Successful Allergen Immunotherapy Research Articles

Easy assessment of the avidity of polyclonal allergen-specific serum antibodies.

Allergen-specific IgE-blocking IgG antibodies contribute to successful allergen immunotherapy (AIT), however, not much is known about their affinity. Since affinity measurements of polyclonal antibodies in serum are technically challenging we evaluated the applicability of acidic disruption of antibody-allergen complexes by a modified ELISA protocol with monoclonal antibodies (mAbs) specific for the relevant major allergens Betv1 and Mald1. Then, AIT-induced blocking and non-blocking Mald1-specific antibodies in sera from individuals with or without reduced apple allergy were compared. After testing their pH stability coated recombinant allergens were incubated with (i) mAbs diluted in PBS or human serum and (ii) sera from individuals after sublingual administration of Mald1 or Betv1 for 16 weeks. Immune complexes were exposed to buffers in the pH range of 6.4-3.4 and residual antibodies were measured. Avidity indexes (AI), defined as the pH removing 50% of antibodies, were compared to the dissociation constants (KD) of mAbs determined by surface plasmon resonance. The selected pH range was applicable to disrupt allergen complexes with highly affine mAbs without compromising allergen integrity. AIs of mAbs accorded with KD values and were unaffected by epitope specificity or the presence of serum proteins. The inter-assay variability was <4% CV. Protective Mald1-specific IgG antibodies from individuals with reduced apple allergy showed a higher collective binding strength than that of the non-protective antibodies of individuals without reduced apple allergy. Acidic disruption of allergen-antibody complexes may be used to estimate the net-binding force of polyclonal serum antibodies and eases the investigation of affinity-related research questions in AIT.

Asthma therapy concepts through the ages.

The development and approval of DMAADs ("disease-modifying anti-asthmatic drugs"), in particular inhaled steroids (alone or in combination with long-acting bronchodilators), biologics and modern allergen immunotherapies, has fundamentally changed the asthma therapy concept from symptom control to symptom prevention. This concept is linked to the new asthma treatment goal of asthma remission: long-term absence of symptoms (good asthma control), absence of exacerbations, and stable lung function, without the use of systemic steroids for asthma therapy. Three types of asthma remission are distinguished: spontaneous remission (e.g., childhood asthma), remission "off treatment" (e.g., after successful allergen immunotherapy), and remission "on treatment" (e.g., during inhaled therapy or biologic therapy). A treat-to-target approach is used, as in rheumatoid arthritis or chronic inflammatory bowel disease: The goal is to achieve asthma remission, through individually tailored treatment with highly effective drugs with minimal side effects. However, this requires precise phenotyping of the patient, including detailed history taking, pulmonary function diagnostics, allergological diagnostics, and measurement of type 2 biomarkers.

Peptide immunotherapy for aeroallergens.

Background: Allergen specific immunotherapy (SIT) has been used for more than a century. Researchers have been working to improve efficacy and reduce the side effects. Objective: We have reviewed the literature about peptides immunotherapy for inhaled allergens. The mechanism of SIT is to induce regulatory T (Treg) cells and to reduce T helper (Th)2 cells to induce class switching from IgE to IgG and induce blocking antibodies to inhibit allergen binding of IgE. Methods: The relevant published literatures on the peptide SIT for aeroallergens have been searched on the medline. Results: Modification of allergens and routes of treatment has been performed. Among them, many researchers were interested in peptide immunotherapy. T-cell epitope peptide has no IgE epitope, that is able to bind IgE, but rather induces Treg and reduces Th2 cells, which was considered an ideal therapy. Results from cellular and animal model studies have been successful. However, in clinical studies, T-cell peptide immunotherapy has failed to show efficacy and caused side effects, because of the high effective rate of placebo and the development of IgE against T-cell epitope peptides. Currently, the modifications of IgE-allergen binding by blocking antibodies are considered for successful allergen immunotherapy. Conclusion: Newly developed hypoallergenic B cell epitope peptides and computational identification methods hold great potential to develop new peptide immunotherapies.

