Major Allergen Content in Allergen Immunotherapy Products: The Limited Value of Numbers.

The prevalence of allergic disorders has increased drastically over the last 50 years to the extent that they can be considered epidemic. At present, allergen-specific immunotherapy (AIT) is the only therapy that targets the underlying cause of allergic disorders, and evidence of its superiority is based on data accumulated from clinical trials and observational studies demonstrating efficacy and safety. However, several aspects remain unresolved, such as harmonization and standardization of manufacturing and quantification procedures across manufacturers, homogeneous reporting of strength, and the establishment of international reference standards for many allergens. This article discusses issues related to the measurement of major allergen content in AIT extracts, raising the question of whether comparison of products from different manufacturers is an appropriate basis for selecting a specific AIT product. Allergen standardization in immunotherapy products is critical for ensuring quality and, thereby, safety and efficacy. However, lack of harmonization in manufacturing processes, allergen quantification (methodologies and references), national regulatory differences, clinical practice, and labeling shows that the comparison of AIT products based solely on major allergen amounts is not rational and, in fact, impossible. Moreover, when rating the information given for a specific product, it is necessary to take into account further inherent characteristics of products and their application in clinical practice, such as the state of extract modification, addition of adjuvant or adjuvant system, route of administration (sublingual/ subcutaneous), and cumulative dose as per posology (including the volume per administration). Finally, only convincing clinical data can serve as the basis for product-specific evaluation and cross-product comparability of individual products.

Major Allergen Content in Allergen Immunotherapy Products: The Limited Value of Numbers

The prevalence of allergic disorders has increased drastically over the last 50 years to the extent that they can be considered epidemic.At present, allergen-specific immunotherapy (AIT) is the only therapy that targets the underlying cause of allergic disorders, and evidenceof its superiority is based on data accumulated from clinical trials and observational studies demonstrating efficacy and safety. However,several aspects remain unresolved, such as harmonization and standardization of manufacturing and quantification procedures acrossmanufacturers, homogeneous reporting of strength, and the establishment of international reference standards for many allergens. Thisarticle discusses issues related to the measurement of major allergen content in AIT extracts, raising the question of whether comparison ofproducts from different manufacturers is an appropriate basis for selecting a specific AIT product. Allergen standardization in immunotherapyproducts is critical for ensuring quality and, thereby, safety and efficacy. However, lack of harmonization in manufacturing processes,allergen quantification (methodologies and references), national regulatory differences, clinical practice, and labeling shows that thecomparison of AIT products based solely on major allergen amounts is not rational and, in fact, impossible. Moreover, when rating theinformation given for a specific product, it is necessary to take into account further inherent characteristics of products and their applicationin clinical practice, such as the state of extract modification, addition of adjuvant or adjuvant system, route of administration (sublingual/subcutaneous), and cumulative dose as per posology (including the volume per administration). Finally, only convincing clinical data canserve as the basis for product-specific evaluation and cross-product comparability of individual products. (AU)

La prevalencia de las enfermedades alérgicas se ha incrementado drásticamente en los últimos 50 años y hoy pueden considerarse unaepidemia. Actualmente, la inmunoterapia específica con alérgenos (ITA) es el único tratamiento dirigido a la causa subyacente de lasenfermedades alérgicas y su superioridad se basa en resultados de ensayos clínicos/estudios observacionales que demuestran su eficaciay seguridad. Pero quedan aspectos sin resolver, como la armonización y estandarización de los procesos de fabricación y cuantificaciónentre fabricantes, la declaración homogénea de la potencia y el establecimiento de estándares internacionales de referencia. En este artículo se discuten aspectos relacionados con la medida del contenido de alérgenos mayores en los extractos de ITA, cuestionandosi, como base para elegir entre productos, es apropiada la comparación entre diferentes fabricantes. La estandarización alergénica escrucial para asegurar la calidad y, por tanto, la seguridad y eficacia de la ITA. Sin embargo, la falta de armonización en los procesos defabricación, la cuantificación alergénica, las diferencias regulatorias, la práctica clínica y el etiquetado, demuestran que comparar productosbasándose únicamente en la cantidad de alérgeno mayor no está justificado y es imposible. Además, cuando se evalúa la informaciónpara un determinado producto, deben tenerse en cuenta las características propias de cada producto y su uso clínico, como el estado dela modificación del extracto, la adición de adyuvantes, la vía de administración y la dosis acumulada. Solo datos clínicos convincentesdeben servir para la evaluación específica de cada producto o como base para la comparación entre productos. (AU)

Allergen-specific immunotherapy: is it vaccination against toxins after all?

