Long-term induction of immune tolerance after blockade of CD40-CD40L interaction in a mouse model of hemophilia A.

A factor VIII-deficient knockout mouse was used as a model for severe hemophilia A to characterize the immune response to recombinant human factor VIII (fVIII) and to study new approaches for induction of immune tolerance to fVIII. Mice initially received periodic injections of fVIII in doses similar to those used for the treatment of human hemophilia A. To induce immune tolerance, a hamster monoclonal antibody specific for murine CD40 ligand (CD40L or CD154) was injected with fVIII. Control mice received fVIII alone or fVIII and hamster immunoglobulin G. After treatment, humoral and cellular immune responses were evaluated. Ninety-five percent of anti-CD40L-treated mice had lower titers of anti-fVIII antibody (less than 1 microg/mL) compared with fVIII-injected control mice (mean, 18 microg/mL). To determine whether anti-CD40L treatment induces long-term immune tolerance, mice were rechallenged 3 times with fVIII alone. At 150 days after treatment, 12 of 22 anti-CD40L-treated mice remained tolerant to fVIII (anti-fVIII antibody titers less than 1 microg/mL). However, tolerant mice immunized with tetanus toxoid (TT) developed high anti-TT antibody, demonstrating that tolerance is fVIII specific. T cells from tolerant mice showed impaired proliferative responses after stimulation with fVIII in vitro and lack of production of the cytokines interleukin-2 (IL-2), IL-4, interferon gamma, and IL-10. These results demonstrate that long-term immune tolerance to fVIII was effectively induced after early blockade of CD40-CD40L interaction. In addition, the lack of tolerance in this model was associated with the expression of a Th2 phenotype.