Lack of Harmonized Adherence Criteria in Allergen Immunotherapy Prevents Comparison of Dosing and Application Strategies: A Scoping Review

Allergen immunotherapy (AIT) involves the application of increasing doses of allergen extract (as oral, sublingual, or subcutaneous immunotherapy) until immunologic tolerance is reached. Successful AIT relies on the consistent administration of allergen extract; therefore, adherence to these treatments is vital for compelling long-term results. Our review discusses the current terminology from adherence research in general, summarizes 25 current studies on adherence research in AIT in a scoping literature review, and delineates recommendations for tools and parameters for adherence research, aiming to improve outcomes in AIT. Almost every adherence study in AIT published to date used different tools, parameters, and data sources for measuring adherence and persistence rates. Unfortunately, an easily accessible, objective parameter or biomarker for monitoring treatment adherence and success has not yet been established for AIT. This situation calls for the development of an international core outcomes set for AIT that defines what is exactly meant by AIT adherence and how AIT adherence should be consistently measured. Therefore, we exemplarily present results and conclusions from adherence research in chronic diseases other than allergology. We aim to facilitate the development of advanced methods, considering the challenging disease specificities of these parameters in a routine care setting of AIT.

Diverse immune mechanisms of allergen immunotherapy for allergic rhinitis with and without asthma

Allergen immunotherapy (AIT) is an effective treatment for allergic rhinitis, inducing long-term clinical tolerance to the sensitizing allergen. Clinical tolerance induction can be achieved when AIT is administered for at least 3 years. AIT is associated with the modulation of innate and adaptive immune systems. This comprises inhibition of IgE-dependent activation of mast cells and basophils in the local target organ, suppression of TH2 cells, immune deviation toward TH1 cells, induction of T and B regulatory cells, and production of allergen-neutralizing antibodies. However, recent developments in their underpinning mechanisms have revealed that AIT, administered subcutaneously or sublingually, induces immune regulation through novel cell targets and molecular mechanisms. This comprehensive review discusses how immune tolerance driven by subcutaneous immunotherapy and sublingual immunotherapy is associated with the induction of a novel regulatory subset of innate lymphoid cells and suppression of proinflammatory TH2, allergen-specific TH2 (TH2A), and T follicular helper cells. Moreover, they are associated with exhaustion of TH2A cells and differential expression of nasal and systemic IgA antibodies. Uncovering the underpinning mechanisms of a successful AIT and immune tolerance induction will allow the development of targeted therapeutics for allergic rhinitis with and without asthma.

The Establishment of EXiLE test for House Dust Mite Allergy

Successful allergen immunotherapy (AIT) is associated with an increase in the production of allergen-specific IgG antibodies including IgG4 which may contribute to IgE-blocking factor (IgE-BF) activity. Recently, in vitro allergy test method was reported as EXiLE (IgE crosslinking-induced luciferase expression) test using a luciferase-reporting humanized rat mast cell line. However, to understand immunomodulation effect after AIT, this functional assay has not been fully validated. This study is aimed to investigate effect of IgE-BFs induced by house dust mite (HDM)-AIT on EXiLE test. HDM allergic rhinitis patients’ sera before- and after-AIT were used. EXiLE test was performed as previously reported (Allergol. Int, 2012). Briefly, RS-ATL8 cells were plated into the wells of a 96 well-plate and incubated with diluted serum overnight. The cells were stimulated with allergen (HDM, 0.1-1000 ng/mL) without washing. Then, luciferase substrate was added to the cells, and chemiluminescence was measured. The data was expressed as luciferase expression (LE) levels which were represented as the fold increase compared with the allergen untreated conditions (baseline), and EC50 value was calculated by fitting a dose response curve. The optimal incubation time was 3hr with HDM (0.1-1000 ng/mL). The LE levels showed significant correlation with the value of Der p specific-IgE (r2>0.95). EC50 of LE levels in sera after-AIT was shifted from that in sera before-AIT (p=0.01) EXiLE test was validated using the HDM allergic rhinitis sera. This test can be utilized for immunomodulation effect induced by AIT.

House dust mite subcutaneous immunotherapy in Chinese patients with allergic asthma and rhinitis.

