MODULATION OF IGE REACTIVITY OF ALLERGENS BY SITE-DIRECTED MUTAGENESIS - POTENTIAL USE OF HYPOALLERGENIC VARIANTS FOR IMMUNOTHERAPY

Specific immunotherapy is an efficient treatment for patients sufferin g from type I allergy. The mechanisms underlying successful immunother apy are assumed to operate at the level of T helper cells, leading to a modulation of the immune response to allergens. During immunotherapy , increasing doses of allergens are given on a regular basis, and the beneficial effects for the patient depend on the concentration of alle rgen used. On the other hand, the risk of IgE-mediated anaphylactic si de effects also increase with the amount of allergen applied per injec tion. Therefore, we have proposed the use of hypoallergenic (low IgE b inding activity) forms of allergens for immunotherapy. We evaluated by site-directed mutagenesis the contributions of individual amino acid residues/positions for IgE binding to Bet v 1, the major allergen of b irch pollen. We found that IgE binding to Bet v 1 depended on at least six amino acid residues/positions. Immunoblot analyses and inhibition experiments showed that the multiple-point Bet v 1 mutant exhibited e xtremely low reactivity with serum IgE from birch pollen-allergic pati ents. In vivo (skin prick) tests showed that the potency of the multip le-point mutant to induce typical urticarial type I reactions in polle n-allergic patients was significantly lower than for wild-type Bet v 1 . Proliferation assays of allergen-specific T cell clones demonstrated that these six amino acid exchanges in the Bet v 1 sequence did not i nfluence T cell recognition. Thus, the Bet v I six-point mutant displa yed significantly reduced IgE binding activity, but conserved T cell a ctivating capacity, which is necessary for immunomodulation. The appro ach described here may be generally applied to produce allergen varian ts to be used in a safe therapy form of immediate-type allergies.