The role of allergen-specific IgE, IgG and IgA in allergic disease.

Immunoglobulin E (IgE)-mediated allergy is the most common hypersensitivity disease affecting more than 30% of the population. Exposure to even minute quantities of allergens can lead to the production of IgE antibodies in atopic individuals. This is termed allergic sensitization, which occurs mainly in early childhood. Allergen-specific IgE then binds to the high (FcεRI) and low-affinity receptors (FcεRII, also called CD23) for IgE on effector cells and antigen-presenting cells. Subsequent and repeated allergen exposure increases allergen-specific IgE levels and, by receptor cross-linking, triggers immediate release of inflammatory mediators from mast cells and basophils whereas IgE-facilitated allergen presentation perpetuates T cell-mediated allergic inflammation. Due to engagement of receptors which are highly selective for IgE, even tiny amounts of allergens can induce massive inflammation. Naturally occurring allergen-specific IgG and IgA antibodies usually recognize different epitopes on allergens compared with IgE and do not efficiently interfere with allergen-induced inflammation. However, IgG and IgA antibodies to these important IgE epitopes can be induced by allergen-specific immunotherapy or by passive immunization. These will lead to competition with IgE for binding with the allergen and prevent allergic responses. Similarly, anti-IgE treatment does the same by preventing IgE from binding to its receptor on mast cells and basophils. Here, we review the complex interplay of allergen-specific IgE, IgG and IgA and the corresponding cell receptors in allergic diseases and its relevance for diagnosis, treatment and prevention of allergy.

Differential induction of allergen-specific IgA responses following timothy grass subcutaneous and sublingual immunotherapy.

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 (per-protocol population, n = 84). TGP-specific IgA1, IgA2, IgG4, IgG, and IgE were measured in nasal fluids by ELISA. TGP-specific IgA1, IgA2, and Phleum pratense (Phl p)1, 2, 4, 5b, 6, 7, 11, and 12 IgE and IgG4 were measured in sera by ELISA and ImmunoCAP, respectively. RESULTS: At years 2 and 3, TGP-IgA1/2 levels in nasal fluid were elevated in SLIT compared with SCIT (4.2- and 3.0-fold for IgA1, 2.0- and 1.8-fold for IgA2, respectively; all P < .01). TGP-IgA1 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-IgG4 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 IgG4 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 IgG4 to TGP components stratified participants according to treatment group and clinical response. CONCLUSIONS: The observed induction of IgA1/2 in SLIT and IgG4 in SCIT suggest key differences in the mechanisms of action.

Treatment Effect of the Tree Pollen SLIT-Tablet on Allergic Rhinoconjunctivitis During Oak Pollen Season.

BACKGROUND: Birch, alder, hazel, and oak are members of the birch homologous group based on cross-reactivity toward the birch pollen allergen Betula verrucosa 1. Theoretically, allergy to these tree pollens may be treated by immunotherapy with one representative allergen extract. OBJECTIVE: To evaluate post hoc whether treatment of birch pollen-induced allergic rhinoconjunctivitis with a standardized tree sublingual immunotherapy (SLIT)-tablet containing birch pollen extract reduces symptoms and symptom-relieving medication use during the oak pollen season (OPS). METHODS: In a randomized, multinational, double-blind trial (EudraCT-2015-004821-15), 634 participants (ages 12-65 years) received daily tree SLIT-tablet (12 SQ-Bet) or placebo before and during tree pollen season (alder/hazel plus birch pollen season [BPS]). Symptom-relieving medication was allowed. The primary end point was the average total combined score (sum of rhinoconjunctivitis daily symptom score and daily medication score) during BPS. Outcomes during the OPS (excluding overlapping BPS days) were analyzed post hoc. RESULTS: Relative improvements in average total combined score, daily symptom score, and daily medication score with the tree SLIT-tablet versus placebo during the OPS were 25%, 22%, and 32%, respectively (all P < .001). Significant correlations were observed between birch and oak serum immunoglobulin E (sIgE) at baseline (r = 0.86) and between birch and oak IgG4 after treatment (r = 0.72). Oak sIgE and IgG4 kinetics in response to tree SLIT-tablet treatment were similar to birch. CONCLUSIONS: The tree SLIT-tablet leads to significant improvement of rhinoconjunctivitis outcomes during the OPS, supporting the clinical relevance of immunological cross-reactivity toward birch and oak allergens.