IgE-mediated allergies, in particular allergic rhinoconjunctivitis and asthma, have reached epidemic proportions, affecting about one-third of the population in developed countries. The most effective treatment for allergies is specific immunotherapy (SIT), which involves the injection of increasing doses of an allergen extract to allergic individuals. The current form of SIT was first introduced in 1911 and recently celebrated its 100th birthday for the treatment of hay fever. The concept of this therapy at the time was straightforward, as it was believed that pollen contained toxins against which the patient could be vaccinated. However, the understanding became blurred with the discovery that IgE antibodies were the effector molecules of the allergic response. Subsequent research focused on the idea that SIT should induce tolerance keeping the IgE antibodies at bay. In this review, we will discuss the various hypotheses for the mechanism of SIT and we will put forward the concept that allergens may be viewed as 'protoxins' which need to be activated by IgE antibodies. Within this framework, protoxin-neutralizing antibodies are the key effector molecules while a shift to Th1 or Treg cells mainly contributes to the efficacy of SIT by helping B cells to produce neutralizing IgG antibodies.

IgG4 but no IgG1 antibody production after intralymphatic immunotherapy with recombinant MAT-Feld1 in human.

Allergy immunotherapy (AIT) mediates protection against allergen exposure in part due to allergen-specific antibodies. While immunization typically stimulated IgG1 and IgG2, AIT is often associated with production of IgG4. Here, twenty cat dander-sensitized patients were randomized to receive three injections of intralymphatic immunotherapy (ILIT) with MAT-Feld1 adsorbed to aluminum hydroxide or just aluminum hydroxide (placebo) in a double-blind setting (ClinicalTrials.gov NCT00718679). Whereas the clinical data, showing benefit of Mat-Feld1 ILIT was published in 2012 (Senti et al., J Allergy Clin Immunol, vol 129(5):1290-1296), the current study investigated the cat allergen-specific antibody responses. Blood was drawn prior to ILIT, as well as 1, 3, and 12 months after first ILIT. The sera were analyzed to characterize all IgG subclasses and IgE antibody responses. ILIT with MAT-Feld1 elicited high levels of total IgG that were maintained for at least 12 months. Interestingly, a strong increase in IgG4 and some increase in IgG2 were observed throughout the study, while production of cat-specific IgG1 and IgG3 was not stimulated by MAT-Feld1 ILIT. The IgE levels remained constant.

Determinants of efficacy and safety in epicutaneous allergen immunotherapy: summary of three clinical trials.

The results of our third trial on epicutaneous allergen-specific immunotherapy (EPIT) will be presented and discussed in the context of our previous trials. This monocentric, placebo-controlled, double-blind phase I/IIa trial included 98 patients with grass pollen rhinoconjunctivitis. Prior to the pollen season 2009, patients received six patches (allergen extract: n = 48; placebo: n = 50) with weekly intervals, administered onto tape-stripped skin. Allergen EPIT produced a median symptom improvement of 48% in 2009 and 40% in the treatment-free follow-up year 2010 as compared to 10% and 15% improvement after placebo EPIT (P = 0.003). After allergen EPIT but not placebo EPIT, conjunctival allergen reactivity was significantly decreased and allergen-specific IgG4 responses were significantly elevated (P < 0.001). In conclusion, our three EPIT trials found that allergen EPIT can ameliorate hay fever symptoms. Overall, treatment efficacy appears to be determined by the allergen dose. Local side-effects are determined by the duration of patch administration, while risk of systemic allergic side-effects is related to the degree of stratum corneum disruption.

Immune regulation by intralymphatic immunotherapy with modular allergen translocation MAT vaccine.

