Grass Pollen Allergens Research Articles

Altered chromatin landscape in circulating T follicular helper and regulatory cells following grass pollen subcutaneous and sublingual immunotherapy

Allergen-specific immunotherapy is a disease-modifying treatment that induces long-term T-cell tolerance. We sought to evaluate the role of circulating CXCR5+PD-1+ T follicular helper (cTFH) and T follicular regulatory (TFR) cells following grass pollen subcutaneous immunotherapy (SCIT) and sublingual immunotherapy (SLIT) and the accompanying changes in their chromatin landscape. Phenotype and function of cTFH cells were initially evaluated in the grass pollen-allergic (GPA) group (n= 28) and nonatopic healthy controls (NAC, n= 13) by mathematical algorithms developed to manage high-dimensional data and cell culture, respectively. cTFH and TFR cells were further enumerated in NAC (n= 12), GPA (n= 14), SCIT- (n= 10), and SLIT- (n= 8) treated groups. Chromatin accessibility in cTFH and TFR cells was assessed by assay for transposase-accessible chromatin sequencing (ATAC-seq) to investigate epigenetic mechanisms underlying the differences between NAC, GPA, SCIT, and SLIT groups. cTFH cells were shown to be distinct from TH2- and TH2A-cell subsets, capable of secreting IL-4 and IL-21. Both cytokines synergistically promoted B-cell class switching to IgE and plasma cell differentiation. Grass pollen allergen induced cTFH-cell proliferation in the GPA group but not in the NAC group (P< .05). cTFH cells were higher in the GPA group compared with the NAC group and were lower in the SCIT and SLIT groups (P< .01). Time-dependent induction of IL-4, IL-21, and IL-6 was observed in nasal mucosa following intranasal allergen challenge in the GPA group but not in SCIT and SLIT groups. TFR and IL-10+ cTFH cells were induced in SCIT and SLIT groups (all, P< .01). ATAC-seq analyses revealed differentially accessible chromatin regions in all groups. For the first time, we showed dysregulation of cTFH cells in the GPA group compared to NAC, SCIT, and SLIT groups and induction of TFR and IL-10+ cTFH cells following SCIT and SLIT. Changes in the chromatin landscape were observed following allergen-specific immunotherapy in cTFH and TFR cells.

Towards middle-up analysis of polyclonal antibodies: subclass-specific N-glycosylation profiling of murine immunoglobulin G (IgG) by means of HPLC-MS

In recent years, advanced HPLC-MS strategies based on intact protein (“top-down”) or protein subunit (“middle-up/middle-down”) analysis have been implemented for the characterization of therapeutic monoclonal antibodies. Here, we assess feasibility of middle-up/middle-down analysis for polyclonal IgGs exhibiting extensive sequence variability. Specifically, we addressed IgGs from mouse, representing an important model system in immunological investigations. To obtain Fc/2 portions as conserved subunits of IgGs, we made use of the bacterial protease SpeB. For this purpose, we initially determined SpeB cleavage sites in murine IgGs. The resulting Fc/2 portions characteristic of different subclasses were subsequently analysed by ion-pair reversed-phase HPLC hyphenated to high-resolution mass spectrometry. This enabled simultaneous relative quantification of IgG subclasses and their N-glycosylation variants, both of which influence IgG effector functions. To assess method capabilities in an immunological context, we applied the analytical workflow to polyclonal antibodies obtained from BALB/c mice immunized with the grass pollen allergen Phl p 6. The study revealed a shift in IgG subclasses and Fc-glycosylation patterns in total and antigen-specific IgGs from different mouse cohorts, respectively. Eventually, Fc/2 characterization may reveal other protein modifications including oxidation, amino acid exchanges, and C-terminal lysine, and may thus be implemented for quality control of functional antibodies.

