Background: Allergen immunotherapy (AIT) is a well-established disease-modifying therapy for allergic rhinitis, yet the fundamental mechanisms underlying its clinical effect remain inadequately understood. Gauging Response in Allergic Rhinitis to Sublingual and Subcutaneous Immunotherapy was a randomized, double-blind, placebo-controlled trial of individuals allergic to timothy grass who received 2 years of placebo (n = 30), subcutaneous immunotherapy (SCIT) (n = 27), or sublingual immunotherapy (SLIT) (n = 27) and were then followed for 1 additional year.Objective: We used yearly biospecimens from the Gauging Response in Allergic Rhinitis to Sublingual and Subcutaneous Immunotherapy study to identify molecular mechanisms of response.Methods: We used longitudinal transcriptomic profiling of nasal brush and PBMC samples after allergen provocation to uncover airway and systemic expression pathways mediating responsiveness to AIT. Trial Registration: ClinicalTrials.gov Identifier: NCT01335139, EudraCT Number: 2010-023536-16. Results: SCIT and SLIT demonstrated similar changes in gene module expression over time. In nasal samples, alterations included downregulation of pathways of mucus hypersecretion, leukocyte migration/activation, and endoplasmic reticulum stress (log2 fold changes -0.133 to -0.640, false discovery rates [FDRs] <0.05). We observed upregulation of modules related to epithelial development, junction formation, and lipid metabolism (log2 fold changes 0.104 to 0.393, FDRs <0.05). In PBMCs, modules related to cellular stress response and type 2 cytokine signaling were reduced by immunotherapy (log2 fold changes -0.611 to -0.828, FDRs <0.05). Expression of these modules was also significantly associated with both Total Nasal Symptom Score and peak nasal inspiratory flow, indicating important links between treatment, module expression, and allergen response.Conclusions: Our results identify specific molecular responses of the nasal airway impacting barrier function, leukocyte migration activation, and mucus secretion that are affected by both SCIT and SLIT, offering potential targets to guide novel strategies for AIT. (J Allergy Clin Immunol 2023;152:1247-60.)

BACKGROUND:Allergen-specific immunotherapy is a disease-modifying treatment that induces long-term T-cell tolerance.OBJECTIVE: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.METHODS: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.RESULTS: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.CONCLUSIONS: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.

Introduction: There is no detailed comparison of allergen-specific immunoglobulin responses following sublingual immunotherapy (SLIT) and subcutaneous immunotherapy (SCIT). Objective: We sought to compare nasal and systemic timothy grass pollen (TGP)-specific antibody responses during 2 years of SCIT and SLIT and 1 year after treatment discontinuation in a double-blind, double-dummy, placebo-controlled trial. Methods: Nasal fluid and serum were obtained yearly (perprotocol population, n = 84). TGP-specific IgA(1), IgA(2), IgG(4), IgG, and IgE were measured in nasal fluids by ELISA. TGPspecific IgA(1), IgA(2), and Phleum pratense (Phl p)1, 2, 4, 5b, 6, 7, 11, and 12 IgE and IgG(4) were measured in sera by ELISA and ImmunoCAP, respectively. Results: At years 2 and 3, TGP-IgA(1/2) levels in nasal fluid were elevated in SLIT compared with SCIT (4.2- and 3.0-fold for IgA(1), 2.0- and 1.8-fold for IgA(2), respectively; all P<.01). TGP-IgA(1) level in serum was elevated in SLIT compared with SCIT at years 1, 2, and 3 (4.6-, 5.1-, and 4.7-fold, respectively; all P<.001). Serum TGP-IgG level was higher in SCIT compared with SLIT (2.8-fold) at year 2. Serum TGP-IgG(4) level was higher in SCIT compared with SLIT at years 1, 2, and 3 (10.4-, 27.4-, and 5.1-fold, respectively; all P<.01). Serum IgG(4) levels to Phl p1, 2, 5b, and 6 were increased at years 1, 2, and 3 in SCIT and SLIT compared with placebo (Phl p1: 11.8- and 3.9-fold; Phl p2: 31.6- and 4.4-fold; Phl p5b: 135.5- and 5.3-fold; Phl p6: 145.4- and 14.7-fold, respectively, all at year 2 when levels peaked; P<.05). IgE to TGP in nasal fluid increased in the SLIT group at year 2 but not at year 3 compared with SCIT (2.8-fold; P = .04) and placebo (3.1-fold; P = .02). IgA to TGP and IgE and IgG(4) to TGP components stratified participants according to treatment group and clinical response. Conclusions: The observed induction of IgA(1/2) in SLIT and IgG(4) in SCIT suggest key differences in the mechanisms of action.