BACKGROUND: Allergen-specific immunotherapy (SIT) faces problems related to side effects and limited efficacy. Direct administration of allergen extracts into lymph nodes induces increased specific IgG production and T-cell responses using significantly lower allergen doses. METHODS: In this study, mechanisms of immune regulation by MAT vaccines in vitro and in allergen-SIT of cat-allergic rhinitis patients, who received 3 inguinal intra-lymph node injections of MAT-Fel d 1 vaccine, were investigated in PBMC and cell cultures for specific T-cell proliferation, Fel d 1-tetramer-specific responses, and multiple immune regulatory molecules. RESULTS: MAT-Fel d 1 vaccine was efficiently internalized by antigen-presenting cells. This was followed by precaspase 1 cleavage to caspase 1 and secretion of IL-1ß, indicating inflammasome activation. Mat-Fel d 1 induced specific T-cell proliferation and an IL-10- and IFN-γ-dominated T-cell responses with decreased Th2 cytokines at 100 times lower doses than Fel d 1. Induction of immune tolerance by MAT-Fel d 1-ILIT involved multiple mechanisms of immune suppression. Early Fel d 1-specific T-cell activation was followed by full T-cell unresponsiveness to allergen after 1 year in the MAT-Fel d 1 group, characterized by increased allergen-specific T regulatory cells, decreased circulating Fel d 1 tetramer-positive cells, increased IL-10 and FOXP3 expression, and change in the HR2/HR1 ratio toward HR2. CONCLUSIONS: This study demonstrates the induction of allergen tolerance after 3 intra-lymph node injections of MAT-Fel d 1 vaccine, mediated by increased cellular internalization of the allergen, activation of inflammasome, and generation of allergen-specific peripheral T-cell tolerance.

A critical appraisal of analyzing nasal provocation test results in allergen immunotherapy trials.

BACKGROUND: The statistical analysis of nasal provocation tests is very complex. We compared the conventional analysis with the maximally selected test statistics and the hierarchical ordered logistic model. METHODS: We re-analyzed data from a trial with 112 patients suffering from grass pollen allergy. The patients had been randomized to receive either intralymphatic immunotherapy (ILIT) or subcutaneous immunotherapy (SCIT). RESULTS: The conventional analysis indicated that the logarithmized ratio between the pre- and the post-treatment threshold concentration was significantly lower for ILIT than for SCIT. The maximally selected test statistics was used to test different threshold symptom scores that would imply positive clinical symptoms at the given allergen concentration. A threshold score of 3 maximised the difference in improvement between the ILIT and the SCIT groups. The hierarchical ordered logistic model does not take threshold allergen concentrations as the basis for analysis, but the single scores measured at each concentration. This approach simultaneously considers the treatment effect (ILIT versus SCIT), the time effect (pre- versus post-treatment), and the dose effect (different allergen concentrations). The hierarchical ordered logistic model revealed that the clinical improvement was greater after ILIT than after SCIT. CONCLUSION: As the choice of method can affect the outcome, guidelines for analysis are highly needed.

Multivalent paediatric allergy vaccines protect against allergic anaphylaxis in mice.

BACKGROUND: Almost a quarter of the world population suffers from IgE-mediated allergies. T cells and IgG-producing B cells can produce protection, but treatment for disease is laborious with unsatisfactory patient compliance. OBJECTIVE: We sought to identify whether paediatric allergy vaccines affected later allergen sensitization and onset of disease when used prophylactically. METHODS: A murine model of anaphylaxis was applied. Mice were first immunized with monovalent or multivalent allergy vaccines that also contained aluminium hydroxide and CpG oligodeoxynucleotide as adjuvants. Later, the mice were sensitized by multiple low-dose injections of aluminium-adsorbed allergen. After a dormant period, the mice were challenged systemically with high-dose allergen, and the clinical signs of anaphylaxis were recorded. Throughout the immunization and sensitization periods, blood was collected for serological testing. RESULTS: Immunization with allergy vaccines produced antigen-specific protection against sensitization as measured by systemic anaphylaxis in mice. The long-term effect was observed both after juvenile (5-6 weeks) and neonatal (7 days) vaccination. Monovalent and pentavalent vaccines were protective to a similar level. Protection was associated with increased secretion of IgG2a and production of IFN-γ. Protection could also be transferred to sensitized mice via serum or via CD25-positive CD4 T cells. CONCLUSION AND CLINICAL RELEVANCE: Prophylactic and multivalent allergy vaccines in juvenile and neonatal mice protected against later sensitization and anaphylaxis. Such treatment may provide a rational measure for future management of allergen-related diseases and their strong socio-economic impact on daily life.

