Mind the gaps: Clinical trial concepts to address unanswered questions in aeroallergen immunotherapy-An NIAID/AHRQ Workshop.

The Agency for Healthcare Research and Quality and the National Institute of Allergy and Infectious Diseases organized a workshop to develop trial concepts that could improve the use and effectiveness of aeroallergen immunotherapy (AAIT). Expert groups were formed to accomplish the following tasks: (1) propose a study design to compare the effectiveness and safety of subcutaneous versus sublingual AAIT; (2) propose a study design to compare the effectiveness and safety of AAIT by using 1 or a few allergens versus all or most allergens to which a patient is sensitized; (3) propose a study design to determine whether AAIT can alter the progression of childhood allergic airways disease; and (4) propose a study design to determine the optimal dose and duration of AAIT to achieve maximal effectiveness with acceptable safety. Study designs were presented by the workgroups, extensively discussed at the workshop, and revised for this report. The proposed trials would be of long duration and require large highly characterized patient populations. Scientific caveats and feasibility matters are discussed. These concepts are intended to help the development of clinical trials that can address some of the major questions related to the practice of AAIT for the management and prevention of allergic airways disease.

In house validation of a high resolution mass spectrometry Orbitrap-based method for multiple allergen detection in a processed model food.

In recent years, mass spectrometry (MS) has been establishing its role in the development of analytical methods for multiple allergen detection, but most analyses are being carried out on low-resolution mass spectrometers such as triple quadrupole or ion traps. In this investigation, performance provided by a high resolution (HR) hybrid quadrupole-Orbitrap™ MS platform for the multiple allergens detection in processed food matrix is presented. In particular, three different acquisition modes were compared: full-MS, targeted-selected ion monitoring with data-dependent fragmentation (t-SIM/dd2), and parallel reaction monitoring. In order to challenge the HR-MS platform, the sample preparation was kept as simple as possible, limited to a 30-min ultrasound-aided protein extraction followed by clean-up with disposable size exclusion cartridges. Selected peptide markers tracing for five allergenic ingredients namely skim milk, whole egg, soy flour, ground hazelnut, and ground peanut were monitored in home-made cookies chosen as model processed matrix. Timed t-SIM/dd2 was found the best choice as a good compromise between sensitivity and accuracy, accomplishing the detection of 17 peptides originating from the five allergens in the same run. The optimized method was validated in-house through the evaluation of matrix and processing effects, recoveries, and precision. The selected quantitative markers for each allergenic ingredient provided quantification of 60-100 µgingred/g allergenic ingredient/matrix in incurred cookies.

Early Introduction of Multi-Allergen Mixture for Prevention of Food Allergy: Pilot Study.

The incidence and prevalence of food allergy (FA) is increasing. While several studies have established the safety and efficacy of early introduction of single allergens in infants for the prevention of FA, the exact dose, frequency, and number of allergens that can be safely introduced to infants, particularly in those at high or low risk of atopy, are still unclear. This 1-year pilot study evaluated the safety of the early introduction of single foods (milk, egg, or peanut) vs. two foods (milk/egg, egg/peanut, milk/peanut) vs. multiple foods (milk/egg/peanut/cashew/almond/shrimp/walnut/wheat/salmon/hazelnut at low, medium, or high doses) vs. no early introduction in 180 infants between 4-6 months of age. At the end of the study, they were evaluated for plasma biomarkers associated with food reactivity via standardized blood tests. Two to four years after the start of the study, participants were evaluated by standardized food challenges. The serving sizes for the single, double, and low dose mixtures were 300 mg total protein per day. The serving sizes for the medium and high dose mixtures were 900 mg and 3000 mg total protein, respectively. Equal parts of each protein were used for double or mixture foods. All infants were breastfed until at least six months of age. The results demonstrate that infants at either high or low risk for atopy were able to tolerate the early introduction of multiple allergenic foods with no increases in any safety issues, including eczema, FA, or food protein induced enterocolitis. The mixtures of foods at either low, medium, or high doses demonstrated trends for improvement in food challenge reactivity and plasma biomarkers compared to single and double food introductions. The results of this study suggest that the early introduction of foods, particularly simultaneous mixtures of many allergenic foods, may be safe and efficacious for preventing FA and can occur safely. These results need to be confirmed by larger randomized controlled studies.

Oral immunotherapy for peanut allergy: The con argument.