The efficacy of allergen immunotherapy (AIT) has been reported with different allergens including house dust mites (HDM). HDM are the most prevalent allergens in patients with asthma and/or rhinitis in China. In addition to improving symptoms, reducing medication need, and improving quality of life, AIT can change the course of allergic disease and induce allergen-specific immune tolerance. To date, the use of AIT is becoming more acceptable in China, and there are many studies about the current clinical practice immunotherapy. In this paper we discuss the main aspects of AIT undertaken in China; including symptom and medication scores, pulmonary function and airway hyperresponsiveness, specific allergen sensitivity, safety evaluation, and mechanisms underlying AIT. This review will provide some important information on AIT treatment strategies to doctors, healthcare professionals and organizations involved in the AIT in China. According to the studies in China, successful AIT may induce antibody responses and cellular reactions in relation to the significant improvement in clinical symptoms, reducing the need for medications and maintenance of stable pulmonary functions.

A Case Report of a Successful Allergen Immunotherapy with Candida Albicans in Patient with Sever Atopic Dermatitis Sensitive to Candida Albicans

Introduction: Atopic dermatitis (AD) is an inflammatory, chronic, relapsing, itchy skin disease with an immunologic basis. This disease is associated with itchy skin lesions (pruritus), dry skin (xerosis) and plaques of eczema. The role of aeroallergens, such as house dust mite (HDM) and food allergens has been proven to exacerbate skin eczema lesions. Alongside drugs such as corticosteroids, topical emollients, antihistamines, tacrolimus, and immune suppressants, phototherapy and subcutaneous immunotherapy (SCIT) also done. SCIT is mostly using for sensitization to mite allergens.Case Presentation: We present a 30 y/o Iranian woman with severe atopic dermatitis and sensitization to Candida allergens. We initiated SCIT with candida allergen and the patient had obvious improvements in her signs and symptoms 3 months after starting SCIT.Conclusion: Although subcutaneous immunotherapy was only approved for mite sensitization in atopic dermatitis, it should be considered in other aeroallergen sensitizations.

Allergoids: Ready for the Future

Improving therapy for people with allergies is a continuously evolving area and, due to the increasing prevalence of allergic diseases, options such as pharmacotherapy and allergen avoidance are inadequate alone to control these diseases. Worldwide, approximately 400 million people are affected by allergic rhinitis (AR) and 300 million people by asthma.1 Unlike anti-allergy medications, a unique feature of allergen immunotherapy (AIT) is that it modifies the underlying cause of disease,2 suggesting that it may be an optimal treatment approach. Guidelines, such as those from the European Medicines Agency (EMA) in 2009,3 provide the basis for optimising trial design for the development of new AIT preparations. A wide range of treatment modalities, including recombinant allergens, have been developed, and results from several studies, some only published in trial registries, provide clarity and insights into optimising clinical trial design even further.4-13 Lessons learned from these studies, which are scientifically informative for the community, were explored in this session. In addition, the latest results were discussed from a dose-finding trial and a Phase III trial of a new allergoid treatment in development for patients with house dust mite (HDM)-induced asthma with or without AR or allergic rhinoconjunctivitis (ARC).14,15 Since AIT is recommended to be administered for 3 years, successful AIT requires adequate patient adherence over the long-term. The last section of this review focusses on strategies to optimise existing AIT and patient care, with a particular emphasis on reducing the number of injections during dose escalation when performing subcutaneous immunotherapy (SCIT) using pollen allergoids.

Allergen immunotherapy in people, dogs, cats and horses - differences, similarities and research needs.

In human patients with seasonal allergic rhinoconjunctivitis sensitized to grass pollen, the first successful allergen immunotherapy (AIT) was reported in 1911. Today, immunotherapy is an accepted treatment for allergic asthma, allergic rhinitis and hypersensitivities to insect venom. AIT is also used for atopic dermatitis and recently for food allergy. Subcutaneous, epicutaneous, intralymphatic, oral and sublingual protocols of AIT exist. In animals, most data are available in dogs where subcutaneous AIT is an accepted treatment for atopic dermatitis. Initiating a regulatory response and a production of "blocking" IgG antibodies with AIT are similar mechanisms in human beings and dogs with allergic diseases. Although subcutaneous immunotherapy is used for atopic dermatitis in cats, data for its efficacy are sparse. There is some evidence for successful treatment of feline asthma with AIT. In horses, most studies evaluate the effect of AIT on insect hypersensitivity with conflicting results although promising pilot studies have demonstrated the prophylaxis of insect hypersensitivity with recombinant antigens of biting midges (Culicoides spp.). Optimizing AIT using allergoids, peptide immunotherapy, recombinant allergens and new adjuvants with the different administration types of allergen extracts will further improve compliance and efficacy of this proven treatment modality.

