Peanut oral immunotherapy increases Tregs and epigenetically modifies FOXP3

The mechanisms contributing to clinical immune tolerance are largely unknown. The objective of Syed et al was to study the changes associated with clinical immune tolerance in antigen induced T cells, basophils, and antibodies in subjects undergoing oral immunotherapy (OIT) for peanut allergy (J Allergy Clin Immunol 2014; 133(2): 500-510). The induction of regulatory T (Treg) cells has been a potential mechanism of maintaining immune tolerance, with Treg deficiencies implicated in the development of allergies.  Fork head box protein 3 (Foxp3) is a transcription factor that regulates Tregs, including natural regulatory Tregs (nTregs) and induced regulatory Tregs (iTregs). Epigenetic modifications to regions within the Foxp3 locus have been associated with stable Foxp3 expression and Treg cell-suppressive function.  

The authors investigated whether antigen-induced Tregs (aiTregs) and humoral and basophil immune markers are induced by OIT in clinically immune tolerant (IT) versus non-tolerant (NT) patients after OIT.  20 participants (with 20 peanut allergic controls undergoing standard avoidance therapy) successfully completed 24 months of OIT, tolerating up to 4g of peanut protein after maintenance therapy.   After 3 months of peanut avoidance, only 7 of 20 participants were defined as IT participants; these 7 avoided peanut for an additional 3 months and only 3 of 7 remained clinically nonreactive (IT).  The peanut induced basophil response was reduced in the OIT participants with a trend of more reduction in the IT group. The IT participants had higher numbers of ai-Treg cells with greater suppressive function and with higher levels of FOXP3 hypomethalation compared with NT and control participants.  The population of ai-Tregs the authors identified during OIT had a marked increase in Foxp3 expression and associated increases in both chemotaxis toward intestinal epithelial cells and suppressive function toward antigen-induced effector T cells (Teff).  Furthermore, dendritic cells (DCs) isolated after OIT therapy significantly decreased the methylation of Foxp3 in Teff cells.

These data suggest that ai-Treg cells are a key regulatory cell type modulating the immune response during OIT and that epigenetic regulation of these T cells might contribute to the induction of such immune tolerance. Although larger phase 2 clinical trials in OIT are justified and feasible, the results may be predictive of a state of operationally defined clinical immune tolerance during peanut OIT and contribute to providing safe and effective therapy for patients with peanut allergy.

Below are some questions posed to the authors regarding their article and the authors’ responses:

Why do there seem to be mixed results about the presence of Treg in food oral immunotherapy in humans?
The gating and definition of Treg is very important. Some labs report CD4+CD25+Foxp3+ cells as Treg but Treg should best be defined by suppression assays on the functional level if there is enough blood sample. In addition, the presence of Foxp3 can indicate an activated CD4+ T cell, not necessarily a Treg and Treg don't have to have Foxp3 to be a Treg.

Is there potential for Type 2 cytokines to be immune markers in IT vs NT participants?

We have examined some Th2 markers by flow cytometry and intracellular staining with no specific trends showing a difference between IT vs NT but further studies are underway.

What if any data is available about ILC2 activation during food allergy?

We are currently examining ILC2 cells in the periphery of food allergic patients.

What could be a potential outcome if patients did not avoid peanut after OIT therapy and included it in their diets?

There are several trials around the world examining this and we will be conducting mechanistic studies to try to understand differences in desensitization vs sustained unresponsiveness long term.

Food allergy: clinical advances and updates

The prevalence of food allergy is on the rise with up to 10% of the population afflicted, though remarkable advances have occurred in understanding and managing food allergies.  In their review article, Sicherer and Sampson focus on advances and updates in epidemiology, pathogenesis, diagnosis and treatment of food allergy (J Allergy Clin Immunol 2014; 133(2): 291-307).  They explain that numerous genetic and environmental risk factors have been identified, with rectifiable risk factors such as vitamin D insufficiency, excess dietary fat, obesity, increased hygiene and timing of exposure to foods as being potential targets to address in prevention for the future.  Interesting clinical insights on route of sensitization, allergen characterization and immune response provide guidance for diagnosis and treatment. Allergen avoidance and emergency treatment remain the current management option for patients, although numerous clinical trials are underway for more definitive therapies.

 A major change in approach to management stems from the observation that many children with milk or egg allergy tolerate heat denatured forms of these foods and that ingestion of these foods by children able to do so may speed allergy recovery.   Furthermore, recommendations about prevention of food allergy and atopic disease through diet have changed radically, with rescinding of many recommendations about extensive and prolonged allergen avoidance. 

There is a wide spectrum of disease caused by food allergy and diagnosis depends on combining a knowledge of pathophysiology and epidemiology with patient history and test results. The authors explain that numerous clinical trials are underway for more definitive therapies, as is basic science research using novel approaches in animal models.  