BACKGROUND: In some countries of the world, peanut allergy represents an important source of anaphylactic reactions. Traditionally treated with the avoidance of responsible allergens, this condition can also be targeted by oral peanut immunotherapy. METHODS: In this study, we review the beneficial and side effects of currently available forms of peanut oral immunotherapy (POIT). We report the discussions resulting from the publication of a meta-analysis that brought to light the downsides of oral immunotherapy for peanuts. RESULTS: In some clinical situations, the risk-benefit ratio can favor peanut oral immunotherapy over avoidance. In many other situations, this is not the case. The decision must be based on the values and preferences of clinicians and patients. Those not ready to accept serious adverse effects from POIT are likely to continue the elimination diet; those motivated to achieving desensitization, and prepared to accept serious adverse effects, may choose to undergo POIT. CONCLUSIONS: Without being prejudiced against peanut oral immunotherapy, we indicate the possible evolution of treatment for this condition is in a rapidly evolving broader scenario. Among the future options, sublingual immunotherapy, parenteral immunotherapy with modified allergens, transcutaneous immunotherapy, and the use of biologics will become important options.

Single-allergen sublingual immunotherapy versus multi-allergen subcutaneous immunotherapy for children with allergic rhinitis.

It has always been controversial whether a single allergen performs better than multiple allergens in polysensitized patients during the allergen-specific immunotherapy. This study aimed to examine the clinical efficacy of single-allergen sublingual immunotherapy (SLIT) versus multi-allergen subcutaneous immunotherapy (SCIT) and to discover the change of the biomarker IL-4 after 1-year immunotherapy in polysensitized children aged 6-13 years with allergic rhinitis (AR) induced by house dust mites (HDMs). The AR polysensitized children (n=78) were randomly divided into two groups: SLIT group and SCIT group. Patients in the SLIT group sublingually received a single HDM extract and those in the SCIT group were subcutaneously given multiple-allergen extracts (HDM in combination with other clinically relevant allergen extracts). Before and 1 year after the allergen-specific immunotherapy (ASIT), the total nasal symptom scores (TNSS), total medication scores (TMS) and IL-4 levels in peripheral blood mononuclear cells (PBMCs) were compared respectively between the two groups. The results showed that the TNSS were greatly improved, and the TMS and IL-4 levels were significantly decreased after 1-year ASIT in both groups (SLIT group: P<0.001; SCIT group: P<0.001). There were no significant differences in any outcome measures between the two groups (for TNSS: P>0.05; for TMS: P>0.05; for IL-4 levels: P>0.05). It was concluded that the clinical efficacy of single-allergen SLIT is comparable with that of multi-allergen SCIT in 6-13-year-old children with HDM-induced AR.

Streamlining the analytical workflow for multiplex MS/MS allergen detection in processed foods.

Allergenic ingredients in pre-packaged foods are regulated by EU legislation mandating their inclusion on labels. In order to protect allergic consumers, sensitive analytical methods are required for detect allergen traces in different food products. As a follow-up to our previous investigations, an optimized, sensitive, label-free LC-MS/MS method for multiplex detection of five allergenic ingredients in a processed food matrix is proposed. A cookie base was chosen as a complex food matrix and home-made cookies incurred with whole egg, skimmed milk, soy flour, ground hazelnut and ground peanut were prepared at laboratory scale. In order to improve the analytical workflow both protein extraction and purification protocols were optimized and finally a sensitive streamlined SRM based analytical method for allergens detection in incurred cookies was devised. The effect of baking on the detection of selected markers was also investigated.

Management of the polyallergic patient with allergy immunotherapy: a practice-based approach.

BACKGROUND: The great majority (60-80 %) of patients consulting specialist physicians for allergic respiratory disease are polysensitized and thus may be potentially clinically polyallergic. However, management approaches to allergen immunotherapy (AIT) in polysensitized and polyallergic patients are not standardized. METHODS: An international group of clinicians with in-depth expertise in AIT product development, clinical trials and clinical practice met to generate up-to-date, unambiguous, pragmatic guidance on AIT in polysensitized and polyallergic patients. The guidance was developed after reviewing (1) the current stance of regulatory bodies and learned societies, (2) the literature data on single- and multi-AIT and (3) the members' confirmed clinical experience with polysensitized patients. RESULTS: AIT is safe and effective in polysensitized and polyallergic patients, and should always be based on the identification of one or more clinically relevant allergens (based on the type and severity of symptoms, the duration of induced symptoms, the impact on quality of life and how difficult an allergen is to avoid). Single-AIT is recommended in polyallergic patients in whom one of the relevant allergens is nevertheless clearly responsible for the most intense and/or bothersome symptoms. Parallel 2-allergen immunotherapy or mixed 2-allergen immunotherapy is indicated in polyallergic patients in whom two causal relevant allergens have a marked clinical and QoL impact. In parallel 2-allergen immunotherapy (whether subcutaneous or sublingual), high-quality, standardized, single-allergen formulations must be administered with an interval of 30 min. Mixing of allergen extracts may be considered, as long as (1) the mixture is technically feasible, (2) the mixture is allowed from a regulatory standpoint, (3) the allergen doses are reduced in proportion to the number of components but are still at concentrations with demonstrated efficacy. CONCLUSIONS: Physicians can prescribe AIT (preferably with high-quality, standardized, single-allergen formulations) with confidence in polysensitized and polyallergic patients by focusing on clinical/QoL relevance and safety.

