Using Food and Nutritional Strategies to Induce Tolerance in Food-Allergic Children

Transcription

1 Allergy Fewtrell MS, Haschke F, Prescott SL (eds): Preventive Aspects of Early Nutrition. Nestlé Nutr Inst Workshop Ser, vol 85, pp 35 53, (DOI: / ) Nestec Ltd., Vevey/S. Karger AG., Basel, 2016 Using Food and Nutritional Strategies to Induce Tolerance in Food-Allergic Children Anna Nowak-Węgrzyn Jaffe Food Allergy Institute, Division of Pediatric Allergy, Mount Sinai School of Medicine, New York, NY, USA Abstract Food allergy is an important and increasing public health problem worldwide, affecting predominantly infants and young children. There is an urgent need to develop effective treatment strategies to restore oral tolerance in food-allergic individuals. Among diverse research approaches, those involving native or heat-modified food proteins are most advanced and are currently being evaluated in clinical trials. Extensively heated (baked) milk and egg diets have already been adopted in clinical practice and benefit the majority of milk- and egg-allergic children. Oral, sublingual and epicutaneous immunotherapy with native foods remain in the sphere of clinical research with encouraging data suggesting that they may induce desensitization in a large proportion of treated patients and potentially permanent tolerance following an adequately long period of treatment. Synbiotics appear to have the most beneficial role in the prevention of food allergy; Lactobacillus rhamnosus GG may promote the development of tolerance to milk in allergic infants Nestec Ltd., Vevey/S. Karger AG, Basel Introduction Food allergy is defined as an aberrant immunologic response towards an ingested food [1]. Food allergy can be mediated via IgE and non-ige mechanisms. The prevalence of food allergy has increased worldwide, initially in the highly developed societies with so-called western lifestyle (US, UK, Canada, Australia and Western Europe), followed by Asia and Latin America [2]. With an estimated

2 prevalence of 8% in US children, food allergy has become a serious public health problem. Currently, there is no cure for food allergy; dietary avoidance and management of accidental reactions remain the cornerstone of therapy. Considering the risk of potentially fatal anaphylaxis, nutritional deficiencies, impaired growth, impaired quality of life and increased financial burden, there is an urgent need to develop effective treatment strategies for food allergy [1]. Oral Tolerance Food allergy is thought to result from the failure to develop tolerance or a breach in oral tolerance. Oral tolerance is defined as a state of selective systemic unresponsiveness to the ingested food antigens [3]. It is mediated by the gut-associated lymphoid tissue and is thought to depend on the generation of antigenspecific T-regulatory cells (Tregs). In some models, T-cell anergy and deletion are also associated with oral tolerance. It is hypothesized that food antigens are captured in the lamina propria and Peyer s patches by CD103+ dendritic cells (DC) and are carried to the mesenteric lymph nodes. CD102+ DC induced Tregs (itregs) by a mechanism dependent on TGF-β, retinoic acid and indoleamine-2,3-dioxygenase. DCs also induce gut-homing IgA-secreting plasma cells through a retinoic acid-dependent mechanism. Gut-homing itregs are expanded in the lamina propria by IL-10-expressing CX3CR1+ macrophages. These itregs are capable of suppressing systemic immune responses, including allergic sensitization, in an antigen-specific manner. Dietary factors (vitamins A and D) and microbial factors ( Clostridium spp. and Bacteroides fragilis polysaccharide A) promote the generation of itregs. In contrast, bacterial adjuvants such as Staphylococcus aureus enterotoxin B and exposure to food allergens via an impaired skin barrier, e.g. in atopic dermatitis or filaggrin gene mutations, promote allergic sensitization. Natural Resolution of Food Allergy In the majority of children, allergy to cow s milk, egg, soybean and wheat resolves with age, a process that is frequently referred to as outgrowing of food allergy. The exact mechanism of the natural tolerance development in foodallergic children is not known. In general, food-specific IgE antibody levels and skin prick test (SPT) reactivity decrease, whereas casein and β-lactoglobulin- IgG 4 antibodies increase. There is an increase in peripheral blood and T cellsecreted IL-10 [4, 5]. 36Nowak-Węgrzyn

3 Allergen specific Allergen nonspecific Native food proteins Efficacy and safety data available Milk, egg and peanut OIT Milk, peanut and hazelnut SLIT Milk and peanut EPIT Modified food proteins Efficacy and safety data available Extensively heated milk and egg diet Efficacy and safety data available Anti-IgE (in peanut allergy) Anti-IL-5 Probiotics Safety data available Chinese herbs (FAHF-2) Safety data available Milk OIT with anti-ige Peanut OIT with anti-ige Multiple-food OIT Safety data available Escherichia coli expressing recombinant modified Ara h 1, 2, 3 rectal vaccine Pilot data available Trichuris suis ova therapy Fig. 1. Diverse approaches to food allergy therapy tested in clinical trials. Reprinted with permission from Albin and Nowak-Węgrzyn [30]. Potential Novel Therapies for Food Allergy Therapies for food allergy aim at restoring oral tolerance. At this time, there is no conclusive evidence that permanent oral tolerance can be induced. The most advanced, diverse strategies for food immunotherapy have been extensively reviewed elsewhere ( fig. 1 ); here, we focus on the strategies for the treatment of IgE-mediated food allergy involving food proteins administered orally, sublingually or via skin patch, hypoallergenic formulas and pre-/probiotics [6]. Baked Milk and Egg Diet It has been shown that high temperature changes protein conformation in cow s milk and hen s egg white; casein and ovomucoid are more heat-stable than whey proteins and ovalbumin [7]. A majority of children with milk and egg allergy tolerate extensively heated (baked) products containing milk and egg [8, 9]. Reactivity to baked milk is a marker of a more severe (higher risk of anaphylaxis) and more persistent milk allergy, whereas reactivity to baked egg is not associated with a more severe phenotype of egg allergy. Introduction of baked milk/egg into the diet is well tolerated and appears to accelerate acquisition of tolerance to unheated milk and egg, compared to children who are strictly avoiding milk/egg, according to the current standard of care [10, 11]. Baked milk diet is associated with decreasing levels of milk- and casein-specific IgE, Strategies to Induce Tolerance 37

