Genetic and T2 biomarkers linked to the efficacy of HDM sublingual immunotherapy in asthma.

BACKGROUND: Hypersensitivity to house dust mite (HDM) allergens is a common cause of allergic asthma symptoms and can be effectively treated with allergy immunotherapy (AIT). OBJECTIVE: To investigate whether genetic and type 2 (T2) inflammatory biomarkers correlate with disease severity in subjects with allergic asthma, and whether this can be modified by AIT. METHODS: MITRA (NCT01433523) was a phase III, randomised, double-blind, placebo-controlled trial of HDM sublingual immunotherapy (SLIT)-tablets in adults with HDM allergic asthma. Post hoc analyses of the study population (N=742) evaluated associations between T2 inflammatory (blood eosinophils, eosinophil cationic protein (ECP), total IgE and tryptase) and genetic (single-nucleotide polymorphisms, SNP) biomarkers (n=582) for the primary study endpoint (time to first moderate/severe asthma exacerbation). SNP associations were verified in HDM-positive subgroup from an independent 3-year Severe Asthma Research Programme (SARP3) subject cohort. RESULTS: An increased asthma exacerbation risk in subjects homozygous for SNP rs7216389 (chromosomal locus 17q12-21) was reduced (p=0.037) by treatment with HDM SLIT (HR=0.37 (95% CI 0.22 to 0.64), p<0.001). The associations between exacerbation risk and 17q12-21 SNPs were replicated in the SARP3 HDM-positive subgroup. High levels of T2 biomarkers were associated with increased risk of asthma exacerbations in the placebo group. HDM SLIT-tablet treatment reduced this risk (blood eosinophils: HR=0.50 (95% CI 0.30 to 0.85); ECP: HR=0.45 (95% CI 0.29 to 0.87); tryptase: HR=0.45 (95% CI 0.25 to 0.80)). The treatment effect was higher (p=0.006) for subjects with a higher number of elevated T2 biomarkers. CONCLUSIONS: HDM SLIT-tablet AIT is efficacious in HDM-sensitised asthma subjects with a genetic asthma predisposition and/or an underlying T2 endotype. TRIAL REGISTRATION NUMBER: NCT01433523.

T cells specific to multiple Bet v 1 peptides are highly cross-reactive toward the corresponding peptides from the homologous group of tree pollens.

Background: Allergens from Fagales trees frequently cause spring allergy in Europe, North America, and some parts of Asia. The definition of the birch homologous group, which includes birch (Bet v), oak (Que a), alder (Aln g), hazel (Cor a), hornbeam (Car b), beech (Fag s), and chestnut (Cas s), is based on high allergen sequence identity and extensive IgE cross-reactivity. Clinical effect was seen during the alder/hazel, birch, and oak pollen seasons after treatment with tree SLIT-tablets containing only birch allergen extract. Here, we characterize T-cell reactivity with respect to epitope specificities and cross-reactivity toward various Bet v 1 family members, (PR-10/group 1 major allergens). This cross-reactivity may be part of the immunological basis of clinical effect or cross-protection when exposed to birch homologous tree species. Method: T-cell lines were generated from 29 birch-allergic individuals through stimulation of peripheral blood mononuclear cells (PBMCs) with birch/Bet v or oak/Que a allergen extracts. T-cell responses to allergen extracts, purified group 1 allergens, and overlapping 20-mer peptides (Bet v 1, Aln g 1, Cor a 1, and Que a 1) were investigated by T-cell proliferation and cytokine production. Cross-reactivity was evaluated based on Pearson's correlations of response strength and further investigated by flow cytometry using tetramer staining for homologous peptide pairs. Results: T-cell reactivity toward extracts and group 1 allergens from across the birch homologous group was observed for birch/Bet v as well as oak/Que a T-cell lines. T-cell lines responded to multiple Bet v 1 homologous peptides from Aln g 1 and Cor a 1 and a subset of Que a 1 peptides. Significant Pearson's correlations between frequently recognized peptides derived from Bet v 1 and the corresponding peptides derived from alder, hazel, and oak strongly supported the T-cell cross-reactivity toward these allergens. Cross-reactivity between birch and birch homologous peptides was confirmed by pMHCII tetramer staining. Conclusion: T cells from birch tree pollen allergic individuals respond to multiple trees within the birch homologous group in accordance with the level of sequence homology between Bet v 1 family members, (PR-10 allergens) from these allergen sources, confirming the basis for clinical cross-protection.

