Sequential class switch recombination to IgE and allergen-induced accumulation of IgE+ plasmablasts occur in the nasal mucosa of local allergic rhinitis patients.

BACKGROUND: The involvement of allergen-specific (s)IgE in local allergic rhinitis (LAR) has been debated. Here, we investigate the effect of nasal allergen challenge with Dermatophagoides pteronyssinus (NAC-DP) in mucosal and peripheral B-cell subpopulations in LAR patients. METHODS: Nine LAR, 5 allergic rhinitis (AR), and 5 non-atopic healthy control (HC) individuals were subjected to a 3-day NAC-DP protocol, and nasal biopsies and blood samples were collected before and after provocation. Nasal biopsies were used for immunohistochemistry and gene expression studies, whereas the frequency of lymphocyte subsets and basophil activation test (BAT) were analyzed in blood samples by flow cytometry. sIgG was measured in sera. RESULTS: NAC-DP induced an increase in IgE+ CD38+ plasmablasts in the nasal mucosa of LAR patients, but not in AR or HC individuals. Markers of sequential recombination to IgE (εCSR) (from IgG) were observed in 33% of LAR, 20% of AR, and 0% of HC subjects. NAC-DP increased the proportion of peripheral CD19+ CD20+ CD38+ plasmablasts in AR and LAR patients, but not in HC. Expression of the mucosal homing receptor CXCR3 in peripheral CD19+ CD20+ CD38+ plasmablasts from LAR, AR, and HC individuals was 7%, 5%, and 0.5%, respectively. In vitro DP stimulation increased proliferating CD19+ CD20+ CD38+ plasmablasts in LAR and AR patients, but not in HC. Serum DP-sIgG was higher in LAR and AR patients as compared to HC. BAT was positive in 33%, 100%, and 0% of LAR, AR, and HC subjects, respectively. CONCLUSION: These results suggest that allergen exposure induces the sequential εCSR of IgG+ CD19+ CD20+ CD38+ plasmablasts in the nasal mucosa of LAR patients.

Polymorphisms in CEP68 gene associated with risk of immediate selective reactions to non-steroidal anti-inflammatory drugs.

Non-steroidal anti-inflammatory drugs (NSAIDs) are the main triggers of drug hypersensitivity reactions. Such reactions can be pharmacologically or immunologically mediated, but in both cases individual susceptibility can be influenced by genetic factors. Polymorphisms in centrosomal protein of 68 kDa (CEP68) have been associated with pharmacologically mediated NSAIDs reactions. Here, we evaluated this gene in immunologically mediated single-NSAID-induced urticaria/angioedema or anaphylaxis (SNIUAA) by analyzing 52 single nucleotide polymorphisms in CEP68 in 176 patients and 363 NSAIDs-tolerant controls. Two intronic variants (rs2241160 and rs2241161) were significantly associated with an increased risk of SNIUAA, suggesting CEP68 to be a key player in both types of NSAIDs hypersensitivity. However, we found no overlap with genetic variants previously associated with pharmacologically mediated hypersensitivity, pointing to a complex role for this gene and its potential use in the development of biomarkers of clinical utility to diagnose patients at risk of these reactions and to differentiate entities.

Eicosanoid mediator profiles in different phenotypes of nonsteroidal anti-inflammatory drug-induced urticaria.

BACKGROUND: The role of arachidonic acid metabolites in NSAID-induced hypersensitivity has been studied in depth for NSAID-exacerbated respiratory disease (NERD) and NSAID-exacerbated cutaneous disease (NECD). However, no information is available for NSAID-induced urticarial/angioedema (NIUA), despite it being the most frequent clinical entity induced by NSAID hypersensitivity. We evaluated changes in leukotriene and prostaglandin metabolites for NIUA patients, using patients with NECD and single-NSAID-induced urticaria/angioedema or anaphylaxis (SNIUAA) for comparison. METHODS: Urine samples were taken from patients with confirmed NSAID-induced urticaria and healthy controls, at baseline and at various time intervals after ASA administration. Eicosanoid measurement was performed using high-performance liquid chromatography-tandem mass spectrometry and gas chromatography-mass spectrometry. RESULTS: No differences were found between groups at baseline. Following ASA administration, LTE4 and 9α,11ß-PGF2 levels were increased in both NIUA and NECD patients compared to baseline, rising initially, before decreasing toward initial levels. In addition, the levels of these metabolites were higher in NIUA and NECD when compared with the SNIUAA and control groups after ASA administration. No changes were found with respect to baseline values for SNIUAA and control groups. CONCLUSIONS: We present for the first time data regarding the role of COX-1 inhibition in NIUA. Patients with this entity show a similar pattern eicosanoid levels following ASA challenge to those with NECD. Further studies will help ascertain the cell populations involved and the underlying molecular mechanisms.

