It is now clear that Th1-type cytokines can suppress IgE production and immediate hypersensitivity. Since dendritic cells (DCs) and macrophages (M s) are potent antigen presenting cells (APCs) for activation of ThI and CD8+ T cells, redirecting allergens for uptake by these APCs could induce immune responses that suppress IgE production. We have shown that targeting antigen (Ag) to CD64 (FcgRI) increases Ag potency 1000-fold in human T cell assays and stimulates a Th1-type response in a transgenic (Tg) mouse with myeloid-specific expression of human CD64. The present application is designed to optimize the potentiation of immunity realized by targeting Ag to CD64, to elucidate the mechanisms by which CD64-targeting induces a Th1-type response and to evaluate the potential for novel genetic constructs of CD64-targeted Ags to induce responses that either prevent or decrease IgE-mediated hypersensitivity. We propose: (1) To elucidate the mechanisms by which CD64 targeting of antigen potentiates and modulates T cell responses. We postulate that different forms of CD64 targeted and non-targeted antigens will be internalized by different pathways, leading to different kinetics, efficiency and/or specificity of peptide processing and insertion into MHC class I and class 11 molecules. We will thus evaluate the influence of Ag size and valency on CD4+ and CD8+ T cell activation and on APC phenotype with respect to expression of costimulatory molecules and production of cytokines in response to targeted and non-targeted Ag. (2) To test the ability of a model CD64-targeted allergen and a novel fusion protein (antiCD64:FcgRI) to stimulate Th1 and CD8+ T cell responses in a mouse model of IgE-mediated hypersensitivity. Anti-CD64:FceRI, in which anti-CD64 is linked to the a-chain of FceRl, is designed to suppress IgE-specific responses even when the Ag is unknown. It is predicted to acutely block binding of Ag-specific IgE to mast cells, as well as to retarget IgE-allergen complexes specifically to DCs and MOs, stimulating a more effective Th1 response to the bound allergen. The induction in vivo of specific cytokines, helper and cytolytic T cells and antibody subclasses, as well as protection from hypersensitivity induced by aerosolized Ag will be measured. (3) To examine CD64-retargeting of the major cat allergen, Fel d1, which induces IgE-mediated hypersensitivity in both a high percentage of humans and in rodents. The Ag has been cloned and dominant peptides mediating human hypersensitivity have been identified. We will examine the ability of targeted Fel d1, as well as native and altered (antagonist) peptides, to induce suppression of Th2 cell activation both in mice and in cell cultures derived from patients allergic to Fel d1. These studies should delineate the mechanisms by which CD64-targeted Ag induces Th1 responses, and lay the foundation for potential use of a novel and potentially powerful approach to prevent or ameliorate human IgE-mediated hypersensitivity.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Allergy and Immunology Study Section (ALY)
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Plaut, Marshall
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Dartmouth College
Schools of Medicine
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