Nanobody-based IgG simultaneously inhibit the allergenic and enzymatic activity of the dominant honeybee venom allergen

Insect venoms can cause severe allergic reactions, including anaphylaxis, in sensitized individuals. In this study, we aim at preventing anaphylaxis mediated by the most abundant and dominant honeybee venom allergen phospholipase A2 (Api m 1) by blocking its interaction with allergic patient IgE. Therefore, we characterize selected Api m 1-specific nanobodies and identify two high-affinity binders with non-overlapping epitopes. Crystal structures of Api m 1/nanobody complexes reveal diametrically opposed epitopes, one of which involves the active site of Api m 1. Based on this background, we develop mono- and bispecific nanobody-human IgG₁ Fc, which exhibits pronounced blocking of IgE binding and effector cell activation in blood samples from honeybee venom allergic patients and reduces systemic reactions in a mouse model of allergen-induced anaphylaxis. This work provides a rationale for using nanobody-based inhibitors to prevent Api m 1-mediated anaphylaxis in honeybee venom allergy.