Early Intervention Improves Clinical Responses to House Dust Mite Immunotherapy in Allergic Rhinitis Patients

Background: Allergen immunotherapy (AIT) is the unique causal treatment for respiratory allergy. As AIT is expensive and of long duration, the availability of a marker predicting AIT responders is of crucial relevance. Objective: To investigate clinical parameters correlated with effective AIT in allergic rhinitis (AR) patients. Methods: This is a prospective, nonrandomized open study in which a total of 284 AR patients who had received house dust mite (HDM) subcutaneous AIT were enrolled from January 2011 to December 2015, and then followed up for 3 consecutive years. Demographic data, clinical history, laboratory tests (specific and total IgE levels), symptoms score, concomitant medication, and adverse reactions during AIT were collected. An AIT responder patient was defined when a visual analog score (assessing global symptoms) had decreased by >30% compared to baseline and concomitant medication was equal to or less than before AIT. Results: Thirty-three patients dropped out, so 251 patients were analyzed; 175 (69.7%) patients were responders. This group had a higher baseline symptom score than the AIT nonresponder group (7.5 vs. 6.9). A significant negative correlation was found between AR symptom duration and the clinical response to AIT. Local reactions (LRs) during AIT had a positive correlation. Other variables such as a family history of atopy, combined asthma history, and the levels of specific and total IgE had no correlations with effective AIT. Conclusion: Early intervention with AIT helps to improve the efficacy of AR treatment. LRs might predict successful AIT. Highly symptomatic AR patients may develop increased clinical responses to AIT.

The regulatory dendritic cell marker C1q is a potent inhibitor of allergic inflammation.

The complement subunit C1q was recently identified as a marker for monocyte-derived regulatory dendritic cells supporting the differentiation of interleukin (IL)-10-secreting CD4+ T cells with a suppressive activity. Furthermore, C1q expression is upregulated in peripheral blood mononuclear cells of allergic patients in the course of successful allergen immunotherapy. Herein, we investigated a potential direct role of C1q in downregulating allergic inflammation. In mice with ovalbumin (OVA) or birch pollen (BP)-induced allergic asthma, C1q is as efficacious as dexamethasone to reduce both airway hyperresponsiveness (AHR), eosinophil, and ILC2 infiltrates in bronchoalveolar lavages, as well as allergen-specific T helper 2 cells in the lungs. Administration of C1q does not expand IL-10+/Foxp3+ regulatory T cells in the lungs, spleen, or in the blood. Depletion of plasmacytoid dendritic cells (pDCs) abrogates the capacity of C1q to reduce AHR and eosinophilic infiltrates in OVA-sensitized mice. Also C1q treatment inhibits the activation of human and mouse pDCs by CpGs, thereby demonstrating a critical role for pDCs in the anti-inflammatory activity of C1q. We conclude that regulatory dendritic cells can mediate a potent direct anti-inflammatory activity via the expression and/or secretion of molecules such as C1q, independently of their capacity to expand the pool of regulatory T cells.

Applications and mechanisms of immunotherapy in allergic rhinitis and asthma.