Allergy to oak pollen in North America.

Background: Oak pollen is an important allergen in North America. The genus Quercus (oak) belongs to the family Fagaceae under the order Fagales. Objective: The objective of this article was to narratively review the oak pollen season, clinical and epidemiologic aspects of allergy to oak pollen, oak taxonomy, and oak allergen cross-reactivity, with a focus on the North American perspective. Methods: A PubMed literature review (no limits) was conducted. Publications related to oak pollen, oak-related allergic rhinitis with or without conjunctivitis, and oak-related allergic asthma were selected for review. Results: Oak species are common throughout the United States and contribute up to 50% to overall atmospheric pollen loads. Mean peak oak pollen counts can reach >2000 grains/m³. The start of the oak pollen season generally corresponds to the seasonal shift from winter to spring based on latitude and elevation, and may begin as early as mid February. The duration of the season can last > 100 days and, in general, is longer at lower latitudes. In the United States, ∼30% of individuals with allergy are sensitized to oak. The oak pollen season correlates with increased allergic rhinitis symptom-relieving medication use and asthma-related emergency department visits or hospitalizations. Oak falls within the birch homologous group. Extensive immunologic cross-reactivity has been demonstrated between oak pollen and birch pollen allergens, and, more specifically, their major allergens Que a 1 and Bet v 1. The cross-reactivity between oak and birch has implications for allergy immunotherapy (AIT) because guidelines suggest selecting one representative allergen within a homologous group for AIT, a principle that would apply to oak. Conclusion: Allergy to oak pollen is common in North America and has a substantial clinical impact. Oak pollen allergens are cross-reactive with birch pollen allergens, which may have implications for AIT.

Basophil sensitivity reflects long-term clinical outcome of subcutaneous immunotherapy in grass pollen-allergic patients.

BACKGROUND: Allergic rhinoconjunctivitis is a public health problem. Allergen Immunotherapy is an effective and safe treatment, that modifies the natural course of allergic disease and induces long-term tolerance. OBJECTIVE: To correlate basophil and antibody biomarkers of subcutaneous immunotherapy to clinical outcomes and cellular changes in target tissue. METHODS: Adults suffering from allergic rhinoconjunctivitis due to grass pollen allergy were randomized to receive subcutaneous immunotherapy (n = 18) or to an open control group (n = 6). Patients reported daily symptom and medication scores and weekly rhinitis related quality of life scores during four pollen seasons. Biomarkers were measured every 3 months for three years treatment and every 6 months in the follow-up year. Nasal and cutaneous allergen challenge tests were performed annually. Leukocyte subsets were assessed in nasal mucosa biopsies at baseline and after treatment. RESULTS: Subcutaneous immunotherapy led to a 447-fold decrease in basophil sensitivity during the first treatment year. This remained 100-fold lower than baseline during the 3 year-treatment period and 10-fold lower during the follow-up year (n = 18, P = .03). Decrease in basophil sensitivity after three weeks of treatment predicted long-term improvement in seasonal combined symptom and medication scores (á¿¥=-0.69, P = .0027) during three years of treatment. AUC of IgE-blocking factor correlated to nasal allergen challenge (á¿¥ = 0.63, P = .0012) and SPT (á¿¥ = 0.45, P = .03). Plasma cell numbers in the nasal mucosa increased during treatment (P = .02). CONCLUSION: Decrease in basophil sensitivity after three weeks of subcutaneous allergen immunotherapy predicted the clinical outcome of this treatment.

Diverse and highly cross-reactive T-cell responses in ragweed allergic patients independent of geographical region.

