Improved diagnosis and vaccination of pet allergy : from molecular mechanisms to clinic

Sammanfattning: Allergy is a disease affecting around 30% of the population worldwide, causing extensive suffering for individual patients and constituting a significant socioeconomic burden to society. The immunological reaction in allergy causes symptoms in the range from mild itching and runny nose to systemic death threatening states. One common cause of allergy is pets, such as dog and horse. Today’s diagnostics of pet allergy is frequently based on extract from dander or hair, however extracts may vary in content of allergens. The only curative treatment is allergen specific immunotherapy (SIT), routinely performed by injections with allergen extracts. For both diagnostics and treatment, the content of allergen component in the extracts is vital. The application of DNA technology has opened opportunities to produce allergens in pure form and at good yield, making the allergen components available for both diagnostics and SIT. Further improvement of SIT could be achieved by the use of adjuvants able to skew the allergic immune response to a non-allergic response. Currently, the adjuvant alum is used in SIT. New adjuvants are needed that more efficiently stimulate regulatory or Th1 type responses. The aim of this thesis was to identify and analyze new sources of pet allergens, to identify new allergen components from horse and dog, and to investigate mechanisms and clinical safety/efficacy of a novel adjuvant candidate based on chitosan, which will possibly be suitable for future use in SIT, as well as for other vaccine applications. In paper I of this thesis we investigated dog saliva as a possible allergen source, and if today’s diagnostic extract could be improved by using saliva. We found that some individuals with negative IgE test to dog dander have IgE to dog saliva, and that the IgE binding is biologically relevant, as dog saliva could activate basophil degranulation. Using immuno-proteomic analyses, four potential IgE binding proteins not previously described as allergens were identified. In paper II we present a thorough analysis of horse allergen components. We identified three novel full-length IgE binding proteins and evaluated the prevalence of IgE reactivity among 100 sera from horse sensitized individuals. All three novel allergens belong to protein families from which allergens from dog, cat or cow have previously been described. The prevalence of sensitization to the new allergens ranged between 34 and 66% and together with three additional known horse allergens all 100 sensitized individuals could be detected. Paper III investigated the adjuvant candidate ViscoGel, composed of chitosan based viscoelastic particles. ViscoGel’s ability to be phagocytosed by, and activate antigen presenting cells was studied in vitro. The antigen presenting cells were able to take up the chitosan particles of 10 and 200μm size, and to stimulate the release of IL-1β in a caspase-1 independent manner. Paper IV describes a clinical phase I/IIa trial evaluating safety of ViscoGel alone and in combination with the model vaccine Act-HIB, and efficacy as an adjuvant for Act-HIB. ViscoGel was well tolerated when injected intramuscularly. No adjuvant effect was observed on the antibody response to Act-HIB, but the IFN response was affected, suggesting that ViscoGel may promote a Th1 type of response. To conclude, the results presented in this thesis have the potential to improve diagnostics of allergy to dog and horse. Moreover, a new potential adjuvant was shown to be safe and exhibited immunological properties that may be favorable for use in SIT.