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Allergen immunotherapy is a cornerstone in the treatment of allergic children. The clinical efficiency relies on a well-defined immunologic mechanism promoting regulatory T cells and downplaying the immune response induced by allergens. Clinical indications have been well documented for respiratory allergy in the presence of rhinitis and/or allergic asthma, to pollens and dust mites. Patients who have had an anaphylactic reaction to hymenoptera venom are also good candidates for allergen immunotherapy. Administration of allergen is currently mostly either by subcutaneous injections or by sublingual administration. Both methods have been extensively studied and have pros and cons. Specifically in children, the choice of the method of administration according to the patient's profile is important. Although allergen immunotherapy is widely used, there is a need for improvement. More particularly, biomarkers for prediction of the success of the treatments are needed. The strength and efficiency of the immune response may also be boosted by the use of better adjuvants. Finally, novel formulations might be more efficient and might improve the patient's adherence to the treatment. This user's guide reviews current knowledge and aims to provide clinical guidance to healthcare professionals taking care of children undergoing allergen immunotherapy.

Local tissue eosinophilia and Th2 cytokines are characteristic features of seasonal allergic rhinitis. Airway remodelling is a feature of asthma whereas evidence for remodelling in allergic rhinitis (AR) is conflicting.

Allergen immunotherapy is a cornerstone in the treatment of allergic children. The clinical efficiency relies on a well-defined immunologic mechanism promoting regulatory T cells and downplaying the immune response induced by allergens. Clinical indications have been well documented for respiratory allergy in the presence of rhinitis and/or allergic asthma, to pollens and dust mites. Patients who have had an anaphylactic reaction to hymenoptera venom are also good candidates for allergen immunotherapy. Administration of allergen is currently mostly either by subcutaneous injections or by sublingual administration. Both methods have been extensively studied and have pros and cons. Specifically in children, the choice of the method of administration according to the patient's profile is important. Although allergen immunotherapy is widely used, there is a need for improvement. More particularly, biomarkers for prediction of the success of the treatments are needed. The strength and efficiency of the immune response may also be boosted by the use of better adjuvants. Finally, novel formulations might be more efficient and might improve the patient's adherence to the treatment. This user's guide reviews current knowledge and aims to provide clinical guidance to healthcare professionals taking care of children undergoing allergen immunotherapy.

Subcutaneous and sublingual immunotherapy are effective for allergic rhinitis. An important question is whether allergen immunotherapy provides a sustained clinical effect after treatment cessation. In view of potential side effects, cost and the necessary patient commitment, long-term benefit is an important consideration for the recommendation of immunotherapy over standard pharmacotherapy.

BACKGROUND:Grass pollen subcutaneous immunotherapy (SCIT) is associated with induction of serum IgG4-associated inhibitory antibodies that prevent IgE-facilitated allergen binding to B cells. OBJECTIVE:We sought to determine whether SCIT induces nasal allergen-specific IgG4 antibodies with inhibitory activity that correlates closely with clinical response. METHODS:In a cross-sectional controlled study, nasal fluid and sera were collected during the grass pollen season from 10 SCIT-treated patients, 13 untreated allergic patients (with seasonal allergic rhinitis [SAR]), and 12 nonatopic control subjects. Nasal and serum IgE and IgG4 levels to Phleum pratense components were measured by using the Immuno Solid Allergen Chip microarray. Inhibitory activity was measured by IgE-facilitated allergen binding assay. IL-10+ regulatory B cells were quantified in peripheral blood by using flow cytometry. RESULTS:Nasal and serum Phl p 1- and Phl p 5-specific IgE levels were increased in patients with SAR compared to nonatopic control subjects (all, P < .001) and SCIT-treated patients (nasal, P < .001; serum Phl p 5, P = .073). Nasal IgG4 levels were increased in the SCIT group compared to those in the SAR group (P < .001) during the pollen season compared to out of season. IgG-associated inhibitory activity in nasal fluid and serum was significantly increased in the SCIT group compared to that in the SAR (both, P < .01). The magnitude of the inhibitory activity was 93% (P < .001) in nasal fluid compared to 66% (P < .001) in serum and was reversed after depletion of IgG. Both nasal fluid (r = -0.69, P = .0005) and serum (r = -0.552, P = .0095) blocking activity correlated with global symptom improvement. IL-10+ regulatory B cells were increased in season compared to out of season in the SCIT group (P < .01). CONCLUSION:For the first time, we show that nasal IgG4-associated inhibitory activity correlates closely with the clinical response to allergen immunotherapy in patients with allergic rhinitis with or without asthma.

