Eva Wollmann

Monitoring Allergen Immunotherapy Effects by Microarray

Opinion statementAllergen-specific immunotherapy (AIT) is the only treatment of IgE-mediated allergies so far that has a sustained effect on clinical symptoms and can modify the course of the disease. It is an allergen-specific treatment and therefore requires the correct identification of the disease-causing allergens. Furthermore, AIT is a time-consuming treatment for which the efficacy is dependent on several factors. Therefore, diagnostic tests and biomarkers are needed that facilitate (1) selection of the correct allergens according to the patient’s individual sensitization profile and (2) to monitor the effects of AIT. This can provide support for the decision to continue, modify, or discontinue vaccination. One significant mechanism of action of AIT is the induction of allergen-specific antibodies that compete with IgE for the binding to allergen molecules, hence referred to as blocking antibodies. It was shown in several studies that the induction of blocking antibodies by AIT, and their specificity can be measured by allergen microarrays. Inhibition of allergen-specific IgE binding by blocking antibodies can also be determined by microarrays and is associated with changes in clinical parameters or other in vivo and in vitro assays demonstrating efficacy of AIT. Furthermore, allergen microarrays allow determination of IgE sensitizations towards a comprehensive set of allergen molecules and therefore are well suited for identifying the disease-causing allergens for correct prescription of AIT. Thus, diagnostic tests based on microarrayed allergens can be useful in determining the correct prescription of AIT and can be used to monitor efficacy of AIT.

Persistence of IgE-Associated Allergy and Allergen-Specific IgE despite CD4+ T Cell Loss in AIDS

The infection of CD4+ cells by HIV leads to the progressive destruction of CD4+ T lymphocytes and, after a severe reduction of CD4+ cells, to AIDS. The aim of the study was to investigate whether HIV-infected patients with CD4 cell counts <200 cells/µl can suffer from symptoms of IgE-mediated allergy, produce allergen-specific IgE antibody responses and show boosts of allergen-specific IgE production. HIV-infected patients with CD4 counts ≤200 cells/µl suffering from AIDS and from IgE-mediated allergy were studied. Allergy was diagnosed according to case history, physical examination, skin prick testing (SPT), and serological analyses including allergen microarrays. HIV infection was confirmed serologically and the disease was staged clinically. The predominant allergic symptoms in the studied patients were acute allergic rhinitis (73%) followed by asthma (27%) due to IgE-mediated mast cell activation whereas no late phase allergic symptoms such as atopic dermatitis, a mainly T cell-mediated skin manifestation, were found in patients suffering from AIDS. According to IgE serology allergies to house dust mites and grass pollen were most common besides IgE sensitizations to various food allergens. Interestingly, pollen allergen-specific IgE antibody levels in the patients with AIDS and in additional ten IgE-sensitized patients with HIV infections and low CD4 counts appeared to be boosted by seasonal allergen exposure and were not associated with CD4 counts. Our results indicate that secondary allergen-specific IgE production and IgE-mediated allergic inflammation do not require a fully functional CD4+ T lymphocyte repertoire.

Natural clinical tolerance to peanut in African patients is caused by poor allergenic activity of peanut IgE.

In Africa, peanuts are frequently consumed, but severe allergic reactions are rare. We investigated immunological patterns of clinical tolerance to peanut in peanut‐sensitized but asymptomatic patients from central Africa compared to peanut‐allergic and peanut‐sensitized but asymptomatic patients from Sweden.