Clinical and immunologic tolerance are hallmarks of successful allergen immunotherapy (AIT). Clinical benefits such as reduced symptoms, pharmacotherapy intake and improvement of quality of life persist following cessation of treatment. Successful AIT is associated with suppression of allergic inflammatory cells such as mast cells, eosinophils and basophils in target organs. Furthermore, AIT down-regulates type 2 innate lymphoid cells and allergen-specific type 2 T-helper (Th2) cells. The immunologic tolerant state following AIT is associated with the induction of distinct phenotypes of regulatory T-cells (T-regs) including interleukin (IL)-10-, IL-35- and transforming growth factor (TGF)-β- producing T-regs and FoxP3+ T-regs. B-cell responses, including the induction of IL-10+ regulatory B-cells (B-regs) and the production of IgG4-associated blocking antibodies are also induced following successful AIT. These events are associated with the suppression of antigen-specific Th2 responses and delayed immune deviation in favour of Th1 type responses. Insight into the mechanisms of AIT has allowed identification of novel biomarkers with potential to predict the clinical response to AIT and also novel therapeutic strategies for more effective and safer AIT.

High-dose bee venom exposure induces similar tolerogenic B-cell responses in allergic patients and healthy beekeepers.

The involvement of B cells in allergen tolerance induction remains largely unexplored. This study investigates the role of B cells in this process, by comparing B-cell responses in allergic patients before and during allergen immunotherapy (AIT) and naturally exposed healthy beekeepers before and during the beekeeping season. Circulating B cells were characterized by flow cytometry. Phospholipase A2 (PLA)-specific B cells were identified using dual-color staining with fluorescently labeled PLA. Expression of regulatory B-cell-associated surface markers, interleukin-10, chemokine receptors, and immunoglobulin heavy-chain isotypes, was measured. Specific and total IgG1, IgG4, IgA, and IgE from plasma as well as culture supernatants of PLA-specific cells were measured by ELISA. Strikingly, similar responses were observed in allergic patients and beekeepers after venom exposure. Both groups showed increased frequencies of plasmablasts, PLA-specific memory B cells, and IL-10-secreting CD73- CD25+ CD71+ BR 1 cells. Phospholipase A2-specific IgG4-switched memory B cells expanded after bee venom exposure. Interestingly, PLA-specific B cells showed increased CCR5 expression after high-dose allergen exposure while CXCR4, CXCR5, CCR6, and CCR7 expression remained unaffected. This study provides the first detailed characterization of allergen-specific B cells before and after bee venom tolerance induction. The observed B-cell responses in both venom immunotherapy-treated patients and naturally exposed beekeepers suggest a similar functional immunoregulatory role for B cells in allergen tolerance in both groups. These findings can be investigated in other AIT models to determine their potential as biomarkers of early and successful AIT responses.

Successful allergen immunotherapy with horse dander allergy

Allergic disease induced by animal exposure is a well-known fact. Exposure to dander of pets such as cats and dogs can lead to allergic manifestations of allergic rhinitis, allergic asthma, and contact urticaria is a well-recognized fact. Allergy to horses can also present with all these symptoms as well as anaphylaxis in the subjects who are previously sensitized. Horse allergy is induced by exposure to the major horse allergens Equ 1 through five with the level of exposure determining the level of severity. Greatest risk of anaphylaxis occurs in those sensitized patients who have large amounts of animal allergen exposure, with maximum chances with animal bite as it exposes the sensitized persons to large quantities of the animal allergen that resides in the saliva. Horse allergy may be successfully treated with allergen specific immunotherapy.

Inhibition Of Human IgE-Allergen Interaction By Antibodies Of Rabbits Immunized With Depigmented-polymerized Allergen Extracts

Successful allergen immunotherapy has been linked to an increase of blocking antibodies, mainly specific IgG. Allergoids have, in comparison with native allergens, a reduced allergenicity, whereas immunogenicity is preserved. Our objetctive was to compare the induction of blocking-IgG antibodies by a native and a depigmented-polymerized (Dpg-Pol) birch pollen extract.