Simplified AIT for allergy to several tree pollens-Arguments from the immune outcome analyses following treatment with SQ tree SLIT-tablet.

BACKGROUND: The SQ tree SLIT-tablet (containing birch extract) proved clinically significant effects during the pollen season for birch as well as alder/hazel. Immune outcomes of this treatment for allergens from multiple birch homologous trees need further investigation. We hypothesize that birch pollen extract AIT modulates a highly cross-reactive immune response and that this may be the basis for the observed clinical cross-protection. METHODS: Blood samples were collected from 397 birch allergic patients during SQ tree SLIT-tablet or placebo treatment (1:1) for up to 40 weeks. Serum IgE and IgG4 specific to birch, and birch homologous tree pollens from alder, hazel, hornbeam, beech and chestnut were measured by ImmunoCAP. IgE-Blocking Factor (IgE-BF) for alder, birch and hazel during treatment was measured by Advia Centaur and blocking effects for birch and all these birch homologous tree pollens were further investigated by basophil activation (BAT). Antibody readouts were investigated in patient subsets. T-cell responses (proliferation) to allergen extracts and peptide pools (group 1 allergens) were investigated in T-cell lines from 29 untreated birch pollen-allergic individuals. RESULTS: Significant Pearson correlations between serum IgE towards birch, alder, hazel, hornbeam and beech were observed (r-values > .86). T-cell reactivity was observed throughout the birch homologous group. Almost identical kinetics for changes in IgE towards birch, alder and hazel were observed during treatment and similar species-specific changes were seen for serum-IgG4 . IgG4 reactivity towards birch and alder, hazel, hornbeam and beech correlated significantly at end-of-treatment (r-values > .72). Treatment resulted in similar IgE-BF kinetics for alder, birch, and hazel and blocking of BAT for multiple trees in most actively treated patients investigated. CONCLUSIONS: Systematic analyses of T-cell and antibody cross-reactivities before and during birch pollen extract AIT provide the immunological basis for the observed clinical effect of SQ tree SLIT-tablet treatment of tree pollen allergy induced by multiple trees in the birch homologous group.

House Dust Mite-Specific Sublingual Immunotherapy Prevents the Development of Allergic Inflammation in a Mouse Model of Experimental Asthma.

BACKGROUND: Evidence regarding sublingual immunotherapy (SLIT) efficacy and its good safety profile has been demonstrated with pollen and house dust mite (HDM) allergens in the treatment of airway allergies. In addition, the use of grass pollen presents a SLIT disease-modifying treatment for respiratory allergies. OBJECTIVES: The aim of this study was to demonstrate the efficacy of HDM-based SLIT in mouse models of allergic airway inflammation and to gain insights into the involved local immunological mechanisms. METHODS: Balb/c mice were sensitized/challenged with Dermatophagoides farinae (Der f) extract and underwent Der f-SLIT in prophylactic and therapeutic settings. The SLIT efficacy was assessed using lung function measurements, analysis of local inflammatory responses by bronchoalveolar lavage cell differentiation and lung histology. Humoral and cellular responses were monitored by ELISA, cytokine bead array and flow cytometry analyses. RESULTS: In a prophylactic setting, Der f-SLIT with 12 development units per dose reduced the eosinophil-dominated inflammatory response in the lung paralleled by a marked reduction in airway hyperresponsiveness. Local Th2 responses were prevented as demonstrated by significantly lower levels of IL-5 and IL-13. Additionally, SLIT-treated mice revealed a lower proportion of CD4-CD8- x03B3;δ cells and a higher frequency of CD8+CD25+IFNx03B3;+ T cells in the lungs compared to sham-treated mice. In a therapeutic setting, Der f-SLIT also resulted in reduced inflammatory responses in the lung. CONCLUSION: The efficacy of Der f-SLIT was demonstrated in prophylactic and therapeutic conditions using experimental mouse models of HDM-induced airway inflammation. A potential role of a so far underestimated lymphocyte subpopulation was also indicated.