Identification of peroxidase-1 and beta-glucosidase as cross-reactive wheat allergens in grass pollen-related wheat allergy

BackgroundSome patients with wheat-dependent exercise-induced anaphylaxis (WDEIA) or wheat allergy showed negative ω-5 gliadin-specific IgE test and high level of grass pollen-specific IgE. It was presumed that these patients developed allergic reaction upon cross-reaction of their IgE antibodies raised against grass pollen allergens to wheat allergens. This study aimed to clarify clinical characteristics and wheat allergens of this phenotype of WDEIA/wheat allergy, which were tentatively diagnosed as grass pollen-related wheat allergy (GPWA). MethodsA total of six patients with GPWA were enrolled, and controls were 17 patients with grass pollen allergy but no episode of wheat allergy, and 29 patients with other wheat allergies: 18 with conventional WDEIA and 11 with hydrolyzed wheat protein allergy. Sensitization to wheat proteins was determined by basophil activation test (BAT). IgE-binding proteins in wheat flour were identified by immunoblotting followed by mass spectrometry. Wheat allergen-specific IgE tests were established by CAP-FEIA system. ResultsAll the six patients with GPWA were sensitized to water-soluble wheat proteins in BAT and IgE-immunoblotting, and peroxidase-1 (35 kDa) and beta-glucosidase (60 kDa) were identified as specific IgE-binding wheat proteins. The binding of patient IgE to these proteins was inhibited by pre-incubation of patient sera with grass pollen. The peroxidase-1- and beta-glucosidase-specific IgE tests identified three and four of six patients with GPWA, respectively, but only two of 29 controls, indicating high specificity of these tests. ConclusionsPeroxidase-1 and beta-glucosidase are specific wheat allergens for GPWA among grass pollen allergy and other types of wheat-induced food allergies.

Immunochemical and Physical Quantitation of Grass and Olive Pollen Allergens: Correlation With Asthma Admissions in Cáceres, Spain.

The association between pollen counts and allergen levels in the air is controversial. Objectives: The aims of the study were to quantify total and major allergen levels of Phleum pratense and Olea europaea and to analyze their correlation with grass and olive pollen counts and the number of asthma attacks attended at Complejo Hospitalario Universitario, Cáceres, Spain. A volumetric air sampler and a Burkard spore trap were used for pollen and aeroallergen collection during April- June 2011. Filters were extracted, and major allergens were quantified using enzyme-linked immunosorbent assay. May was the main grass pollination period, with a maximum peak of 1362 grains/m3 (May 13). The main pollination period for olive was April 30-May 20, with a maximum peak of 851 grains/m3 (May 11). A moderate correlation was observed between asthma exacerbations and grass pollen counts or Phleum total allergen levels; this became stronger when a 3-day offset was introduced. A significant association was observed between asthma exacerbations and total olive allergen or olive pollen grain levels when a 1-day offset was introduced. The maximum correlation (moderate-high) was observed 4 days and 6 days away from the maximum olive pollen peak and the maximum Ole e 1 peak level, respectively. This study reveals a significant correlation between grass and olive pollination and an increase in the number of visits to the emergency room for asthma attacks. The aerobiological pattern of allergen levels in the air is similar to that of pollen counts during the grass and olive pollination periods.

Polyvalent Mechanical Bacterial Lysate Administration Improves the Clinical Course of Grass Pollen–Induced Allergic Rhinitis in Children: A Randomized Controlled Trial

Recent studies highlight the immunoregulatory potential of bacterial lysates, indicating their potential use in the prevention and treatment of allergic diseases. To investigate the clinical efficacy of polyvalent mechanical bacterial lysates (PMBLs) in children with grass pollen-induced allergic rhinitis. Seventy children with seasonal allergic rhinitis were enrolled to this study and were randomly assigned to the PMBL and placebo groups. Severity of seasonal allergic rhinitis symptoms was assessed by the total nasal symptom score, total ocular symptom score, and visual analogue scale. During 3 visits, peak nasal inspiratory flow was measured, and nasal smears for the presence of eosinophils and nasal lavage fluids for the presence of allergen-specific IgE against timothy grass pollen allergens were sampled. A statistically significant decrease in total nasal symptom score (P= .001), total ocular symptom score (P=.04), and visual analogue scale score for nasal and eye symptoms (P < .001 and P < .001, respectively) and an increase in peak nasal inspiratory flow (P= .04) were observed in the PMBL group versus the placebo group. During the grass pollen season, an increase and then a decrease in the number of eosinophils in nasal smears was observed in both groups; however, the number of eosinophils was significantly lower in the PMBL group versus the placebo group. No significant changes in allergen-specific IgE concentrations were observed in the PMBL group, whereas in the placebo group a statistically significant increase in allergen-specific IgE concentration was observed. Sublingual administration of PMBLs during the grass pollen season offers significant efficacy in alleviating seasonal allergic rhinitis symptoms in children sensitized to grass pollen allergens. PMBLs probably affect mucosal immunity, weakening the response of TH2 cells.

