Clinical management of plant food allergy in patients sensitized to lipid transfer proteins is heterogeneous: identifying the gaps.

BACKGROUND AND OBJECTIVE: Patients sensitized to lipid transfer protein (LTP) present a wide clinical variability. The lack of practical diagnostic and therapeutic guidelines complicate their management. The aim of the study was to describe the clinical approach of Spanish allergists to this pathology using a survey designed by PICO method and subsequent Delphi approach validation. METHODS: Designed survey was answered by 224 allergists (75% women; 57.1% with >20 years of professional experience). Homogeneity regarding clinical practice on the main points of LTP allergy diagnosis was observed, except for patients with suspected NSAID hypersensitivity (44.6% frequently include LTP skin testing). Oral food challenges were not frequently performed (63.6% occasionally to never), and they were generally (75.5%) used to confirm tolerance. It was common to recommend fruit skins avoidance (77.2%) and maintaining consumption of foods to which patients are sensitised but tolerant (99.1%). RESULTS: There was heterogeneity on other dietary indications, modifications due to co-factors, or traces avoidance. Peach sublingual immunotherapy (SLIT) was considered very/quite effective by 55.9% of allergists. The majority (79.5%) consider SLIT indicated in <25% of LTP allergic patients, based on severity (95.2%), frequency of reactions (99.4%), allergy to multiple food families (97.4%), and the quality of life/nutrition impairment (91.5%). There was different practice on SLIT prescription based on co-factor involvement. CONCLUSION: These data suggest that there is a need to increase evidence to reduce the clinical practice heterogeneity in the management of LTP allergy.

Unraveling the Diagnosis of Kiwifruit Allergy: Usefulness of Current Diagnostic Tests.

OBJECTIVES: To determine the usefulness of the in vitro and in vivo methods used in the diagnosis of kiwifruit allergy and to specifically assess the impact of seed proteins on sensitivity. METHODS: We performed skin prick tests (SPTs) using various commercial extracts, homemade pulp, and seed extracts and prick-prick tests with kiwifruit on 36 allergic patients. The presence of specific IgE (sIgE) was assessed using the ImmunoCAP (kiwifruit extract), ELISA (Act d 1, Act d 2), ISAC, and FABER assays. Immunoblotting of seed extract was carried out, and a single-blind oral food challenge was performed with whole seeds in seed-sensitized individuals. RESULTS: The prick prick test with kiwifruit demonstrated the highest diagnostic capacity (81.8% sensitivity and 94.1% specificity) among the in vivo tests. The sIgE levels measured using ImmunoCAP (kiwifruit extract) showed a similar sensitivity to that of global ISAC and FABER (63.9%, 59.5%, and 58.3%, respectively). Act d 1 was the major allergen. Sensitization to Act d 1 was associated with positive sIgE results to whole kiwifruit extract detected by ImmunoCAP (P<.000). A positive SPT result to kiwifruit seeds was associated with severe symptoms induced by kiwifruit (P=.019) as a marker of advanced disease, but not with clinically relevant sensitization. Challenge testing with kiwifruit seeds performed on 8 seed-sensitized patients yielded negative results. CONCLUSION: Sensitization to Act d 1 is associated with a positive result in conventional diagnostic techniques, whereas kiwifruit seed sensitization does not increase the sensitivity of the diagnostic techniques evaluated.

Comparable actions of omalizumab on mast cells and basophils.

BACKGROUND: Omalizumab (OmAb) has recently been approved for the treatment of diseases other than allergic asthma, including chronic urticaria. The exploration of the use of OmAb in chronic urticaria was based on the presence of IgE autoantibodies against autoantigens such as anti-IgE, anti-FcεRI, and IgE antibodies against thyroid peroxidase in certain patients with chronic urticaria. OmAb recognizes and sequesters free IgE to prevent its interaction with FcεRI. However, OmAb is equally and rapidly effective against autoimmune and non-autoimmune urticaria, suggesting the possible involvement of additional mechanisms of IgE. OBJECTIVES: We sought to investigate the in vitro mechanism of action of OmAb in mast cells and basophils. METHODS: Both LAD2 human mast cell line, previously sensitized with IgE, and ex vivo basophils were incubated with OmAb at different doses, analysing its effect on IgE-dependent events (e.g., degranulation, phosphorylation-mediated signalling, and eicosanoid release). RESULTS: We found that OmAb dissociates pre-bound IgE from mast cells and basophils, resulting in a reduction of proximal phosphorylation-mediated signalling events (Syk, PLCγ, and LAT) and in a decrease in degranulation and leukotriene synthesis. CONCLUSION: Our data prove the existence of common mechanisms of action of OmAb in mast cells and basophils that would explain its effectiveness and rapid effect in chronic urticaria and provide a basis for its use in other diseases mediated by these cells.