4 basophil reactivity and milk SPT wheal diameter, and increasing levels of casein-specific IgG 4. Tolerance to baked milk is associated with increased numbers of circulating FoxP3+ (forkhead box protein 3+) Tregs; these changes are similar to the immune changes occurring during oral immunotherapy (OIT) [12]. Oral Immunotherapy OIT utilizes the pathways underlying oral tolerance [6]. The aim of food allergy therapy is first to achieve desensitization and then to reestablish permanent oral tolerance. Desensitization is a state of temporary antigen hyporesponsiveness that depends on the regular ingestion of the food. When dosing is interrupted or discontinued, the protective effect of desensitization is lost, and augmentation factors (e.g. viral infection, exercise, use of nonsteroidal anti-inflammatory drugs or menstruation) can trigger reactions to the previously tolerated maintenance dose. The immunologic mechanism of desensitization is not known, but it is associated with decreased reactivity of mast cells (measured by SPT reactivity) and basophils, and increased food-specific serum and salivary IgG 4 and IgA antibodies, and initially increased but eventually decreased serum food-specific IgE antibodies. Similarly, the mechanism of permanent tolerance is not known, but it presumably involves the generation of Tregs followed by anergy and/or deletion of effector T cells. Specific Tregs increase and peak at around 12 months, with a subsequent decrease over time. Increased antigen-induced regulatory T- cell function is associated with hypomethylation of FoxP3. To date, no development of permanent tolerance (or even long-term desensitization) due to OIT as opposed to natural acquisition has been conclusively demonstrated. Individualized dosing and longer duration of OIT is associated with increased rates of sustained unresponsiveness [13, 14]. While permanent oral tolerance is the ultimate goal of OIT, desensitization is perceived as beneficial by many patients and parents. Results from two uncontrolled studies suggest that OIT may improve some aspects of life quality, such as dietary and social limitations, risk of accidental exposure and anxiety. The true impact of OIT on quality of life remains to be determined. Dosing Schedule During OIT, food is mixed with a vehicle and eaten in gradually increasing doses. Dose escalations occur in a controlled setting. Most studies include an initial rapid dose escalation day that is followed by further dose escalation in the clinic on a biweekly schedule until the maintenance dose is achieved. Daily ingestion of tolerated doses during the buildup and maintenance phases occurs at home. The need for daily dosing raises concerns regarding patient 38Nowak-Węgrzyn

5 Tolerance food challenge Buildup phase Maintenance phase Discontinuationelimination diet SPT wheal diameter, basophil activation Specific IgE level Regulatory T cells Specific IgG 4 level Allergy Tolerance Fig. 2. OIT time course and immunologic changes occurring during OIT. Reprinted with permission from Albin and Nowak-Węgrzyn [30]. adherence over long periods of time, as well as unintentional dosing interruptions due to illness or travel. In a small clinical trial of milk OIT, twice weekly maintenance dosing was as safe and efficacious as daily maintenance dosing [15]. Response to Food Oral Immunotherapy Figure 2 summarizes the different phases of OIT, as well as the general trends of immunologic changes seen in OIT. Desensitization failure may be associated with the most severe food allergy phenotype and high baseline food IgE levels, as opposed to desensitization success that may be associated with a milder, transient phenotype and higher chances of spontaneous resolution of food allergy. Clinical Trials of Oral Immunotherapy for Food Allergy Many studies of food OIT have been published to date, but only a few had a rigorous design [16 18]. There is a significant heterogeneity in the study design, inclusion criteria, dosing schedule, duration of therapy and approach to evaluating the persistence of the protection following discontinuation of OIT. Selected relevant clinical trials are presented in detail in table 1. Strategies to Induce Tolerance 39

6 Table 1. Selected clinical trials in food OIT Study/subjects Success rate Immunologic changes Side effects/comments Peanut OIT Blumchen et al. [31], 2010 (n = 23; age 3 14 years) Subjects underwent a rush protocol for 1 week and then received OIT for 8 weeks Goal maintenance dose: 0.5 g peanut protein daily 22 of 23 (96%) subjects finished the initial rush protocol, but 17 of these subjects did not achieve the goal dose of 0.5 g peanut with rush therapy (they continued on individualized long-term buildup protocols to reach 0.5 g) Only 14 of 22 subjects (64%) reached a maintenance dose of at least 0.5 g of peanut and completed OIT The median tolerated dose after completion of OIT was 1 g of peanut After OIT, all subjects had reductions in IL-5, IL-4 and IL-2 (p < 0.001); after discontinuing OIT for 2 weeks, peanut-induced IL-5, IL-4 and IL-2 secretion was stable in most of the subjects After OIT, there was a small reduction in peanut SPT wheal diameter (p < 0.05), but after discontinuing OIT for 2 weeks, this reduction was no longer evident After OIT, peanut-specific IgG4 levels increased (p < 0.001); after discontinuing OIT for 2 weeks, there was a small decrease in specific IgG4 levels (p < 0.001) Peanut-specific IgE levels as well as total IgE levels did not change with OIT The dropout rate of this study was 35% (8 of 23 subjects); in 4 subjects, OIT was discontinued by the study physician because of adverse allergic reactions; 2 subjects dropped out because of compliance: 1 because of a severe infection, and 1 was a partial responder who did not qualify for the final challenge Adverse reactions were frequent, particularly during the rush protocol; of 317 total doses of OIT during the rush protocol, 25 doses (7.9%) were associated with objective symptoms; of 6,137 doses of OIT during the long-term buildup protocol, 160 doses (2.6%) were associated with objective symptoms Varshney et al. [32], 2011 (n = 28; age 1 16 years) Subjects were randomized 19:9 to OIT or placebo for 48 weeks Goal maintenance dose: 4 g peanut protein daily During the initial day of rapid dose escalation, 26 of 28 (93%) subjects reached the maximum cumulative dose of 12 mg of peanut protein or placebo Sixteen subjects received the full course of peanut OIT and all tolerated 5 g of peanut protein following OIT Compared to placebo, the peanut OIT group showed reductions in SPT size (p < 0.001), IL-5 (p = 0.01) and IL-13 (p = 0.02), and increases in peanutspecific IgG4 (p < 0.001) The ratio of FoxP3 high /FoxP3 intermediate CD4+ CD25+ T cells increased at the time of OFC (p = 0.04) in peanut OIT subjects Adverse reactions were common, but most reactions were mild In the active arm, 9 of 19 subjects (47%) experienced adverse reactions during the initial day escalation During the buildup phase, adverse reactions occurred following 1.2% of 407 buildup doses None in the placebo group required treatment during initial day escalation or buildup dosing, but 3 (33%) were treated with epinephrine during the final peanut DBPCFC 40Nowak-Węgrzyn