BACKGROUND: Ragweed frequently causes seasonal allergies in North America and Europe. In the United States, several related ragweed species with diverse geographical distribution cause allergic symptoms. Cross-reactivity towards related ragweed species of IgE and treatment-induced IgG4 has been demonstrated previously. However, less is known about the underlying T-cell cross-reactivity. METHODS: The allergen content of ragweed extracts was determined by mass spectrometry and related to T-cell epitopes of Amb a allergens (group 1, 3, 4, 5, 8 and 11) in 20 American ragweed allergic patients determined by FluoroSpot and proliferation assays. T-cell responses to 50 frequently recognized Amb a-derived T-cell epitopes and homologous peptides from western ragweed (Amb p), giant ragweed (Amb t) and mugwort (Art v) were investigated in an additional 11 American and 14 Slovakian ragweed allergic donors. RESULTS: Ragweed extracts contained all known allergens and isoallergens thereof. Donor T-cell responses were diverse and directed against all Amb a 1 isoallergens and to most minor allergens investigated. Similar response patterns were seen in American and Slovakia donors. Several epitopes were cross-reactive between isoallergens and ragweed species, some even including mugwort. T-cell cross-reactivity generally correlated with allergen sequence homology. CONCLUSION: T-cell epitopes of multiple allergens/isoallergens are involved in the diverse T-cell responses in ragweed allergic individuals. T-cell lines were highly cross-reactive to epitopes of related ragweed species without any apparent geographical response bias. These data support that different ragweed species can be considered an allergen homology group with Amb a as the representative species regarding diagnosis as well as allergy immunotherapy.

Persistent regulatory T-cell response 2 years after 3 years of grass tablet SLIT: Links to reduced eosinophil counts, sIgE levels, and clinical benefit.

BACKGROUND: In the first 2 years of grass tablet sublingual immunotherapy treatment, we have previously demonstrated a progressive development of a regulatory T-cell response, which was preceded by an early decrease in the frequency of both IL-4+ cells and sIgE levels. A progressive increase in sIgG4 levels and FAB blockage were also found. METHODS: By monitoring immunological kinetics during 3 years of active treatment + 2 years of follow-up, we aimed to identify key immunological parameters that could explain sustained clinical benefit of grass tablet sublingual immunotherapy. RESULTS: Thirty patients completed the 5-year clinical trial protocol. Although individual responses were heterogeneous, reduction in both sIgE and circulating IL-4+ cells compared to the initial 1- to 4-month peak was maintained throughout the 3-year treatment period and for 2 years after discontinuation. Meanwhile, after a 2-year increase in sIgG4, the levels were stabilized during the third year and decreased post-therapy. FAB inhibition remained significantly inhibited throughout the study compared to preimmunotherapy in 83% of patients. A sustained regulatory T-cell response, after IT cessation, occurs in two-thirds of the patients. There was a statistical association between this regulatory response, the maintenance of lower eosinophil counts during grass pollen seasons, and sIgE titers lower than before immunotherapy treatment, and the latter were significantly associated with clinical response. CONCLUSION: Our results suggest that the immunological mechanisms underlying the sustained response after 2 years of cessation of immunotherapy (3-year treatment period) are linked to the acquisition and maintenance of a regulatory T-cell response.

Synchronous immune alterations mirror clinical response during allergen immunotherapy.

BACKGROUND: Three years of treatment with either sublingual or subcutaneous allergen immunotherapy has been shown to be effective and to induce long-term tolerance. The Gauging Response in Allergic Rhinitis to Sublingual and Subcutaneous Immunotherapy (GRASS) trial demonstrated that 2 years of treatment through either route was effective in suppressing the response to nasal allergen challenge, although it was insufficient for inhibition 1 year after discontinuation. OBJECTIVE: We sought to examine in the GRASS trial the time course of immunologic changes during 2 years of sublingual and subcutaneous immunotherapy and for 1 year after treatment discontinuation. METHODS: We performed multimodal immunomonitoring to assess allergen-specific CD4 T-cell properties in parallel with analysis of local mucosal cytokine responses induced by nasal allergen exposure and humoral immune responses that included IgE-dependent basophil activation and measurement of serum inhibitory activity for allergen-IgE binding to B cells (IgE-facilitated allergen binding). RESULTS: All 3 of these distinct arms of the immune response displayed significant and coordinate alterations during 2 years of allergen desensitization, followed by reversal at 3 years, reflecting a lack of a durable immunologic effect. Although frequencies of antigen-specific TH2 cells in peripheral blood determined by using HLA class II tetramer analysis most closely paralleled clinical outcomes, IgE antibody-dependent functional assays remained inhibited in part 1 year after discontinuation. CONCLUSION: Two years of allergen immunotherapy were effective but insufficient for long-term tolerance. Allergen-specific TH2 cells most closely paralleled the transient clinical outcome, and it is likely that recurrence of the T-cell drivers of allergic immunity abrogated the potential for durable tolerance. On the other hand, the persistence of IgE blocking antibody 1 year after discontinuation might be an early indicator of a protolerogenic mechanism.

Immunoproteomic analysis of house dust mite antigens reveals distinct classes of dominant T cell antigens according to function and serological reactivity.

