Novas metodologias de diagnóstico e de imunoterapia para alergias baseadas no proteoma de Alternaria alternata

The main goal of the Path&D proposal is to identify proteome-based solutions to increase the accuracy of Alternaria alternata respiratory allergies diagnosis and to select proteins with a strong potential to be involved in the triggering of the host cellular responses that lead to colonization/infection, type I hypersensitivity and human respiratory allergic disease. These studies will clarify the impact of A. alternata proteins in the disease from early interaction with human host cells until the development of a severe allergic reaction. This project will strongly contribute for a better diagnosis of A. alternata respiratory allergies and will open windows of opportunities to design drugs to prevent adhesion and new targets for the formulation of innovative therapeutics and preventive strategies. Based on the current knowledge in the scientific area and on the proposal’s scientific team preliminary results, the specific objectives proposed are: 1. Based on the proteomic profiles of several forms of A. alternata to which individuals can be exposed (conidia, germinated conidia, hyphae, EVs), to elaborate novel protocols to be used in the preparation of extracts to be used in Alternaria respiratory allergies diagnosis; 2. To evaluate the allergenic and adhesive properties of proteins identified in the extracts using an in silico approach; 3. To evaluate the function of A. alternata and proteins selected in the in silico analysis, in what regards an allergic immune response, or associated with adhesion and type I hypersensitivity response, in human respiratory epithelial cells and to innate immune cells, using an in vitro approach; 4. To detect specific IgEs in allergic patients’ sera, providing information on proteins with high allergenic potential, not identified before, and that can constitute novel allergens; 5. To obtain monoclonal antibodies against the selected protein, to be used as immunotherapeutics; 5. To evaluate both in vivo the contribution of the selected proteins/allergens in inducing sensitization to A. alternata 6. To evaluate in vivo the potential of selected proteins/allergens and of the respective monoclonal antibodies as immunotherapeutics.