IgG-mediated down-regulation of IgE bound to mast cells: a potential novel mechanism of allergen-specific desensitization.

BACKGROUND: Allergen-specific IgGs are known to inhibit IgE-mediated mast cell degranulation by two mechanisms, allergen-neutralization and engagement of the inhibitory FcγRIIB recruiting the phosphatase SHIP-1. Here we unravel an additional mechanism of IgG-mediated mast cell desensitization in mice: down-regulation of allergen-specific IgE. METHODS: Mast cells were loaded in vitro and in vivo with monoclonal IgE antibodies specific for Fel d1 and exposed to immune complexes consisting of Fel d1-specific IgG antibodies recognizing different epitopes. Down regulation of IgE was followed by flow cytometry. RESULTS: Mast cells loaded with 2 different IgE antibodies efficiently internalized the IgE antibodies if exposed to recombinant Feld d1. In contrast, no down-regulation occurred if mast cells were loaded with IgE antibodies exhibiting a single specificity before stimulation with recombinant Fel d1 [corrected]. Interestingly, however, IgEs of a single specificity were rapidly down-regulated in vitro and in vivo in the presence of Fel d1-specific monoclonal IgGs recognizing another epitope on Fel d1. Despite FceRI-internalization, little calcium flux or mast cell degranulation occurred. FcγRIIB played a dual role in the process since it enhanced IgE internalization and prevented cellular activation as documented by the inhibited calcium flux and mast cell degranulation. Similar observations were made in the presence of low concentrations of IgEs recognizing several epitopes on Fel d1. CONCLUSION: We demonstrate here that Fel d1-specific IgG antibodies interact with FcγRIIB which (i) promotes IgE internalization; and (ii) inhibits mast cell activation. These results broaden our understanding of allergen-specific desensitization and may provide a mechanism for long-term desensitization of mast cells by selective removal of long-lived IgE antibodies on mast cells.

[Immunotherapy of allergic rhinitis without allergens? : new options for immunomodulation by vaccination with virus-like particles and CpG motifs]. / Immuntherapie der allergischen Rhinitis ohne Allergene? : Neue Möglichkeiten einer Immunmodulation durch Vakzinierung mittels "virus-like particles" und CpG-Motiven.

BACKGROUND: Allergen-specific immunotherapy is generally accepted as the only causal therapy for allergic rhinitis. Up to now there has been a dogma in immunotherapy for inhalation allergies that at least allergen components are necessary for effective therapy. This dogma could, however, be swayed by current results of effective immunotherapy without allergens. Virus-like particles (VLP) represent a novel and interesting aspect for immunotherapy for inhalation allergies. AIM: Initial experiences with successful immunotherapy without allergens are available. This article describes the currently available clinical experiences with bacteriophage VLPs filled with oligonucleotides which contain CpG motifs with tumor antigens on the surface for the treatment of allergic rhinitis. RESULTS: Vaccination with VLPs was found to be well tolerated, immunogenic and effective for prophylactic treatment of various infections. Bacteriophage VLPs filled with CpG motifs with tumor antigens on the surface led clinically to an induction of specific T-cells in tumor patients and were successfully implemented as adjuvants during prophylactic vaccinations. CONCLUSION: The VLP technique in combination with the use of CpG motifs could contribute to the causal treatment of complex diseases, such as malignancies, autoimmune diseases and allergies.

The contact sensitizer diphenylcyclopropenone has adjuvant properties in mice and potential application in epicutaneous immunotherapy.