Multi-allergen detection in food by micro high-performance liquid chromatography coupled to a dual cell linear ion trap mass spectrometry.

There is a raising demand for sensitive and high throughput MS based methods for screening purposes especially tailored to the detection of allergen contaminants in different food commodities. A challenging issue is represented by complex food matrices where the antibody-based kits commercially available might encounter objective limitations consequently to epitope masking phenomena due to a multitude of interfering compounds arising from the matrix. The performance of a method duly optimized for the extraction and simultaneous detection of soy, egg and milk allergens in a cookie food matrix by microHPLC-ESI-MS/MS, is herein reported. Thanks to the innovative configuration and the versatility shown by the dual cell linear ion trap MS used, the most intense and reliable peptide markers were first identified by untargeted survey experiment, and subsequently employed to design an ad hoc multi-target SRM method, based on the most intense transitions recorded for each selected precursor peptide. A sample extraction and purification protocol was optimized also including an additional step based on sonication, which resulted in a considerable improvement in the detection of milk allergen peptides. Data Dependent™ Acquisition scheme allowed to fill out a tentative list of potential peptide markers, which were further filtered upon fulfilling specific requirements. A total of eleven peptides were monitored simultaneously for confirmation purposes of each allergenic contaminant and the two most sensitive peptide markers/protein were selected in order to retrieve quantitative information. Relevant LODs were found to range from 0.1µg/g for milk to 0.3µg/g for egg and 2µg/g for soy.

Orbitrap™ monostage MS versus hybrid linear ion trap MS: application to multi-allergen screening in wine.

Food allergen research has made giant steps in the last years thanks to the features offered by the latest technology of mass analyzers placed on the market allowing multiplex sensitive detection of proteins. Potentials and features of two mass analyzers namely a linear ion trap capable of performing a data dependent or selected reaction monitoring analysis and an Orbitrap(TM) stand-alone MS enabling a broadband fragmentation without mass selection at highest mass resolving power are herein described and applied to the multiplex screening of allergens in a type of wine chosen as a reference matrix. Quantitative and confirmative capabilities of both platforms were assessed on the specific case study, the multiple detection of egg and milk -related proteins, typically employed in white wines as fining agents. Commercial bioinformatic tools used for a quick allergen identification will be also discussed.

Single-allergen Sublingual Immunotherapy versus Multi-allergen Subcutaneous Immunotherapy for Children with Allergic Rhinitis / 华中科技大学学报（医学）（英德文版）

It has always been controversial whether a single allergen performs better than multiple allergens in polysensitized patients during the allergen-specific immunotherapy.This study aimed to examine the clinical efficacy of single-allergen sublingual immunotherapy (SLIT) versus multi-allergen subcutaneous immunotherapy (SCIT) and to discover the change of the biomarker IL-4 after 1-year immunotherapy in polysensitized children aged 6-13 years with allergic rhinitis (AR) induced by house dust mites (HDMs).The AR polysensitized children (n=78) were randomly divided into two groups:SLIT group and SCIT group.Patients in the SLIT group sublingually received a single HDM extract and those in the SCIT group were subcutaneously given multiple-allergen extracts (HDM in combination with other clinically relevant allergen extracts).Before and 1 year after the allergen-specific immunotherapy (ASIT),the total nasal symptom scores (TNSS),total medication scores (TMS) and IL-4 levels in peripheral blood mononuclear cells (PBMCs) were compared respectively between the two groups.The results showed that the TNSS were greatly improved,and the TMS and IL-4 levels were significantly decreased after 1-year ASIT in both groups (SLIT group:P＜0.001;SCIT group:P＜0.001).There were no significant differences in any outcome measures between the two groups (for TNSS:P＞0.05;for TMS:P＞0.05;for IL-4 levels:P＞0.05).It was concluded that the clinical efficacy of single-allergen SLIT is comparable with that of multi-allergen SCIT in 6-13-year-old children with HDM-induced AR.