Extracorporeal IgE Immunoadsorption in Allergic Asthma: Safety and Efficacy

Background Prevention of IgE-binding to cellular IgE-receptors by anti-IgE (Omalizumab) is clinically effective in allergic asthma, but limited by IgE threshold-levels. To overcome this limitation, we developed a single-use IgE immunoadsorber column (IgEnio). IgEnio is based on a recombinant, IgE-specific antibody fragment and can be used for the specific extracorporeal desorption of IgE. Objective To study safety and efficacy of IgEnio regarding the selective depletion of IgE in a randomized, open-label, controlled pilot trial in patients with allergic asthma and to investigate if IgEnio can bind IgE-Omalizumab immune complexes. Methods Fifteen subjects were enrolled and randomly assigned to the treatment group (n = 10) or to the control group (n = 5). Immunoadsorption was done by veno-venous approach, processing the twofold calculated plasma volume during each treatment. A minimum average IgE-depletion of 50% after the last cycle in the intention-to-treat population was defined as primary endpoint. Safety of the treatment was studied as secondary endpoint. In addition, possible changes in allergen-specific sensitivity were investigated, as well as clinical effects by peak flow measurement and symptom-recording. The depletion of IgE-Omalizumab immune complexes was studied in vitro. The study was registered at clinicaltrials.gov (NCT02096237) and conducted from December 2013 to July 2014. Results IgE immunoadsorption with IgEnio selectively depleted 86.2% (± 5.1% SD) of IgE until the end of the last cycle (p < 0.0001). Removal of pollen allergen-specific IgE was associated with a reduction of allergen-specific basophil-sensitivity and prevented increases of allergen-specific skin-sensitivity and clinical symptoms during pollen seasons. IgEnio also depleted IgE-Omalizumab immune complexes in vitro. The therapy under investigation was safe and well-tolerated. During a total of 81 aphereses, 2 severe adverse events (SAE) were recorded, one of which, an episode of acute dyspnea, possibly was related to the treatment and resolved after administration of antihistamines and corticosteroids. Conclusions This pilot study indicates that IgE immunoadsorption with IgEnio may be used to treat patients with pollen-induced allergic asthma. Furthermore, the treatment could render allergic patients with highly elevated IgE-levels eligible for the administration of Omalizumab and facilitate the desorption of IgE-Omalizumab complexes. This study was funded by Fresenius Medical Care Deutschland GmbH, Bad Homburg, Germany.

Comparison of the Specificities of IgG, IgG-Subclass, IgA and IgM Reactivities in African and European HIV-Infected Individuals with an HIV-1 Clade C Proteome-Based Array

A comprehensive set of recombinant proteins and peptides of the proteome of HIV-1 clade C was prepared and purified and used to measure IgG, IgG-subclass, IgA and IgM responses in HIV-infected patients from Sub-Saharan Africa, where clade C is predominant. As a comparison group, HIV-infected patients from Europe were tested. African and European patients showed an almost identical antibody reactivity profile in terms of epitope specificity and involvement of IgG, IgG subclass, IgA and IgM responses. A V3-peptide of gp120 was identified as major epitope recognized by IgG1>IgG2 = IgG4>IgG3, IgA>IgM antibodies and a C-terminal peptide represented another major peptide epitope for the four IgG subclasses. By contrast, gp41-derived-peptides were mainly recognized by IgG1 but not by the other IgG subclasses, IgA or IgM. Among the non-surface proteins, protease, reverse transcriptase+RNAseH, integrase, as well as the capsid and matrix proteins were the most frequently and strongly recognized antigens which showed broad IgG subclass and IgA reactivity. Specificities and magnitudes of antibody responses in African patients were stable during disease and antiretroviral treatment, and persisted despite severe T cell loss. Using a comprehensive panel of gp120, gp41 peptides and recombinant non-surface proteins of HIV-1 clade C we found an almost identical antibody recognition profile in African and European patients regarding epitopes and involved IgG-sublass, IgA- and IgM-responses. Immune recognition of gp120 peptides and non-surface proteins involved all four IgG subclasses and was indicative of a mixed Th1/Th2 immune response. The HIV-1 clade C proteome-based test allowed diagnosis and monitoring of antibody responses in the course of HIV-infections and assessment of isotype and subclass responses.

Possible effect of landscape design on IgE recognition profiles of two generations revealed with micro‐arrayed allergens

The aim of this study was to investigate possible effects of landscape design on the IgE sensitization profile toward inhalant allergens in patients with respiratory allergy from Uzbekistan where green areas have been changed during the last two decades by a State program. Sera from two different generations of Uzbek (n=58) and, for control purposes, from two generations of Austrian (n=58) patients were analyzed for IgE reactivity to 112 different micro‐arrayed allergen molecules by ImmunoCAP ISAC technology. Changes in molecular IgE sensitization profiles to pollen allergens in the young vs the middle‐aged Uzbek population were associated with replanting, whereas those in the Vienna populations reflected natural changes in plant growth. Our data indicate that anthropologic as well as natural changes in the biome may have effects on IgE sensitization profiles already from one to another generation.

EFIS-EJI African International Conference on Immunity (AICI).