BACKGROUND: Epicutaneous vaccination has gained increasing interest during the past decade as it offers a safe, needle-free, and patient-friendly alternative to invasive vaccine administrations. Recently, the safety and early efficacy of epicutaneous immunotherapy were also demonstrated in patients with hay fever, as an alternative to conventional subcutaneous allergen-specific immunotherapy (SCIT). One major challenge to epicutaneous vaccination is the barrier function of the stratum corneum, which must be overcome either by abrasive methods or by hydration. Such barrier function of the stratum corneum also hampers the use of common adjuvants used to enhance the efficacy of vaccination. METHODS: In a mouse model of allergy, we tested the adjuvant potential of diphenylcyclopropenone (DCP), a strong contact sensitizer, which is currently used for the treatment of a T cell-mediated hair loss disease (alopezia areata). RESULTS: Diphenylcyclopropenone enhanced antigen-specific IgG2a antibody responses as well as IL-10 cytokine production after epicutaneous immunization with ovalbumin (OVA). Epicutaneous allergen-specific immunotherapy (EPIT) with OVA and DCP also protected sensitized mice from anaphylaxis and asthma. The protective effect was more robust than that of conventional SCIT, which did not significantly alleviate the symptoms of allergy in the murine models of anaphylaxis and asthma. CONCLUSIONS: This preclinical study confirmed previous clinical data that have demonstrated the potential of the skin as a target for allergen immunotherapy. The study also suggests that epicutaneous immunization or immunotherapy can be improved when an appropriate adjuvant such as DCP is used.

Immunotherapy concepts under investigation.

Allergen-specific immunotherapy (SIT) is effective, but used by only 5% of allergy patients, partly because it requires several years. Intralymphatic immunotherapy (ILIT) administers allergen directly into a subcutaneous lymph node and requires only three injections and a lower allergen dose than subcutaneous administration. ILIT was further improved using a recombinant allergen with enhanced uptake and decreased degradation by antigen-presenting cells. Another interesting route is transcutaneous/epicutaneous immunotherapy (TCIT, EPIT), which administers allergen by a skin patch. Clinical trials show that TCIT, EPIT is safe and comparably effective to conventional immunotherapy. Implementation of these new methods would increase the spectrum of SIT options, allowing greater individual choice of SIT.

[Allergen specific immunotherapy: new approaches]. / Allergenspezifische Immuntherapie: Auf neuen Wegen in die Zukunft.

Specific immunotherapy (SIT) is the only disease-modifying and causal treatment of IgE mediated allergic diseases. Soon this treatment will turn 100 years old. Subcutaneous immunotherapy is still considered to be the gold standard of SIT. With the intention to improve efficacy, safety and desirability for patients, new strategies such as epicutaneous immunotherapy, i.e. administration of allergens using a skin patch, are under investigation in clinical trials at the Zurich University Hospital.

Use of A-type CpG oligodeoxynucleotides as an adjuvant in allergen-specific immunotherapy in humans: a phase I/IIa clinical trial.

BACKGROUND: B-type CpG oligodeoxynucleotides (ODN) is currently used in clinical trials because of its prolonged half-life, which is due to its phosphorothioate backbone. A-type CpG ODN is a stronger inducer of IFN but has an unstable phosphodiester backbone that has so far prohibited its clinical use. However, upon association with virus-like particles (VLP) consisting of the bacteriophage Qbeta coat protein, A-type CpG ODN can be stabilized and can become an efficient adjuvant in mice. Therefore, the phase I/IIa study presented represents the first test of A-type CpGs in humans. OBJECTIVE: To test the safety, tolerability and clinical efficacy of QbG10 as an adjuvant for subcutaneous immunotherapy with a house dust mite (HDM) allergen extract in allergic patients. METHODS: A single centre, open-label phase I/IIa study evaluated the safety, tolerability and clinical efficacy of QbG10 as an adjuvant to immunotherapy with a subcutaneous HMD allergen extract in 20 patients suffering from HDM allergy. Twenty-one patients were enrolled between March and July 2005. Individual immunotherapy lasted 10 weeks. Clinical end-points included questionnaires, conjunctival provocation, skin prick tests and the measurement of allergen-specific IgG and IgE. RESULTS: QbG10 was well tolerated. Almost complete tolerance to the allergen was observed in conjunctival provocation testing after treatment with QbG10, and symptoms of rhinitis and allergic asthma were significantly reduced. Within 10 weeks of therapy, patients were nearly symptom-free and this amelioration lasted for at least 38 weeks post-treatment. Following injections of QbG10 and HDM allergen extract, allergen-specific IgG increased, while there was a transient increase in allergen-specific IgE titres. Skin reactivity to HDM was reduced. CONCLUSION: The subcutaneous application of HDM allergen, together with A-type CpG ODN packaged into VLP, was safe. All patients achieved practically complete alleviation of allergy symptoms after 10 weeks of immunotherapy. This promising clinical outcome calls for larger placebo-controlled phase II studies.

