Leanne M. Gardner

Induction of T ‘regulatory’ cells by standardized house dust mite immunotherapy: an increase in CD4+CD25+ interleukin‐10+ T cells expressing peripheral tissue trafficking markers

Background Clinically effective subcutaneous allergen‐specific immunotherapy (SIT) is associated with altered circulating T cell cytokine production and altered local cytokine responses with increased IL‐10 following allergen challenge in target organs.

Allergen-related approaches to immunotherapy.

Allergic diseases, including asthma, rhinitis and eczema, represent a major health burden worldwide. Mainstay treatments are allergen avoidance where feasible and pharmacotherapy for symptom relief. For selected patients, allergen-specific immunotherapy (SIT) offers the prospect of long lasting clinical efficacy. SIT involves the administration of allergen extract using a standardized regimen, usually subcutaneously or increasingly sublingually. However, application of this potentially curative treatment is restricted, largely due to the risk of serious adverse events, especially in asthmatics and for potent allergens such as peanut, seafood and latex. New insights into immunological mechanisms underlying effective SIT and molecular characterization of allergens and their recognition by the immune system suggest strategies for refinement of SIT. Selective targeting of allergen-specific T cells, especially regulatory T cells, is likely to be pivotal for efficacy. Recombinant allergens lacking IgE reactivity and small T cell epitope-based peptides are being trialled clinically with evidence of efficacy without serious IgE-mediated adverse reactions. Adjuvants, either co-administered or incorporated into a recombinant allergen vaccine to target tolerogenic dendritic cells may also increase efficacy. The safer sublingual route of allergen administration is attracting interest and different allergen forms may be optimal for inducing tolerance by this route. Defined allergen-derived molecules or peptides offer ease of standardization and, coupled with appropriate targeting of immunoregulatory mechanisms, will result in more widespread clinical use of SIT. Adjunct therapies such as anti-IgE antibody and corticosteroids may minimize the likelihood of adverse reactions in those with severe allergic disease who would most benefit from this treatment.

High Dose Allergen Stimulation of T Cells from House Dust Mite-Allergic Subjects Induces Expansion of IFN-γ+ T Cells, Apoptosis of CD4+IL-4+ T Cells and T Cell Anergy

Background: During clinically effective allergen-specific immunotherapy a shift in cytokine dominance from IL-4, IL-5 predominant to IFN-γ predominant has been observed. As antigen concentration influences Th cell priming, this study aimed to determine the effect of different allergen concentrations on human house dust mite (HDM)-specific T cell production of IL-4 and IFN-γ, proliferation and apoptosis. Methods: HDM-allergic donor PBMC were cultured for 14 days with different concentrations of HDM extract (1, 10 and 100 µg/ml). T cell intracellular IL-4 and IFN-γ, division (CFSE labelling) and apoptosis (active caspase-3 staining) were analysed by flow cytometry. Proliferation was assessed by 3H-thymidine incorporation. Results: Increased CD4+IFN-γ+ and CD8+IFN-γ+ T cell numbers were observed in high allergen concentration cultures compared with low concentration cultures whereas there were no differences in CD4+IL-4+ and CD8+IL-4+ T cell numbers. CFSE cell labelling revealed that high allergen concentration favours the expansion of IFN-γ-producing CD4+ T cells. The proportion of apoptotic cells increased with allergen concentration and there was preferential apoptosis of CD4+IL-4+ T cells. HDM-induced proliferation was decreased in high allergen concentration cultures; this was reversible by IL-2 consistent with anergy. Conclusion: These results show that T cell division and apoptosis contribute to the cytokine skewing to predominant IFN-γ production by T cells observed at high allergen concentration. Thus the use of hypoallergenic T cell reactive preparations which can be given safely at higher doses than natural extracts may enhance efficacy of allergen-specific immunotherapy.

Functional regulatory T cells and allergen immunotherapy

Purpose of reviewBurgeoning literature ascribing roles for regulatory T cells (Treg) in controlling clinical allergy frequently relies on phenotypic markers rather than functional suppression assays. This review examines evidence for functional Treg in controlling immune responses to allergen with particular focus on allergen-specific immunotherapy. Recent findingsOver the last year, use of recently identified Treg markers has facilitated more precise delineation and isolation of Treg subtypes. Advanced flow cytometry allows better discrimination and tracking of effector T cells (Teff) and Treg in co-culture assays for Treg function. These approaches enable critical appraisal of the evidence for Treg mediation of the observed changes in immune reactivity to allergen accompanying immunotherapy. Recent studies suggest that different mechanisms of immune regulation may occur at different times of immunotherapy, with Treg playing a more important role early in treatment. Additionally, advanced imaging techniques reveal increased Treg numbers in allergen-exposed mucosal tissue after therapy consistent with an enhanced local functional role. SummaryThese findings support a mechanistic role for Treg early in allergen immunotherapy. Research is required to clarify the precise Treg subsets involved, their specificity and how their activity can be enhanced during immunotherapy by appropriate allergen form, optimal site of administration and use of adjuvants and adjunct therapies.