7 Table 1. Continued Study/subjects Success rate Immunologic changes Side effects/comments Vickery et al. [13], 2014 (n = 24; age 1 16 years) Subjects were treated for up to 5 years with peanut OIT, and then treatment was stopped for 1 month to test for sustained unresponsiveness Goal maintenance dose: the protocol was modified over time to permit dose increases to a maximum of 4 g peanut protein daily Of the 39 subjects originally enrolled, 24 (62%) completed the protocol and had evaluable outcomes All subjects completing the study successfully ingested 5 g of peanut protein without symptoms during desensitization challenge Only 12 of 24 (50%) successfully ingested 5 g of peanut protein 1 month after stopping OIT and achieved sustained unresponsiveness Of those who failed the 5-gram challenge, the median amount of peanut protein ingested cumulatively before the development of symptoms was 3.75 g (range, g) In those who achieved sustained unresponsiveness, subjects had smaller skin test results, lower IgE levels specific for peanut, Ara h 1 and 2, and lower ratios of peanut-specific IgE/total IgE compared to those who did not achieve sustained unresponsiveness There were no differences in peanut IgG4 levels or FoxP3 CD4+ CD25+ T cells between these groups This study is the first to demonstrate sustained unresponsiveness after peanut OIT Anagnostou et al. [33], 2014 (n = 99; age 7 16 years) Phase 1: subjects were randomized 1:1 to OIT or control (standard of care food avoidance) for 26 weeks Phase 2: crossover from the control group to OIT for 26 weeks Goal maintenance dose: 0.8 g peanut protein daily 84% of subjects receiving OIT were able to ingest 0.8 g of peanut daily for 26 weeks in the first phase, and 91% of subjects from the crossover control group were able to ingest 0.8 g of peanut daily for 26 weeks during the second phase 24 of 39 subjects (62%) receiving OIT tolerated 1.4 g of peanut at the end of the first phase of the study, as compared to 0 of 46 control subjects (p < 0.001) After OIT, there was a small reduction in peanut SPT wheal diameter After 24 weeks of OIT, there was an increase in peanut-specific IgE No significant within-patient differences were identified after treatment for basophil activation (although there was a reduction in mean fluorescence intensity and proportion of CD63 at lower peanut concentrations after OIT) During the first phase of the study, 1 subject discontinued and 5 withdrew from the OIT arm; 4 subjects could not achieve target maintenance dose at 6 months in the OIT arm; in the control group, 4 subjects withdrew and 1 discontinued The number and nature of adverse events was similar in both groups after treatment, and most events were mild; the most common adverse event was oral itching, which occurred after 6.3% of all doses Strategies to Induce Tolerance 41

8 Table 1. Continued Study/subjects Success rate Immunologic changes Side effects/comments Egg OIT Burks et al. [34], 2012 (n = 55; age 5 11 years) Subjects were part of a multicenter double-blind placebo-controlled clinical trial Goal maintenance dose: 2 g egg white powder daily At the 10-month challenge, 22 of 40 subjects (55%) receiving OIT tolerated 5 g of egg white powder, and 0 of 15 subjects (0%) of the placebo group passed (p < 0.001) At the 22-month challenge, 30 of 40 subjects (75%) receiving OIT tolerated 10 g of egg white powder (were considered desensitized) At the 24-month challenge (6 8 weeks off OIT), 11 of 29 subjects (28%) tolerated 10 g of egg white powder plus whole egg (were considered to have sustained unresponsiveness); according to the intention-to-treat analysis, 11 of the 40 subjects (28%) in the OIT group passed the OFC at 24 months (p = 0.03 vs. placebo) Compared with subjects who received placebo, those who received OIT had a decreased wheal size on SPT, reduced egg-induced basophil activation and increased egg-specific IgG4 antibody levels over time, whereas no change in egg-specific IgE antibody levels was noted All 55 subjects completed the initial-day dose escalation, but 7 subjects withdrew before the maintenance phase (5 in the OIT group and 2 in the placebo group) There were no severe adverse events, and rates were highest during the first 10 months of OIT Adverse events were associated with 25% of 11,860 doses of OIT and 3.9% of 4,018 doses of placebo The results of this clinical trial raise concerns about the long-term protection (sustained unresponsiveness) afforded by OIT; it remains to be determined whether a higher maintenance dose and longer duration of OIT might improve the rates of sustained unresponsiveness Milk OIT Staden et al. [35], 2007 (n = 45; age <1 12 years) Subjects were randomized to OIT or elimination diet; oral tolerance for all subjects was evaluated after a median 21 months (range months) Goal maintenance dose: doses were increased (according to individual tolerance) to a maximum dose of 8.25 g cow s milk protein or 2.8 g hen s egg protein; the maintenance phase mandated a minimum daily maintenance dose of 3.3 g cow s milk protein and 1.6 g hen s egg protein 25 subjects were randomized to milk or egg OIT, and 14 underwent milk OIT 20 subjects were randomized to an elimination diet, 10 of which were avoiding milk In the OIT group, 16 of 25 subjects (64%) were able to integrate the food allergen into their diet; 9 of 25 subjects (36%) showed sustained tolerance after a secondary elimination diet, 3 of 25 (12%) showed tolerance with regular intake, and 4 of 25 (16%) were partial responders who never met the planned full maintenance dose In contrast, only 7 of 20 subjects (35%) showed tolerance (p = 0.05) Allergen-specific IgE levels decreased significantly both in subjects who developed natural tolerance during the elimination diet (p < 0.05) and in those treated with OIT (p < 0.001) All subjects in the OIT group had adverse events to some extent: 21 had mild symptoms and 4 had moderate symptoms In the control group, 5 subjects had mild-to-moderate adverse events due to inadvertent ingestion of food allergens 42Nowak-Węgrzyn

