A novel bispecific DARPin targeting Fc gamma RIIB and Fc epsilon RI-bound IgE inhibits allergic responses

作者：Zellweger F; Gasser P; Brigger D; Buschor P; Vogel M; Eggel A^*

来源：Allergy, 2017, 72(8): 1174-1183.

DOI：10.1111/all.13109

• allergy
• bispecific DARPin
• FccRIIb
• FceRIa
• IgE

Background: Binding of allergen-specific IgE to its high-affinity receptor Fc epsilon RI on basophils and mast cells is a central event in the development of allergies. Exposure of these cells to allergens induces the release of soluble mediators causing allergic symptoms. The inhibitory low-affinity IgG Fc-receptor Fc gamma RIIB is co-expressed on allergic effector cells and has been implicated in negative regulation of immediate hypersensitivity responses. In order to harvest the inhibitory function of this receptor, we aimed to select specific binders against Fc gamma RIIB and to generate a bispecific molecule simultaneously targeting Fc gamma RIIB and Fc epsilon RI-bound IgE on the surface of allergic effector cells.
Methods: We selected Fc gamma RIIB-specific binding molecules from a library of designed ankyrin repeat proteins using ribosome display technology. The bispecific binding modality was generated by molecular cloning and recombinant protein expression. We determined binding characteristics on molecular and cellular levels using SPR, ELISA, and flow cytometry. The inhibitory potential of the newly described molecules was assessed in different cellular degranulation assays ex vivo and in a mouse model of passive systemic anaphylaxis.
Results: We demonstrate that the selected DARPin (R) proteins recognize Fc gamma RIIB with high affinity. Furthermore, the bispecific binding protein successfully interferes with allergen-induced cell degranulation and efficiently inhibits systemic anaphylaxis in vivo. Mechanistically, we report that Fc gamma RIIB-mediated inhibition of effector cell activation requires direct ligation to an activating Fc epsilon RI receptor.
Conclusion: The described bispecific DARPin protein has the ability to co-ligate Fc gamma RIIB with Fc epsilon RI-bound IgE on allergic effector cells and represents an efficient dual-modality to interfere with allergic hypersensitivity reactions.

• 出版日期2017-8