Omalizumab for the Treatment of Multiple Food Allergies.

BACKGROUND: Food allergies are common and are associated with substantial morbidity; the only approved treatment is oral immunotherapy for peanut allergy. METHODS: In this trial, we assessed whether omalizumab, a monoclonal anti-IgE antibody, would be effective and safe as monotherapy in patients with multiple food allergies. Persons 1 to 55 years of age who were allergic to peanuts and at least two other trial-specified foods (cashew, milk, egg, walnut, wheat, and hazelnut) were screened. Inclusion required a reaction to a food challenge of 100 mg or less of peanut protein and 300 mg or less of the two other foods. Participants were randomly assigned, in a 2:1 ratio, to receive omalizumab or placebo administered subcutaneously (with the dose based on weight and IgE levels) every 2 to 4 weeks for 16 to 20 weeks, after which the challenges were repeated. The primary end point was ingestion of peanut protein in a single dose of 600 mg or more without dose-limiting symptoms. The three key secondary end points were the consumption of cashew, of milk, and of egg in single doses of at least 1000 mg each without dose-limiting symptoms. The first 60 participants (59 of whom were children or adolescents) who completed this first stage were enrolled in a 24-week open-label extension. RESULTS: Of the 462 persons who were screened, 180 underwent randomization. The analysis population consisted of the 177 children and adolescents (1 to 17 years of age). A total of 79 of the 118 participants (67%) receiving omalizumab met the primary end-point criteria, as compared with 4 of the 59 participants (7%) receiving placebo (P<0.001). Results for the key secondary end points were consistent with those of the primary end point (cashew, 41% vs. 3%; milk, 66% vs. 10%; egg, 67% vs. 0%; P<0.001 for all comparisons). Safety end points did not differ between the groups, aside from more injection-site reactions in the omalizumab group. CONCLUSIONS: In persons as young as 1 year of age with multiple food allergies, omalizumab treatment for 16 weeks was superior to placebo in increasing the reaction threshold for peanut and other common food allergens. (Funded by the National Institute of Allergy and Infectious Diseases and others; ClinicalTrials.gov number, NCT03881696.).

Targeting inhibitory Siglec-3 to suppress IgE-mediated human basophil degranulation.

BACKGROUND: Sialic acid-binding immunoglobulin-like lectin-3 (Siglec-3 [CD33]) is a major Siglec expressed on human mast cells and basophils; engagement of CD33 leads to inhibition of cellular signaling via immunoreceptor tyrosine-based inhibitory motifs. OBJECTIVE: We sought to inhibit human basophil degranulation by simultaneously recruiting inhibitory CD33 to the IgE-FcεRI complex by using monoclonal anti-IgE directly conjugated to CD33 ligand (CD33L). METHODS: Direct and indirect basophil activation tests (BATs) were used to assess both antigen-specific (peanut) and antigen-nonspecific (polyclonal anti-IgE) stimulation. Whole blood from donors with allergy was used for direct BAT, whereas blood from donors with nonfood allergy was passively sensitized with plasma from donors with peanut allergy in the indirect BAT. Blood was incubated with anti-IgE-CD33L or controls for 1 hour or overnight and then stimulated with peanut, polyclonal anti-IgE, or N-formylmethionyl-leucyl-phenylalanine for 30 minutes. Degranulation was determined by measuring CD63 expression on the basophil surface by flow cytometry. RESULTS: Incubation for 1 hour with anti-IgE-CD33L significantly reduced basophil degranulation after both allergen-induced (peanut) and polyclonal anti-IgE stimulation, with further suppression after overnight incubation with anti-IgE-CD33L. As expected, anti-IgE-CD33L did not block basophil degranulation due to N-formylmethionyl-leucyl-phenylalanine, providing evidence that this inhibition is IgE pathway-specific. Finally, CD33L is necessary for this suppression, as monoclonal anti-IgE without CD33L was unable to reduce basophil degranulation. CONCLUSIONS: Pretreating human basophils with anti-IgE-CD33L significantly suppressed basophil degranulation through the IgE-FcεRI complex. The ability to abrogate IgE-mediated basophil degranulation is of particular interest, as treatment with anti-IgE-CD33L before antigen exposure could have broad implications for the treatment of food, drug, and environmental allergies.