Influence of Pore Size in Protein G'-Grafted Mesoporous Silica Nanoparticles as a Serum Pretreatment System for In Vitro Allergy Diagnosis.

Particles with the capacity to bind to immunoglobulin G (IgG) can be used for the purification of IgG or to process clinical samples for diagnostic purposes. For in vitro allergy diagnosis, the high IgG levels in serum can interfere with the detection of allergen-specific IgE, the main diagnostic biomarker. Although commercially available, current materials present a low IgG capture capacity at large IgG concentrations or require complex protocols, preventing their use in the clinic. In this work, mesoporous silica nanoparticles are prepared with different pore sizes, to which IgG-binding protein G' is grafted. It is found that for one particular optimal pore size, the IgG capture capacity of the material is greatly enhanced. The capacity of this material to efficiently capture human IgG in a selective way (compared to IgE) is demonstrated in both solutions of known IgG concentrations as well as in complex samples, like serum, from healthy controls and allergic patients using a simple and fast incubation protocol. Interestingly, IgG removal using the best-performing material enhances in vitro IgE detection in sera from patients allergic to amoxicillin. These results highlight the great translation potential of this strategy to the clinic in the context of in vitro allergy diagnosis.

Deep sequencing of prostaglandin-endoperoxide synthase (PTGE) genes reveals genetic susceptibility for cross-reactive hypersensitivity to NSAID.

BACKGROUND AND PURPOSE: Cross-reactive hypersensitivity to nonsteroidal anti-inflammatory drugs (NSAIDs) is a relatively common adverse drug event caused by two or more chemically unrelated drugs and that is attributed to inhibition of the COX activity, particularly COX-1. Several studies investigated variations in the genes coding for COX enzymes as potential risk factors. However, these studies only interrogated a few single nucleotide variations (SNVs), leaving untested most of the gene sequence. EXPERIMENTAL APPROACH: In this study, we analysed the whole sequence of the prostaglandin-endoperoxide synthase genes, PTGS1 and PTGS2, including all exons, exon-intron boundaries and both the 5' and 3' flanking regions in patients with cross-reactive hypersensitivity to NSAIDs and healthy controls. After sequencing analysis in 100 case-control pairs, we replicated the findings in 540 case-control pairs. Also, we analysed copy number variations for both PTGS genes. KEY RESULTS: The most salient finding was the presence of two PTGS1 single nucleotide variations, which are significantly more frequent in patients than in control subjects. Patients carrying these single nucleotide variations displayed a significantly and markedly lower COX-1 activity as compared to non-carriers for both heterozygous and homozygous patients. CONCLUSION AND IMPLICATIONS: Although the risk single nucleotide variations are present in a small proportion of patients, the strong association observed and the functional effect of these single nucleotide variations raise the hypothesis of genetic susceptibility to develop cross-reactive NSAID hypersensitivity in individuals with an impairment in COX-1 enzyme activity.

Genetic Variants in Cytosolic Phospholipase A2 Associated With Nonsteroidal Anti-Inflammatory Drug-Induced Acute Urticaria/Angioedema.