In silico Design of Phl p 6 Variants With Altered Fold-Stability Significantly Impacts Antigen Processing, Immunogenicity and Immune Polarization.

Introduction: Understanding, which factors determine the immunogenicity and immune polarizing properties of proteins, is an important prerequisite for designing better vaccines and immunotherapeutics. While extrinsic immune modulatory factors such as pathogen associated molecular patterns are well-understood, far less is known about the contribution of protein inherent features. Protein fold-stability represents such an intrinsic feature contributing to immunogenicity and immune polarization by influencing the amount of peptide-MHC II complexes (pMHCII). Here, we investigated how modulation of the fold-stability of the grass pollen allergen Phl p 6 affects its ability to stimulate immune responses and T cell polarization.Methods: MAESTRO software was used for in silico prediction of stabilizing or destabilizing point mutations. Mutated proteins were expressed in E. coli, and their thermal stability and resistance to endolysosomal proteases was determined. Resulting peptides were analyzed by mass spectrometry. The structure of the most stable mutant protein was assessed by X-ray crystallography. We evaluated the capacity of the mutants to stimulate T cell proliferation in vitro, as well as antibody responses and T cell polarization in vivo in an adjuvant-free BALB/c mouse model.Results: In comparison to wild-type protein, stabilized or destabilized mutants displayed changes in thermal stability ranging from −5 to +14°. While highly stabilized mutants were degraded very slowly, destabilization led to faster proteolytic processing in vitro. This was confirmed in BMDCs, which processed and presented the immunodominant epitope from a destabilized mutant more efficiently compared to a highly stable mutant. In vivo, stabilization resulted in a shift in immune polarization from TH2 to TH1/TH17 as indicated by higher levels of IgG2a and increased secretion of TNF-α, IFN-γ, IL-17, and IL-21.Conclusion: MAESTRO software was very efficient in detecting single point mutations that increase or reduce fold-stability. Thermal stability correlated well with the speed of proteolytic degradation and presentation of peptides on the surface of dendritic cells in vitro. This change in processing kinetics significantly influenced the polarization of T cell responses in vivo. Modulating the fold-stability of proteins thus has the potential to optimize and polarize immune responses, which opens the door to more efficient design of molecular vaccines.

IL-31 transgenic mice show reduced allergen-induced lung inflammation.

Interleukin‐31 (IL‐31) is a Th2 cell–derived cytokine that has been closely linked to pruritic skin inflammation. More recently, enhanced IL‐31 serum levels have also been observed in patients with allergic rhinitis and allergic asthma. Therefore, the main aim of this study was to unravel the contribution of IL‐31 to allergen‐induced lung inflammation. We analyzed lung inflammation in response to the timothy grass (Phleum pratense) pollen allergen Phl p 5 in C57BL/6 wild‐type (wt) mice, IL‐31 transgenic (IL‐31tg) mice, and IL‐31 receptor alpha‐deficient animals (IL‐31RA−/−). IL‐31 and IL‐31RA levels were monitored by qRT‐PCR. Cellular infiltrate in bronchoalveolar lavage fluid (BALF) and lung tissue inflammation, mucus production as well as epithelial thickness were measured by flow cytometry and histomorphology. While allergen challenge induced IL‐31RA expression in lung tissue of wt and IL‐31tg mice, high IL‐31 expression was exclusively observed in lung tissue of IL‐31tg mice. Upon Phl p 5 challenge, IL‐31tg mice showed reduced numbers of leukocytes and eosinophils in BALF and lung tissue as well as diminished mucin expression and less pronounced epithelial thickening compared to IL‐31RA−/− or wt animals. These findings suggest that the IL‐31/IL‐31RA axis may regulate local, allergen‐induced inflammation in the lungs.