Is Microarray Analysis Really Useful and Sufficient to Diagnose Nut Allergy in the Mediterranean Area?

BACKGROUND: Component-based diagnosis on multiplex platforms is widely used in food allergy but its clinical performance has not been evaluated in nut allergy. OBJECTIVE: To assess the diagnostic performance of a commercial protein microarray in the determination of specific IgE (sIgE) in peanut, hazelnut, and walnut allergy. METHODS: sIgE was measured in 36 peanut-allergic, 36 hazelnut-allergic, and 44 walnut-allergic patients by ISAC 112, and subsequently, sIgE against available components was determined by ImmunoCAP in patients with negative ISAC results. ImmunoCAP was also used to measure sIgE to Ara h 9, Cora 8, and Jug r 3 in a subgroup of lipid transfer protein (LTP)-sensitized nut-allergic patients (positive skin prick test to LTP-enriched extract). sIgE levels by ImmunoCAP were compared with ISAC ranges. RESULTS: Most peanut-, hazelnut-, and walnut-allergic patients were sensitized to the corresponding nut LTP (Ara h 9, 66.7%; Cor a 8, 80.5%; Jug r 3, 84% respectively). However, ISAC did not detect sIgE in 33.3% of peanut-allergic patients, 13.9% of hazelnut-allergic patients, or 13.6% of walnut-allergic patients. sIgE determination by ImmunoCAP detected sensitization to Ara h 9, Cor a 8, and Jug r 3 in, respectively, 61.5% of peanut-allergic patients, 60% of hazelnut-allergic patients, and 88.3% of walnut-allergic patients with negative ISAC results. In the subgroup of peach LTP-sensitized patients, Ara h 9 sIgE was detected in more cases by ImmunoCAP than by ISAC (94.4% vs 72.2%, P < .05). Similar rates of Cora 8 and Jug r 3 sensitization were detected by both techniques. CONCLUSIONS: The diagnostic performance of ISAC was adequate for hazelnut and walnut allergy but not for peanut allergy. sIgE sensitivity against Ara h 9 in ISAC needs to be improved.

Is the ISAC 112 Microarray Useful in the Diagnosis of Pollinosis in Spain?

BACKGROUND: Multiple sensitization is frequent among pollen-allergic patients. The goal of this study was to determine the diagnostic accuracy of the ImmunoCAP ISAC 112 (ISAC112) microarray in allergy to pollen from several taxa and its clinical utility in a Spanish population. METHODS: Specific IgE was determined in 390 pollen-allergic patients using the ISAC112 microarray. Diagnostic accuracy (sensitivity, specificity, predictive values, and area under the ROC curve) was calculated for the diagnosis of allergy to pollen from grass (n=49), cypress (n=75), olive tree (n=33), plane tree (n=63), and pellitory of the wall (n=17) and compared with that of the singleplex ImmunoCAP immunoassay. RESULTS: The sensitivity of the ISAC112 microarray ranged from 68.2% for allergy to plane tree pollen to 93.9% for allergy to grass pollen. The specificity was >90%. The AUC for the diagnosis of allergy to plane tree pollen was 0.798, whereas the AUC for the remaining cases was ≥0.876. The accuracy of ISAC112 was higher than that of ImmunoCAP for plane tree pollen and similar for the remaining pollens. The frequency of sensitization to most species-specific allergenic components and profilins varied between the different geographical regions studied. A total of 73% of pollen-allergic patients were sensitized to species-specific components of more than 1 pollen type. CONCLUSIONS: The ISAC112 microarray is an accurate tool for the diagnosis of allergy to pollen from grass, cypress, olive tree, plane tree, and pellitory of the wall. The features of the ISAC112 microarray are similar or superior (in the case of plane tree pollen) to those of ImmunoCAP. This microarray is particularly useful for the etiologic diagnosis of pollinosis in patients sensitized to multiple pollen species whose pollination periods overlap.

