Background Oral immunotherapy (OIT) is an effective experimental food allergy treatment that is limited by treatment withdrawal and the frequent reversibility of desensitization if interrupted. Newly ...diagnosed preschool children may have clinical and immunological characteristics more amenable to treatment. Objective We sought to test the safety, effectiveness, and feasibility of early OIT (E-OIT) in the treatment of peanut allergy. Methods We enrolled 40 children aged 9 to 36 months with suspected or known peanut allergy. Qualifying subjects reacted to peanut during an entry food challenge and were block-randomized 1:1 to receive E-OIT at goal maintenance doses of 300 or 3000 mg/d in a double-blinded fashion. The primary end point, sustained unresponsiveness at 4 weeks after stopping early intervention oral immunotherapy (4-SU), was assessed by double-blinded, placebo-controlled food challenge either upon achieving 4 prespecified criteria, or after 3 maintenance years. Peanut-specific immune responses were serially analyzed. Outcomes were compared with 154 matched standard-care controls. Results Of 40 consented subjects, 3 (7.5%) did not qualify. Overall, 29 of 37 (78%) in the intent-to-treat analysis achieved 4-SU (300-mg arm, 17 of 20 85%; 3000 mg, 12 of 17 71%, P = .43) over a median of 29 months. Per-protocol, the overall proportion achieving 4-SU was 29 of 32 (91%). Peanut-specific IgE levels significantly declined in E-OIT-treated children, who were 19 times more likely to successfully consume dietary peanut than matched standard-care controls, in whom peanut-specific IgE levels significantly increased (relative risk, 19.42; 95% CI, 8.7-43.7; P < .001). Allergic side effects during E-OIT were common but all were mild to moderate. Conclusions At both doses tested, E-OIT had an acceptable safety profile and was highly successful in rapidly suppressing allergic immune responses and achieving safe dietary reintroduction.
The Wnt signaling pathways play pivotal roles in carcinogenesis. Modulation of the cell-surface abundance of Wnt receptors is emerging as an important mechanism for regulating sensitivity to Wnt ...ligands. Endocytosis and degradation of the Wnt receptors Frizzled (Fzd) and lipoprotein-related protein 6 (LRP6) are regulated by the E3 ubiquitin ligases zinc and ring finger 3 (ZNRF3) and ring finger protein 43 (RNF43), which are disrupted in cancer. In a genome-wide small interfering RNA screen, we identified the deubiquitylase ubiquitin-specific protease 6 (USP6) as a potent activator of Wnt signaling. USP6 enhances Wnt signaling by deubiquitylating Fzds, thereby increasing their cell-surface abundance. Chromosomal translocations in nodular fasciitis result in USP6 overexpression, leading to transcriptional activation of the Wnt/β-catenin pathway. Inhibition of Wnt signaling using Dickkopf-1 (DKK1) or a Porcupine (PORCN) inhibitor significantly decreased the growth of USP6-driven xenograft tumors, indicating that Wnt signaling is a key target of USP6 during tumorigenesis. Our study defines an additional route to ectopic Wnt pathway activation in human disease, and identifies a potential approach to modulate Wnt signaling for therapeutic benefit.
Taken together, these results suggest that Ah2 STALs induce antigen-specific tolerance toward the major peanut allergen, severely blunting Ah2 sIgE and sIgG1 levels and reactions upon intraperitoneal ...challenge with Ah2. Because Ah2 is only 1 of several related antigens in peanuts and sensitization was done with WPE, we also examined the impact of Ah2 STALs on sIgE levels to WPE (Fig 2, A) and to another peanut allergen, Ara h 1 (Ah1). Plates were coated with 20 μg/mL WPE, 5 μg/mL Ah2, 5 μg/mL Ah1, or 5 μg/mL CT in carbonate-bicarbonate buffer at pH 9.6 for 1 hour at 37°C. Plates were blocked with PBS containing 0.05% Tween 20 and 2% BSA for 2 hours at 37°C. Serum samples were added for 1 hour at 37°C. Detection of IgE was performed using sheep antimouse IgE (0.5 μg/mL; Binding Site, Birmingham, United Kingdom), followed by biotinylated donkey antisheep IgG (0.5 μg/mL; Accurate Chemical, Westbury, NY) and neutravidin-horseradish peroxidase (HRP; 0.2 μg/mL; Pierce) for 1 hour at 37°C. IgG1 was detected by HRP-conjugated goat anti-mouse IgG1 (Southern Biotech, Birmingham, Ala) used at 1:40,000 for 1 hour at 37°C. HRP activity was measured by blue color development of Sure Blue TMB Microwell Peroxidase Substrate (KPL, Gaithersburg, Md).