Desensitization and remission after peanut sublingual immunotherapy in 1- to 4-year-old peanut-allergic children: A randomized, placebo-controlled trial.

BACKGROUND: Prior studies of peanut sublingual immunotherapy (SLIT) have suggested a potential advantage with younger age at treatment initiation. OBJECTIVE: We studied the safety and efficacy of SLIT for peanut allergy in 1- to 4-year-old children. METHODS: Peanut-allergic 1- to 4-year-old children were randomized to receive 4 mg peanut SLIT versus placebo. Desensitization was assessed by double-blind, placebo-controlled food challenge (DBPCFC) after 36 months of treatment. Participants desensitized to at least 443 mg peanut protein discontinued therapy for 3 months and then underwent DBPCFC to assess for remission. Biomarkers were measured at baseline and longitudinally during treatment. RESULTS: Fifty participants (25 peanut SLIT, 25 placebo) with a median age of 2.4 years were enrolled across 2 sites. The primary end point of desensitization was met with actively treated versus placebo participants having a significantly greater median cumulative tolerated dose (4443 mg vs 143 mg), higher likelihood of passing the month 36 DBPCFC (60% vs 0), and higher likelihood of demonstrating remission (48% vs 0). The highest rate of desensitization and remission was seen in 1- to 2-year-olds, followed by 2- to 3-year-olds and 3- to 4-year-olds. Longitudinal changes in peanut skin prick testing, peanut-specific IgG4, and peanut-specific IgG4/IgE ratio were seen in peanut SLIT but not placebo participants. Oropharyngeal itching was more commonly reported by peanut SLIT than placebo participants. Skin, gastrointestinal, upper respiratory, lower respiratory, and multisystem adverse events were similar between treatment groups. CONCLUSION: Peanut SLIT safely induces desensitization and remission in 1- to 4-year-old children, with improved outcomes seen with younger age at initiation.

A mutation in Themis contributes to anaphylaxis severity following oral peanut challenge in CC027 mice.

BACKGROUND: The development of peanut allergy is due to a combination of genetic and environmental factors, although specific genes have proven difficult to identify. Previously, we reported that peanut-sensitized Collaborative Cross strain CC027/GeniUnc (CC027) mice develop anaphylaxis upon oral challenge to peanut, in contrast to C3H/HeJ (C3H) mice. OBJECTIVE: This study aimed to determine the genetic basis of orally induced anaphylaxis to peanut in CC027 mice. METHODS: A genetic mapping population between CC027 and C3H mice was designed to identify the genetic factors that drive oral anaphylaxis. A total of 356 CC027xC3H backcrossed mice were generated, sensitized to peanut, then challenged to peanut by oral gavage. Anaphylaxis and peanut-specific IgE were quantified for all mice. T-cell phenotyping was conducted on CC027 mice and 5 additional Collaborative Cross strains. RESULTS: Anaphylaxis to peanut was absent in 77% of backcrossed mice, with 19% showing moderate anaphylaxis and 4% having severe anaphylaxis. There were 8 genetic loci associated with variation in response to peanut challenge-6 associated with anaphylaxis (temperature decrease) and 2 associated with peanut-specific IgE levels. There were 2 major loci that impacted multiple aspects of the severity of acute anaphylaxis, at which the CC027 allele was associated with worse outcome. At one of these loci, CC027 has a private genetic variant in the Themis gene. Consistent with described functions of Themis, we found that CC027 mice have more immature T cells with fewer CD8+, CD4+, and CD4+CD25+CD127- regulatory T cells. CONCLUSIONS: Our results demonstrate a key role for Themis in the orally reactive CC027 mouse model of peanut allergy.

Different airborne particulates trigger distinct immune pathways leading to peanut allergy in a mouse model.