9 Table 1. Continued Study/subjects Success rate Immunologic changes Side effects/comments Skripak et al. [36], 2008 (n = 20; age 6 17 years) Subjects were randomized 2:1 to OIT or placebo for 13 weeks Goal maintenance dose: 0.5 g cow s milk protein daily 19 subjects completed treatment (12 receiving OIT and 7 receiving placebo) After OIT, the median cumulative dose of milk inducing a reaction in the active group increased from 40 to 5,140 mg, but there was no change in the placebo group (p = ) Milk-specific IgE levels did not change in either group Milk IgG levels increased significantly in the OIT group, with a predominant milk-igg4 increase End-point titration milk SPT wheal decreased over time (p = vs. baseline) Among 2,437 active OIT doses, there were 1,107 reactions (45.4%) Among 1,193 placebo doses, there were 134 reactions (11.2%) Local symptoms were most common Narisety et al. [37], 2009 (n = 15; age 6 16 years) Open-label follow-up to Skripak study, which occurred over weeks (median 17 weeks) 14 subjects were able to escalate daily doses, with maximum doses ranging from 1 to 16 g (median, 7 g) After weeks of open-label dosing, challenges were conducted on 13 subjects; 6 tolerated 16 g with no reaction, and 7 reacted at 3 16 g Milk-specific IgE levels decreased IgG4 levels increased over the followup period from 3 to 17 months Challenges were not performed in 2 subjects because of ongoing reactions with home dose escalations Adverse reactions were common and unpredictable, but overall rates of reaction decreased over time. In the 2,465 home doses recorded, there were 419 local reactions (17% of doses) Epinephrine was required for 6 (0.2%) doses in 4 subjects Of note, 1 subject developed symptoms consistent with eosinophilic esophagitis Pajno et al. [15], 2013 (n = 32; age 4 13 years) Subjects who were successfully desensitized with OIT were randomized to two maintenance regimens for 1 year: group A had to ingest ml milk daily, and group B ml milk twice weekly 29 subjects completed the 12-month study: 15 randomized to daily milk and 14 randomized to twice weekly milk None of the subjects permanently discontinued their maintenance diet based on adverse events There were no significant differences in milk-specific IgE, IgG4 and SPT reactivity between groups A and B Three subjects and their families withdrew during maintenance because of personal problems not related to the study procedures Adverse events included asthma, oral itching, urticaria, rhinitis and abdominal pain, usually associated with concomitant illness or exercise There were 8 events in group A and 9 events in group B (no significant difference between the groups: p = 0.08) Strategies to Induce Tolerance 43

10 Table 1. Continued Study/subjects Success rate Immunologic changes Side effects/comments Multiple-food OIT Begin et al. [38], 2014 (n = 40; age 4 46 years) Pilot phase I study for multiple-food OIT (15 subjects on peanut OIT, 25 subjects on multiple-food OIT) Goal maintenance dose: 4 g of food protein per allergen daily Rates of reaction per dose did not differ significantly between the two groups (median of 3.3 and 3.7% in multi- and single-food OIT group, respectively; p = 0.31) In both groups, most reactions were mild but two 2 reactions requiring epinephrine occurred in each group Those on multiple-food OIT took longer to reach equivalent doses per food (median +4 months; p < ) In the multiple-food group, there was an increase in peanut-specific IgG4 similar to the monotherapy group Peanut-specific IgE levels were stable after 1 year This was a proof-of-concept phase I safety study, and the primary endpoint of the study was occurrence of allergic reactions Over the study period, there were 5 dropouts for reasons which included noncompliance with study medication and change of residence One subject in the multiple-food OIT group was unable to increase doses due to eczema flares and was categorized a treatment failure OFC = Oral food challenge. 44Nowak-Węgrzyn

11 Oral Immunotherapy Combined with Anti-IgE Monoclonal Antibody Anti-IgE monoclonal antibody is a non-allergen-specific treatment that has been successfully tested as monotherapy for peanut allergy. In one study, subjects treated with the highest dose of anti-ige showed an increased dose threshold response for peanut after 4 months [19]. In a logical next step, pretreatment with anti-ige monoclonal antibody has also been shown to be effective and improve safety of food OIT, likely by lowering circulating IgE and downregulating expression of the IgE receptor on antigen-presenting cells ( table 2 ). The preliminary data on pretreatment with omalizumab prior to OIT are encouraging, and large clinical trials are testing the efficacy of such approach. Oral Immunotherapy with Multiple Foods Considering that approximately 30% of children undergoing OIT are allergic to multiple foods and that for aeroallergens, specific immunotherapy frequently combines multiple allergens, a safety trial of a multiple-food OIT was conducted in parallel to peanut OIT in subjects with peanut allergy ( table 2 ). Pretreatment with omalizumab may also be an option for multiple-food OIT. These preliminary results suggest that multiple-food OIT may be a practical and safe option for patients with multiple food allergy, but these findings need validation by large and rigorous clinical trials. Safety of Food Immunotherapy OIT is associated with acute adverse allergic reactions in virtually all patients, which are more common during dose escalation than during maintenance. Most of the reactions are mild and limited to the oropharynx. However, systemic reactions are seen and may occur at previously tolerated doses in the presence of augmentation factors. Outside of acute allergic reactions, many patients complain of gastrointestinal symptoms that may lead to study withdrawal. Gastrointestinal side effects affect as many as 50% of patients, and those undergoing OIT may be at higher risk of developing eosinophilic esophagitis than the general allergic population [20]. Sublingual Immunotherapy In sublingual immunotherapy (SLIT), a food allergen extract is kept in the mouth for 2 3 min and then spit out or swallowed. It is generally better tolerated and utilizes significantly lower doses as compared to OIT, but appears to have inferior clinical effects of desensitization [6]. Clinical trials of food SLIT have been reported for milk, peanut, hazelnut and peach ( table 3 ). Some of these trials evaluated efficacy in patients with symptoms of pollen food syndrome rather than systemic food allergy. SLIT is generally better Strategies to Induce Tolerance 45

