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.

Paediatric perioperative hypersensitivity: the performance of the current consensus formula and the effect of uneventful anaesthesia on serum tryptase.

Background: Paired sampling of acute (aST) and basal (bST) serum tryptase has been recommended when investigating patients with a suspected perioperative hypersensitivity (POH) reaction. In the current consensus formula, an aST value exceeding (1.2×bST+2) confirms mast cell activation. The current consensus formula has been validated in adults but not in children. Methods: We prospectively included 96 children who underwent uneventful anaesthesia and sampled serum tryptase at baseline and 60-90 min after induction. Tryptase changes were then compared with those in 94 children with suspected POH who were retrospectively included from four reference centres in Belgium, France, and Denmark. Results: We observed a median decrease in serum tryptase during uneventful anaesthesia of 0.41 µg L-1 (-15.9%; P<0.001). The current consensus formula identified mast cell activation in 31.9% of paediatric POH patients. After generating receiver operating characteristic curves through 100 repeated five-fold cross-validation, aST>bST+0.71 was identified as the optimal cut-off point to identify mast cell activation. This new paediatric formula has higher sensitivity than the current consensus formula (53.2% vs 31.9%, P<0.001) with a specificity of 96.9%. Analysis in the subpopulation where a culprit was identified and in grade 3-4 reactions similarly yielded higher sensitivity for the new paediatric formula when compared with the current consensus formula (85.3% vs 61.8%; P=0.008 and 78.0% vs 48.8%; P<0.001, respectively). Internally cross-validated sensitivity and specificity were 53.3% and 93.3%, respectively. Conclusions: This is the first study suggesting the need for an adjusted formula in children to identify perioperative mast cell activation as tryptase is significantly lowered during uneventful anaesthesia. We propose a new formula (aST>bST+0.71) which performs significantly better than the current consensus formula in our multicentric paediatric population.

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.

Mastocytosis and related entities: a practical roadmap.

Mastocytosis is a complex heterogenous multisystem disorder that is characterized by pathologic activation or accumulation of neoplastic mast cells (MCs) in one or more organs. This clonal MC expansion is often associated with a somatic gain-of-function mutation (D816V in most of the cases) in the KIT gene, encoding for the MC surface receptor KIT (CD117), a stem cell growth factor receptor. Based on clinical and biochemical criteria, the World Health Organization (WHO) divided mastocytosis into different subclasses. The exact prevalence of mastocytosis remains elusive, but it is estimated that the disease affects approximately 1 in 10,000 persons. The clinical presentation of mastocytosis varies significantly, ranging from asymptomatic patients to a life-threatening disease with multiple organ involvement, potentially leading to cytopenia, malabsorption, hepatosplenomegaly, lymphadenopathy, ascites or osteolytic bone lesions with pathological fractures. Patients with mastocytosis may experience symptoms related to release of MC mediators, such as flushing or diarrhea or even more severe symptoms such as anaphylaxis. Recently, a new genetic trait, hereditary alpha tryptasemia (HaT), was described which involves a copy number variation in the TPSAB1-gene. Its role as standalone multisystem syndrome is heavily debated. There is emerging evidence suggesting there might be a link between HaT and due to the increased prevalence of HaT in patients with SM. The aim of this review is to provide a practical roadmap for diagnosis and management of mastocytosis and its associated entities, since there are still many misconceptions about these topics.Abbreviations: AdvSM: Advanced systemic mastocytosis; ASM: Aggressive systemic mastocytosis; aST: acute serum tryptase; BM: Bone marrow; BMM: Bone marrow mastocytosis; bST: baseline serum tryptase; CM: Cutaneous mastocytosis; DCM: Diffuse cutaneous mastocytosis; HVA: Hymenoptera venom allergy; HaT: Hereditary alpha tryptasemia; ISM: Indolent systemic mastocytosis; MC: Mast cell; MCA: Mast cell activation; MCAS: Mast cell activation syndrome; MCL: Mast cell leukemia; MIS: Mastocytosis in the skin; MMAS: Monoclonal mast cell activation syndrome; MPCM: Maculopapular cutaneous mastocytosis; SM: Systemic mastocytosis; SM-AHN: Systemic mastocytosis with associated hematological neoplasm; SSM: Smouldering systemic mastocytosis; VIT: Venom immunotherapy.

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.

Long-term hematopoietic stem cells as a parasite niche during treatment failure in visceral leishmaniasis.

Given the discontinuation of various first-line drugs for visceral leishmaniasis (VL), large-scale in vivo drug screening, establishment of a relapse model in rodents, immunophenotyping, and transcriptomics were combined to study persistent infections and therapeutic failure. Double bioluminescent/fluorescent Leishmania infantum and L. donovani reporter lines enabled the identification of long-term hematopoietic stem cells (LT-HSC) as a niche in the bone marrow with remarkably high parasite burdens, a feature confirmed for human hematopoietic stem cells (hHSPC). LT-HSC are more tolerant to antileishmanial drug action and serve as source of relapse. A unique transcriptional 'StemLeish' signature in these cells was defined by upregulated TNF/NF-κB and RGS1/TGF-ß/SMAD/SKIL signaling, and a downregulated oxidative burst. Cross-species analyses demonstrated significant overlap with human VL and HIV co-infected blood transcriptomes. In summary, the identification of LT-HSC as a drug- and oxidative stress-resistant niche, undergoing a conserved transcriptional reprogramming underlying Leishmania persistence and treatment failure, may open therapeutic avenues for leishmaniasis.