IgE-Mediated Cannabis Allergy and Cross-Reactivity Syndromes: A Roadmap for Correct Diagnosis and Management.

PURPOSE OF THE REVIEW: With increased access and decriminalization of cannabis use, cases of IgE-dependent cannabis allergy (CA) and cross-reactivity syndromes have been increasingly reported. However, the exact prevalence of cannabis allergy and associated cross-reactive food syndromes (CAFS) remains unknown and is likely to be underestimated due to a lack of awareness and insufficient knowledge of the subject among health care professionals. Therefore, this practical roadmap aims to familiarize the reader with the early recognition and correct management of IgE-dependent cannabis-related allergies. In order to understand the mechanisms underlying these cross-reactivity syndromes and to enable personalized diagnosis and management, special attention is given to the molecular diagnosis of cannabis-related allergies. RECENT FINDINGS: The predominant signs and symptoms of CA are rhinoconjunctivitis and contact urticaria/angioedema. However, CA can also present as a life-threatening condition. In addition, many patients with CA also have distinct cross-reactivity syndromes, mainly involving fruits, vegetables, nuts and cereals. At present, five allergenic components of Cannabis sativa (Can s); Can s 2 (profilin), Can s 3 (a non-specific lipid protein), Can s 4 (oxygen-evolving enhancer protein 2 oxygen), Can s 5 (the Bet v 1 homologue) and Can s 7 (thaumatin-like protein) have been characterized and indexed in the WHO International Union of Immunological Sciences (IUIS) allergen database. However, neither of them is currently readily available for diagnosis, which generally starts by testing crude extracts of native allergens. The road to a clear understanding of CA and the associated cross-reactive food syndromes (CAFS) is still long and winding, but well worth further exploration.

Mast cell versus basophil activation test in allergy: Current status.

In the past two decades, we witnessed the evolution of the basophil activation test (BAT) from mainly research applications to a potential complementary diagnostic tool to document IgE-dependent allergies. However, BAT presents some technical weaknesses. Around 10%-15% of tested patients are non-responders, BAT can be negative immediately post-reaction and the use of fresh basophils, ideally analysed within 4 h of collection, restricts the number of tests that can be performed per sample. The need for fresh basophils is especially limiting when conducting batch analyses and interlaboratory comparisons to harmonize BAT methodology. These limitations significantly hinder the wider application of BAT and urge the development of alternative testing, such as the mast cell activation test (MAT). The essential difference between BAT and MAT is the heterogeneity of the starting material used to perform the assays. Mast cells are tissue-resident, so cannot be easily accessed. Current alternative sources for functional studies are generating primary human mast cells, differentiated from donor progenitor cells, or using immortalized mast cell lines. Hence, the methodological approaches for MAT are not only vastly different from BAT, but also different among MAT protocols. This review summarizes the advantages and disadvantages of BAT and MAT assays, dedicating special attention to elucidating the key differences between the cellular sources used and provides an overview of studies hitherto performed comparing BAT and MAT in the diagnosis of IgE-mediated food and drug allergies.

Comparison of the passive mast cell activation test with the basophil activation test for diagnosis of perioperative rocuronium hypersensitivity.

BACKGROUND: Rocuronium is a major cause of perioperative hypersensitivity (POH). Skin tests (STs) and quantification of specific immunoglobulin E antibodies (sIgEs) can yield incongruent results. In such difficult cases, the basophil activation test (BAT) can be helpful. Here, we evaluated the passive mast cell activation test (pMAT) as a substitute of BAT as part of the diagnostic tests for rocuronium allergy. METHODS: Sera from patients with a suspected POH reaction potentially related to rocuronium were included. All patients had a complete diagnostic investigation, including STs, quantification of sIgEs to morphine and rocuronium, and BAT. For execution of pMAT, human mast cells were generated from healthy donor peripheral blood CD34+ progenitor cells and sensitised overnight with patient sera. RESULTS: In total, 90 sera were studied: 41 from ST+sIgE+ patients, 13 from ST-sIgE- patients, 20 from ST+sIgE- patients, and 16 from ST-sIgE+ patients. According to BAT results, patients were further allocated into subgroups. Of the 38 BAT+ patients, 25 (66%) showed a positive pMAT as well. Of the 44 BAT- patients, 43 (98%) also showed a negative pMAT. Mast cells that were not passively sensitised did not respond to rocuronium. CONCLUSIONS: We show that the pMAT, in many cases, can substitute for BAT in the diagnosis of rocuronium hypersensitivity and advance diagnosis in difficult cases with uncertain ST or sIgE results when BAT is not locally available.