BACKGROUND: Environmental exposure to peanut through non-oral routes is a risk factor for peanut allergy. Early-life exposure to air pollutants, including particulate matter (PM), is associated with sensitization to foods through unknown mechanisms. We investigated whether PM promotes sensitization to environmental peanut and the development of peanut allergy in a mouse model. METHODS: C57BL/6J mice were co-exposed to peanut and either urban particulate matter (UPM) or diesel exhaust particles (DEP) via the airways and assessed for peanut sensitization and development of anaphylaxis following peanut challenge. Peanut-specific CD4+ T helper (Th) cell responses were characterized by flow cytometry and Th cytokine production. Mice lacking select innate immune signaling genes were used to study mechanisms of PM-induced peanut allergy. RESULTS: Airway co-exposure to peanut and either UPM- or DEP-induced systemic sensitization to peanut and anaphylaxis following peanut challenge. Exposure to UPM or DEP triggered activation and migration of lung dendritic cells to draining lymph nodes and induction of peanut-specific CD4+ Th cells. UPM- and DEP-induced distinct Th responses, but both stimulated expansion of T follicular helper (Tfh) cells essential for peanut allergy development. MyD88 signaling was critical for UPM- and DEP-induced peanut allergy, whereas TLR4 signaling was dispensable. DEP-induced peanut allergy and Tfh-cell differentiation depended on IL-1 but not IL-33 signaling, whereas neither cytokine alone was necessary for UPM-mediated sensitization. CONCLUSION: Environmental co-exposure to peanut and PM induces peanut-specific Tfh cells and peanut allergy in mice.

Open-label study of the efficacy, safety, and durability of peanut sublingual immunotherapy in peanut-allergic children.

BACKGROUND: Studies on the efficacy of peanut sublingual immunotherapy (SLIT) are limited. The durability of desensitization after SLIT has not been well described. OBJECTIVE: We sought to evaluate the efficacy and safety of 4-mg peanut SLIT and persistence of desensitization after SLIT discontinuation. METHODS: Challenge-proven peanut-allergic 1- to 11-year-old children were treated with open-label 4-mg peanut SLIT for 48 months. Desensitization after peanut SLIT was assessed by a 5000-mg double-blind, placebo-controlled food challenge (DBPCFC). A novel randomly assigned avoidance period of 1 to 17 weeks was followed by the DBPCFC. Skin prick test results immunoglobulin levels, basophil activation test results, TH1, TH2, and IL-10 cytokines were measured longitudinally. Safety was assessed through patient-reported home diaries. RESULTS: Fifty-four participants were enrolled and 47 (87%) completed peanut SLIT and the 48-month DBPCFC per protocol. The mean successfully consumed dose (SCD) during the DBPCFC increased from 48 to 2723 mg of peanut protein after SLIT (P < .0001), with 70% achieving clinically significant desensitization (SCD > 800 mg) and 36% achieving full desensitization (SCD = 5000 mg). Modeled median time to loss of clinically significant desensitization was 22 weeks. Peanut skin prick test; peanut-specific IgE, IgG4, and IgG4/IgE ratio; and peanut-stimulated basophil activation test, IL-4, IL-5, IL-13, IFN-γ, and IL-10 changed significantly compared with baseline, with changes seen as early as 6 months. Median rate of reaction per dose was 0.5%, with transient oropharyngeal itching being the most common, and there were no dosing symptoms requiring epinephrine. CONCLUSIONS: In this open-label, prospective study, peanut SLIT was safe and induced clinically significant desensitization in most of the children, lasting more than 17 weeks after discontinuation of therapy.

A single priming event prevents oral tolerance to peanut.

BACKGROUND: Indoor dust (ID) is a source of peanut proteins and immunostimulatory adjuvants (e.g. LPS) that can promote airway sensitization to peanut. We aimed to determine whether a single airway exposure to peanut plus adjuvant is sufficient to prevent oral tolerance. METHODS: To determine the effect of a single priming event, C57BL/6J mice were exposed once to peanut plus adjuvant through the airway, followed by either airway or low-dose oral exposure to peanut, and assessed for peanut allergy. Oral tolerance was investigated by feeding high-dose peanut followed by airway sensitization. To determine whether a single priming could prevent oral tolerance, the high-dose peanut regimen was applied after a single airway exposure to peanut plus adjuvant. Peanut-specific IgE and IgG1 were quantified, and mice were challenged to peanut to assess allergy. Peanut-specific CD4+ memory T cells (CD4+ TCRß+ CD44hi CD154+ ) were quantified in mediastinal lymph nodes following airway priming. RESULTS: Mice co-exposed to peanut with LPS or ID through the airway were primed to develop peanut allergy after subsequent low-dose oral or airway exposures to peanut. Oral tolerance was induced in mice fed high-dose peanut prior to airway sensitization. In contrast, mice fed high-dose peanut following a single airway exposure to peanut plus adjuvant led to allergy. Peanut-specific CD4+ memory T cells were detected as early as 7 days after the single airway priming with peanut plus adjuvant, however, delaying peanut feeding even 1 day following priming led to allergy, whereas peanut feeding the same day as priming led to tolerance. CONCLUSIONS: A single airway exposure to peanut plus adjuvant is sufficient to prime the immune system to develop allergy following subsequent high-dose oral exposure. These results highlight the importance of introducing peanut as early as possible to prevent sensitization through a non-oral priming event.