Meteorological factors driving airborne grass pollen concentration in central Iberian Peninsula

Grasses are a taxonomic group of considerable environmental importance, playing a major ecological role as well as making a significant contribution to the world’s economy, since they include valuable crop and pasture species. Grass pollen allergens are also among the main causes of respiratory allergies worldwide. The interpretation of airborne grass pollen concentrations is a particularly complex task, given the marked diversity of pollen-emitting species and the influence of weather-related variables. This paper examines the influence of meteorological variables on flowering in the species contributing most to airborne grass pollen concentrations. This study was carried out in the surroundings of the city of Toledo (Spain), a Mediterranean city located in the center of the Iberian Peninsula. Temperature was the variable most influencing flowering onset, which was also affected by relative humidity, rainfall and hours of sunlight. The flowering period of grass species that flower at the start of spring (Bromus rubens and Hordeum leporinum) began earlier in years with higher mean winter temperatures, while the species that flower from mid to late spring (Trisetaria panicea and Dactylis glomerata subsp. hispanica), the flowering period began earlier in years with higher cumulative rainfall in winter and spring, which were also the years with the highest mean temperatures. Research into the influence of weather-related variables on grass phenology can shed important light on variations in airborne pollen concentrations, which determine the potential period of exposure for allergy sufferers.

Cost-effectiveness of grass pollen allergen immunotherapy in adults.

Major scientific societies, such as the EAACI or the AAAAI, do not express any suggestion on which form of allergen immunotherapy (AIT) is to be preferred (subcutaneous immunotherapy, SCIT, vs sublingual immunotherapy, SLIT). This choice could depend on their relative pharmacoeconomic value. To assess the cost-effectiveness of AIT for grass pollen, administered as SCIT or SLIT. We created a Markovian Model, to evaluate, in a hypothetical cohort of adult patients suffering from moderate-to-severe rhino-conjunctivitis with or without allergic asthma, the cost-effectiveness of SLIT (tablets, Grazax® and Oralair® ) or SCIT (various currently available products, plus indirect nonmedical costs, such as travel and productivity costs) in addition to pharmacological therapy, assuming a 9-year horizon to capture AIT long-term effects. The incremental cost-effectiveness ratio (ICER) was calculated assuming pharmacological therapy as the reference comparator. In the base case, SCIT was slightly more expensive, but more effective than SLIT, being the most cost-effective option (ICER for SCIT, €11418; ICER for SLIT, €15212). ICERs greater than €120000 for both SCIT and SLIT were demonstrated in a scenario assuming that low treatment persistence rates, which are common in real-life, lead to absence of long-term AIT clinical benefit. Considering indirect nonmedical costs SLIT resulted more cost-effective than SCIT (ICER for SCIT, €17318; ICER for SLIT, €15212). In daily practice, AIT for grass pollens may be a cost-effective option only in patients with low discontinuation rates. SCIT, which is less affected by this limitation than SLIT, seems the most cost-effective AIT form.

Is grass pollen allergen immunotherapy cost effective?

Is grass pollen allergen immunotherapy cost effective?

Sensitization to grass pollen allergen molecules in a birth cohort—natural Phl p 4 as an early indicator of grass pollen allergy