BACKGROUND: House dust mite (HDM) allergens are a common cause of allergy and allergic asthma. A comprehensive analysis of proteins targeted by T cells, which are implicated in the development and regulation of allergic disease independent of their antibody reactivity, is still lacking. OBJECTIVE: To comprehensively analyse the HDM-derived protein targets of T cell responses in HDM-allergic individuals, and investigate their correlation with IgE/IgG responses and protein function. METHODS: Proteomic analysis (liquid chromatography-tandem mass spectrometry) of HDM extracts identified 90 distinct protein clusters, corresponding to 29 known allergens and 61 novel proteins. Peripheral blood mononuclear cells (PBMC) from 20 HDM-allergic individuals were stimulated with HDM extracts and assayed with a set of ~2500 peptides derived from these 90 protein clusters and predicted to bind the most common HLA class II types. 2D immunoblots were made in parallel to elucidate IgE and IgG reactivity, and putative function analyses were performed in silico according to Gene Ontology annotations. RESULTS: Analysis of T cell reactivity revealed a large number of T cell epitopes. Overall response magnitude and frequency was comparable for known and novel proteins, with 15 antigens (nine of which were novel) dominating the total T cell response. Most of the known allergens that were dominant at the T cell level were also IgE reactive, as expected, while few novel dominant T cell antigens were IgE reactive. Among known allergens, hydrolase activity and detectable IgE/IgG reactivity are strongly correlated, while no protein function correlates with immunogenicity of novel proteins. A total of 106 epitopes accounted for half of the total T cell response, underlining the heterogeneity of T cell responses to HDM allergens. CONCLUSIONS AND CLINICAL RELEVANCE: Herein, we define the T cell targets for both known allergens and novel proteins, which may inform future diagnostics and immunotherapeutics for allergy to HDM.

Polarized Airway Epithelial Models for Immunological Co-Culture Studies.

Epithelial cells line all cavities and surfaces throughout the body and play a substantial role in maintaining tissue homeostasis. Asthma and other atopic diseases are increasing worldwide and allergic disorders are hypothesized to be a consequence of a combination of dysregulation of the epithelial response towards environmental antigens and genetic susceptibility, resulting in inflammation and T cell-derived immune responses. In vivo animal models have long been used to study immune homeostasis of the airways but are limited by species restriction and lack of exposure to a natural environment of both potential allergens and microflora. Limitations of these models prompt a need to develop new human cell-based in vitro models. A variety of co-culture systems for modelling the respiratory epithelium exist and are available to the scientific community. The models have become increasingly sophisticated and specific care needs to be taken with regard to cell types, culture medium and culture models, depending on the aim of the study. Although great strides have been made, there is still a need for further optimization, and optimally also for standardization, in order for in vitro co-culture models to become powerful tools in the discovery of key molecules dictating immunity and/or tolerance, and for understanding the complex interplay that takes place between mucosa, airway epithelium and resident or infiltrating immune cells. This review focuses on current knowledge and the advantages and limitations of the different cell types and culture methods used in co-culture models of the human airways.

Grass pollen immunotherapy: where are we now.

During allergen immunotherapy (AIT), the allergic patient is exposed to the disease-inducing antigens (allergens) in order to induce clinical and immunological tolerance and obtain disease modification. Large trials of grass AIT with highly standardized subcutaneous and sublingual tablet vaccines have been conducted to document the clinical effect. Induction of blocking antibodies as well as changes in the balance between T-cell phenotypes, including induction of regulatory T-cell subtypes, have been demonstrated for both treatment types. These observations increase the understanding of the immunological mechanism behind the clinical effect and may make it possible to use the immunological changes as biomarkers of clinical effect. The current review describes the recent mechanistic findings for subcutaneous immunotherapy and sublingual immunotherapy/tablet treatment and discusses how the observed immunological changes translate into a scientific foundation for the observed clinical effects of grass pollen immunotherapy and lead to new treatment strategies for grass AIT.

Effect of Polarization on Airway Epithelial Conditioning of Monocyte-Derived Dendritic Cells.