Do Skin Prick Test and In Vitro Techniques Diagnose Sensitization to Peach Lipid Transfer Protein and Profilin Equally Well in Allergy to Plant Food and Pollen?

OBJECTIVE: To compare the skin prick test (SPT) with in vitro techniques (single and multiplex fluorescence enzyme-immunoassay [FEIA]) for detecting sensitization to profilin and lipid transfer protein (LTP). METHODS: We retrospectively studied 181 patients with pollen and/or plant food allergy and 61 controls. SPT was performed with date palm profilin (Pho d 2) and peach LTP (Pru p 3), and specific IgE (sIgE) to Phl p 12 and Pru p 3 was analyzed using single FEIA and microarray. RESULTS: Fifteen of 201 patients with negative results for LTP in the SPT were sensitized to this allergen in the in vitro tests, and 18 of 41 patients with positive results for LTP in the SPT were not sensitized according to the in vitro tests. Seventeen of 186 patients with negative results for profilin in the SPT were sensitized to Phl p 12 by serum sIgE, and 30 out of 56 patients with positive results for profilin in SPT were not sensitized to Phl p 12 according to the other tests. Moderate agreement was observed between the 3 techniques studied. CONCLUSIONS: SPT is a sensitive technique for detecting sensitization to LTP and profilin. Its results are similar to those of in vitro techniques, especially in patients with negative SPT results for peach LTP and palm tree profilin.

The usefulness of plasma histamine and different tryptase cut-off points in the diagnosis of peranaesthetic hypersensitivity reactions.

UNLABELLED: Anaesthetic hypersensitivity reactions can be IgE- or not IgE-mediated and are a challenge to find the causal agent. Histamine and tryptase determination are classically considered useful in the diagnosis of these reactions. The aim of our study was to assess the diagnostic usefulness of plasma histamine and different cut-off points of serum tryptase. METHODS: Patients suffering a reaction suggestive of hypersensitivity during general anaesthesia in Clínica Universidad de Navarra (2008-2012) were included. Serum tryptase and plasma histamine were measured at the time of the reaction and 2 h later. Baseline tryptase was also determined. Four to eight weeks after the reaction an allergological study was performed to all the drugs or products involved in the reaction. RESULTS: Sixty-five patients suffered an immediate hypersensitivity reaction during the period of the study. Thirty-seven patients (20 male) with median age 48 years (12-79) were included because they completed allergological study, and histamine and tryptase were correctly obtained. Elevated plasma histamine was observed in 34 cases (92%). Tryptase exceeded twice the basal values in 10 patients (31%). Using different cut-off points of tryptase, the number of patients with elevated tryptase would be 15 patients (41%) for a cut-off point of 5 µg/L; 12 patients (32%) for a cut-off point of 8.23 µg/L; nine patients (24%) for 10.5 µg/L; and eight patients (22%) for 11.4 µg/L. The median tryptase level for the IgE-mediated reactions was 9.0 µg/L (2-70 µg/L) and 4.0 µg/L (3-13 µg/L) in non-IgE-mediated reactions (P < 0.01). Median tryptase levels were higher in more severe reactions (grade 2 or 3) in comparison with grade 1. The best ratio for serum-tryptase-during-reaction/basal-serum-tryptase to discriminate between IgE and non-IgE reactions was 2.0. CONCLUSION: The best criterion for discriminating IgE- and non IgE-mediated hypersensitivity reactions in anaesthesia was a tryptase value exceeding twice the basal one.

In vivo and in vitro testing with rAni s 1 can facilitate diagnosis of Anisakis simplex allergy.

