX mice significantly shortened activated partial thromboplastin time and partially restored thrombin generation capacity. Using a 2-factor factorial design, 55 mice were assigned to 4 treatment groups: (A) FVIII prophylaxis, (B) on-demand FVIII, (C) emicizumab with FVIII prophylaxis, and (D) emicizumab with on-demand FVIII. On-demand groups received weekly knee joint puncture injuries to simulate bleeding episodes throughout the experimental period. Although most animals developed high-titer inhibitors, limiting the detection of intergroup differences in inhibitor development rates, inhibitor titers varied significantly across treatment groups (P = .0098), with notably lower levels in emicizumab-treated mice (P = .0006). In addition, emicizumab treatment preserved joint architecture, evidenced by reduced synovitis. These findings demonstrate that emicizumab therapy is associated with attenuated inhibitor titers and enhanced joint preservation against bleeding in HemoAhfIX/hfX mice, establishing this model as a valuable platform for preclinical evaluation of emicizumab-based therapeutic strategies.