12 Table 2. Clinical trials with combination food OIT Study/subjects Success rate Immunologic changes Side effects/comments OIT and SLIT safety and efficacy study Keet et al. [22], 2012 (n = 30; age 6 17 years) Randomized clinical trial comparing milk OIT and SLIT with challenge performed after 12 and 60 weeks of maintenance Goal maintenance dose (milk protein/day) SLIT: 7 mg daily Low-dose OIT: 1 g High-dose OIT: 2 g 1 of 10 subjects in the SLIT group, 6 of 10 subjects in the SLIT/low-dose OIT group, and 8 of 10 subjects in the SLIT/high-dose OIT group passed the 8-gram milk protein challenge (p = 0.002, SLIT vs. OIT) After avoidance, 6 of 15 subjects (3 subjects in each OIT group) regained reactivity, 2 after only 1 week off therapy Titrated milk SPT wheal diameter and basophil activity decreased in all groups Milk-specific IgG4 increased in all groups Milk-specific IgE and spontaneous histamine release decreased only in the OIT group OIT was more efficacious for desensitization than SLIT alone but was accompanied by more systemic side effects.there were symptoms with 1,802 (29%) of 6,246 SLIT doses and 2,402 (23%) of 10,645 OIT doses However, OIT had significantly more multisystem, upper/lower respiratory tract and gastrointestinal symptoms than SLIT OIT and anti-ige-safety studies Nadeau et al. [39], 2011 (n = 11; age 7 17 years) Pilot phase I study using omalizumab in combination with oral milk desensitization (desensitization performed 9 weeks after the start of omalizumab treatment) Goal maintenance dose: 2 g milk protein/day The mean frequency for total reactions reported by week 24 was 1.6% (32 reactions of 2,199 doses in total for all 11 subjects) All subjects experienced some adverse events, though most reactions were defined as mild and needed no treatment 1 dose of epinephrine was given during rush desensitization, and 2 subjects received epinephrine at home during the maintenance phase 9 of the 11 subjects tolerated desensitization to a dose of 2 g milk within a period of 7 11 weeks Within 1 week of treatment, the CD4+ T-cell response to milk was nearly eliminated Over the following 3 months, the CD4+ T-cell response returned, characterized by a shift from IL-4 to IFN-γ Milk IgE decreased Milk IgG4 increased 15-fold This was a phase I study and the primary objectives were to examine the safety of this approach and to determine whether subjects could be dosed up to 2 g milk within 7 11 weeks of initiating desensitization 46Nowak-Węgrzyn

13 Table 2. Continued Study/subjects Success rate Immunologic changes Side effects/comments Schneider et al. [40], 2013 (n = 13; age 8 16 years) Pilot study using omalizumab in combination with oral peanut desensitization (desensitization performed 12 weeks after the start of omalizumab treatment) Goal maintenance dose: 4 g peanut protein/ day All 13 subjects (100%) reached the 0.5-gram peanut desensitization dose on the 1st day (cumulative dose, 992 mg), which was the primary outcome of the study, with minimal or no symptoms 12 of the 13 subjects (92%) reached the 4-gram maintenance dose, which was a secondary outcome of the study, requiring a median time of 8 weeks 12 weeks after stopping omalizumab (week 32), 12 subjects underwent a DBPCFC (cumulative dose, 8 g peanut 11 of the subjects (85%) tolerated this challenge, and the 12th subject later passed an open challenge of 8 g peanut Not reported There were a total of 72 reactions during the study (2.0% of the 3,502 total peanut doses ingested) During the study, 6 of the 13 subjects experienced mild or no allergic reactions, 5 subjects had grade 2 reactions, and 2 subjects had grade 3 reactions, all of which responded rapidly to treatment Begin et al. [41], 2014 (n = 25; age 4 15) Phase 1 study using omalizumab with rush multiple-food OIT (omalizumab given 8 weeks prior to and 8 weeks following initiation of rush multiple-food OIT) Goal maintenance dose: 4 g of food protein per allergen daily After pretreatment with omalizumab, 19 subjects tolerated all 6 steps of the initial escalation day (up to 1,250 mg of combined food proteins) The remaining 6 were started on their highest tolerated dose as their initial daily home doses Subjects reached their maintenance dose of 4 g of protein per allergen at a median of 18 weeks, and all subjects reached maintenance dose by 9 months After 52 weeks of therapy, peanut-specific IgE did not change significantly, but peanut-specific IgG4 antibody levels showed median increases of 8.23 mga/l (p < ). Peanut SPT decreased by a median of 8 mm (p < ) During initial escalation of rush multiple-food OIT, 13 participants (52%) experienced some symptoms However, subjects required minimal or no rescue therapy Subjects reported 401 reactions per 7,530 home doses (5.3%) with a median of 3.2 reactions per 100 doses 94% of reactions were mild, although there was one severe reaction requiring epinephrine Strategies to Induce Tolerance 47