Nonsteroidal anti-inflammatory drugs (NSAIDs) are among the main triggers of drug hypersensitivity reactions, probably due to their high consumption worldwide. The most frequent type of NSAID hypersensitivity is NSAID cross-hypersensitivity, in which patients react to NSAIDs from different chemical groups in the absence of a specific immunological response. The underlying mechanism of NSAID cross-hypersensitivity has been linked to cyclooxygenase (COX)-1 inhibition causing an imbalance in the arachidonic acid pathway. Despite NSAID-induced acute urticaria/angioedema (NIUA) being the most frequent clinical phenotype, most studies have focused on NSAID-exacerbated respiratory disease. As NSAID cross-hypersensitivity reactions are idiosyncratic, only appearing in some subjects, it is believed that individual susceptibility is under the influence of genetic factors. Although associations with polymorphisms in genes from the AA pathway have been described, no previous study has evaluated the potential role of cytosolic phospholipase A2 (cPLA2) variants. This enzyme catalyzes the initial hydrolysis of membrane phospholipids to release AA, which can be subsequently metabolized into eicosanoids. Here, we analyzed for the first time the overall genetic variation in the cPLA2 gene (PLA2G4A) in NIUA patients. For this purpose, a set of tagging single nucleotide polymorphisms (tagSNPs) in PLA2G4A were selected using data from Europeans subjects in the 1,000 Genomes Project, and genotyped with the iPlex Sequenom MassArray technology. Two independent populations, each comprising NIUA patients and NSAID-tolerant controls, were recruited in Spain, for the purposes of discovery and replication, comprising a total of 1,128 individuals. Fifty-eight tagSNPs were successfully genotyped in the discovery cohort, of which four were significantly associated with NIUA after Bonferroni correction (rs2049963, rs2064471, rs12088010, and rs12746200). These polymorphisms were then genotyped in the replication cohort: rs2049963 was associated with increased risk for NIUA after Bonferroni correction under the dominant and additive models, whereas rs12088010 and rs12746200 were protective under these two inheritance models. Our results suggest a role for PLA2G4A polymorphisms in NIUA. However, further studies are required to replicate our findings, elucidate the mechanistic role, and evaluate the participation of PLA2G4A variants in other phenotypes induced by NSAID cross-hypersensitivity.

Lack of Major Involvement of Common CYP2C Gene Polymorphisms in the Risk of Developing Cross-Hypersensitivity to NSAIDs.

Cross-hypersensitivity to non-steroidal anti-inflammatory drugs (NSAIDs) is a relatively common, non-allergic, adverse drug event triggered by two or more chemically unrelated NSAIDs. Current evidence point to COX-1 inhibition as one of the main factors in its etiopathogenesis. Evidence also suggests that the risk is dose-dependent. Therefore it could be speculated that individuals with impaired NSAID biodisposition might be at increased risk of developing cross-hypersensitivity to NSAIDs. We analyzed common functional gene variants for CYP2C8, CYP2C9, and CYP2C19 in a large cohort composed of 499 patients with cross-hypersensitivity to NSAIDs and 624 healthy individuals who tolerated NSAIDs. Patients were analyzed as a whole group and subdivided in three groups according to the main enzymes involved in the metabolism of the culprit drugs as follows: CYP2C9, aceclofenac, indomethacin, naproxen, piroxicam, meloxicam, lornoxicam, and celecoxib; CYP2C8 plus CYP2C9, ibuprofen and diclofenac; CYP2C19 plus CYP2C9, metamizole. Genotype calls ranged from 94 to 99%. No statistically significant differences between patients and controls were identified in this study, either for allele frequencies, diplotypes, or inferred phenotypes. After patient stratification according to the enzymes involved in the metabolism of the culprit drugs, or according to the clinical presentation of the hypersensitivity reaction, we identified weak significant associations of a lower frequency (as compared to that of control subjects) of CYP2C8*3/*3 genotypes in patients receiving NSAIDs that are predominantly CYP2C9 substrates, and in patients with NSAIDs-exacerbated cutaneous disease. However, these associations lost significance after False Discovery Rate correction for multiple comparisons. Taking together these findings and the statistical power of this cohort, we conclude that there is no evidence of a major implication of the major functional CYP2C polymorphisms analyzed in this study and the risk of developing cross-hypersensitivity to NSAIDs. This argues against the hypothesis of a dose-dependent COX-1 inhibition as the main underlying mechanism for this adverse drug event and suggests that pre-emptive genotyping aiming at drug selection should have a low practical utility for cross-hypersensitivity to NSAIDs.

Platelet-Adherent Leukocytes Associated With Cutaneous Cross-Reactive Hypersensitivity to Nonsteroidal Anti-Inflammatory Drugs.