Airway epithelial cells (AECs) form polarized barriers that interact with inhaled allergens and are involved in immune homeostasis. We examined how monocyte-derived dendritic cells (MDDCs) are affected by contact with the airway epithelium. In traditional setups, bronchial epithelial cell lines were allowed to polarize on filter inserts, and MDDCs were allowed to adhere to the epithelial basal side. In an optimized setup, the cell application was reversed, and the culture conditions were modified to preserve cellular polarization and integrity. These two parameters were crucial for the MDDCs' immunoregulatory properties; thus, previous observations obtained using traditional setups should be considered with caution. Using the optimized setup, AEC conditioning of MDDCs led to increased expression of programmed death 1 ligand 1, immunoglobulin-like transcript 3, CD40, CD80, and CD23. This increased expression was accompanied by decreased secretion of monocyte chemotactic protein 1 and eotaxin and donor-variable effects on IL-12 and IL-10 secretion. Conditioning varied between maturation states and depended partly on direct contact between AECs and MDDCs. The setup allowed MDDCs on the basal side of the epithelium to sample allergens administered to the apical side. Allergen uptake depended on polarization and the nature of the allergen. AEC conditioning led to decreased birch allergen-specific proliferation of autologous T cells and a trend toward decreased secretion of the Th2-specific cytokines IL-5 and IL-13. In conclusion, we determined that AEC conditioning favoring cellular integrity leads to a tolerogenic MDDC phenotype, which is likely to be important in regulating immune responses against commonly inhaled allergens.

Grass tablet sublingual immunotherapy downregulates the TH2 cytokine response followed by regulatory T-cell generation.

BACKGROUND: Sublingual administration of Phleum pratense allergen immunotherapy (SLIT) tablets is a clinically efficient treatment for grass pollen-induced rhinoconjunctivitis. This immunotherapy downregulates TH2 immune responses, induces tolerogenic pathways, and increases regulatory T cells. However, associated immune response markers of allergen desensitization remain undefined. OBJECTIVE: We sought to characterize the kinetics of individual changes in the immunologic response to grass tablet SLIT. METHODS: We evaluated the systemic effects of SLIT in a longitudinal analysis of humoral and cellular immune parameters in peripheral blood samples. RESULTS: Grass tablet SLIT administration induced a 2-phase systemic humoral and cellular response. The TH2 response was initially exacerbated and detected as increased allergen-specific IgE (sIgE) and IgG4 (sIgG4) levels and an increase in IL-4-producing cells, followed by downregulation of the TH2 response with a shift toward a TH1 cytokine profile. T cells with a regulatory phenotype were also elicited. Statistical correlations between immunologic measurements for each patient throughout therapy indicated that TH2 response downregulation and reduction of the immediate SLIT-induced IgE response were associated with increased allergen-specific IgG4 synthesis early in therapy. TH2 response downregulation by month 4 correlated with increased frequency of CD4(+) T cells with a regulatory phenotype by 12 months. CONCLUSION: Changes in sIgE levels after therapy were linked to a specific IgG4 response, and production of blocking antibodies correlated with TH2 response downregulation. Reduced IL-4(+) cell frequency was linked to an increase in the frequency of CD4(+) T cells with a regulatory phenotype. Changes in sIgE levels and reduced IL-4 and blocking antibody levels could thus be used as indicators of a patient's immune response to therapy.

Birch pollen immunotherapy results in long-term loss of Bet v 1-specific TH2 responses, transient TR1 activation, and synthesis of IgE-blocking antibodies.

BACKGROUND: Early events of specific immunotherapy (SIT) are induction of allergen-specific IL-10-producing T(R)1 cells and production of IgG antibodies, but there is little knowledge about the long-term immune mechanisms responsible for sustained allergen tolerance. OBJECTIVE: Bet v 1-specific immune responses of 16 patients with birch pollen allergy were characterized up to 54 months at defined time points before, during, and after a 3-year period of SIT. METHODS: We sought to analyze allergen-specific T- and B-cell responses. Bet v 1-specific IL-5-, IFN-γ-, and IL-10-secreting T cells were quantified in peripheral blood, and birch pollen-specific IgE and IgG antibody levels were determined in serum. Furthermore, the inhibitory capacity of SIT-induced IgG was evaluated by blocking allergen binding to IgE and inhibition of facilitated allergen presentation. RESULTS: Seasonal increases in Bet v 1-specific T(H)2 cell numbers ceased to appear after the first year of SIT without deviation to a T(H)1-dominated immune response. Furthermore, the frequency of IL-10-producing T(R)1 cells, which had increased during the first year of SIT, returned to pretreatment levels in the second year. In contrast, allergen-specific IgG antibody concentrations continuously increased during SIT but started to decrease after cessation of treatment. Functional analysis confirmed the ability of the IgG antibodies to inhibit IgE-allergen interactions, which peaked at the end of SIT but then slowly started to decrease. CONCLUSION: Long-term allergen tolerance achieved by SIT is associated with the development of peripheral T-cell tolerance characterized by decreased reactivity of Bet v 1-specific T(H)2 cells and enriched allergen-specific IgG competing with IgE antibodies for allergen binding.