The EFIS-EJI African International Conference on Immunity (AICI: www.aici2011.org) took place from November 3 to November 7, 2011 in Victoria Falls, Zimbabwe, a meeting that is sponsored in part by the EFIS-EJI partnership beyond its typical European remit [1]. The central theme of the meeting was “Immunity in infectious diseases and allergy—diagnosis, treatment and prevention” and it was a follow-up to two earlier conferences at the same venue, one in 2007 whose theme was “Immunity: From deficit to excess” and the Fifth Congress of the Federation of African Immunological Societies (FAIS: www.faisoc.org) in 2003. The latter meeting was attended by representatives of the International Union of Immunological societies (IUIS) and its regional affiliates, the European Federation of Immunological Societies (EFIS), the Federation of Immunological Societies of Asia (FIMSA), and the Latin American Federation of Immunological Societies (ALAI). There is no doubt that the slogan of the IUIS, “Immunology without borders,” has been a driving force behind the meetings organized in Africa, which are intended to create a platform where international immunologists have the chance to discuss with those directly working in the field some of the most important immunologically mediated diseases that have high prevalence in this continent. In sub Saharan Africa, the world’s major infectious diseases have found a safe haven, causing untold human suffering, death, and economic devastation. The big three infectious diseases, HIV/AIDS, tuberculosis, and malaria, continue to be the commonest causes of illness and death. In addition to these, are diarrheal illnesses and respiratory infections other than tuberculosis. Furthermore, the prevalence rates of asthma, rhinitis, and allergic diseases are also increasing exponentially in this area, and traditional tropical infections such as schistosomiasis have not gone away. New infections including the pandemic influenza infections add to the disease burdens of these countries whose human and financial resources are limited. The thinking behind the EFIS-EJI AICI was to bring leading immunologists of the world together with health personnel who deal with the effects of these infectious diseases in order to create a platform that allowed the participants to appreciate the challenges and the opportunities that can accrue from juxtaposing the human aspects, the research and perhaps the financial capabilities of the established, usually northern and western, centers with those of the heavily disease-burdened centers in the South. Already at the first AICI, which was held in 2007, it turned out that this mix, and the reciprocal

In-print of the environment on the molecular sensitisation profile towards pollen allergens revealed by allergen micro-array

Background The green area in Tashkent-city in Uzbekistan has been re-organized during the last two decades. It covers approximately 35% of the total area and includes classical vegetation present in Central Asia such as saltwort and newly planted species such as Bermuda grass. The knowledge of the molecular sensitization profiles of allergic patients is essential for the correct treatment with allergen specific immunotherapy but has not been established for allergic patients in the Central Asian area. Objective The aim of this study was to determine the IgE-sensitization profile towards pollen allergens in patients with respiratory allergy from Tashkent. Methods Fifty adult patients with allergic rhinitis and/or asthma were tested using an allergen micro-array containing 112 different allergen molecules (ImmunoCAP ISAC; Thermo Scientific). Results We found that the major Bermuda grass pollen allergen, Cyn d 1 and the major saltwort allergen Sal k 1 were most frequently recognized allergens. More than 42% of the patients displayed IgE reactivity to Cyn d 1 and the percentage was even higher in the group suffering from allergic asthma (n=12) (i.e., 66%). Sal k 1 was recognized by 46% of the patients and by 50% of the patients with asthma. The other pollen allergens were recognized less frequently. Interestingly, more than 20% of patients showed IgE cross-reactivity with profilins from grass pollen, weed pollen and birch pollen whereas no patient mounted IgE reactivity to the major birch pollen allergen, Bet v 1. Conclusion The results of the IgE profiling identify grass pollen and in particular Bermuda grass and salkwort as the most important pollen allergen sources in Tashkent. Although approximately 20% of the patients reacted with birch pollen profilin, none of the patients reacted with the major birch pollen allergen, Bet v 1, indicating that these patients had no genuine sensitization to birch. These data show that the molecular sensitization profile towards pollen allergens in Central Asia is an in-print of the local flora and indicates how important allergen micro-array analysis is for the selection of the correct immunotherapy treatment. This study was supported by the Austrian Science Fund (FWF), American Austrian Foundation (AAF) and was performed in the framework of International Network University for Molecular Allergology and Immunology.