Grass pollen allergy is one of the most common allergies worldwide. The aim of this study was to evaluate the usefulness of grass pollen allergen molecules for prediction of grass pollen allergy during childhood and up to adolescence. Questionnaire data and sera obtained from the study subjects at the ages of 4, 8, and 16 years from the population-based Barn/Children Allergy Milieu Stockholm Epidemiology birth cohort were used. Sera from 763 representative subjects with serum samples available at all 3 ages were analyzed for IgE reactivity to 8 Phleum pratense (Phl p) allergens (MeDALL [Mechanisms for the Development of Allergies] chip) and to timothy grass extract (ImmunoCAP). Allergic rhinitis to grass pollen (ARg) was defined as upper airway symptoms during grass pollen exposure. The prevalence of sensitization to any Phl p molecule was higher compared with that to timothy extract at all 3 ages: at the age of 4 years, 9.7% versus 6.8%; at the age of 8 years, 28.4% versus 15.3%; and at the age of 16 years, 37.1% versus 27.1%. General estimating equations analyses revealed that among children sensitized at the age of 4 years, the overall odds ratio (OR) of later ARg (up to 16 years) was increased only for IgE reactivity to Phl p 1 (OR= 4.9) and natural Phl p 4 (OR= 6.9). The likelihood of later symptoms increased with the number of allergen molecules; at the age of 4 years, 2 or more molecules predicted ARg to 78% and 3 or more molecules predicted ARg to 95%. Apositive test result for timothy extract predicted ARg to 70%. Natural Phl p 4 is a hitherto unrecognized early indicator of grass pollen allergy, in addition to Phl p 1. To identify grass pollen sensitization and predict later ARg, allergen molecules are of added value to timothy extract alone and may help clinicians improve prediction of grass pollen allergy.

Biogeographical variation in specific IgE recognition of temperate and subtropical grass pollen allergens in allergic rhinitis patients.

ObjectiveGlobally, grass pollens (GP) are major aeroallergen triggers of allergic rhinitis (AR) and asthma. However, patterns of allergic sensitisation to pollen of temperate (Pooideae: Lolium perenne) and subtropical (Chloridoideae: Cynodon dactylon and Panicoideae: Paspalum notatum) subfamilies in diverse climates remain unclear. This study aims to evaluate the level of allergic sensitisation and IgE specificity for major GP allergens representing the three subfamilies in biogeographically distinct regions.MethodsParticipants (GP‐allergic with AR, 330; non‐atopic, 29; other allergies, 54) were recruited in subtropical: Queensland, and temperate: New South Wales, Western and South Australia, regions. Clinical history, skin prick test (SPT), total and specific IgE to GP and purified allergens (ImmunoCAP) were evaluated. Cross‐inhibition of sIgE with Pas n 1, Cyn d 1 and Lol p 1 by GP extracts was investigated.ResultsQueensland participants showed higher sensitisation to P. notatum and C. dactylon than L. perenne GP. sIgE was higher to Pas n 1 and Cyn d 1, and sIgE to Pas n 1 and Cyn d 1 was inhibited more by Panicoideae and Chloridoideae, respectively, than Pooideae GP. Conversely, participants from temperate regions showed highest sensitisation levels to L. perenne GP and Lol p 1, and sIgE to Lol p 1 was inhibited more by Pooideae than other GP.ConclusionLevels and patterns of sensitisation to subtropical and temperate GP in AR patients depended on biogeography. Knowledge of the specificity of sensitisation to local allergens is important for optimal diagnosis and choice of allergen‐specific immunotherapy to maximise benefit.

Allergen-specific IgG+ memory B cells are temporally linked to IgE memory responses

IgE is the least abundant immunoglobulin and tightly regulated, and IgE-producing B cells are rare. The cellular origin and evolution of IgE responses are poorly understood. The cellular and clonal origin of IgE memory responses following mucosal allergen exposure by sublingual immunotherapy (SLIT) were investigated. In a randomized double-blind, placebo-controlled, time course SLIT study, PBMCs and nasal biopsy samples were collected from 40 adults with seasonal allergic rhinitis at baseline and at 4, 8, 16, 28, and 52 weeks. RNA was extracted from PBMCs, sorted B cells, and nasal biopsy samples for heavy chain variable gene repertoire sequencing. Moreover, mAbs were derived from single B-cell transcriptomes. Combining heavy chain variable gene repertoire sequencing and single-cell transcriptomics yielded direct evidence of a parallel boost of 2 clonally and functionally related B-cell subsets of short-lived IgE+ plasmablasts and IgG+ memory B cells. Mucosal grass pollen allergen exposure by SLIT resulted in highly diverse IgE and IgGE repertoires. These were extensively mutated and appeared relatively stable as per heavy chain isotype, somatic hypermutations, and clonal composition. Single IgGE+ memory B-cell and IgE+ preplasmablast transcriptomes encoded antibodies that were specific for major grass pollen allergens and able to elicit basophil activation at very low allergen concentrations. For the first time, we have shown that on mucosal allergen exposure, human IgE memory resides in allergen-specific IgG+ memory B cells. These cells rapidly switch isotype, expand into short-lived IgE+ plasmablasts, and serve as a potential target for therapeutic intervention.