With a deeper understanding of genetics and the microbiome, incorporation of bioinformatics and numerous approaches to treatment, the next several years show promise of a revolution in the clinical approach to food allergy. 

Questions for the authors:

What impact does the modern world have on food allergy, such as the American diet, lack of exercise, and stress that aren’t discussed in this review?  Are there clinical trials in place that use a more natural approach to improving outcomes?

There are a number of studies underway looking at a more natural approach to introducing foods to the diet (earlier).  The review discusses how modern living may affect allergy outcomes (e.g., lower vitamin D due to sunscreens and indoor activities, obesity, etc., and a study showing a “healthy diet” may be important to reduce allergy.  The role of stress has not been extensively studied but a review of this topic is in press:

Schreier HM, Wright RJ. Stress and food allergy: mechanistic considerations. Ann Allergy Asthma Immunol. 2013 Aug 28. pii: S1081-1206(13)00561-9. doi: 10.1016/j.anai.2013.08.002. [Epub ahead of print]

Insights provided by mouse models relating to food allergy

Food allergy is a growing public health concern due to its increasing prevalence and life threatening potential.  Mouse models of food allergy have become useful tools for identifying the mechanisms involved in the sensitization of food allergens which are normally harmless as well as delineating the critical immune components of the effector phase of allergic reactions to food. In their review, Oyoshi et al have summarized the importance of animal models in food allergy research contrary to concerns regarding the relevance murine models have in understanding human disease (J Allergy Clin Immunol 2014; 133(2): 309-317). 

Mouse models have been exceedingly useful in the study of atopic dermatitis (AD) and asthma, paving the way for food allergy research.  For example, allergic sensitization or tolerance can be induced to specific allergens under controlled environmental conditions within defined genetic backgrounds that cannot be matched in human studies. The importance of Treg cells in the development of tolerance has been established in both mouse and humans in which deficiency of forkhead box protein 3 (foxp3)+ T cells leads to increased allergic disorders such as AD and food allergy.  These T cells have been found to be reduced in antibiotic treated mice which exhibit a predisposition towards allergic sensitization.  Furthermore, administration of commensal microbiota to these mice promoted Treg cells and limited allergic responses to foods. 

The authors explain how animal models of food allergy are invaluable tools for dissecting etiology, mechanisms and preventive strategies, as well as assisting in the identification, validation and development of therapies before they progress towards patients. While the application of animal models to human disease requires careful and thorough consideration and interpretation, their utility in facilitating truly translational discoveries has been demonstrated repeatedly and on many levels. Particularly in the setting of food allergy, where risks of adverse reactions to therapy are a major issue for patients, animal models will be indispensable to effectively and ethically develop new treatments. Mechanistically, the recent discoveries of the  roles of microbiota on the etiology of food allergy, derived from studies of animal models, provides an excellent example of how lessons learned from experimental animals can provide new breakthrough areas that educate future studies of host factors in human patients with food allergy. 

Question for the authors:

What effects on food allergy research do you anticipate from the significant cuts in basic science research funding that may reduce overall animal research?

Short-time decisions of the budget cuts in basic research funding in a time of financial crisis may lead to an abrupt slow of growth of basic science with severe long-term consequences. Particularly in the area of food allergy, where animal models are indispensible to perform mechanistic studies that are often not feasible in humans because of ethical, technical, and even financial reasons, cuts in basic science funding will leave researchers with fewer opportunities to try novel and innovative ideas that could have a high return. In addition, many of young basic scientists with limited budgets cannot survive even short term cuts. The annual costs to pediatric food allergy have been estimated at $25 billion. In today’s environment, I believe that the continued careful and coordinative work of basic and clinical science with strong support will lead to effective development of new treatment for food allergy.

Maternal psychological distress during pregnancy affects wheezing in preschool children

Mechanisms such as social, behavioral, and environmental factors during pregnancy that may affect the risk of childhood asthma have been studied, but only few studies have assessed the relationship between maternal psychological distress and childhood wheezing.  Guxens et al performed a population-based study among children from birth to 6 years of age while assessing maternal and paternal psychological distress during pregnancy (J Allergy Clin Immunol 2014; 133(1): 59-67).  This large study of 4,848 children was embedded within the Generation R study, a population-based prospective cohort from fetal life onwards in Rotterdam, The Netherlands.  Maternal and paternal distress data was collected at 20 weeks gestation and again at 2 and 6 months, and 3 years after delivery by using the Brief Symptom Inventory questionnaire. Wheezing was annually assessed in the children up to 4 years of age. Physician diagnosed ever asthma was reported by parents at age 6 years. 

