Co-Administration Therapy Modulates Immunity to FVIII In Hemophilia A Mice

Abstract 1219

Our long-term objective is the development of novel therapies for the safe and effective treatment of hemophilia. In particular a safe and economic prophylactic modality to prevent inhibitors development is therefore highly desirable. To this end we have explored strategies to modulate inhibitor development based on the co-administration of FVIII with other proteins, specifically albumin and IVIG. We hypothesise that such co-administration may modulate immunity to FVIII and result in a safe and cost-effective therapy. Based on some published evidence that FIX and vWF may have a modulatory effect on FVIII, we reasoned that perhaps albumin might also play an immunomodulatory effect on FVIII. We have developed a protocol for infusing recombinant FVIII in hemophilia A mice modelled on the current therapy of hemophiliacs, and in response to this protocol hemophilia mice develop high titre of inhibitors. Thus, this model can be used to assess potential immununomodulatory strategies.

Groups of hemophilia A mice (n=5) were infused weekly IV with 2IU rFVIII for a total of 4 weeks. Three of these groups of mice were also administered human albumin (either 25mg IV, 25mg IP, or 500mg IV) at the same time as FVIII. Mice were bled before the treatment and weekly for the 4 weeks, and total antibodies to FVIII were detected by ELISA. Importantly, while mice receiving FVIII had high antibody titer and were in good health, there was significant mortality in the mice treated with FVIII and albumin. Interestingly, the surviving mice had a very low antibody titre, statistically significant from mice receiving only rFVIII.

Given the mortality, another experiment was designed to test the effect of reduced doses of albumin. Groups of hemophilia A mice (n=5) were infused IV with a weekly injection of 2IU rFVIII for a total of 4 weeks supplemented with either 25μg, 50μg or 100μg of human albumin. Mice were bled at weekly intervals until the end of the experiment (day 28), and the titre of anti-FVIIII antibodies was determined. Most All mice survived the entire treatment and were found in good health. Further, the FVIII antibody titre in the group that received albumin was detectable, but significantly reduced with respect to the mice that received only FVIII. Marked variability among individual mice was observed.

A group of hemophilia A mice (n=5) received weekly IP injections of 2IU rFVIII. Another group of mice received the same regimen of FVIII in addition to human IVIG (1g/kg). A third group of hemophilia mice received 2IU rFVIII and a higher dose of human IVIG (2g/kg). These doses mimic the established human IVIG regimen. Mice were bled before treatment and at weekly intervals thereafter. The titre of anti-FVIII antibodies in mice was determined by ELISA. By day 21, the antibody titre in mice injected with FVIII was high, as expected. In contrast, the titre in mice receiving FVIII and IVIG was low, and comparable to that of all the mice prior to the treatment (na•ve mice), strongly suggesting an immune modulatory effect by IVIG (Figure 1). Our data supports the limited use of IVIG to treat inhibitors.

Figure 1.

View largeDownload slide

FVIII / IVIG co-administration

Figure 1.

View largeDownload slide

FVIII / IVIG co-administration

Close modal

Our findings suggest that co-administration of FVIII with albumin and IVIG can modulate the humoral immune response to recombinant human FVIII in hemophilia mice. Further, the observed effect appears to be dependent on the dose of albumin co-administered. It is hypothesized that the observed modulation is due to antigen competition, as previously speculated. Importantly, while IVIG was well tolerated by mice, very serious adverse effects were observed in mice treated with high doses of albumin. Interestingly, even the high dose used represents a small fraction of the physiological levels of albumin in human plasma, which is 3–5 g/dL. IVIG, in contrast, is a known modulator of the immune response in humans, and the study of this modulatory effect in a hemophilia mouse may lead to novel therapeutic strategies, relevant for the management of hemophilia.