TGFβ1 mimetic peptide modulates immune response to grass pollen allergens in mice.

BackgroundTransforming growth factor β1 (TGFβ1) is a cytokine that exerts immunosuppressive functions, as reflected by its ability to induce regulatory T (Treg) cell differentiation and inhibit Th1 and Th2 responses. Hence, peptides that mimic the active core domain of TGFβ1 may be promising candidates for modulation of the allergic response. This study aimed to investigate a synthetic TGFβ1 mimetic peptide (TGFβ1‐mim) for its ability to modulate the immune response during allergic sensitization to grass pollen allergens.MethodsThe in vitro action of TGFβ1‐mim was evaluated in human lung epithelial cells, Jurkat cells, and rat basophilic leukemia cells. The in vivo action was evaluated in a murine model of Phl p 5 allergic sensitization. Additionally, the Th2 modulatory response was evaluated in IL‐4 reporter mice.ResultsIn vitro, TGFβ1‐mim downregulated TNF‐α production, IL‐8 gene expression, and cytokine secretion, upregulated IL‐10 secretion, and inhibited Phl p 5‐induced basophil degranulation. During Phl p 5 sensitization in mice, TGFβ1‐mim downregulated IL‐2, IL‐4, IL‐5, IL‐13, and IFN‐γ, upregulated IL‐10, and induced Treg cell production. Furthermore, mice treated with TGFβ1‐mim had lower levels of IgE, IgG1, IgG2a and higher levels of IgA antibodies than control mice. In a reporter mouse, the mimetic inhibited Th2 polarization.ConclusionThe TGFβ1‐mim efficiently modulated various important events that exacerbate the allergic microenvironment, including the production of main cytokines that promote Th1 and Th2 differentiation, and the induction of allergen‐specific regulatory T cells, highlighting its potential use in therapeutic approaches to modulate the immune response toward environmental allergens.

Mast cells and γδ T cells are largely dispensable for adaptive immune responses after laser-mediated epicutaneous immunization

BackgroundThe skin resembles an attractive target for vaccination due to its accessibility and abundance of resident immune cells. Cells like γδ T cells and mast cells (MCs) are part of the first line of defence against exogenous threats. Despite being important mediators for eliciting TH2 immune responses after epithelial stress, γδ T cell and MC functions still remain to be completely understood. Here, we aimed to characterize their roles in shaping adaptive immune responses after laser-mediated epicutaneous immunization (EPI). Methodsγδ T cell knock out, MC-depleted, and wildtype control mice were immunized with mannan-conjugated grass pollen allergen Phl p 5 (P5-MN) by laser-mediated EPI. After 2–3 immunizations, cytokine expression, T helper polarization, and antigen-specific IgG1/IgE levels were analysed. Furthermore, the local cytokine/chemokine milieu after laser microporation was determined. ResultsThe majority of inflammatory chemokines and cytokines induced by laser treatment were not affected by the presence of γδ T cells or MCs. However, RANTES was elevated in γδ T cell knock out mice and GROα, TSLP, and IL-33 were significantly decreased after MC depletion. The absence of γδ T cells or depletion of MCs had no substantial effect on adaptive immune responses after laser-mediated EPI, except for slightly reduced IgG1 and effector T cell levels in MC-depleted mice. Conclusionsγδ T cells did not play a pivotal role in shaping the humoral and cellular adaptive immune response after laser-mediated EPI. MC depletion decreased the numbers of effector T cells, indicating a potential role of MCs in the activation and maturation of T cells after EPI.