Targeting CD22 on memory B cells to induce tolerance to peanut allergens.

BACKGROUND: Circulating IgE and subsequent severe allergic reactions to peanut are sustained and propagated by recall of peanut allergen-specific memory B cells. OBJECTIVES: This study aimed to determine whether targeting mouse and human CD22 on peanut-specific memory B cells induces tolerance to peanut allergens. METHODS: Siglec-engaging tolerance-inducing antigenic liposomes (STALs) codisplaying peanut allergens (Ara h 1, Ara h 2, or Ara h 3) and high-affinity CD22 ligand (CD22L-STALs) were employed in various mouse models (BALB/cJ, C57BL/6, human CD22 transgenic, and NSG) of peanut allergy. To investigate memory B cells, a conferred memory model was used in which splenocytes from peanut-sensitized mice were transferred into naive animals. Reconstituted mice received either CD22L-STALs or an immunogenic liposome control, followed by a peanut allergen boost and later a challenge with individual peanut allergens. To assess the effects of CD22L-STALs on human B cells, PBMCs were injected into NSG mice, followed by administration of human CD22L-STALs (hCD22L-STALs) and later a whole peanut extract boost. Blood was collected to quantify WPE- and Ara h 1-, 2-, and 3-specific immunoglobulins. RESULTS: Mouse CD22L-STALs (mCD22L-STALs) significantly suppressed systemic memory to Ara h 1, Ara h 2, and Ara h 3 in BALB/cJ and C57BL/6 mice, as demonstrated by reduced allergen-specific IgE, IgG1, and anaphylaxis on challenge. Importantly, 2 doses of mCD22L-STALs led to prolonged tolerance for at least 3 months. hCD22L-STALs displayed similar suppression in mice expressing human CD22 on B cells. Finally, human B cells were tolerized in vivo in NSG mice by hCD22L-STALs. CONCLUSIONS: Antigen-specific exploitation of CD22 on memory B cells can induce systemic immune tolerance.

Kinetics of basophil hyporesponsiveness during short-course peanut oral immunotherapy.

BACKGROUND: Oral immunotherapy (OIT) leads to suppression of mast cell and basophil degranulation along with changes in the adaptive immune response. OBJECTIVES: This study aimed to determine how rapidly these effects occur during OIT and more broadly, the kinetics of basophil and mast cell suppression throughout the course of therapy. METHODS: Twenty participants, age 4 to 12 years, were enrolled in a peanut OIT trial and assessed for desensitization and sustained unresponsiveness after 9 months of therapy. Blood was collected 5 times in the first month and then intermittently throughout to quantify immunoglobulins and assess basophil activation by CD63, CD203c, and phosphorylated SYK (pSYK). RESULTS: Twelve of 16 participants that completed the trial were desensitized after OIT, with 9 achieving sustained unresponsiveness after discontinuing OIT for 4 weeks. Basophil hyporesponsiveness, defined by lower CD63 expression, was detected as early as day 90. pSYK was correlated with CD63 expression, and there was a significant decrease in pSYK by day 250. CD203c expression remained unchanged throughout therapy. Interestingly, although basophil activation was decreased across the cohort during OIT, basophil activation did not correlate with individual clinical outcomes. Serum peanut-specific IgG4 and IgA increased throughout therapy, whereas IgE remained unchanged. CONCLUSIONS: Suppression of basophil activation occurs within the first 90 days of peanut OIT, ultimately leading to suppression of signaling through pSYK.