BACKGROUND: Traditional diagnostic tests such as skin prick tests (SPT) and specific IgE (slgE) against whole Anisakis simplex extract have low specificity. Consequently, allergy to A simplex is overdiagnosed. OBJECTIVE: Our aim was to compare tests used in component-resolved diagnosis. METHODS: We evaluated 34 patients with allergy to A simplex, 15 patients with acute urticaria who were sensitized to A simplex but had no clinical history of allergy to A simplex, and 10 patients allergic to seafood. SPT, slgE (ELISA and ISAC-I 12), and the basophil activation test (BAT) were performed with A simplex whole extract and the molecular components rAni s 1, rAni s 3, and nPen m 1. Sensitivity and specificity were calculated and compared with different cutoffs. RESULTS: With the A simplex whole extract, SPT, slgE, and BAT yielded specificity values of 72%, 68%, and 70%, respectively, with a cutoff (wheal size) of 11.2 mm, an slgE value of 7.9 kUAIL, and a stimulation index of 1.9. Specificity increased to 100% using the molecular component rAni s 1 with SPT, slgE by ELISA, and ISAC-112. Neither rAni s 3 sensitization nor cross-reactivity with Pen m 1 was observed in patients sensitized to A simplex. CONCLUSION: rAni s 1 is recognized by 100% of our patients and is able to distinguish between patients allergic to A simplex and patients with acute urticaria who are sensitized to A simplex but have no clinical history of allergy to this parasite.

Early skin testing is effective for diagnosis of hypersensitivity reactions occurring during anesthesia.

Allergic skin tests have to be performed 4-6 weeks after an allergic anesthetic reaction. Patients with allergic reactions during anesthesia were prospectively included (n = 44). Skin tests were performed in two stages: (i) Stage 1 (S1), 0-4 days after the reaction; and (ii) Stage 2 (S2), 4-8 weeks after. Five (11.5%) surgical procedures were suspended due to the reaction. Positive skin tests were obtained in 25/44 patients (57%). Allergic diagnosis was carried out at S1 in 15/25 (60%) and at S2 in 10/25 (40%). Three patients resulted positive only in S1. Overall agreement among S1 and S2 skin tests was 70.45%. The kappa statistic was 0.41 (P-value = 0.002). Odds ratio of obtaining a false negative in S1 (compared with S2) was 3.33. Early allergological study is useful, could minimize false negatives, but should be considered as a complement to late skin tests.

Recommendations for the use of in vitro methods to detect specific immunoglobulin E: are they comparable?

Total and specific immunoglobulin (Ig) E can be detected in vitro using several commercially available methods. The largest share of the global market for these methods is held by the ImmunoCAP technique (Thermo Fisher, previously Phadia), Immulite (Siemens), and Hytec-288 (Hycor). Most comparative studies examine Immulite and ImmunoCAP, which differ methodologically but use similar units of measurement relative to the same standard of total IgE (WHO IgE Standard 75/502). Despite their similarity, these kits differ in their quantification of specific IgE, which varies depending on the allergen studied.Thus, specific IgE results obtained with ImmunoCAP and Immulite are not interchangeable. It is important to bear this in mind, especially when determining cutoff points as predictors of a response to oral challenge with specific food allergens. The method used in practice must be the same as the one in the publication guiding clinical decision making. We analyze differences between ImmunoCAP and ISAC microarray, 2 methods from the same manufacturer used to detect IgE to specific proteins (purified or recombinant).The results show that the IgE values obtained with ImmunoCAP are not equivalent to the corresponding values obtained with the ISAC microarray system.

Local allergic rhinitis in children: Clinical characteristics and role of basophil activation test as a diagnostic tool.

BACKGROUND: Local allergic rhinitis (LAR) is a condition involving a localized nasal allergic response in absence of systemic atopy. Most studies on LAR have been performed in adults. We aimed to describe clinical characteristics of LAR pediatric patients, its clinical evolution over a 7-year follow-up period and to study the role of basophil activation test (BAT), for its diagnosis. METHODS: Forty-four children with non-allergic rhinitis (NAR) were included (24 males, 20 females, aged under 15 years). Nasal allergen provocation test (NAPT) and BAT were performed with Dermatophagoides pteronyssinus and Phleum pratense. RESULTS: Seven patients (16%) were diagnosed of LAR. Six reacted to D pteronyssinus and one to P pratense. All LAR and 86% of NAR patients presented perennial symptoms. Fifty-seven percent of NAR and LAR patients referred persistent symptoms. Around half of NAR and LAR patients reported mild-moderate clinical manifestations. Three LAR patients associated conjunctival symptoms, proportionally more than NAR patients (19%, 7 out of 37). NAR patients presented bronchial asthma (n = 10) more frequently than LAR children (n = 1). More than half of LAR and NAR patients presented family history of atopy. BAT was negative in all LAR patients. On follow-up, 3 LAR patients and 10 of the 25 NAR patients who agreed to be retested, presented systemic sensitization. Dust mites were the most frequent allergen involved. CONCLUSIONS: LAR should be ruled out in children with NAR. Almost half of children with LAR develop systemic sensitization over time. BAT shows low sensitivity for the diagnosis of LAR in children.

