Search

IgE sensitization to cockroach allergens is associated with development of allergic diseases, such as asthma. To understand the relevance of different cockroach allergens for diagnosis and immunotherapy, a comprehensive analysis of IgE antibody levels and T cell reactivity to an expanded set of cockroach allergens and their relationship to disease was performed in a cohort of USA cockroach sensitized patients. IgE antibody levels to recombinant chitinase and hemocyanin were measured for 23 subjects by custom-made ImmunoCAPs and compared with IgE levels to eight cockroach allergens we previously reported for the same cohort. Ex vivo T cell activation (Ox40/PDL-1 expression) of PBMCs stimulated with peptide pools derived from 11 German cockroach proteins, including nine official cockroach allergens, plus chitinase and vitellogenin, was determined by flow cytometry. IgE prevalences to chitinase (17%) and hemocyanin (44%) were comparable to values for the other eight allergens that we previously reported (21-57%). Hemocyanin (Bla g 3), was a major allergen (one to which more than 50% of patients with an allergy to its source react) for a sub-group of 15 highly cockroach-sensitized subjects (IgE > 3.5 kU A /L: 53%). Chitinase was officially named as new allergen Bla g 12. Cockroach-specific IgE levels in plasma showed excellent correlation with the sum of 10 allergen-specific IgE (r = 0.94, p < 0.001). T cell reactivity to 11 proteins was highly variable among subjects, the highest being for vitellogenin, followed by Bla g 3. The main finding was that cockroach allergen-specific IgE and T cell reactivity patterns were unique per subject, and lacked immunodominant allergens and correlation with clinical phenotype/disease severity in the studied cohort. Knowing the subject-specific B/T cell reactivity profiles to a comprehensive panel of cockroach allergens will contribute to diagnosis of cockroach allergy and will be important for planning and assessing allergen immunotherapy outcomes, according to the allergen content in therapeutic cockroach extracts., Competing Interests: AP is an employee of Indoor Biotechnologies, Inc. and the contact principal investigator of the NIH R01 Award that funded the study. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Pomés, Schulten, Glesner, da Silva Antunes, Sutherland, Bacharier, Beigelman, Busse, Frazier and Sette.)

Background: Allergy to German cockroach (CR) is common in urban environments and is an important allergen in children with asthma., Objective: We hypothesize that the evolution of allergic sensitization and clinical disease is associated with distinct patterns of allergen-specific T cell reactivity. To test this hypothesis, a subset of high-risk inner-city children participating in the URECA (Urban Environment and Childhood Asthma) birth cohort were selected to evaluate CR-specific T cell reactivity from three distinct groups based on acquisition of aeroallergen sensitivity from ages 2 to 10: low atopy with minimal to no sensitivity (n = 26), early-onset allergic sensitization (n = 25) and late-onset allergic sensitization (n = 25)., Methods: Using pools of previously identified CR-derived T cell epitopes, we characterized the allergen-specific T cell response in these 76 subjects from blood samples obtained at age 10. CR-specific production of IL-5, IFNγ and IL-10 was measured by ELISPOT following two-week in vitro culture with CR extract., Results: T cell responses were significantly higher in the early-onset atopy group compared to low atopy (P = 0.01), and a trend for higher cytokine production in the late onset compared to the low atopy cohort was also observed (P = 0.06). T cell responses were similar between early- and late-onset cohorts. Furthermore, a comparison of T cell reactivity between asthmatic and non-asthmatic individuals revealed significantly higher cytokine production in asthmatics compared to non-asthmatics (P = 0.02) within both the CR-allergic and non-allergic cohorts., Conclusions and Clinical Relevance: In conclusion, the present study reports that higher T cell reactivity is associated with allergen sensitization and asthma. Interestingly, no significant difference in T cell reactivity was observed in allergic children with early-onset versus late-onset atopy., (© 2019 John Wiley & Sons Ltd.)