Manipulating the microbiome to enhance oral tolerance in food allergy.

Loss of oral tolerance (OT) to food antigens results in food allergies. One component of achieving OT is the symbiotic microorganisms living in the gut (microbiota). The composition of the microbiota can drive either pro-tolerogenic or pro-inflammatory responses against dietary antigens though interactions with the local immune cells within the gut. Products from bacterial fermentation, such as butyrate, are one of the main communication molecules involved in this interaction, however, this is released by a subset of bacterial species. Thus, strategies to specifically expand these bacteria with protolerogenic properties have been explored to complement oral immunotherapy in food allergy. These approaches either provide digestible biomolecules to induce beneficial bacteria species (prebiotics) or the direct administration of live bacteria species (probiotics). While this combined therapy has shown positive outcomes in clinical trials for cow's milk allergy, more research is needed to determine if this therapy can be extended to other food allergens.

The airway as a route of sensitization to peanut: An update to the dual allergen exposure hypothesis.

Food allergies have increased at an alarming rate over the past 2 decades, indicating that environmental factors are driving disease progression. It has been postulated that sensitization to foods, in particular, peanut, occurs through impaired skin. Peanut allergens have been quantified in household dust and may be the culprit source. Indeed, TH2 cell-skewing innate cytokines can be driven by application of food antigens on both intact and impaired skin of mice, resulting in antigen-specific IgE production and anaphylaxis following allergen exposure. However, allergy induction through the skin can be prevented by induction of oral tolerance before skin exposure. These observations led to the dual allergen exposure hypothesis, according to which oral exposure to food antigens leads to tolerance and antigen exposure on impaired skin leads to allergy. Here, we propose the airway as an alternative route of sensitization in the dual allergen exposure hypothesis that leads to food allergy. Specifically, we will provide evidence from mouse models and human cell-based studies that together implicate the airway as a plausible route of sensitization.

Mechanisms of oral immunotherapy.

Food allergy presents a significant global health concern with up to 10% of the population affected in developed nations and a steadily increasing prevalence. In many cases, particularly with peanut, tree nut and shellfish, food allergy is a lifelong and potentially life-threatening diagnosis. While no 'cure' for IgE-mediated food allergy exists, oral immunotherapy (OIT) is a promising treatment modality with the peanut OIT drug Palforzia (Aimmune Therapeutics) the only treatment for food allergy that is currently approved by the United States Food and Drug Administration. OIT primarily induces a state of desensitization with only a minority of subjects achieving sustained unresponsiveness, a state of limited clinical remission that appears to be immunologically distinct from natural tolerance. Early humoural changes during OIT include an initial increase in allergen-specific IgE, which eventually decreases to below baseline levels as OIT progresses, and a gradual increase in allergen-specific IgA and IgG4 that continues throughout the course of OIT. Basophil hyporesponsiveness and decreased skin prick test wheal size are observed within the first year of OIT, and persistence after completion of therapy has been associated with sustained unresponsiveness. In the T-cell compartment, there is an initial expansion followed by a decline in the number and activity of T helper 2 (TH 2) cells, the latter of which may be dependent on an expansion of IL-10-producing cells, including regulatory T-cells. Our understanding of the immunomodulatory effects of OIT continues to evolve, with new technologies such as single-cell transcriptional profiling and antibody epitope analysis allowing for more detailed study of T-cell and B-cell responses to OIT. In this review, we present evidence to illustrate what is currently known about the immunologic changes induced by OIT, explore potential mechanisms and emphasize knowledge gaps where future research is needed.

Long-term sublingual immunotherapy for peanut allergy in children: Clinical and immunologic evidence of desensitization.