Performance of different in vitro techniques in the molecular diagnosis of peanut allergy.

INTRODUCTION: Peanut allergy is an increasingly serious disorder with a heterogeneous pattern of sensitization across different countries. In vitro diagnostic techniques may help in establishing these patterns. OBJECTIVES: To analyze the usefulness of determining specific immunoglobulin E (sIgE) with the ImmunoCAP fluorescence enzyme immunoassay (FEIA), the ImmunoCAP ISAC CRD103 microarray (ISAC), and the basophil activation test (BAT) in the molecular diagnosis of peanut allergy. METHODS: In 26 peanut-allergic patients, sIgE antibodies against allergic components were measured with FEIA, ISAC, and BAT. RESULTS: The major peanut component in our population wasAra h 9.The detection of sIgE toAra h 9 using FEIA and BAT with this allergen yielded a sensitivity of 92% and 88% and a specificity of 95% and 100%, respectively. Overall diagnosis of peanut allergy by ISAC showed a sensitivity of 11% but a specificity of 95% since Ara h 9 was not present in the microarray version used. There was diagnostic agreement between the 3 techniques for the peanut allergens studied. CONCLUSIONS: The determination of sIgE to Ara h 9 using FEIA and BAT offers high sensitivity and specificity in the diagnosis of peanut allergy in the Spanish population. The CRD103 version of ISAC is not of value in our region as it does not include the most common allergen, Ara h 9.

Differences in Linear Epitopes of Ara h 9 Recognition in Peanut Allergic and Tolerant, Peach Allergic Patients.

Background: Peanut-allergic patients from the Mediterranean region are predominantly sensitized to the lipid transfer protein (LTP) Ara h 9, and the peach LTP Pru p 3 seems to be the primary sensitizer. However, LTP sensitization in peanut allergy is not a predictive marker for clinically relevant symptoms. Objective: We aimed to identify sequential epitopes of IgE and IgG4 from Pru p 3 and Ara h 9 in peach-allergic patients sensitized to peanuts. We also sought to determine the differences in IgE and IgG4 binding between patients who had developed peanut allergy and those tolerating peanuts. Methods: A total of 46 peach-allergic patients sensitized to peanuts were selected. A total of 35 patients were allergic to peanuts (peanut-allergic group) and 11 were tolerant to peanuts (peanut-tolerant group). We measured sIgE and sIgG4 in peanut, peach, and their recombinant allergen (Ara h 1, Ara h 2, Ara h 3, Ara h 8, and Ara h 9) with fluorescence enzyme immunoassay. We examined the IgE and IgG4 binding to sequential epitopes using a peptide microarray corresponding to linear sequences of the LTPs Ara h 9 and Pru p 3 with a library of overlapping peptides with a length of 20 amino acids (aa) and an offset of 3 aa. Results: The frequency and the intensity of IgE recognition of Ara h 9 and Pru p 3 peptides were higher in the peanut-tolerant group than in the peanut-allergic group. We found four Ara h 9 peptides (p4, p14, p21, and p25) and four Pru p 3 peptides (p1, p3, p21, and p24) with a significantly elevated IgE recognition in peanut-tolerant patients. Only one peptide of Ara h 9 (p4) recognized by IgG4 was significantly elevated in the peanut-tolerant group. The IgG4/IgE ratio of Ara h 9 peptide 4 was significantly higher in peanut-tolerant patients than in peanut-allergic patients, while no significant differences were observed in the IgG4/IgE ratio of this peptide in Pru p 3. Conclusion: Although we found significant differences in IgE and IgG4 recognition of Ara h 9 and Pru p 3 between peanut-tolerant and peanut-allergic patients (all of whom were allergic to peach), polyclonal IgE peptide recognition of both LTPs was observed in peach-allergic patients tolerating peanuts. However, the IgG4 blocking antibodies against Ara h 9 peptide 4 could provide an explanation for the absence of clinical reactivity in peanut-tolerant peach-allergic patients. Further studies are needed to validate the usefulness of IgG4 antibodies against Ara h 9 peptide 4 for peanut allergy diagnosis.