A quarter of a century fundamental and translational research in perioperative hypersensitivity and anaphylaxis at the Antwerp university hospital, a Belgian Centre of Excellence of the World Allergy Organization.

Perioperative hypersensitivity constitutes an important health issue, with potential dramatic consequences of diagnostic mistakes. However, safe and correct diagnosis is not always straightforward, mainly because of the application of incorrect nomenclature, absence of easy accessible in-vitro/ex-vivo tests and uncertainties associated with the non-irritating skin test concentrations. In this editorial we summarize the time line, seminal findings, and major realizations of 25 years of research on the mechanisms, diagnosis, and management of perioperative hypersensitivity.

Allergenic and Mas-Related G Protein-Coupled Receptor X2-Activating Properties of Drugs: Resolving the Two.

Since the seminal description implicating occupation of the Mas-related G protein-coupled receptor X2 (MRGPRX2) in mast cell (MC) degranulation by drugs, many investigations have been undertaken into this potential new endotype of immediate drug hypersensitivity reaction. However, current evidence for this mechanism predominantly comes from (mutant) animal models or in vitro studies, and irrefutable clinical evidence in humans is still missing. Moreover, translation of these preclinical findings into clinical relevance in humans is difficult and should be critically interpreted. Starting from our clinical priorities and experience with flow-assisted functional analyses of basophils and cultured human MCs, the objectives of this rostrum are to identify some of these difficulties, emphasize the obstacles that might hamper translation from preclinical observations into the clinics, and highlight differences between IgE- and MRPGRX2-mediated reactions. Inevitably, as with any subject still beset by many questions, alternative interpretations, hypotheses, or explanations expressed here may not find universal acceptance. Nevertheless, we believe that for the time being, many questions remain unanswered. Finally, a theoretical mechanistic algorithm is proposed that might advance discrimination between MC degranulation from MRGPRX2 activation and cross-linking of membrane-bound drug-reactive IgE antibodies.

Basophil Activation Test Shows Poor Sensitivity in Immediate Amoxicillin Allergy.

BACKGROUND: In light of the pandemic of spurious penicillin allergy, correct diagnosis of amoxicillin (AX) allergy is of great importance. The diagnosis of immediate hypersensitivity reactions relies on skin tests and specific IgE, and although reliable, these are not absolutely predictive. Therefore, drug challenges are needed in some cases, which contain the risk of severe reactions. Safe in vitro diagnostics as an alternative for the drug challenge in the diagnostic workup of AX allergy would be more than welcome to fill this gap. In this respect, the basophil activation test (BAT) has shown potential, but its clinical reliability is doubtful. OBJECTIVE: To investigate the reliability of the BAT to AX and determining its exact place in the diagnostic algorithm of AX allergy. METHODS: BAT for AX was performed in 70 exposed control individuals and 66 patients diagnosed according to the European Academy of Allergy and Clinical Immunology guidelines for AX allergy. Upregulation of both CD63 and CD203c was flow-cytometrically assessed. RESULTS: Analyses revealed that 1370 µmol/L and 685 µmol/L were the most discriminative stimulation concentrations for CD63 and CD203c upregulation, respectively, and a diagnostic threshold of 9% for positivity for both markers was identified. At these concentrations, sensitivity and specificity for CD63 upregulation were 13% and 100%, respectively, and for CD203c upregulation, 23% and 98%. CONCLUSIONS: BAT with dual analysis of CD63 and CD203c is of poor performance to document AX allergy. The sensitivity is too low to let it occupy a prominent role in the diagnostic algorithm.

