Induction Of Tolerance To FVIII Using Nanoparticles In a Murine Model Of Hemophilia A

Anti-FVIII inhibitor development is currently considered the most serious side effect in the treatment of hemophilia A patients. In this study, we sought to address this problem by determining the efficacy of a novel tolerogenic protocol utilizing nanoparticle formulations provided by Selecta Biosciences. Thus, hemophilia A mice (FVIII KO, n = 6-8) were treated weekly for five weeks through intravenous route with 1 µg recombinant human FVIII (rFVIII), plus immune modulating agent coupled nanoparticles or control nanoparticles (coded F1 or F2 or F3). An additional control group was included, in which the mice were treated on the same schedule with high dose IVIG + rFVIII. As expected, all the mice in the control group (later revealed to be F3) developed high levels anti-FVIII IgG after the five treatments, with the average levels at 21.2 ± 4.4 µg/ml. Both the F1 and F2 nanoparticle treatments effectively prevented the anti-FVIII inhibitory antibody development throughout the course of the 5-week treatment. One week after the final treatment, the majority (5/7) of both F1 and F2 groups remained undetectable for anti-FVIII IgG antibodies. Consistent with this pattern, the anti-FVIII inhibitor titers for the F1, F2, F3 and IVIG groups were 42.3 ± 9.9, 14.3 ± 7.2, 92.9 ± 2.5, and 30.3 ± 5.6 Bethesda Units (BU)/ml, respectively after another 7 days (p < 0.01 for F1 and F2 vs. F3). To determine the duration of the FVIII-tolerance, the mice were challenged twice with rFVIII (1µg) 29 and 53 days after the final nanoparticle treatment. After the first FVIII challenge, the inhibitor titers for the F1, F2, F3 and IVIG groups were 81.3 ± 10.8, 33.1 ± 13.2, 84.1 ± 2.2 (p = 0.41 for F1 vs. F3; and p < 0.01 for F2 vs. F3), and 50.6 ± 9.7 BU/ml, respectively. Remarkably, the FVIII-tolerance in F2 group was sustained even after additional FVIII challenges ∼ 2 months later, when the effect in IVIG group was completely lost. A therapeutic protocol using mice pre-immunized with FVIII also led to substantial reductions in anti-FVIII titers. In conclusion, the concomitant Selecta F2 nanoparticle treatments not only significantly prevented the anti-FVIII inhibitory antibody development during the treatments, but also resulted in long-term FVIII-tolerance, suggesting a potential clinical application. (Supported in part by a grant-in-aid from Selecta Bioscience and by HL061883)