IgG-Blocking Antibodies Inhibit IgE-Mediated Anaphylaxis in Vivo Through Both Antigen Interception and FcγRIIb Cross-linking

Strait RT, Morris SC, Finkelman FD. J Clin Invest. 2006;116:833–841 PURPOSE OF THE STUDY. It has been hypothesized that at least part of the mechanism of successful allergen immunotherapy is the induction of specific immunoglobulin (Ig) G that can “block” IgE-dependent responses by competitively preventing IgE binding and/or by signaling via inhibitory FcγRIIb on basophils and mast cells, thereby downregulating FcεRI-dependent signaling. In vivo evidence of this blocking function has been lacking. The authors of this study carefully addressed whether and how specific IgG can inhibit IgE-mediated anaphylaxis by using a murine model. METHODS. The authors had previously established a model of anaphylaxis by immunizing BALB/c mice with goat anti-mouse IgD antibody (GαMD), which elicits a strong T-helper 2 response with high levels of IgE, IgG1, and mastocytosis. In this study they very cleverly modified the model by conjugating GαMD with the hapten trinitrophenyl to generate a trinitrophenyl-specific IgG and IgE response to allow for more detailed evaluation of the role of different antibody isotypes with the same epitope specificity. They used a number of tools including antibody blocking of FcγRs, FcRIII-deficient knockout mice, and pharmacologic inhibition to discriminate between IgE- and IgG-dependent anaphylaxis. Assessment of anaphylaxis was by temperature drop and hemoconcentration. IgG-trinitrophenyl antibody complexes were measured by enzyme-linked immunosorbent assay (ELISA). In some experiments, interleukin 4 secretion was measured from whole blood by ELISA. RESULTS. In immunized mice, the authors showed that IgE-dependent anaphylaxis is primarily inhibited by IgG preventing antigen-induced cross-linking of cell-associated IgE. In FcγRIII-deficient mice (ie, those capable of only IgE-dependent anaphylaxis), blockade of the FcγRIIb inhibitory receptors did not exacerbate antigen-induced anaphylaxis, and IgG–antigen complexes could be detected in whole blood within 5 minutes of antigen administration. Furthermore, animals passively sensitized with IgE in the absence of IgG reacted with 1000-fold lower doses of antigen. Epitope density was inversely correlated with threshold dose for inducing reactivity. Finally, by adding back increasing amounts of IgG to the passively IgE-sensitized animals, they showed that FcRIIb does inhibit IgE responses when IgG levels are low. CONCLUSIONS. Antigen-specific IgG does block IgE-mediated reactions in vivo by both direct competition and signaling via the inhibitory IgG receptor, FcγRIIb. REVIEWER COMMENTS. This was the first in vivo demonstration of specific IgG's ability to block IgE-dependent reactions, which has been suggested for many years by the frequently observed rise in allergen-specific IgG titer during successful immunotherapy. There are potentially important large differences between the mouse model and IgG induction in the context of allergen immunotherapy, including the fact that IgG-dependent anaphylaxis is well established in mice but uncertain to even exist in humans. It is also unclear from the article how the ratio of IgG to IgE in this model compares to the ratio of IgG and IgE specific to a particular allergen in human patients. However, the authors carefully and elegantly demonstrated the capacity and mechanistic details of IgG blocking antibody in vivo and strongly support the concept of blocking IgG in the context of immunotherapy.

Increased frequency of large local reactions among systemic reactors during subcutaneous allergen immunotherapy

Large local reactions are not uncommon during allergen immunotherapy (AIT). Dosage adjustments after large local reactions are commonly instituted despite literature that suggests individual large local reactions do not seem to predict subsequent systemic reactions. To investigate the relationship between large local reactions and the risk of systemic reactions to AIT. Retrospective review of the AIT database of a large, multicenter allergy practice group was conducted between June 1, 2003, and May 31, 2005. Numbers of large local reactions in 258 patients who experienced systemic reactions to AIT were compared with 299 age-, sex-, and sensitivity-matched control patients who did not experience systemic reactions during AIT. A total of 283 systemic reactions occurred in 258 patients during the surveillance period, which included 661,123 patient visits for 1,108,621 allergy injections. The systemic reaction rate was 0.043% of visits and 0.025% of injections. The large local reaction rate was 35.2% of visits and 19.5% of injections among systemic reactors compared with 8.9% of visits and 5.3% of injections in the controls (P < .001 each). Thus, the odds of experiencing large local reactions were significantly increased among systemic reactors. Although AIT is a safe and effective immunomodulatory therapeutic option for the treatment of allergic diseases, patients with increased frequency of large local reactions may have increased risk for future systemic reactions. Identifying additional risk factors remains viable. Recognizing the relevance of frequent large local reactions is important for designing safer protocols for successful AIT in these patients.

