Characterization of a factor VIII/immunoglobulin heavy chain μ double-knockout mouse model of hemophilia A for long-term exposure to factor VIII proteins

Deficiency of coagulation factor (F)VIII is the key characteristic of hemophilia A. The FVIII knockout mouse model is a valuable tool for investigating disease mechanisms and evaluating the pharmacokinetics (PK) and efficacy of therapeutic agents. However, its utility for long-term studies, particularly those focused on prophylaxis, is limited by the development of anti-FVIII antibodies following repeated FVIII administration.

To develop a FVIII knockout model that does not generate antibodies against FVIII but is not severely immunosuppressed.

Established FVIII single-knockout (FVIII^-/-; SKO) mice were crossed with the immunoglobulin heavy chain μ (Ighm) knockout strain to generate FVIII/Ighm double-knockout (FVIII^-/-/Ighm^-/-; DKO) mice. Both SKO and DKO mice received 4 intravenous doses of recombinant human FVIII on days 0, 7, 14, and 21. Peripheral blood, bone marrow, and spleen samples were collected to assess (i) the immune response to FVIII, (ii) FVIII PK after repeated dosing, and (iii) overall immune status.

DKO mice did not develop detectable anti-FVIII antibodies. The formation of FVIII-specific antibody-secreting cells was abrogated, and the presence of FVIII-specific T cells was substantially reduced. PK analysis performed after 3 weeks of FVIII exposure showed variable reductions in FVIII recovery and half-life in SKO mice. In contrast, PK parameters in DKO mice remained consistently within the expected physiological range.

The DKO mouse model can be used for long-term studies of FVIII products and other hemostatic proteins, enabling the evaluation of repeated-dose PK and prophylactic efficacy without interference from inhibitor formation.