EcN-Arah2-Sia

Oral immunotherapies for food allergies remain limited by safety concerns and inconsistent efficacy.

We developed a probiotic-based strategy using Escherichia coli Nissle 1917 (EcN) engineered to codisplay the major peanut allergen Ara h 2 and the inhibitory sialic acid-binding immunoglobulin-like lectin (Siglec) ligand Sia(α2-6)Gal(β1-4)Glc to dampen allergen-specific B-cell responses.

Expression of Siglec ligand and Ara h 2 on EcN was confirmed by Western blot and flow cytometry. C3H/HeJ mice were sensitized by intraperitoneal injections and treated orally with bacteria. B-cell responses were assessed by flow cytometry after in vitro stimulation of splenocytes. Serum antibody and mast cell protease levels were measured by ELISA. Anaphylaxis protection was evaluated through hypothermia and symptom scores.

In vitro stimulation with peanut extract increased B-cell activation in sensitized splenocytes, and wild-type EcN further enhanced this activation. In contrast, EcN displaying Ara h 2 and Siglec ligands significantly reduced CD23 upregulation and restored CD86 and CD69 expression. Oral administration of EcN displaying Ara h 2 and Siglec ligands to peanut-sensitized mice reduced allergen-specific B-cell activation, decreased serum IgE and IgG levels, and significantly attenuated anaphylactic responses. EcN codisplaying Ara h 2 and Siglec ligand conferred protection, highlighting the requirement for antigen-specific engagement of inhibitory B-cell pathways. While this study focused on Ara h 2, including multiple allergens could extend protection.

Our findings establish proof of concept for orally delivered antigen-specific immune tolerance using engineered probiotics, providing a scalable, noninvasive alternative to intravenous nanoparticle therapies. This approach could be extended to other allergies or autoimmune conditions.