Background: Cockroach allergens are an important cause of IgE-mediated sensitization in inner-city asthmatic patients. However, cockroach extracts used for diagnosis and immunotherapy are not standardized., Objective: We sought to determine the allergen content of nonstandardized German cockroach extracts and the levels of sensitization to an expanded set of cockroach allergens as determinants of in vitro extract potency for IgE reactivity., Methods: Twelve German cockroach extracts were compared for allergen content and potency of IgE reactivity. Bla g 1, Bla g 2, and Bla g 5 were measured by using immunoassays. IgE antibody levels to 8 purified recombinant allergens from groups 1, 2, 4, 5, 6, 7, 9, and 11 were measured by using ImmunoCAP. IgE antibody binding inhibition assays were performed to assess extract in vitro potencies (concentration inhibiting 30% of the total IgE antibody-binding inhibition) relative to an arbitrarily selected reference extract in 5 patients with cockroach allergy., Results: Allergen levels were highly variable. Three new major allergens (groups 6, 9, and 11), were identified among highly cockroach-sensitized subjects (CAP class ≥ 3). Sensitization profiles were unique per subject without immunodominant allergens. The sum of IgE to 8 allergen components showed a good correlation with cockroach-specific IgE levels (r = 0.88, P < .001). In vitro potencies varied among different extracts per subject and among subjects for each extract., Conclusions: The in vitro potency of German cockroach extracts for IgE reactivity depends on allergen content and allergen-specific IgE titers of patients with cockroach allergy. These factors are relevant for selection of potent extracts to be used for immunotherapy and for the design and interpretation of data from immunotherapy trials., (Copyright © 2018 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

German cockroach extract is used clinically to evaluate allergen-specific sensitization and for subcutaneous allergen-specific immunotherapy, though there are no guidelines for standardization in its manufacture. We performed an immunological evaluation of 12 different cockroach extracts prepared from different sources and their potency to induce allergen-specific T cell reactivity. PBMC from 13 cockroach allergic donors were expanded in vitro with 12 different German cockroach extracts. After culture expansion, cells were re-stimulated with the different extracts and T cell responses were assessed by FluoroSpot (IL-5, IFNγ and IL-10 production). In parallel to the extracts, single allergen peptide pools for allergens from groups 1, 2, 4, 5, and 11 were tested to determine allergen immunodominance. Furthermore, to assess allergy specificity, PBMC from 13 non-allergic donors were also tested with the most potent extract and T cell responses were compared to the allergic cohort. Dramatic variations in T cell reactivity were observed to the different cockroach extract batches. Response magnitudes varied over 3 logs within a single donor. IL-5 production in the allergic cohort was significantly higher compared to the non-allergic cohort (p=0.004). Allergen content determination by ELISA detected much lower concentrations of Bla g 5 compared to Bla g 1 and 2. Mass spectrometric analysis revealed that Bla g 5 was present in similar amounts to Bla g 1 and 2 in extracts made from whole body, whereas it was not detected in extracts made from fecal matter, suggesting that Bla g 5 is not excreted into feces. Different donors exhibit different response patterns to different extracts, potentially dependent on the donor-specific T cell allergen immunodominance pattern and the allergen content of the extract tested. These findings have dramatic implications for the selection of potent extracts used for diagnostic purposes or allergen-specific immunotherapy.

Background and Objective: The allergenicity of several German cockroach (Bla-g) antigens at the level of IgE responses is well established. However, less is known about the specificity of CD4+ TH responses, and whether differences exist in associated magnitude or cytokine profiles as a function of disease severity., Methods: Proteomic and transcriptomic techniques were used to identify novel antigens recognized by allergen-specific T cells. To characterize different TH functionalities of allergen-specific T cells, ELISPOT assays with sets of overlapping peptides covering the sequences of known allergens and novel antigens were employed to measure release of IL-5, IFNγ, IL-10, IL-17 and IL-21., Results: Using these techniques, we characterized TH responses in a cohort of adult Bla-g-sensitized subjects, either with (n = 55) or without (n = 17) asthma, and nonsensitized controls (n = 20). T cell responses were detected for ten known Bla-g allergens and an additional ten novel Bla-g antigens, representing in total a 5-fold increase in the number of antigens demonstrated to be targeted by allergen-specific T cells. Responses of sensitized individuals regardless of asthma status were predominantly TH 2, but higher in patients with diagnosed asthma. In asthmatic subjects, Bla-g 5, 9 and 11 were immunodominant, while, in contrast, nonasthmatic-sensitized subjects responded mostly to Bla-g 5 and 4 and the novel antigen NBGA5., Conclusions: Asthmatic and nonasthmatic cockroach-sensitized individuals exhibit similar TH 2-polarized responses. Compared with nonasthmatics, however, asthmatic individuals have responses of higher magnitude and different allergen specificity., (© 2015 John Wiley & Sons Ltd.)