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

## TL;DR

Researchers created a new mouse model for hemophilia A that avoids antibody formation, allowing long-term study of factor VIII treatments.

## Contribution

A novel FVIII/Ighm double-knockout mouse model was developed to enable long-term prophylactic studies without inhibitor formation.

## Key findings

- DKO mice did not develop anti-FVIII antibodies after repeated dosing.
- Pharmacokinetic parameters in DKO mice remained stable and physiological.
- The DKO model supports long-term evaluation of hemostatic proteins without immune interference.

## Abstract

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.

•The hemophilia A mouse model is not suitable for prophylaxis studies due to inhibitor formation.•We developed a new model lacking the immunoglobulin heavy chain μ.•After 3 weeks of exposure, mice did not develop inhibitors and had stable pharmacokinetics.•This mouse model can be used to study long-term factor prophylaxis.

The hemophilia A mouse model is not suitable for prophylaxis studies due to inhibitor formation.

We developed a new model lacking the immunoglobulin heavy chain μ.

After 3 weeks of exposure, mice did not develop inhibitors and had stable pharmacokinetics.

This mouse model can be used to study long-term factor prophylaxis.