The Suppressive Activities via FoxP3 Demethylation in Oral, epicutaneous, and Sublingual Peanut Immunotherapies

Katie Litman

Introduction. Food allergies in the US increased dramatically over the past decade, affecting 8% of Americans. Peanut allergies are a lifelong and life-threatening problem.1 They progress quickly (<2hrs) and have become the leading cause of anaphylaxis/death related to food allergy in the US.1 Methods. 6-8 week old BALB/c(H-2d) and BALB/c/Rag1-/-(H-2d) were used. Single cell suspensions pooled lymph nodes were stained using MAB to foxp3, CD4, CD8 and analyzed with flow cytometry. In-vitro assays of Treg function were preformed by isolating APCs from pooled LN using magnetic beads. CFSE-labeled Teff cells and an equal number of gamma irradiated APCs were added to wells with serial diluted WT or Mbd2-/- Treg cells starting at a 1:1 ratio. siRNA knockdown was done using lipofectamine 2000 and siRNA specific for Mbd2. Data was analyzed with ANOVA. Results. Eosinophils control CD103+ DC activation and migration from the intestine to lymph nodes which is integral for Th2 priming.2 For Ara h 1, DCs are primed by the binding of glycan moieties to C-type lectin receptors. This results in a dendritic cell phenotype that has an increased capacity for Th2 skewing3. FoxP3 Tcells upregulate expression of tissue-specific homing receptors for Th2 (CCR4&CCR8) and upregulation for Th-17 cells (CCR4, CCr6 CCR2 & CxCR3). In patients receiving OIT Th2 reactivity persisted despite desensitization/modulation of allergen-specific immune mediators (IgG4 antibodies & Th1 skewing). Potential mechanism of immunotherapy-induced tolerance is the increase in allergen specific blocking of IgG antibody levels.4 Desensitization following OIT protects against the risk for an IgE-mediated reactivity by suppressing Th2. This occurs through demethylation of Foxp3 CpG sites in aiTregs5. In OIT we see an increase in Tregs mediated by increasing FoxP3 hypomethylation. Mbd2 is a key mediator promoting demethylation of FoxP3 and upregulating Treg suppressive function against immune responses to ara h1.6 Demethylation of FoxP3 is reversible. Most regained sensitivity along with increased methylation of Foxp3 Cpg sites in the aiTreg cells.7 Data suggests epicutaneous(EPIT) and sublingual(SLIT) immunotherapies elicit different Treg responses. Foxp3+ was induced by all treatment routes but was greatest in EPIT8. Conclusions. Various forms of immunotherapy can drive production of ai-Tregs by decreasing methylation of FoxP3 through Mbd2.9,10 OIT can promote desensitization to peanut allergen. The next focus for immunotherapy is driving longevity. One important mechanism is the demethylation of FoxP3 and preventing the reversibility of demethylation of this modulator, producing long-term protection.

 

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  9. Wang M, Yang IV, Davidson EJ, et al. Forkhead box protein 3 demethylation is associated with tolerance induction in peanut-induced intestinal allergy. J Allergy Clin Immunol. 2018;141(2):659–670.e2. doi:10.1016/j.jaci.2018.04.020
  10. Wang L, Liu Y, Han R, et al. Mbd2 promotes foxp3 demethylation and T-regulatory-cell function. Mol Cell Biol. 2013;33(20):4106–4115. doi:10.1128/MCB.00144-13