The authors point out that the association between maternal psychological distress during pregnancy and childhood asthma might be explained by developmental adaptations of the hypothalamic-pituitary-adrenal axis, the autonomic nervous system, the lung structure and function, and immune responses in the children but also other mechanisms such as social, behavioral, or environmental factors.  To assess the role of different mechanisms, the authors assessed the associations both for the mother (unique intrauterine mechanism) and father (social, behavioral, or environmental factors), all taking into account many socioeconomic and lifestyle variables, and maternal psychological distress after the child was born. Children ultimately included in the analysis were more frequently from parents with a higher education level, and their parents had less psychological distress during pregnancy compared to those lost in follow-up. 

Guxens concludes that mothers with psychological distress during pregnancy had increased odds of overall wheezing in their children from 1 to 4 years of life, or of asthma at 6 years of age, compared to mothers that did not.  This result was similar in children of mothers with a history of asthma and atopy, compared to those without.  The authors go on to show that the robust result is independent of paternal psychological distress during pregnancy and maternal and paternal psychological distress after delivery.  These results suggest a direct intrauterine programming effect of maternal psychological distress on respiratory morbidity. To study these potential underlying biological mechanisms, markers of stress (diurnal cortisol rythm) and changes in the immune system (types of lymphocytes) could be measured. Our study has these data in a limited number of children only. Data for a genetic underlying mechanism, including interaction with environmental factors, are available and will be explored in the future. 

Implications of race and ethnicity on genetic profiles for personalized medicine

It is well known that frequencies and severity of disease can differ among races.  The mixing of African, European, and Native American ancestries has resulted in a variety of different ethnic groups with varied diversity. There is robust evidence from various clinical trials that different ethnic groups have variable responses to specific therapeutic agents.  The study of pharmacogenetics is used to personalize therapies specific to individuals from different ethnic or racial groups and it has historically been composed of mostly non-Hispanic whites of European descent. 

Ortega and Meyers summarize the genetic and epidemiologic basis for the variable genetic backgrounds observed between different, recently admixed ethnic groups such as African American or Hispanic (J Allergy Clin Immunol 2014; 133(1): 16-26). Rare genetic variants appear to be more frequent among individuals of African ancestry and could account for inter-ethnic differences in drug responses especially for rare, adverse events. Variability in phenotypes important in determining asthma severity between different ethnic groups could impact therapeutic responses to commonly used therapies.  For example, individuals with higher degrees of African ancestry have lower baseline lung function and increased risk for asthma exacerbations . In addition, African Americans are an ethnic group with an increased risk for adverse, possibly life-threatening, responses to long acting B2-adrenergic receptor agonists compared to non-Hispanic whites. 

The authors go on to explain that the genetic complexity of admixed ethnic groups is further challenged by representing only a minority of subjects enrolled in studies, cultural and habitual differences, and increased genetic diversity at an individual level.  These complex variables will require a combination of methods including genome-wide association studies, admixture-based analytical methods, and next-generation sequencing in larger populations to find and study genetic differences among ethnic groups and contribute to future pharmacogenetic studies.  

Treatment of peanut allergy with omalizumab

Peanut allergy is a well known food allergy estimated to include 3-4% of the US population and accounts for a disproportionate number of severe allergic reactions. The vast majority of food allergy death is related to peanut allergy and is often ingested accidentally despite strict food avoidance. Peanut allergy sensitivity often fails to diminish over time compared to other food allergens causing a lifetime of anxiety and food avoidance for patients and families. The only effective treatment option for this epidemic other than food avoidance is ready access to injectable epinephrine. 

Recent clinical trials using double blind, placebo controlled food challenges (DBPCFC) have reported success with allergen immunotherapy and desensitization with common food allergens including peanut. Although long term tolerance can be achieved with daily intake, most patients experienced mild to severe symptoms including anaphylaxis which occurred in up to 25% of patients with a high peanut specific IgE. Nevertheless, these trials demonstrate that oral food challenge is a useful method for treating food allergies by increasing the threshold for tolerance with possible resolution.

Schneider et al hypothesized that treatment with an anti-IgE monoclonal antibody (mAb) such as omalizumab may contribute to a more rapid desensitization with greater success (Journal of Allergy and Clinical Immunology 2013; 132(6): 1368-1374). Omalizumab binds free IgE which inhibits allergic reactions and is currently approved for older children and adults with moderate to severe asthma. 