Targeting the MHC class II pathway of antigen presentation enhances immunogenicity and safety of allergen immunotherapy.

BACKGROUND: Current s.c. allergen-specific immunotherapy (SIT) leads to amelioration of IgE-mediated allergy, but it requires numerous allergen injections over several years and is frequently associated with severe side-effects. The aim of this study was to test whether modified recombinant allergens can improve therapeutic efficacy in SIT while reducing allergic side-effects. METHODS: The major cat allergen Fel d 1 was fused to a TAT-derived protein translocation domain and to a truncated invariant chain for targeting the MHC class II pathway (MAT-Fel d 1). The immunogenicity was evaluated in mice, while potential safety issues were assessed by cellular antigen stimulation test (CAST) using basophils from cat-dander-allergic patients. RESULTS: MAT-Fel d 1 enhanced induction of Fel d 1-specific IgG2a antibody responses as well as the secretion of IFN-gamma and IL-2 from T cells. Subcutaneous allergen-specific immunotherapy of mice using the modified Fel d 1 provided stronger protection against anaphylaxis than SIT with unmodified Fel d 1, and MAT-Fel d 1 caused less degranulation of human basophils than native Fel d 1. CONCLUSION: MAT-Fel d 1 allergen enhanced protective antibody and Th1 responses in mice, while reducing human basophil degranulation. Immunotherapy using MAT-Fel d 1 allergen therefore has the potential to enhance SIT efficacy and safety, thus, shortening SIT. This should increase patient compliance and lower treatment costs.

Medication with antihistamines impairs allergen-specific immunotherapy in mice.

BACKGROUND: Histamine released from activated mast cells and basophils is an important mediator in allergy. Therefore, antihistamines are efficiently and widely used to suppress allergic symptoms. OBJECTIVE: This study evaluated the role of antihistamines in sensitization against allergens and in the efficiency of allergen-specific immunotherapy. METHODS: CBA mice were sensitized and de-sensitized with bee venom allergen extracts and the major allergen phospholipase A2. Clemastine was used to test the effect of a histamine-1 receptor antagonist on the immune responses to phospholipase A2. RESULTS: The results demonstrated that sensitization against bee venom was strongly enhanced during treatment with antihistamines. Clemastine increased IgE production while decreasing IgG2a production against bee venom. This T-helper type 2 shift of the humoral response appeared to be caused by reduced IFN-gamma and enhanced IL-4 secretion from allergen-specific T cells. We also found reduced TNF-alpha, IL-6 and major histocompatibility complex class-II expression by macrophages. In sensitized mice, the efficiency of allergen-specific immunotherapy was reduced by clemastine treatment. CONCLUSION: Antihistamines may enhance allergic sensitization and reduce the efficiency of allergen-specific immunotherapy. Future studies will need to demonstrate to what extent pre-medication with antihistamine also affects allergen-specific immunotherapy in humans.

Analysis of the relationship between pollinosis and date of birth in Switzerland.

BACKGROUND: The first months of life may represent a vulnerable period in the development of atopic diseases. The objective of this study was to examine the relationship between the month of birth and the development of birch and grass pollen allergy in the Swiss population. METHODS: Data from the Swiss Study on Air Pollution and Lung Diseases in Adults(SAPALDIA) as well as the Swiss Study on Childhood Allergy and Respiratory Symptoms with Respect to Air Pollution and Climate (SCARPOL) were used. A logistic regression was calculated with grass and birch pollen sensitisation (positive skin prick test) or allergy (positive skin prick test and allergic symptoms) as outcome variables and the season of birth as predictor variable. The contribution of the season of birth on pollinosis was further adjusted for well-known risk factors and potential confounding variables. RESULTS: The logistic regression revealed a significant effect of the season of birth on birch pollen sensitisation and an effect of borderline significance on birch pollen allergy, i.e. subjects born in the pollen season (March to April) showed an increased risk of being sensitised/allergic to birch pollen. The results also indicated a tendency towards an increased risk for subjects born in the grass pollen season (May to June) to develop grass pollen allergy. CONCLUSION: Our results support the hypothesis that the first few months of life constitute a sensitive period, during which inhalative exposure to certain allergens may predispose to the subsequent development of atopic respiratory disease.