BACKGROUND: Peanut sublingual immunotherapy (SLIT) for 1 year has been shown to induce modest clinical desensitization in allergic children. Studies of oral immunotherapy, epicutaneous immunotherapy, and SLIT have suggested additional benefit with extended treatment. OBJECTIVE: We sought to investigate the safety, clinical effectiveness, and immunologic changes with long-term SLIT in children with peanut allergy. METHODS: Children with peanut allergy aged 1 to 11 years underwent extended maintenance SLIT with 2 mg/d peanut protein for up to 5 years. Subjects with peanut skin test wheals of less than 5 mm and peanut-specific IgE levels of less than 15 kU/L were allowed to discontinue therapy early. Desensitization was assessed through a double-blind, placebo-controlled food challenge (DBPCFC) with up to 5000 mg of peanut protein after completion of SLIT dosing. Sustained unresponsiveness was further assessed by using identical DBPCFCs after 2 to 4 weeks without peanut exposure. RESULTS: Thirty-seven of 48 subjects completed 3 to 5 years of peanut SLIT, with 67% (32/48) successfully consuming 750 mg or more during DBPCFCs. Furthermore, 25% (12/48) passed the 5000-mg DBPCFC without clinical symptoms, with 10 of these 12 demonstrating sustained unresponsiveness after 2 to 4 weeks. Side effects were reported with 4.8% of doses, with transient oropharyngeal itching reported most commonly. Side effects requiring antihistamine treatment were uncommon (0.21%), and no epinephrine was administered. Peanut skin test wheals, peanut-specific IgE levels, and basophil activation decreased significantly, and peanut-specific IgG4 levels increased significantly after peanut SLIT. CONCLUSION: Extended-therapy peanut SLIT provided clinically meaningful desensitization in the majority of children with peanut allergy that was balanced with ease of administration and a favorable safety profile.

Genetic diversity between mouse strains allows identification of the CC027/GeniUnc strain as an orally reactive model of peanut allergy.

BACKGROUND: Improved animal models are needed to understand the genetic and environmental factors that contribute to food allergy. OBJECTIVE: We sought to assess food allergy phenotypes in a genetically diverse collection of mice. METHODS: We selected 16 Collaborative Cross (CC) mouse strains, as well as the classic inbred C57BL/6J, C3H/HeJ, and BALB/cJ strains, for screening. Female mice were sensitized to peanut intragastrically with or without cholera toxin and then challenged with peanut by means of oral gavage or intraperitoneal injection and assessed for anaphylaxis. Peanut-specific immunoglobulins, T-cell cytokines, regulatory T cells, mast cells, and basophils were quantified. RESULTS: Eleven of the 16 CC strains had allergic reactions to intraperitoneal peanut challenge, whereas only CC027/GeniUnc mice reproducibly experienced severe symptoms after oral food challenge (OFC). CC027/GeniUnc, C3H/HeJ, and C57BL/6J mice all mounted a TH2 response against peanut, leading to production of IL-4 and IgE, but only the CC027/GeniUnc mice reacted to OFC. Orally induced anaphylaxis in CC027/GeniUnc mice was correlated with serum levels of Ara h 2 in circulation but not with allergen-specific IgE or mucosal mast cell protease 1 levels, indicating systemic allergen absorption is important for anaphylaxis through the gastrointestinal tract. Furthermore, CC027/GeniUnc, but not C3H/HeJ or BALB/cJ, mice can be sensitized in the absence of cholera toxin and react on OFC to peanut. CONCLUSIONS: We have identified and characterized CC027/GeniUnc mice as a strain that is genetically susceptible to peanut allergy and prone to severe reactions after OFC. More broadly, these findings demonstrate the untapped potential of the CC population in developing novel models for allergy research.

Model of Walnut Allergy in CC027/GeniUnc Mice Recapitulates Key Features of Human Disease.

Tree nut allergies affect 1% of the United States population, are often severe in nature and rarely outgrown. Despite the severity and prevalence, there are no FDA-approved treatments for tree nut allergy. Development of a therapeutic would be expedited by having a mouse model that mimics the human disease. We utilized the CC027/GeniUnc mouse strain, which was previously identified as an orally reactive model of peanut allergy, to develop a model of walnut allergy. Mice were sensitized with walnut and cholera toxin for 4 weeks and subsequently challenged by oral gavage. Blood samples were collected to measure serum IgE. Walnut-sensitized mice produced high levels of walnut-IgE and were cross-sensitized to pecan. Oral challenges with walnut resulted in severe anaphylaxis and accompanying allergic symptoms. Importantly, pecan challenges also led to severe allergic reactions, indicating cross-reactivity to pecan. Overall, this novel mouse model reproduces key characteristics of human walnut allergy, which provides a platform to develop novel therapies and better understand sensitization mechanisms.