Nonsteroidal anti-inflammatory drugs (NSAIDs) are among the most highly consumed drugs worldwide and the main triggers of drug hypersensitivity reactions. The most frequent reaction, named cross-reactive NSAID-hypersensitivity, is due to the pharmacological activity of these drugs by blocking the cyclooxygenase-1 enzyme. Such inhibition leads to cysteinyl-leukotriene synthesis, mainly LTE4, which are responsible for the reaction. Although the complete molecular picture of the underlying mechanisms remains elusive, the participation of platelet-adherent leukocytes (CD61+) and integrins have been described for NSAID-exacerbated respiratory disease (NERD). However, there is a lack of information concerning NSAID-induced urticaria/angioedema (NIUA), by far the most frequent clinical phenotype. Here we have evaluated the potential role of CD61+ leukocytes and integrins (CD18, CD11a, CD11b, and CD11c) in patients with NIUA, and included the other two phenotypes with cutaneous involvement, NSAID-exacerbated cutaneous disease (NECD) and blended reactions (simultaneous skin and airways involvement). A group NSAID-tolerant individuals was also included. During the acute phase of the reaction, the three clinical phenotypes showed increased frequencies of CD61+ neutrophils, eosinophils, and monocytes compared to controls, which correlated with urinary LTE4 levels. However, no correlation was found between these variables at basal state. Furthermore, increased expressions of CD18 and CD11a were found in the three CD61+ leukocytes subsets in NIUA, NECD and blended reactions during the acute phase when compared with CD61-leukocyte subpopulations. During the acute phase, CD61+ neutrophils, eosinophils and monocytes showed increased CD18 and CD11a expression when compared with CD61+ leukocytes at basal state. No differences were found when comparing controls and CD61+ leukocytes at basal state. Our results support the participation of platelet-adherent leukocytes and integrins in cutaneous cross-hypersensitivity to NSAIDs and provide a link between these cells and arachidonic acid metabolism. Our findings also suggest that these reactions do not involve a systemic imbalance in the frequency of CD61+ cells/integrin expression or levels of LTE4, which represents a substantial difference to NERD. Although further studies are needed, our results shed light on the molecular basis of cutaneous cross-reactive NSAID-hypersensitivity, providing potential targets for therapy through the inhibition of platelet-leukocyte interactions.

An Update on the Immunological, Metabolic and Genetic Mechanisms in Drug Hypersensitivity Reactions.

Drug hypersensitivity reactions (DHRs) represent a major burden on the healthcare system since their diagnostic and management are complex. As they can be influenced by individual genetic background, it is conceivable that the identification of variants in genes potentially involved could be used in genetic testing for the prevention of adverse effects during drug administration. Most genetic studies on severe DHRs have documented HLA alleles as risk factors and some mechanistic models support these associations, which try to shed light on the interaction between drugs and the immune system during lymphocyte presentation. In this sense, drugs are small molecules that behave as haptens, and currently three hypotheses try to explain how they interact with the immune system to induce DHRs: the hapten hypothesis, the direct pharmacological interaction of drugs with immune receptors hypothesis (p-i concept), and the altered self-peptide repertoire hypothesis. The interaction will depend on the nature of the drug and its reactivity, the metabolites generated and the specific HLA alleles. However, there is still a need of a better understanding of the different aspects related to the immunological mechanism, the drug determinants that are finally presented as well as the genetic factors for increasing the risk of suffering DHRs. Most available information on the predictive capacity of genetic testing refers to abacavir hypersensitivity and anticonvulsants-induced severe cutaneous reactions. Better understanding of the underlying mechanisms of DHRs will help us to identify the drugs likely to induce DHRs and to manage patients at risk.

Pharmacogenomics of Prostaglandin and Leukotriene Receptors.

Individual genetic background together with environmental effects are thought to be behind many human complex diseases. A number of genetic variants, mainly single nucleotide polymorphisms (SNPs), have been shown to be associated with various pathological and inflammatory conditions, representing potential therapeutic targets. Prostaglandins (PTGs) and leukotrienes (LTs) are eicosanoids derived from arachidonic acid and related polyunsaturated fatty acids that participate in both normal homeostasis and inflammatory conditions. These bioactive lipid mediators are synthesized through two major multistep enzymatic pathways: PTGs by cyclooxygenase and LTs by 5-lipoxygenase. The main physiological effects of PTGs include vasodilation and vascular leakage (PTGE2); mast cell maturation, eosinophil recruitment, and allergic responses (PTGD2); vascular and respiratory smooth muscle contraction (PTGF2), and inhibition of platelet aggregation (PTGI2). LTB4 is mainly involved in neutrophil recruitment, vascular leakage, and epithelial barrier function, whereas cysteinyl LTs (CysLTs) (LTC4, LTD4, and LTE4) induce bronchoconstriction and neutrophil extravasation, and also participate in vascular leakage. PTGs and LTs exert their biological functions by binding to cognate receptors, which belong to the seven transmembrane, G protein-coupled receptor superfamily. SNPs in genes encoding these receptors may influence their functionality and have a role in disease susceptibility and drug treatment response. In this review we summarize SNPs in PTGs and LTs receptors and their relevance in human diseases. We also provide information on gene expression. Finally, we speculate on future directions for this topic.