14 Table 3. Food SLIT SLIT Goal daily maintenance dose; range from published studies Dosing interval Allergen uptake Safety/side effects Desensitization success rates Long-term unresponsiveness Humoral immunologic changes Basophil activation with food allergen T-cell responses Milk: 1 ml or 7 mg Peanut: 165 2,000 μg, 3,696 μg in one crossover group Hazelnut: 22 mg Peach: 10 μg of Pru p 3 Milk, peanut, hazelnut: daily Peach: 3 times per week Oral mucosal uptake by Langerhans cells Local, mild oropharyngeal Milk: 10 70% Keet et al. [22], % receiving SLIT were desensitized 60% receiving SLIT/low-dose OIT were desensitized 80% receiving SLIT/high-dose OIT were desensitized Peanut: 44 70% Kim et al. [10], 2011 After 12 months, subjects receiving SLIT safely ingested a median cumulative dose of 1,710 mg of peanut protein (equivalent to 6 7 peanuts), which was significant compared to the placebo group who safely ingested a median cumulative dose of 85 mg (<1 peanut) Hazelnut: Enrique et al. [42], % ingested the highest planned dose (20 g) after 8 12 weeks of SLIT 71% ingested the highest planned dose after 4 12 months of SLIT Peach: Fernández-Rivas et al. [43], 2009 Actively treated patients tolerated at least 3 times (3 9 times) more peach in the DBPCFC after 6 months of SLIT Undetermined Milk, peanut, hazelnut, peach: Variable changes in serum IgE Increased serum IgG 4 Decreased SPT wheal diameters Milk: No changes Peanut: Decreased Peanut: Decreased IL-5 production in PBMCs Decreased basophil activation No difference in Tregs Hazelnut: Increased IL-10 levels PBMCs = Peripheral blood mononuclear cells. 48Nowak-Węgrzyn

15 tolerated with lower rates of systemic reactions as compared to OIT, which is partially due to 100- to 1,000-fold-lower starting doses in SLIT compared to OIT. In addition, the oral mucosa contains a limited number of proinflammatory cells, such as mast cells. However, the degree of desensitization appears to be inferior compared to OIT. A retrospective comparison of SLIT versus OIT for peanut showed that peanut OIT resulted in greater changes in peanut-specific IgE, IgG 4 and basophil activation as compared to peanut SLIT, and eliciting dose thresholds were lower during double-blind placebocontrolled food challenge (DBPCFC) at 12 months in patients who underwent SLIT [21]. One study compared these two routes of immunotherapy together in a rigorous, single-center clinical trial. Thirty milk-allergic children (aged 6 17 years) were randomized to SLIT or SLIT followed by low- or high-dose milk OIT [22]. Following therapy, 1 of 10 subjects in the SLIT group, 6 of 10 subjects in the SLIT/low-dose OIT group, and 8 of 10 subjects in the SLIT/high-dose OIT group passed the 8-gram desensitization challenge (p = 0.002, SLIT vs. OIT). Systemic reactions were more common during OIT escalation and maintenance (0.43% with high-dose and 0.08% with low-dose OIT) than during SLIT maintenance (0.02%). Milk OIT was more efficacious for desensitization than SLIT alone, but it was associated with more systemic side effects. Epicutaneous Immunotherapy Skin has a high capacity for pleiotropic immune responses. The epidermis contains large numbers of antigen-presenting Langerhans cells and is not vascularized, reducing the risk of systemic side effects. The epicutaneous route for allergen immunotherapy has been introduced at the beginning of the 20th century, with allergen extracts being applied through scarified skin, with reported treatment success rates of 80% for co-seasonal treatment with aeroallergens. The interest in epicutaneous immunotherapy (EPIT) reemerged in the past decade, spurred by evidence that transcutaneous vaccination can be effective against infectious diseases ( Helicobacter pylori ) and cancer. The initial randomized double-blind placebo-controlled study with grass pollen EPIT utilized patches with allergen that were applied weekly to tape-stripped skin [23]. Tape stripping enhances allergen penetration by removing stratum corneum and by activating keratinocytes to secrete IL-1, IL-6, IL-8, TNF-α and IFN-γ, which stimulate maturation and migration of DCs to the draining lymph nodes. Actively treated subjects had 70% improvement in allergic rhinitis symptoms compared with 20% in placebo-treated subjects. Local eczematous reactions under the tape were frequent, but no systemic reactions were observed. These results were confirmed in larger trials in grass-allergic adults and children [24, 25]. Strategies to Induce Tolerance 49

16 Instead of tape stripping, penetration of the allergens through the epidermis can be enhanced by skin hydration, by application of a patch that leads to accumulation of sweat. This approach has been taken in EPIT studies in food allergy. In a clinical pilot study, an epicutaneous delivery system of milk (Viaskin patches) was applied to children with milk allergy [26]. In a small pilot study, 18 children (mean age 3.8 years, range 10 months to 7.7 years) with cow s milk allergy were randomized 1: 1 to receive active EPIT or placebo. Children received three 48-hour applications (1 mg skimmed milk powder or 1 mg glucose as placebo) via the skin patch per week for 3 months. EPIT-treated children had a trend toward increased threshold doses at the follow-up oral milk challenge, from a mean of 1.8 ml at baseline to 23.6 ml at 3 months; there was no change in the placebo group. There were no severe systemic reactions; however, 1 child had repeated episodes of diarrhea following EPIT. Large clinical trials are currently underway for peanut and milk allergy. EPIT is applied by the patient at home, rotating the application site, and is generally better tolerated and more convenient than OIT. Synbiotics and Hypoallergenic Formulas Synbiotics include pro- and prebiotics. A probiotic is defined as a live microorganism with potential health benefit, and a prebiotic is a nondigestible food ingredient that can stimulate the growth and metabolic activity of beneficial bacteria in the gut. Children with allergy have been shown to have different gut microflora as compared to healthy controls (e.g. higher levels of clostridia and lower levels of bifidobacteria), and some hypothesize that a beneficial gut microflora supports immunoregulation by promoting the development of Tregs. Preand probiotics have been studied extensively in the prevention of food allergy, with inconclusive results mostly due to a significant heterogeneity in the study design, including specific strains of bacteria used, and timing and duration of treatment [27]. In a mouse model of food allergy, treatment with a probiotic mixture was effective in the protection against anaphylaxis and caused a redirection of allergen-specific Th2-polarized immune responses towards Th1-Treg responses. Clostridia have been associated with protective immune responses. A first randomized, controlled trial of probiotic supplementation ( Lactobacillus casei CRL431 and Bifidobacterium lactis Bb-12) in the setting of cow s milk allergy did not demonstrate accelerated tolerance in infants [28]. A recent nonrandomized clinical trial demonstrated that supplementation of an extensively hydrolyzed casein formula (ehcf) with a different probiotic ( Lactobacillus GG) accelerated the development of tolerance to milk in infants with milk allergy as confirmed by DBPCFC. Berni Canani et al. [29] prospectively evaluated tolerance to milk 50Nowak-Węgrzyn