Unique and Cross-Reactive T Cell Epitope Peptides of the Major Bahia Grass Pollen Allergen, Pas n 1

Background: Bahia grass pollen (BaGP) is a major cause of allergic rhinitis. Subcutaneous allergen-specific immunotherapy is effective for grass pollen allergy, but is unsuitable for patients with moderate to severe asthma due to the risk of anaphylaxis. T cell-reactive but IgE nonreactive peptides provide a safer treatment option. This study aimed to identify and characterize dominant CD4+ T cell epitope peptides of the major BaGP allergen, Pas n 1. Methods: Pas n 1-specific T cell lines generated from the peripheral blood of BaGP-allergic subjects were tested for proliferative and cytokine response to overlapping 20-mer Pas n 1 peptides. Cross-reactivity to homologous peptides from Lol p 1 and Cyn d 1 of Ryegrass and Bermuda grass pollen, respectively, was assessed using Pas n 1 peptide-specific T cell clones. MHC class II restriction of Pas n 1 peptide T cell recognition was determined by HLA blocking assays and peptide IgE reactivity tested by dot blotting. Results: Three Pas n 1 peptides showed dominant T cell reactivity; 15 of 18 (83%) patients responded to one or more of these peptides. T cell clones specific for dominant Pas n 1 peptides showed evidence of species-specific T cell reactivity as well as cross-reactivity with other group 1 grass pollen allergens. The dominant Pas n 1 T cell epitope peptides showed HLA binding diversity and were non-IgE reactive. Conclusions: The immunodominant T cell-reactive Pas n 1 peptides are candidates for safe immunotherapy for individuals, including those with asthma, who are allergic to Bahia and possibly other grass pollens.

Increased allergen concentration enhances IFN-γ production by allergic donor T cells expressing a peripheral tissue trafficking phenotype

Background:  Clinically effective allergen‐specific immunotherapy correlates with decreased circulating allergen‐specific IL‐4+ T cells but increased IFN‐γ+ cells at sites of allergen challenge. Whether immunotherapy promotes trafficking of IFN‐γ+ T cells to peripheral tissues is unknown. As aeroallergen is administered at higher concentrations during immunotherapy than those encountered naturally, the effect of allergen concentration on adhesion molecule (CD62L and CD49d) and chemokine receptor (CCR3 and CCR5) expression by peripheral‐blood T cells was analysed in parallel with cytokine production.

The hevein domain of the major latex‐glove allergen Hev b 6.01 contains dominant T cell reactive sites

Background Sensitization to natural rubber latex (Hevea brasiliensis) is a major cause of occupational asthma and rhinitis affecting frequent latex‐glove users. Hev b 6.01, a known major latex allergen, is cleaved naturally into hevein (4.7 kDa) and a C‐terminal fragment (14 kDa). Hevein is an abundant protein in latex‐glove extracts. As the immune response to allergens is initiated by activation of allergen‐specific CD4+ T cells, identification of dominant T cell epitopes is crucial for the development of specific immunotherapy.

T Cell Targeted Strategies for Improved Efficacy and Safety of Specific Immunotherapy for Allergic Disease

Allergic diseases including asthma, rhinitis and eczema are known to be a major health and economic burden worldwide. Specific immunotherapy (SIT) is potentially curative but restricted in use, e.g. for asthmatics, due to risk of serious adverse events. Safer, effective SIT preparations require elucidation of mechanisms and immunoregulatory factors. Allergen-specific T cells play a pivotal role. For allergic individuals, allergen-stimulated T cells largely secrete IL-4, IL-5 and IL-13 (Th2-type cytokines), whereas non-allergics show predominant IFN-γ secretion (Th1-type). Clinically successful SIT is accompanied by altered allergen-specific T cell response, with decreased Th2/Th1 ratio, enhanced IL-10 secretion and regulatory T cell induction. Contributing factors include allergen concentration and form, adjuvant and antigen presenting cell type. In conventional SIT, high dose unfractionated allergen extracts are injected incrementally via the subcutaneous route. To avoid adverse IgE-mediated events but retain efficacy, hypoallergenic T cell-reactive allergen derivatives can be used. These include peptides containing dominant T cell epitopes of allergens, chemically-modified allergens, and recombinant whole or mutant allergens. Such approaches have been evaluated successfully in animal models and early phase clinical trials. Adjuvants and carriers including bacterial and viral components, liposomes and DNA vaccines also promote repolarisation of T cell response and regulatory T cell induction. However caution is needed as excessive IFN- γ secretion may invoke pathogenic inflammation. Sublingual administration has fewer adverse events and is gaining popularity for respiratory allergens, and other routes including intranasal and oral are under evaluation. T cell targeted strategies will facilitate wider clinical application of SIT and reliable laboratory assays for monitoring treatment.