Evaluation of serum IgG4 antibodies specific to grass pollen allergen components in the follow up of allergic patients undergoing subcutaneous and sublingual immunotherapy

Background A number of reports suggest that induction of IgG ‘blocking antibodies’ may be important for successful allergen immunotherapy. Nevertheless, a significant increase in specific IgG and IgG4 antibodies has not been consistently demonstrated for sublingual immunotherapy (SLIT). Methods The present observation included three groups of grass pollen allergic patients all submitted to three different allergen immunotherapeutic regimens in an open, non-placebo controlled clinical study: (i) 16 patients underwent a modified ‘cluster’ regimen of weekly injections of a standardized aluminium-absorbed Phleum pratense extract for 5 weeks, followed by 3 weeks of maintenance injections. (ii) Fifteen patients were treated with standardized timothy grass pollen-allergen oral vaccine. In the first session of a SLIT protocol without up-dosing, each patient received 2.4 ml of sublingual vaccine containing about 57 μg of Phl p 5 and received a maintenance dose of 24 μg of Phl p 5 once a day for 120 days. (iii) Fourteen subjects were treated with a standardized allergen extract containing 5-grass pollen mixture; a SLIT protocol was performed without up-dosing, administering a dose corresponding to about 10.0 μg/ml grass-pollen Group 5 and a maintenance dose of 4 μg of grass-pollen Group 5 once a day for 135 days. Patients’ sera were characterized in detail by determining IgG4 antibodies to rPhl p 1, 2, 5, 6, 7, 11, 12 and nPhl p 4 and eosinophil cationic protein before the start of immunotherapy and during the peak of pollen season. Results No relevant systemic side effects were registered in patients treated with the modified cluster subcutaneous immunotherapy (SCIT) protocol and the two SLIT protocols without build-up phase. After SCIT all patients had high titres of serum allergen-specific IgG4 antibodies. High-dose SLIT led to an IgG4 increase comprising 25% of the SCIT level, while low-dose SLIT increased to 4% of the SCIT. Furthermore, the increase of specific IgG antibodies corresponds to a decrease of serum ECP during allergen exposure. Conclusions These preliminary data seem to indicate that: (i) pre-seasonal high-dose SLIT protocol without build-up phase is safe and well-tolerated by allergic patients; (ii) compared to IgG4 levels induced by SCIT, only a high-dose SLIT regimen results in an appreciable serum specific IgG4 increase.

Osteopontin: A potential biomarker for successful allergen immunotherapy

Rationale Both biomarkers and regulatory mechanisms related to successful allergen immunotherapy remain undefined. To clarify these further, we have investigated patterns of gene expression in peripheral blood mononuclear cells (PBMCs) of venom allergic patients pre and post venom immunotherapy (VIT). Methods Gene-chip array analysis was performed to identify differential gene expression in the PBMCs of 3 study groups: (1): 3 subjects not on VIT (no-VIT); (2): 3 subjects on VIT for <1 year (VIT); (3): 3 subjects who completed 5-7 yrs of VIT and discontinued VIT for 4-6 yrs (disc-VIT). Patterns of gene expression were identified using high-order bioinformatics tools and GeneSpring software. Serum osteopontin (OPN) was measured by EIA. Results Gene clustering analysis of expression profiles identified a unique cluster of genes with lower levels of expression for OPN in the “no VIT” group as compared to the “VIT”, and particularly the “disc-VIT” groups. These osteopontin expression profiles results were confirmed by quantitative RT-PCR. A significantly higher level of serum OPN was measured in the “disc-VIT” group as compared to the “no VIT” group (n=17 in each group; p<0.001). For comparative purpose, 8 subjects receiving ragweed immunotherapy [dose: Amb1 4.8-24μg/injection, duration: 4 years] had been analyzed for serum OPN levels and similar results were obtained. Conclusions OPN is reportedly involved in Th1-associated immune responses. Our data support OPN's use as a biomarker and possibly a regulatory molecule in allergen immunotherapy.