Protocol for a randomised, double-blind, placebo-controlled study of grass allergen immunotherapy tablet for seasonal allergic rhinitis: time course of nasal, cutaneous and immunological outcomes.

BACKGROUND: Seasonal Allergic Rhinitis is characterised by inflammation of the nasal mucosa upon exposure to common aeroallergens, affecting up to 20-25 % of the population. For those patients whose symptoms are not controlled by standard medical treatment, allergen specific immunotherapy is a therapeutic alternative. Although several studies have shown changes in immunologic responses as well as long term tolerance following treatment with a sublingual allergy immunotherapy tablet, a detailed time course of the early mechanistic changes of local and systemic T and B cell responses and the effects on B cell repertoire in the nasal mucosa have not been fully examined. METHODS/DESIGN: This is a randomized, double-blind, single-centre, placebo controlled, two arm time course study based in the United Kingdom comparing sublingual allergy immunotherapy tablet (GRAZAX(®), ALK-Abello Horsholm, Denmark) plus standard treatment with placebo plus standard treatment. Up to 50 moderate to severe grass pollen allergic participants will be enrolled to ensure randomisation of at least 44. Further, we shall enrol 20 non-atopic volunteers. Screening will be completed before eligible atopic participants are randomised to one of the two treatment arms in a 1 to 1 ratio. The primary endpoint will be the total nasal symptom score assessed over 60 min following grass pollen nasal allergen challenge after 12 months of treatment. Clinical assessments and/or mechanistic analyses on blood, nasal fluid, brushing and biopsies will be performed at baseline at 1, 2, 3, 4 (coinciding with the peak pollen season), 6 and 12 months of treatment. After 12 months of treatment, unblinding will take place. Those atopic participants receiving active treatment will continue therapy for another 12 months followed by a post treatment phase of 12 months. Assessments and collection of biologic samples from these participants will take place again at 24 and at 36 months from the start of treatment. The 20 healthy, non-atopic controls will undergo screening and one visit only coinciding with the 12 month visit for the atopic participants. DISCUSSION: The trial will end in April 2017. The trial is registered with ClinicalTrials.gov and the trial identifying number is NCT02005627. TRIAL REGISTRATION: Primary Registry: ClinicalTrials.gov, Trial Identifying number: NCT02005627, Secondary identifying numbers: EudraCT number: 2013-003732-72 REC: 13/EM/0351, Imperial College London (Sponsor): 13IC0847, Protocol Version 6.0, Date: 16.05.2014.

Rational identification of an optimal antibody mixture for targeting the epidermal growth factor receptor.

The epidermal growth factor receptor (EGFR) is frequently dysregulated in human malignancies and a validated target for cancer therapy. Two monoclonal anti-EGFR antibodies (cetuximab and panitumumab) are approved for clinical use. However, the percentage of patients responding to treatment is low and many patients experiencing an initial response eventually relapse. Thus, the need for more efficacious treatments remains. Previous studies have reported that mixtures of antibodies targeting multiple distinct epitopes are more effective than single mAbs at inhibiting growth of human cancer cells in vitro and in vivo. The current work describes the rational approach that led to discovery and selection of a novel anti-EGFR antibody mixture Sym004, which is currently in Phase 2 clinical testing. Twenty-four selected anti-EGFR antibodies were systematically tested in dual and triple mixtures for their ability to inhibit cancer cells in vitro and tumor growth in vivo. The results show that targeting EGFR dependent cancer cells with mixtures of antibodies is superior at inhibiting their growth both in vitro and in vivo. In particular, antibody mixtures targeting non-overlapping epitopes on domain III are efficient and indeed Sym004 is composed of two monoclonal antibodies targeting this domain. The superior growth inhibitory activity of mixtures correlated with their ability to induce efficient EGFR degradation.