9ABSTRACTCandidate: Name: Marian L Denson Degree Title: PhDUniversity: The University of Manchester Date: January 2012Thesis Title: Rational Design of Immunotherapy to treat Fungal AllergyBackgroundAsthma affects 5.4 million people in the UK. Asthma subgroups are also susceptible to inhalation of fungal spores (Aspergillus fumigatus) and development of pulmonary fungal aspergilloma; presenting a life threatening but poorly understood condition. NHS costs for corticosteroids, bronchodilators and antifungal agents that are only partially effective continue to rise. Allergy immunotherapy development is of great interest as it is specific to the allergen and can harnesses key adaptive immune T-cells to downregulate inflammatory responses. Immunotherapy has been used with varying degrees of success for treatment of grass, pollen, venom, cat and dog allergens however to date has not been directed to fungal allergens. The study aims were: 1) to further understand the A. fumigatus allergens and the protein epitopes responsible for generating immune responses. 2) To genotype participating ABPA/SAFS patients to observe any HLA associations.Methods: 37 subjects with fungal sensitivity were recruited to the study which received permission from the local ethics committee (UHSM LREC). Computer bioinformatic predictions using Propred software identified several potential fungal T cell peptide epitopes; of which 8 peptides were soluble and tested in vitro for specific T-cell proliferation responses by flow cytometric analysis. Skin prick tests determined subject responses to fungal allergens including A. fumigatus, and DNA analysis determined subject HLA type.Results5 of 8 soluble peptides were Aspergillus fumigatus derived and 3 from Alternaria alternata. All 8 peptides induced higher CD4 proliferative responses in ABPA/SAFS patients, compared to healthy controls from highest significance to lowest as follows: peptide 1.1>9.1>8.1>2.1>9.1.1>4.1>4.1.1 and 10.1.173% subjects elicited skin responses to A. fumigatus. DNA HLA typing identified alleles associated with ABPA/SAFS but not all allele sub types.DiscussionThe ABPA/SAFS group consistently raised T-cell responses to fungal peptides compared to controls. This demonstrates peripheral CD4s retain memory for fungal specificity and clearly respond when challenged with fungal epitopes in vitro. This concept underpins the rationale to further characterize the responding CD4 cells and pursuing bioinformatics approaches for immunotherapy investigations for fungal allergy.