17 after 12 months of treatment with one of the following formulas: ehcf, ehcf with Lactobacillus rhamnosus GG (LGG), hydrolyzed rice formula (HRF), soy or amino acid. After 12 months of treatment, the rate of cow s milk tolerance was significantly higher (p < 0.05) in the groups receiving ehcf (43.6%) or ehcf + LGG (78.9%) compared with the other groups, i.e. HRF (32.6%), soy formula (23.6%) and amino acid-based formula (18.2%). Subjects with persistent cow s milk allergy were challenged again after 12 months of treatment, and 60% receiving ehcf alone compared to 45% receiving ehcf + LGG had a positive DBPCFC. It should be noted that there is a natural tendency to outgrow milk allergy over time, particularly in non-ige-mediated milk allergy, which comprised the majority of subjects in each treatment arm. This study also suggested that if tolerated, formulas with more intact food protein (ehcf or HRF) may have a more beneficial effect on the development of tolerance. However, considering the nonrandomized design, there might have been bias, such as the infants with the most severe milk allergy phenotype were treated with the amino acid-based formula because they did not tolerate more allergenic formulas. Conclusions Food allergy is an important and increasing public health problem worldwide, affecting predominantly infants and young children. There is an urgent need to develop effective treatment strategies to restore oral tolerance in food-allergic individuals. Among diverse research approaches, those involving native or heatmodified food proteins are most advanced and are currently being evaluated in clinical trials. Extensively heated (baked) milk and egg diets have already been adopted in clinical practice and have been of benefit in the majority of milk- and egg-allergic children. OIT, SLIT and EPIT with native foods remain in the sphere of clinical research with encouraging data suggesting that they may induce desensitization in the large proportion of treated subjects and potentially permanent tolerance following an adequately long period of treatment. Synbiotics appear to have the most beneficial role in the prevention of food allergy; LGG may promote the development of milk tolerance in allergic infants. Disclosure Statement The author received grant support from NIH NIAID, FARE, Nestle, Nutricia, DBV and royalties from UpToDate. She is a member of the Merck and Nutricia advisory boards and speaker for Annenberg and Nestle. Strategies to Induce Tolerance 51

18 References 1 Sicherer SH, Sampson HA: Food allergy: epidemiology, pathogenesis, diagnosis, and treatment. J Allergy Clin Immunol 2014; 133: ; quiz Prescott SL, Pawankar R, Allen KJ, et al: A global survey of changing patterns of food allergy burden in children. World Allergy Organ J 2013; 6: Berin MC, Sampson HA: Mucosal immunology of food allergy. Curr Biol 2013; 23:R389 R Sommanus S, Kerddonfak S, Kamchaisatian W, et al: Cow s milk protein allergy: immunological response in children with cow s milk protein tolerance. Asian Pac J Allergy Immunol 2014; 32: Fishbein AB, Qamar N, Erickson KA, et al: Cytokine responses to egg protein in previously allergic children who developed tolerance naturally. Ann Allergy Asthma Immunol 2014; 113: 667.e4 670.e4. 6 Nowak-Wegrzyn A, Albin S: Oral immunotherapy for food allergy: mechanisms and role in management. Clin Exp Allergy 2015; 45: Bloom KA, Huang FR, Bencharitiwong R, et al: Effect of heat treatment on milk and egg proteins allergenicity. Pediatr Allergy Immunol 2014; 25: Nowak-Wegrzyn A, Bloom KA, Sicherer SH, et al: Tolerance to extensively heated milk in children with cow s milk allergy. J Allergy Clin Immunol 2008; 122: , 347.e1 347.e2. 9 Lemon-Mule H, Sampson HA, Sicherer SH, et al: Immunologic changes in children with egg allergy ingesting extensively heated egg. J Allergy Clin Immunol 2008; 122: 977.e e1. 10 Kim JS, Nowak-Wegrzyn A, Sicherer SH, et al: Dietary baked milk accelerates the resolution of cow s milk allergy in children. J Allergy Clin Immunol 2011; 128: 125.e2 131.e2. 11 Leonard SA, Sampson HA, Sicherer SH, et al: Dietary baked egg accelerates resolution of egg allergy in children. J Allergy Clin Immunol 2012; 130: 473.e1 480.e1. 12 Leonard SA, Caubet JC, Kim JS, et al: Baked milk- and egg-containing diet in the management of milk and egg allergy. J Allergy Clin Immunol Pract 2015; 3: 13 23; quiz Vickery BP, Scurlock AM, Kulis M, et al: Sustained unresponsiveness to peanut in subjects who have completed peanut oral immunotherapy. J Allergy Clin Immunol 2014; 133: Syed A, Garcia MA, Lyu SC, et al: Peanut oral immunotherapy results in increased antigeninduced regulatory T-cell function and hypomethylation of forkhead box protein 3 (FOXP3). J Allergy Clin Immunol 2014; 133: Pajno GB, Caminiti L, Salzano G, et al: Comparison between two maintenance feeding regimens after successful cow s milk oral desensitization. Pediatr Allergy Immunol 2013; 24: Fisher HR, du Toit G, Lack G: Specific oral tolerance induction in food allergic children: is oral desensitisation more effective than allergen avoidance? A meta-analysis of published RCTs. Arch Dis Child 2011; 96: Nurmatov U, Devereux G, Worth A, et al: Effectiveness and safety of orally administered immunotherapy for food allergies: a systematic review and meta-analysis. Br J Nutr 2014; 111: Nurmatov U, Venderbosch I, Devereux G, et al Allergen-specific oral immunotherapy for peanut allergy. Cochrane Database Syst Rev 2012; 9:CD Leung DY, Sampson HA, Yunginger JW, et al: Effect of anti-ige therapy in patients with peanut allergy. N Engl J Med 2003; 348: Wood RA, Sampson HA: Oral immunotherapy for the treatment of peanut allergy: is it ready for prime time? J Allergy Clin Immunol Pract 2014; 2: Chin SJ, Vickery BP, Kulis MD, et al: Sublingual versus oral immunotherapy for peanutallergic children: a retrospective comparison. J Allergy Clin Immunol 2013; 132: 476.e e2. 22 Keet CA, Frischmeyer-Guerrerio PA, Thyagarajan A, et al: The safety and efficacy of sublingual and oral immunotherapy for milk allergy. J Allergy Clin Immunol 2012; 129: , 455.e1 455.e5. 23 Senti G, Graf N, Haug S, et al: Epicutaneous allergen administration as a novel method of allergen-specific immunotherapy. J Allergy Clin Immunol 2009; 124: Nowak-Węgrzyn