Reduced in vitro T-cell responses induced by glutaraldehyde-modified allergen extracts are caused mainly by retarded internalization of dendritic cells.

Although allergen-specific immunotherapy is a clinically effective therapy for IgE-mediated allergic diseases, the risk of IgE-mediated adverse effects still exists. For this reason, chemically modified allergoids have been introduced, which may destroy IgE-binding sites while T-cell activation should be retained. The aim of the study was to analyse the differences between intact allergens and differently modified/aggregated allergoids concerning their internalization as well as T-cell and basophil activation. For this purpose human monocyte-derived immature dendritic cells (DC) were incubated with Phleum pratense or Betula verrucosa pollen extract or with the corresponding allergoids, modified with formaldehyde or glutaraldehyde. After an additional maturation process, the antigen-loaded mature DC were co-cultured with autologous CD4(+) T cells. Allergenicity was tested by leukotriene release from basophils. In addition, the uptake of intact allergens and allergoids by immature DC was analysed. The proliferation of, as well as the interleukin-4 (IL-4), IL-10, IL-13 and interferon-γ production by, CD4(+) T cells which had been stimulated with glutaraldehyde allergoid-treated DC was reduced compared with CD4(+) T cells stimulated with intact allergen-treated or formaldehyde allergoid-treated DC. In line with this, glutaraldehyde-modified allergoids were more aggregated and were internalized more slowly. Furthermore, only the allergoids modified with glutaraldehyde induced a decreased leukotriene release by activated basophils. These findings suggest that IgE-reactive epitopes were destroyed more efficiently by modification with glutaraldehyde than with formaldehyde under the conditions chosen for these investigations. Glutaraldehyde-modified allergoids also displayed lower T-cell stimulatory capacity, which is mainly the result of greater modification/aggregation and diminished uptake by DC.

SQ-standardized sublingual grass immunotherapy: confirmation of disease modification 2 years after 3 years of treatment in a randomized trial.

BACKGROUND: The main aim of specific immunotherapy is sustained effect due to changes in the immune system that can be demonstrated only in long-term trials. OBJECTIVE: To investigate sustained efficacy and disease modification in a 5-year double-blind, placebo-controlled trial, including 2 years of blinded follow-up after completion of a 3-year period of treatment, with the SQ-standardized grass allergy immunotherapy tablet, Grazax (Phleum pratense 75,000 SQ-T/2,800 BAU,(∗) ALK, Denmark) or placebo. METHODS: A randomized, double-blind, placebo-controlled, multinational, phase III trial included adults with a history of moderate-to-severe grass pollen-induced allergic rhinoconjunctivitis, with or without asthma, inadequately controlled by symptomatic medications. Two hundred thirty-eight participants completed the trial. End points included rhinoconjunctivitis symptom and medication scores, combined scores, asthma symptom and medication scores, quality of life, days with severe symptoms, immunologic end points, and safety parameters. RESULTS: The mean rhinoconjunctivitis daily symptom score was reduced by 25% to 36% (P ≤ .004) in the grass allergy immunotherapy tablet group compared with the placebo group over the 5 grass pollen seasons covered by the trial. The rhinoconjunctivitis DMS was reduced by 20% to 45% (P ≤ .022 for seasons 1-4; P = .114 for season 5), and the weighted rhinoconjunctivitis combined score was reduced by 27% to 41% (P ≤ .003) in favor of active treatment. The percentage of days with severe symptoms during the peak grass pollen exposure was in all seasons lower in the active group than in the placebo group, with relative differences of 49% to 63% (P ≤ .0001). Efficacy was supported by long-lasting significant effects on the allergen-specific antibody response. No safety issues were identified. CONCLUSION: The results confirm disease modification by SQ-standardized grass allergy immunotherapy tablet in addition to effective symptomatic treatment of allergic rhinoconjunctivitis.