Importance Sublingual immunotherapy and subcutaneous immunotherapy are effective in seasonal allergic rhinitis. Three years of continuous treatment with subcutaneous immunotherapy and sublingual immunotherapy has been shown to improve symptoms for at least 2 years following discontinuation of treatment. Objective To assess whether 2 years of treatment with grass pollen sublingual immunotherapy, compared with placebo, provides improved nasal response to allergen challenge at 3-year follow-up. Design, Setting, and Participants A randomized double-blind, placebo-controlled, 3–parallel-group study performed in a single academic center, Imperial College London, of adult patients with moderate to severe seasonal allergic rhinitis (interfering with usual daily activities or sleep). First enrollment was March 2011, last follow-up was February 2015. Interventions Thirty-six participants received 2 years of sublingual immunotherapy (daily tablets containing 15 µg of major allergen Phleum p 5 and monthly placebo injections), 36 received subcutaneous immunotherapy (monthly injections containing 20 µg of Phleum p 5 and daily placebo tablets) and 34 received matched double-placebo. Nasal allergen challenge was performed before treatment, at 1 and 2 years of treatment, and at 3 years (1 year after treatment discontinuation). Main Outcomes and Measures Total nasal symptom scores (TNSS; range; 0 [best] to 12 [worst]) were recorded between 0 and 10 hours after challenge. The minimum clinically important difference for change in TNSS within an individual is 1.08. The primary outcome was TNSS comparing sublingual immunotherapy vs placebo at year 3. Subcutaneous immunotherapy was included as a positive control. The study was not powered to compare sublingual immunotherapy with subcutaneous immunotherapy. Results Among 106 randomized participants (mean age, 33.5 years; 34 women [32.1%]), 92 completed the study at 3 years. In the intent-to-treat population, mean TNSS score for the sublingual immunotherapy group was 6.36 (95% CI, 5.76 to 6.96) at pretreatment and 4.73 (95% CI, 3.97 to 5.48) at 3 years, and for the placebo group, the score was 6.06 (95% CI, 5.23 to 6.88) at pretreatment and 4.81 (95% CI, 3.97 to 5.65) at 3 years. The between-group difference (adjusted for baseline) was −0.18 (95% CI, −1.25 to 0.90; [P = .75]). Conclusions and Relevance Among patients with moderate to severe seasonal allergic rhinitis, 2 years of sublingual grass pollen immunotherapy was not significantly different from placebo in improving the nasal response to allergen challenge at 3-year follow-up. Trial Registration clinicaltrials.gov Identifier: NCT01335139; EudraCT Number: 2010-023536-16

A randomised, double-blind, double-placebo, controlled-trial of subcutaneous (Alutard SQ® Phleum Pratense ALK, Denmark) and sublingual (GRAZAX® ALK) grass pollen immunotherapy (105 patients,randomized 1:1:1). We previously reported the results of 2-years treatment and 1-year follow-up (year 3) using the total nasal symptom score (TNSS) at 0-10 hours after nasal allergen challenge as the primary outcome (Scadding G et al. Allergy 2016: 71, S102;3). Here we evaluate the relationship between the response to nasal allergen challenge and seasonal clinical outcomes. Post-hoc correlation of response to out of season nasal allergen challenge TNSS and seasonal outcomes including weekly visual analogue score (VAS), rhinitis quality of life (miniRQLQ), and end of season global evaluation of hay fever severity at baseline, years 1, 2 and 3. Positive correlations were seen between response to nasal challenge and each of VAS, miniRQLQ and global evaluation at year-1 of treatment (r=0.22, p=0.02; r=0.25, p=0.01 and r=0.24, p=0.01, respectively), becoming stronger at year 2(r=0.32, p=0.002; r=0.33, p=0.002 and r=0.31, p=0.002). Correlations persisted at year 3, off treatment, for all three outcomes(r=0.22, p=0.04; r=0.25, p=0.01 and r=0.42, p<0.001). No significant correlations were observed at baseline, possibly because overall spread of nasal challenge results was constrained by initial dose titration to achieve a minimum entry criterion of TNSS ≥7/12 for study inclusion. Total nasal symptom score following nasal allergen challenge out of season correlated with seasonal outcomes during allergen immunotherapy. These data support the use of nasal allergen challenge as a useful clinical surrogate in immunotherapy trials.

Dysregulation of TGF-b and activin signalling play fundamental roles in lower airways remodelling. This study focuses on characterising remodelling changes seen in CRSwNP and accompanying alteration in TGF-b signalling. Immunohistochemical staining was performed on inferior turbinate and nasal polyp biopsy specimens measuring TGF-beta1, activin-A and its receptor ALK-4, and phosphorylated SMAD2in subjects with CRSwNP (n=10) and healthy controls (n=19). Staining for D2-40 and CD34 were used to define lymphatic and vascular remodelling; smooth muscle actin and HSP-47 to study collagen synthesis and myofibroblast transformation. Matrix metalloprotease7/9 with their inhibitor TIMP-1 were enumerated. Basement membrane thickness was defined using Siris red stain. Basement membrane zone was markedly thinned in both polyp and turbinates of CRSwNP (p<0.01 versus controls). Turbinates show increased lymphatic and vascular remodelling with polyps nearly devoid of glands and possessing very little blood vessels as demonstrated by differences in total CD31/ D2-40 cell counts (p<0.01, p=0.01), blood vessels (p<0.01, p=0.02), vessel size (p<0.01, p=0.04), or vascularity (p=0.02, p=0.25). HSP-47 expression is elevated in polyps (p=0.09) whilst SMA is increased in turbinates (p=0.03). MMP7/9: TIMP-1 ratios are elevated in turbinates. TGF-b expression is increased in polyps (p<0.01) with ALK-4 elevated in polyps and turbinates (p=0.02 and p=0.03). Activin levels tend to be higher in polyps CRSwNP (p=0.08). This data demonstrates remodelling alterations in both polyps and turbinates of CRSwNP. It is possible that dysregulated TGF-b signalling in inflammatory polyps drives chronic changes in turbinate architecture thereby resulting in characteristic remodelling in this nasal disease.