19 24 Senti G, von Moos S, Tay F, et al: Epicutaneous allergen-specific immunotherapy ameliorates grass pollen-induced rhinoconjunctivitis: a double-blind, placebo-controlled dose escalation study. J Allergy Clin Immunol 2012; 129: Agostinis F, Forti S, Di Berardino F: Grass transcutaneous immunotherapy in children with seasonal rhinoconjunctivitis. Allergy 2010; 65: Dupont C, Kalach N, Soulaines P, et al: Cow s milk epicutaneous immunotherapy in children: a pilot trial of safety, acceptability, and impact on allergic reactivity. J Allergy Clin Immunol 2010; 125: Jensen MP, Meldrum S, Taylor AL, et al: Early probiotic supplementation for allergy prevention: long-term outcomes. J Allergy Clin Immunol 2012; 130: 1209.e e5. 28 Hol J, van Leer EH, Elink Schuurman BE, et al: The acquisition of tolerance toward cow s milk through probiotic supplementation: a randomized, controlled trial. J Allergy Clin Immunol 2008; 121: Berni Canani R, Nocerino R, Terrin G, et al: Formula selection for management of children with cow s milk allergy influences the rate of acquisition of tolerance: a prospective multicenter study. J Pediatr 2013; 163: 771. e1 777.e1. 30 Albin S, Nowak-Węgrzyn A: Potential treatments for food allergy. Immunol Allergy Clin North Am 2015; 35: Blumchen K, Ulbricht H, Staden U, et al: Oral peanut immunotherapy in children with peanut anaphylaxis. J Allergy Clin Immunol 2010; 126: 83.e1 91.e1. 32 Varshney P, Jones SM, Scurlock AM, et al: A randomized controlled study of peanut oral immunotherapy: clinical desensitization and modulation of the allergic response. J Allergy Clin Immunol 2011; 127: Anagnostou K, Islam S, King Y, et al: Assessing the efficacy of oral immunotherapy for the desensitisation of peanut allergy in children (STOP II): a phase 2 randomised controlled trial. Lancet 2014; 383: Burks AW, Jones SM, Wood RA, et al: Oral immunotherapy for treatment of egg allergy in children. N Engl J Med 2012; 367: Staden U, Rolinck-Werninghaus C, Brewe F, et al: Specific oral tolerance induction in food allergy in children: efficacy and clinical patterns of reaction. Allergy 2007; 62: Skripak JM, Nash SD, Rowley H, et al: A randomized, double-blind, placebo-controlled study of milk oral immunotherapy for cow's milk allergy. J Allergy Clin Immunol 2008; 122: Narisety SD, Skripak JM, Steele P, et al: Open-label maintenance after milk oral immunotherapy for IgE-mediated cow's milk allergy. J Allergy Clin Immunol 2009; 124: Bégin P, Winterroth LC, Dominguez T, et al: Safety and feasibility of oral immunotherapy to multiple allergens for food allergy. Allergy Asthma Clin Immunol 2014; 10: Nadeau KC, Schneider LC, Hoyte L, et al: Rapid oral desensitization in combination with omalizumab therapy in patients with cow's milk allergy. J Allergy Clin Immunol 2011; 127: Schneider LC, Rachid R, LeBovidge J, et al: A pilot study of omalizumab to facilitate rapid oral desensitization in high-risk peanut-allergic patients. J Allergy Clin Immunol 2013; 132: Bégin P, Dominguez T, Wilson SP, et al: Phase 1 results of safety and tolerability in a rush oral immunotherapy protocol to multiple foods using omalizumab. Allergy Asthma Clin Immunol 2014; 10: Enrique E, Pineda F, Malek T, et al: Sublingual immunotherapy for hazelnut food allergy: a randomized, double-blind, placebocontrolled study with a standardized hazelnut extract. J Allergy Clin Immunol 2005; 116: Fernández-Rivas M, Garrido Fernández S, Nadal JA, et al: Randomized double-blind, placebo-controlled trial of sublingual immunotherapy with a Pru p 3 quantified peach extract. Allergy 2009; 64: Strategies to Induce Tolerance 53

20