Two years of treatment with the recombinant grass pollen allergy vaccine BM32 induces a continuously increasing allergen-specific IgG4 response.

BackgroundBM32, a grass pollen allergy vaccine containing four recombinant fusion proteins consisting of hepatitis B-derived PreS and hypoallergenic peptides from the major timothy grass pollen allergens adsorbed on aluminium hydroxide has been shown to be safe and to improve clinical symptoms of grass pollen allergy upon allergen-specific immunotherapy (AIT). We have investigated the immune responses in patients from a two years double-blind, placebo-controlled AIT field trial with BM32.MethodsBlood samples from patients treated with BM32 (n = 27) or placebo (Aluminium hydroxide) (n = 13) were obtained to study the effects of vaccination and natural allergen exposure on allergen-specific antibody, T cell and cytokine responses. Allergen-specific IgE, IgG, IgG1 and IgG4 levels were determined by ImmunoCAP and ELISA, respectively. Allergen-specific lymphocyte proliferation by 3H thymidine incorporation and multiple cytokine responses with a human 17-plex cytokine assay were studied in cultured peripheral blood mononuclear cells (PBMCs).FindingsTwo years AIT comprising two courses of 3 pre-seasonal injections of BM32 and a single booster after the first pollen season induced a continuously increasing (year 2 > year 1) allergen-specific IgG4 response without boosting allergen-specific IgE responses. Specific IgG4 responses were accompanied by low stimulation of allergen-specific PBMC responses. Increases of allergen-specific pro-inflammatory cytokine responses were absent. The rise of allergen-specific IgE induced by seasonal grass pollen exposure was partially blunted in BM32-treated patients.InterpretationAIT with BM32 is characterised by the induction of a non-inflammatory, continuously increasing allergen-specific IgG4 response (year 2 > year1) which may explain that clinical efficacy was higher in year 2 than in year 1. The good safety profile of BM32 may be explained by lack of IgE reactivity and low stimulation of allergen-specific T cell and cytokine responses.FundingsGrants F4605, F4613 and DK 1248-B13 of the Austrian Science Fund (FWF).

Grass pollen season in selected cities of Poland in 2019

The pollen of grass is the primary trigger of pollen allergies during the summer months. It causes some of the most severe and difficult-to-treat symptoms. During the flowering of grass, over 90% of patients allergic to grass pollen suffer from allergic conditions. The symptoms caused by grass pollen allergens appear in some patients after exceeding the concentration of 20 grains in m3 of air. The aim of this study was to compare Poaceae pollen seasons in 2019 in 11 cities located in different regions of Poland. Pollen monitoring was conducted in Bialystok, Bydgoszcz, Sosnowiec, Lublin, Olsztyn, Szczecin, Piotrkow Trybunalski, Warsaw, Opole, Zielona Gora and Wroclaw. Airborne pollen was monitored by the volumetric method using Burkard or Lanzoni pollen samplers. Pollen season length was determined by the 98% method, where the start of the season was defined as the date when 1% of the seasonal pollen total was trapped and the end of the season as the date, when 99% of the seasonal pollen total was reached.The pollen season start date was recorded earliest in Zielona Gora (April 27th), latest in Szczecin (May 25th). The highest grass pollen concentration values were found in Lublin, Piotrkow Trybunalski and Warsaw, whereas the lowest ones in Bialystok. The annual grass pollen sum was highest in Lublin (4715 pollen grains), whereas it was lowest in Bialystok (1256 pollen grains). A relatively high pollen count was also recorded in Warsaw (4106 pollen grains) and Zielona Gora (4077 pollen grains). The longest grass pollination season was recorded in Zielona Gora (139 days) and the shortest in Szczecin (110 days). In the remaining measuring points, grass pollen in the air maintained from 112 to 131 days. The highest grass pollen allergen risk occurred in Lublin, Olsztyn and Zielona Gora and the lowest in Bialystok.