The authors administered the drug prior to and during oral peanut desensitization to 13 children who failed initial DBPCFC at low doses (< 100 mg of peanut flour). After pretreatment, all subjects tolerated initial desensitization doses given on the first day, including the maximum dose of 500 mg of peanut flour and 12 subjects reached the maximum daily dose of 4000mg/day within a median time of 8 weeks, at which point omalizumab was discontinued.  The 12 subjects continued the 4000mg/day of peanut flour and subsequently tolerated a challenge of 8000 mg which is up to 400 times the dose that was tolerated before desensitization.  Of the 13 subjects, 6 experienced mild or no allergic symptoms, 6 had a grade 2 reaction, and 2 had a grade 3 reaction which all responded rapidly to treatment. 

These results suggest that omalizumab can facilitate a more rapid oral desensitization in high risk patients with high peanut specific IgE. Schneider’s results provide strong evidence that omalizumab can effectively reduce allergic reactions and expedite rapid oral desensitization.  Larger studies are currently under way to confirm the beneficial role of omalizumab in facilitating oral peanut desensitization.

Questions for the authors:

Could this treatment be used for other severe allergic diseases not yet indicated?  Could longer treatment with omalizumab prevent or reduce other side effects such as eosinophil esophagitis?

Further studies would be needed to determine whether this approach can be applied to other severe allergic diseases.  We don't know if  longer treatment with omalizumab would reduce reactions during desensitization since this was not part of the study.

Atopic disease and the herpes microbiome

Unlike bacteria or fungi, herpes viruses establish life-long infection in the human host through latent genomic persistence within the host cells nuclei and are thus considered part of the human microbiome. The ability of the virus to interact with the human genome influences allergic and atopic disease due to the bias these patients have towards a Th2 profile. Dr. David Dreyfus examined the role of common human herpes viruses on the microbiome of atopic patients, who have more severe and atypical disease when infected (J Allergy Clin Immunol 2013; 132(6): 1278-1286).

The herpes virus Epstein Barr (EBV or mononucleosis) expresses latency in lymphocytes and has co-evolved with humans long enough to encode for a protein that resembles the cytokine IL-10, as well as other cytokines by activating host transcription factors. The author explains that EBV and other herpes viruses encode for microRNAs that cause immunomodulation of distant cells. This lead to studies suggesting that EBV infection early in life is protective against atopic disease compared to infection later in life, where there is a predisposition to atopic disease. Atopic patients that have a primary infection of EBV can have a more severe non-specific rash that can be mistaken for allergic diseases or other infectious diseases and can be misdiagnosed.

Another ubiquitous herpes virus, VZV or shingles, becomes latent in neuronal cells and reactivation risk increases with age as the natural antibodies decline over time. The current VZV vaccine has been successful for a decade, reducing the VZV associated morbidity and mortality especially in the elderly. However, studies suggest that children who receive the vaccine have an increased incidence for atopic disease compared to children that are infected naturally with the virus, suggesting there could be a protective benefit to natural infection. Like VZV, Human Simplex Virus 1 and 2 (HSV1 and HSV 2) become latent in the neuron after primary infection of subcutaneous and pulmonary epithelial cells. The author explains that infants and others that are immunodeficient are at high risk of pulmonary syndromes such as bronchitis and pneumonia, and neurologic syndromes such as encephalitis. The most common atypical presentation of the virus is a severe skin rash called eczema herpeticum among atopic patients. This is similar to the severe drug reaction with eosinophilia and systemic symptoms (DRESS syndrome) seen with the reactivation of HSV6, also referred to as roseola, which is present in both atopic and non-atopic patients. Furthermore, another related herpes virus, cytomegalovirus (CMV) can cause cutaneous symptoms similar to EBV, as well as chronic inflammation and cardiovascular disease due to its latency in macrophages that are associated with vascular inflammation by directly inducing Th2 cytokines.

Molecular evidence suggests that herpes viruses have been co-evolving with the human immune system since before the origin of the adaptive immune system. The herpes microbiome along with environmental factors such as improved global hygiene that prevents the exposure to a variety of components interact with each other to create the atopic phenotype. Dr. Dreyfus emphasizes that this information can positively impact the care for the allergic and atopic patient populations. Clinicians must have a heightened awareness of the various presentations of herpes viruses to avoid unnecessary testing and treatments for drug allergy or autoimmune disease. 

Questions for the author:

What are the effects of the herpes microbiome on other allergic diseases?  Is there an increase in Th2 dependent disease in patients who express viral lesions and reactivation of herpes viruses? 

Regarding the question of effects of herpes on atopic disease, in addition to the references cited in my article particularly regarding an increase in atopic disease in pediatric patients who receive the varicella vaccine vs wild type virus there is a more recent article just published with some more experimental observations on this question:

Sohlberg, E. et al (2013) Cytomegalovirus-Seropositive Children Show Inhibition of an In Vitro EBV infection That Is Associated with CD8+ CD57+ T-cell Enrichment and IFN-gamma. Journal of Immunology 191:5669