Indoor dust acts as an adjuvant to promote sensitization to peanut through the airway.

BACKGROUND: There is growing evidence that environmental peanut exposure through non-oral routes, including the skin and respiratory tract, can result in peanut sensitization. Environmental adjuvants in indoor dust can promote sensitization to inhaled antigens, but whether they contribute to peanut allergy development is unclear. OBJECTIVE: We investigated whether indoor dust promotes airway sensitization to peanut and peanut allergy development in mice. METHODS: Female and male C57BL/6J mice were exposed via the airways to peanut, indoor dust extract, or both for 2 weeks. Mice were then challenged with peanut and assessed for anaphylaxis. Peanut-specific immunoglobulins, peanut uptake by lung conventional dendritic cells (cDCs), lung innate cytokines, and T cell differentiation in lung-draining lymph nodes were quantified. Innate cytokine production by primary human bronchial epithelial cells exposed to indoor dust was also determined. RESULTS: Inhalational exposure to low levels of peanut in combination with indoor dust, but neither alone, resulted in production of peanut-specific IgE and development of anaphylaxis upon peanut challenge. Indoor dust triggered production of innate cytokines in murine lungs and in primary human bronchial epithelial cells. Additionally, inhaled indoor dust stimulated maturation and migration of peanut-laden lung type 1 cDCs to draining lymph nodes. Inhalational exposure to peanut and indoor dust induced peanut-specific T helper 2 cell differentiation and accumulation of T follicular helper cells in draining lymph nodes, which were associated with increased B cell numbers and peanut-specific immunoglobulin production. CONCLUSIONS & CLINICAL RELEVANCE: Indoor dust promotes airway sensitization to peanut and development of peanut allergy in mice. Our findings suggest that environmental adjuvants in indoor dust may be determinants of peanut allergy development in children.

Blocking antibodies induced by peanut oral and sublingual immunotherapy suppress basophil activation and are associated with sustained unresponsiveness.

BACKGROUND: Oral and sublingual immunotherapies for peanut allergy have demonstrated efficacy in small clinical trials; however, mechanisms and biomarkers correlating with clinical outcomes remain elusive. Previous studies have demonstrated a role for IgG in post-OIT plasma in the suppression of IgE-mediated mast cell reactions. OBJECTIVE: The aim of this study was to characterize the role that peanut oral and sublingual immunotherapy-induced plasma factors play in the inhibition of ex vivo basophil activation and whether inhibitory activity is associated with clinical outcomes. METHODS: Plasma samples from subjects on placebo, peanut oral immunotherapy (OIT) or peanut sublingual immunotherapy (SLIT), and IgG-depleted plasma or the IgG fraction were incubated with sensitized basophils, and the inhibition of basophil activation following stimulation with peanut extract was measured. Basophil inhibition results were compared between the two routes of immunotherapy, time on treatment and clinical outcomes. RESULTS: Plasma from subjects after 12 months of active peanut OIT, but not placebo, inhibits basophil activation ex vivo. Depletion of IgG abrogated the blocking effect of OIT plasma, while the IgG fraction substantially blocked basophil activation. Basophils are inhibited to a similar extent by undiluted OIT and SLIT plasma; however, diluted OIT plasma from the time of desensitization challenge inhibited basophils more than diluted SLIT plasma from time of desensitization challenge. Plasma from subjects who experienced sustained unresponsiveness following OIT inhibited basophils to a greater extent than plasma from subjects who were desensitized, but this was not true for SLIT. CONCLUSIONS AND CLINICAL RELEVANCE: Peanut immunotherapy induces IgG-dependent functional changes in plasma that are associated with OIT but not SLIT clinical outcomes. Understanding the mechanisms of peanut OIT and SLIT may help derive informative biomarkers.

High- and low-dose oral immunotherapy similarly suppress pro-allergic cytokines and basophil activation in young children.

