Arbeiten aus dem Paul-Ehrlich-Institut (Bundesamt fur Sera und Impfstoffe) zu Frankfurt a.M最新文献

Allergen immunotherapy was first introduced in the early part of the twentieth century. It is widely practiced despite having specific limitations. Considerable effort has been devoted to developing new modified allergens that, compared with conventional allergen immunotherapy, improve efficacy, decrease the time required to achieve effect, reduce inconvenience, and enhance safety. Increased understanding of allergic respiratory inflammation has led to the development of therapeutic modalities that potentially arrest the disease process in asthma or allergic rhinitis. This paper addresses an adjuvant approach in which highly active immunostimulatory phosphorothioate oligodeoxyribonucleotide sequence (i.e. immunostimulatory DNA) are conjugated to the principal allergenic moiety of a relevant aeroallergen. We have recently completed the first human safety studies in patients with allergic rhinitis with Amb a 1-immuno-stimulatory oligonucleotide conjugate (AIC) --a novel therapeutic vaccine comprised of Amb a 1, the principal allergenic protein of ragweed, conjugated specific immunostimulatory oligonucleotides (ISS). The results demonstrate that AIC was several hundred-fold less reactive than a standardized ragweed extract when evaluated by quantitative intradermal skin titration methodology. Furthermore, AIC reduced histamine release from basophils to a similar degree. The DNA vaccine induced IgG antibody production in treated patients. AIC compared with standardized aqueous ragwood exhibited fewer local reactions on skin testing, a finding that suggests that AIC offers the potential for an improved safety profile for immunotherapy. Additional trials to further evaluate the safety, immunologic effect, and therapeutic efficacy of AIC for ragwood-induced allergic rhinitis and asthma are ongoing.

Biological standardization of allergen extracts is one of the steps in the characterization of an extract. The gold standard for determination of biological potency is the skin prick test, but histamine release (HR) has been used as a convenient ex vivo method for analyzing a large number of samples. We describe the use of rabbit basophils as a tool in biological standardization. Using peanut as a model allergen, it is described how rabbits immunized for production of antiserum may become sensitized and their basophils used for histamine release experiments. It is also possible to use rabbit antiserum to passively sensitize basophils derived from naive rabbits, but the sensitivity of this method is so far 100-1000 times lower than the direct histamine release. The rabbit histamine release results are compared to an ELISA developed by means of the same antisera and by passive sensitization of human basophils using serum from a strongly sensitized peanut-allergic patient. The overall sensitivity of the methods were ELISA > HR-human cells > HR-sensitized rabbit cells > HR-passively sensitized rabbit cells. The use of rabbit basophils for biological standardizations will allow for the use of rabbit antisera.

Induction of specific unresponsiveness (tolerance) in peripheral T cells by IL-10 and/or TGF-beta and recovery by cytokines from the tissue microenvironment represent two key steps in specific immunotherapy of allergy and natural exposure to allergens in healthy individuals. IL-10 and TGF-beta elicit tolerance in T cells and thereby control the suppression and development of antigen-specific immunity. Both cytokines also play an important role on the generation of a non-inflammatory IgG4 and IgA type of allergen--specific antibodies during the course of specific immunotherapy. Histamine plays an important role in upper and lower airway inflammation. In addition to its well-characterized effects in the acute inflammatory and allergic responses, histamine regulates several aspects of antigen-specific immune response development. Histamine affects the maturation of dendritic cells and alters their T cell polarizing capacity. Histamine regulates antigen specific Th1 and Th2 cells as well as related antibody isotype responses. Histamine and four different known histamine receptors (HR) display a very complex system and their expression changes according to the stage of cell differentiation as well as microenvironmental influences.

Rationale: Hazelnut allergy ranks among the most frequently observed food allergies. Clinical symptoms range from the oral allergy syndrome to life threatening anaphylaxis. Diagnosis of hazelnut allergy partially relies on in vivo testing by means of skin prick testing (SPT). The aim of this study was to characterize hazelnut SPT extracts both in vitro and in vivo.

Methods: Hazelnut SPT extracts were investigated for protein concentration and composition. The major hazelnut allergen Cor a 1, lipid transfer protein (LTP) and thaumatin-like-protein (TLP) were monitored by competitive RIA and immunoblotting. SPT extracts (n = 6) were analyzed for skin reactivity and the correlation between the SPT extract protein concentration and the mean skin reactivity (HEIC) was determined in a group of hazelnut-allergic patients (n = 30). For one SPT extract, the threshold level for Cor a 1 was determined in Cor a 1-monosensitized patients (n = 5).

Results: Protein concentrations ranged from 0.2-14 mg/ml. Although some proteins were present in most extracts (bands at 10, 22-28, 32 and around 48 kDa), clear differences in composition were observed (both intra- and inter-variability). The concentration of the major hazelnut allergen Cor a 1 differed up to a factor 50 (0.6-32 micrograms/ml). LTP was virtually absent in 3/9 SPT extracts and variable quantities of TLP were detected by immunoblotting. Some patients (6/30) had a false-negative SPT with 3/6 SPT extracts. There was a clear correlation between the protein concentration and the mean HEIC (RPearson = 0.87). The threshold level for Cor a 1 was +/- 3.2 ng/ml as assessed with one of the products investigated.

Conclusions: Heterogeneous protein concentration/composition of SPT extracts results in variable skin test responses. The absence of potentially severe allergens like LTP may lead to false-negative SPT results that jeopardize a patient's safety. From these results it can be concluded that there is a strong need for standardization of products for SPT.