Comparison of Six Different Allergen Extracts for Subcutaneous Specific Immunotherapy in Children: An Open-Labelled, Prospective, Controlled Observational Trial

Background: Numerous products are available for subcutaneous (SCIT) and sublingual allergen-specific immunotherapy, but there are no information about the direct comparability regarding efficacy, safety, and tolerability of the different extracts. Aims: The aim of this open-labelled, prospective, controlled observational trial was to test the feasibility of a comparison of different products for SCIT in children. Methods: Pediatrician practices recruited patients with a confirmed diagnosis of a seasonal allergic rhinoconjunctivitis (AR) with or without asthma and an allergic sensitization against grass pollen allergen. Every patient was offered SCIT with one out of six allergen extracts: ALK SQ Depot, ALK Avanz, Allergovit, Depigoid, Purethal, Pollinex Quattro. Scores for symptoms and medications were calculated and the difference between treatment years and baseline were recorded. Results: In total, 284 were recruited and 255 children (89.8%; mean age 10.4, SD 3.54 years; 65% males) participated in this trial. Overall, 49,649 patient days were recorded in the electronic database (mean 183.2 days/patient). There was no significant difference in the AR and asthma symptom score or the medication score between the six different SCIT preparations. Similarly, no differences were observed in terms of safety and tolerability. Conclusion: The comparison of different SCIT products using an online tool is feasible. Based on our preliminary data, all extracts indicated efficacy; however, larger groups would be necessary to demonstrate superiority or non-inferiority of one specific SCIT product.

Strong dose response after immunotherapy with PQ grass using conjunctival provocation testing

BackgroundPollinex Quattro Grass (PQ Grass) is an effective, well-tolerated, short pre-seasonal subcutaneous immunotherapy to treat seasonal allergic rhinoconjunctivitis (SAR) due to grass pollen. In this Phase II study, 4 cumulative doses of PQ Grass and placebo were evaluated to determine its optimal cumulative dose. MethodsPatients with grass pollen-induced SAR were randomised to either a cumulative dose of PQ Grass (5100, 14400, 27600 and 35600 SU) or placebo, administered as 6 weekly subcutaneous injections over 31–41 days (EudraCT number 2017-000333-31). Standardized conjunctival provocation tests (CPT) using grass pollen allergen extract were performed at screening, baseline and post-treatment to determine the total symptom score (TSS) assessed approximately 4 weeks after dosing. Three models were pre-defined (Emax, logistic, and linear in log-dose model) to evaluate a dose response relationship. ResultsIn total, 95.5% of the 447 randomized patients received all 6 injections. A highly statistically significant (p < 0.0001), monotonic dose response was observed for all three pre-specified models. All treatment groups showed a statistically significant decrease from baseline in TSS compared to placebo, with the largest decrease observed after 27600 SU (p < 0.0001). The full course of 6 injections was completed by 95.5% of patients. Treatment-emergent adverse events were similar across PQ Grass groups, and mostly mild and transient in nature. ConclusionsPQ Grass demonstrated a strong curvilinear dose response in TSS following CPT without compromising its safety profile.

Interplay between Affinity and Valency in Effector Cell Degranulation: A Model System with Polcalcin Allergens and Human Patient-Derived IgE Antibodies.

An allergic reaction is rapidly generated when allergens bind and cross-link IgE bound to its receptor FcεRI on effector cells, resulting in cell degranulation and release of proinflammatory mediators. The extent of effector cell activation is linked to allergen affinity, oligomeric state, valency, and spacing of IgE-binding epitopes on the allergen. Whereas most of these observations come from studies using synthetic allergens, in this study we have used Timothy grass pollen allergen Phl p 7 and birch pollen allergen Bet v 4 to study these effects. Despite the high homology of these polcalcin family allergens, Phl p 7 and Bet v 4 display different binding characteristics toward two human patient-derived polcalcin-specific IgE Abs. We have used native polcalcin dimers and engineered multimeric allergens to test the effects of affinity and oligomeric state on IgE binding and effector cell activation. Our results indicate that polcalcin multimers are required to stimulate high levels of effector cell degranulation when using the humanized RBL-SX38 cell model and that multivalency can overcome the need for high-affinity interactions.