Can component-based microarray replace fluorescent enzimoimmunoassay in the diagnosis of grass and cypress pollen allergy?

BACKGROUND: Few data on the diagnostic accuracy in pollinosis of the microarray ISAC of allergens are available. OBJECTIVE: We aim to comparatively analyse ISAC CRD103 with the whole-extract ImmunoCAP in grass and cypress pollen allergy, evaluating the suitability of the manufacturer's recommended cut-off points for both techniques. METHODS: We studied 120 atopic patients grouped into grass and cypress pollen-allergic patients and controls based on clinical history and skin prick tests. Specific IgE against Phleum pratense and Cupressus arizonica by ImmunoCAP and ISAC CRD103 were performed on all subjects. RESULTS: In the grass pollen group (43 allergic/26 controls), both microarray and CAP showed high sensitivity (Se) and specificity (Sp) values (ISAC: Se 97.7, Sp 92.3; CAP: Se 95.3, Sp 96.1) for recommended cut-off points. Comparing the optimal (ISAC: 0.4 ISU; CAP: 0.33 kU/L) with the recommended cut-off points within the same technique, diagnostic agreement was observed in both techniques. Thus, CAP and ISAC showed similar diagnostic performance in grass pollen allergy when using recommended cut-off points. In cypress pollen group (12 allergic/92 controls), the microarray (Se: 91.7, Sp 91.3) showed similar Se but significantly higher Sp (P=0.034) than CAP (Se: 91.7, Sp: 80.4) using recommended cut-off points. However, although diagnostic performance of the microarray did not change when comparing the optimal (0.82 ISU) with the recommended cut-off point, CAP improved diagnosis of cypress pollen allergy, when applying the optimal (0.66 kU/L)(CAP Se: 91.7, Sp: 89.1) instead of the manufacturer's recommended cut-off point. Thus, when the most suitable cut-off point for both techniques (ISAC: 0.3 ISU; CAP: 0.66 kU/L) is selected, microarray and CAP provide equivalent diagnoses. CONCLUSIONS AND CLINICAL RELEVANCE: Component-based microarray ISAC CRD103 and whole-allergen CAP showed high Se and Sp diagnosing equally grass and cypress pollen allergy. The cut-off point for each allergen should be properly applied for both techniques.

A new rapid desensitization protocol for chemotherapy agents.

BACKGROUND: Desensitization has been used for some decades to treat patients with the allergenic drug when an alternative drug with similar efficacy and safety is not available. We present the results from a series of oncology patients desensitized at our hospital during the last 2 years. OBJECTIVE: To assess the efficacy of a new desensitization protocol in patients allergic to chemotherapy drugs. METHODS: We performed an observational retrospective study of 11 women (6 breast cancer and 5 ovarian cancer) who underwent our desensitization protocol. Four patients had immediate reactions to carboplatin, 3 to docetaxel, 3 to paclitaxel, and 1 to both docetaxel and paclitaxel. Premedication was administered in all cases. A 5-step protocol based on 5 different dilutions of the drugs was used. RESULTS: We performed 39 desensitization procedures: 14 to carboplatin, 3 to oxaliplatin, 16 to docetaxel, and 6 to paclitaxel. Eight patients tolerated the full dose in 36 procedures. One patient suffered an anaphylactic reaction to carboplatin that reverted with treatment. One patient had dyspnea after a paclitaxel cycle. One patient experienced dyspnea due to chronic pulmonary thromboembolism related to her disease. CONCLUSION: Desensitization is a useful procedure in patients who are allergic to their chemotherapy agents.