BACKGROUND: Mechanisms underlying oral immunotherapy (OIT) are unclear and the effects on immune cells at varying maintenance doses are unknown. OBJECTIVE: We aimed to determine the immunologic changes caused by peanut OIT in preschool aged children and determine the effect on these immune responses in groups ingesting low or high-dose peanut OIT (300 mg or 3000 mg, respectively) as maintenance therapy. METHODS: Blood was drawn at several time-points throughout the OIT protocol and PBMCs isolated and cultured with peanut antigens. Secreted cytokines were quantified via multiplex assay, whereas Treg and peanut-responsive CD4 T cells were studied with flow cytometry. Basophil activation assays were also conducted. RESULTS: Th2-, Th1-, Th9- and Tr1-type cytokines decreased over the course of OIT in groups on high- and low-dose OIT. There were no significant differences detected in cytokine changes between the high- and low-dose groups. The initial increase in both the number of peanut-responsive CD4 T cells and the number of Tregs was transient and no significant differences were found between groups. Basophil activation following peanut stimulation was decreased over the course of OIT and associated with increased peanut-IgG4/IgE ratios. No differences were found between high- and low-dose groups in basophil activation at the time of desensitization or sustained unresponsiveness oral food challenges. CONCLUSIONS AND CLINICAL RELEVANCE: Peanut OIT leads to decreases in pro-allergic cytokines, including IL-5, IL-13, and IL-9 and decreased basophil activation. No differences in T cell or basophil responses were found between subjects on low or high-dose maintenance OIT, which has implications for clinical dosing strategies.

IgE binding to linear epitopes of Ara h 2 in peanut allergic preschool children undergoing oral Immunotherapy.

BACKGROUND: For patients with peanut allergy, there are currently no methods to predict who will develop sustained unresponsiveness (SU) after oral immunotherapy (OIT). OBJECTIVE: Assess IgE binding to peanut (PN), Ara h 2, and specific linear epitopes of Ara h 2 as predictors of the important clinical parameters: eliciting dose threshold and attainment of SU following OIT. METHODS: Samples and clinical data were collected from children undergoing OIT. PN- and Ara h 2-sIgE were quantified by ImmunoCAP® . IgE binding to linear peptides of Ara h 2 and Ara h 6 was measured with peptide microarrays. RESULTS: Values of PN-sIgE correlated with eliciting dose (P = .001) and with a higher likelihood of achieving SU (P < .0001), but these relationships were lost at higher values for PN-sIgE (≥14 kIU for eliciting dose and ≥35 kIU/L for SU). In subjects with PN-sIgE ≥ 14 kIU/L, binding of IgE to epitopes 5 and 6 of Ara h 2 was associated with a lower eliciting dose at baseline challenge (P < .001; Pc  < .02). In subjects with PN-sIgE ≥ 35 kIU/L, a combined model of IgE binding to epitopes 1, 5 and 6 with PN-sIgE was highly predictive of attainment of SU (AUC of 0.86; P = .0067). CONCLUSION: In young patients with peanut allergy, measurement of PN-sIgE and IgE binding to specific linear epitopes of Ara h 2 in baseline samples may allow stratification of patients regarding sensitivity to challenge and outcome of OIT.

Biomarkers in Food Allergy Immunotherapy.

PURPOSE OF REVIEW: Investigational allergen immunotherapies (AITs) including oral immunotherapy (OIT), sublingual immunotherapy (SLIT), and epicutaneous immunotherapy (EPIT) have proven to increase allergen thresholds required to elicit an allergic reaction in a majority of subjects. However, these studies lack consistent biomarkers to predict therapy outcomes. Here, we will review biomarkers that are currently being investigated for AIT. RECENT FINDINGS: The mechanisms underlying the therapeutic benefit of AIT involve various cell types, including mast cells, basophils, T cells, and B cells. Skin prick and basophil activation tests assess effector cell sensitivity to allergen and are decreased in subjects on AIT. Allergen-specific IgE increases initially and decreases with continued therapy, while allergen-specific IgG and IgA increase throughout therapy. Allergen-induced regulatory T cells (Tregs) increase throughout therapy and were found to be associated with sustained unresponsiveness after OIT. Subjects on OIT and SLIT have decreased Th2 cytokine production during therapy. Although trends have been reported, a common limitation of these biomarkers is that none are able to reproducibly predict prognosis during AIT. Further studies are needed to expand the currently available biomarker repertoire to provide personalized approaches to AIT.