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.
Necrotizing enterocolitis (NEC) is a devastating, multifactorial disease mainly affecting the intestine of premature infants. Recent discoveries have significantly enhanced our understanding of risk ...factors, as well as, cellular and genetic mechanisms of this complex disease. Despite these advancements, no essential, single risk factor, nor the mechanism by which each risk factor affects NEC has been elucidated. Nonetheless, recent research indicates that maternal factors, antibiotic exposure, feeding, hypoxia, and altered gut microbiota pose a threat to the underdeveloped immunity of preterm infants. Here we review predisposing factors, status of unwarranted immune responses, and microbial pathogenesis in NEC based on currently available scientific evidence. We additionally discuss novel techniques and models used to study NEC and how this research translates from the bench to the bedside into potential treatment strategies.
Oral immunotherapy (OIT) leads to suppression of mast cell and basophil degranulation along with changes in the adaptive immune response.
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.
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).
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.
Suppression of basophil activation occurs within the first 90 days of peanut OIT, ultimately leading to suppression of signaling through pSYK.
Improved animal models are needed to understand the genetic and environmental factors that contribute to food allergy.
We sought to assess food allergy phenotypes in a genetically diverse collection ...of mice.
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.
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.
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.
B-cell receptors (BCRs) play a critical role in adaptive immunity as they generate highly diverse immunoglobulin repertoires to recognize a wide variety of antigens. To better understand immune ...responses, it is critically important to establish a quantitative and rapid method to analyze BCR repertoire comprehensively. Here, we developed "Bcrip", a novel approach to characterize BCR repertoire by sequencing millions of BCR cDNA using next-generation sequencer. Using this method and quantitative real-time PCR, we analyzed expression levels and repertoires of BCRs in a total of 17 peanut allergic subjects' peripheral blood samples before and after receiving oral immunotherapy (OIT) or placebo. By our methods, we successfully identified all of variable (V), joining (J), and constant (C) regions, in an average of 79.1% of total reads and 99.6% of these VJC-mapped reads contained the C region corresponding to the isotypes that we aimed to analyze. In the 17 peanut allergic subjects' peripheral blood samples, we observed an oligoclonal enrichment of certain immunoglobulin heavy chain alpha (IGHA) and IGH gamma (IGHG) clones (P = 0.034 and P = 0.027, respectively) in peanut allergic subjects after OIT. This newly developed BCR sequencing and analysis method can be applied to investigate B-cell repertoires in various research areas, including food allergies as well as autoimmune and infectious diseases.
Food allergy is a potentially fatal disease affecting 8% of children and has become increasingly common in the past two decades. Despite the prevalence and severe nature of the disease, the ...mechanisms underlying sensitization remain to be further elucidated. The Collaborative Cross is a genetically diverse panel of inbred mice that were specifically developed to study the influence of genetics on complex diseases. Using this panel of mouse strains, we previously demonstrated CC027/GeniUnc mice, but not C3H/HeJ mice, develop peanut allergy after oral exposure to peanut in the absence of a Th2-skewing adjuvant. Here, we investigated factors associated with sensitization in CC027/GeniUnc mice following oral exposure to peanut, walnut, milk, or egg. CC027/GeniUnc mice mounted antigen-specific IgE responses to peanut, walnut and egg, but not milk, while C3H/HeJ mice were not sensitized to any antigen. Naïve CC027/GeniUnc mice had markedly lower total fecal IgA compared to C3H/HeJ, which was accompanied by stark differences in gut microbiome composition. Sensitized CC027/GeniUnc mice had significantly fewer CD3
T cells but higher numbers of CXCR5
B cells and T follicular helper cells in the mesenteric lymph nodes compared to C3H/HeJ mice, which is consistent with their relative immunoglobulin production. After oral challenge to the corresponding food, peanut- and walnut-sensitized CC027/GeniUnc mice experienced anaphylaxis, whereas mice exposed to milk and egg did not. Ara h 2 was detected in serum collected post-challenge from peanut-sensitized mice, indicating increased absorption of this allergen, while Bos d 5 and Gal d 2 were not detected in mice exposed to milk and egg, respectively. Machine learning on the change in gut microbiome composition as a result of food protein exposure identified a unique signature in CC027/GeniUnc mice that experienced anaphylaxis, including the depletion of
. Overall, these results demonstrate several factors associated with enteral sensitization in CC027/GeniUnc mice, including diminished total fecal IgA, increased allergen absorption and altered gut microbiome composition. Furthermore, peanuts and tree nuts may have inherent properties distinct from milk and eggs that contribute to allergy.
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