Recent Knowledge and Insights on the Mechanisms of Immediate Hypersensitivity and Anaphylaxis: IgE/FcεRI- and Non-IgE/FcεRI-Dependent Anaphylaxis.

Immediate hypersensitivity reactions can pose a clinical and diagnostic challenge, mainly because of the multifarious clinical presentation and distinct underlying - frequently uncertain - mechanisms. Anaphylaxis encompasses all rapidly developing and life-threatening signs and may cause death. Evidence has accumulated that immediate hypersensitivity and anaphylaxis do not necessarily involve an allergen-specific immune response with cross-linking of specific IgE (sIgE) antibodies bound to their high-affinity IgE receptor (FcεRI) on the surface of mast cells (MCs) and basophils. Immediate hypersensitivity and anaphylaxis can also result from alternative specific and nonspecific MC and basophils activation and degranulation, such as complementderived anaphylatoxins and off-target occupancy of MC and/or basophil surface receptors such as the Masrelated G protein-coupled receptor X2 (MRGPRX2). Degranulation of MCs and basophils results in the release of inflammatory mediators, which can be, depending on the underlying trigger, in a different spatiotemporal manner. In addition, hypersensitivity and anaphylaxis can occur entirely independently of MC and basophil degranulation, as observed in hypersensitivity to nonsteroidal anti-inflammatory drugs (NSAIDs) that divert normal arachidonic acid metabolism by inhibiting the cyclooxygenase (COX)-1 isoenzyme. Finally, one should remember that anaphylaxis might be part of the phenotype of particular - sometimes poorly recognizable - conditions such as clonal MC diseases (e.g. mastocytosis) and MC activation syndrome. This review provides a status update on the molecular mechanisms involved in both sIgE/FcεRI- and non-sIgE/FcεRI-dependent immediate hypersensitivity and anaphylaxis. In conclusion, there is increasing evidence for alternative pathophysiological hypersensitivity and anaphylaxis endotypes that are phenotypically and biologically indistinguishable, which are frequently difficult to diagnose, mainly because of uncertainties associated with diagnostic tests that might not enable to unveil the underlying mechanism.

Establishing diagnostic strategies for cannabis allergy.

INTRODUCTION: Cannabis is the most widely consumed illicit drug in the world and carries a risk of severe IgE-mediated allergic reactions, requiring appropriate diagnostic management. Currently available diagnostics are still relatively limited and require careful interpretation of results to avoid harmful over- and underdiagnosis. AREAS COVERED: This review focuses on the most up-to-date understandings of cannabis allergy diagnosis, starting with the main clinical features of the disease and the allergenic characteristics of Cannabis sativa, and then providing insights into in vivo, in vitro, and ex vivo diagnostic tests. EXPERT OPINION: At present, the diagnosis of IgE-mediated cannabis allergy is based on a three-step approach that starts with accurate history taking and ends with a confirmation of sensitization to the whole extract and, finally, molecular components. Although much has been discovered since its first description in 1971, the diagnosis of cannabis allergy still has many unmet needs. The lack of commercial standardized and validated extracts and in vitro assays makes a harmonized workup of cannabis allergy difficult. Furthermore, the epidemiological characteristics, and clinical implications of sensitization to different molecular components are not yet fully known. Future research will complete the picture and likely result in an individualized and standardized approach.

Flow-based allergen testing: Can mast cells beat basophils?

The basophil activation test (BAT) has emerged as a reliable complementary diagnostic to document IgE-dependent allergies and to study cross-reactivity between structural homologues. However, the BAT has some weaknesses that hinder a wider application. The BAT requires fresh blood samples and is lost as a diagnostic in patients showing a non-responder status of their cells. The BAT is difficult to standardize mainly because of the difficulty to perform batch analyses. In contrast, mast cell activation tests (MATs), using passively sensitized mast cells (MCs) with patients' sera (henceforth indicated as passive MAT; pMAT), use serum samples that can be frozen, stored, and shipped to a reference center experienced in MC lines and/or cultures and capable of offering batch testing. With the recent recognition of the Mas-related G protein-coupled receptor X2 (MRGPRX2) occupation as a putative mechanism of immediate drug hypersensitivity reactions, the MAT has another advantage compared to the BAT. MCs, in contrast to resting basophils, express the MRGPRX2 and can therefore be used to study this IgE-independent mechanism. This review provides a status update of pMAT in the diagnosis of allergic IgE-mediated hypersensitivity and speculates how direct activation of MCs via the MRGPRX2 receptor could advance paradigms for this non-allergic hypersensitivity.

