Suppression of anti-drug antibody formation against coagulation factor VIII by oral delivery of anti-CD3 monoclonal antibody in hemophilia A mice

Abstract

Active tolerance to ingested dietary antigens forms the basis for oral immunotherapy to food allergens or autoimmune self-antigens. Alternatively, oral administration of anti-CD3 monoclonal antibody can be effective in modulating systemic immune responses without T cell depletion. Here we assessed the efficacy of full length and the F(ab')₂ fragment of oral anti-CD3 to prevent anti-drug antibody (ADA) formation to clotting factor VIII (FVIII) protein replacement therapy in hemophilia A mice. A short course of low dose oral anti-CD3 F(ab')₂ reduced the production of neutralizing ADAs, and suppression was significantly enhanced when oral anti-CD3 was timed concurrently with FVIII administration. Tolerance was accompanied by the early induction of FoxP3⁺LAP^-, FoxP3⁺LAP⁺, and FoxP3^-LAP⁺ populations of CD4⁺ T cells in the spleen and mesenteric lymph nodes. FoxP3⁺LAP⁺ Tregs expressing CD69, CTLA-4, and PD1 persisted in spleens of treated mice, but did not produce IL-10. Finally, we attempted to combine the anti-CD3 approach with oral intake of FVIII antigen (using our previously established method of using lettuce plant cells transgenic for FVIII antigen fused to cholera toxin B (CTB) subunit, which suppresses ADAs in part through induction of IL-10 producing FoxP3^-LAP⁺ Treg). However, combining these two approaches failed to improve suppression of ADAs. We conclude that oral anti-CD3 treatment is a promising approach to prevention of ADA formation in systemic protein replacement therapy, albeit via mechanisms distinct from and not synergistic with oral intake of bioencapsulated antigen.