Overexpression of FcεRI on Bone Marrow Mast Cells, but Not MRGPRX2, in Clonal Mast Cell Disorders With Wasp Venom Anaphylaxis.

Background: Uncertainties remain about the molecular mechanisms governing clonal mast cell disorders (CMCD) and anaphylaxis. Objective: This study aims at comparing the burden, phenotype and behavior of mast cells (MCs) and basophils in patients with CMCD with wasp venom anaphylaxis (CMCD/WVA+), CMCD patients without anaphylaxis (CMCD/ANA-), patients with an elevated baseline serum tryptase (EBST), patients with wasp venom anaphylaxis without CMCD (WVA+) and patients with a non-mast cell haematological pathology (NMHP). Methods: This study included 20 patients with CMCD/WVA+, 24 with CMCD/ANA-, 19 with WVA+, 6 with EBST and 5 with NMHP. We immunophenotyped MCs and basophils and compared baseline serum tryptase (bST) and both total and venom specific IgE in the different groups. For basophil studies, 13 healthy controls were also included. Results: Higher levels of bST were found in CMCD patients with wasp venom anaphylaxis, CMCD patients without anaphylaxis and EBST patients. Total IgE levels were highest in patients with wasp venom anaphylaxis with and without CMCD. Bone marrow MCs of patients with CMCD showed lower CD117 expression and higher expression of CD45, CD203c, CD63, CD300a and FcεRI. Within the CMCD population, patients with wasp venom anaphylaxis showed a higher expression of FcεRI as compared to patients without anaphylaxis. Expression of MRGPRX2 on MCs did not differ between the study populations. Basophils are phenotypically and functionally comparable between the different patient populations. Conclusion: Patients with CMCD show an elevated burden of aberrant activated MCs with a significant overexpression of FcεRI in patients with a wasp venom anaphylaxis.

Mast cell activation test: A new asset in the investigation of the chlorhexidine cross-sensitization profile.

BACKGROUND: Insights into the IgE cross-sensitization and possible cross-reactivity patterns of sera reactive to chlorhexidine (CHX) are still incomplete and are likely to benefit from a functional exploration using a passive mast cell activation test (pMAT). Therefore, we want to study whether the pMAT with CHX-specific IgE (sIgE) enables to depict effector cell degranulation in response to alexidine (ALX), octenidine (OCT) and/or polyhexamethylene biguanide (PHMB) indicative of cross-reactivity between these compounds and CHX. METHODS: Serum of 10 CHX-allergic patients, nine individuals with an isolated sIgE CHX and five healthy controls were included. Human cultured mast cells (MCs) were, before and after sensitization, challenged with CHX, ALX, OCT or PHMB. Degranulation was measured via quantification of upregulation of CD63. RESULTS: Mast cell responsiveness to ALX and OCT was demonstrable with 4/10 and 3/10 of the sera of CHX-allergic patients respectively. Percentage of degranulation varied between 12 and 34% for ALX-reactive MCs and between 4 and 22% for OCT-reactive MCs. No reactivity to ALX or OCT was demonstrable when using sera obtained from individuals with an isolated sIgE CHX or from healthy controls. Unlike CHX, ALX and OCT, PHMB turned out to be a direct MC activator via occupation of MRGPRX2. PHMB-reactive sIgEs were demonstrable in some patients with an isolated sIgE CHX but were unable to trigger PHMB-induced degranulation in MRGPRX2 knockdown MCs. CONCLUSION: Mast cells constitute an attractive tool to explore cross-reactivity between structurally similar compounds. Along with the identification of safe alternatives for the individual patient, the pMAT can advance our insights into sIgE cross-reactivity patterns including assessment of molecules not yet approved for human use.