Adriana Ariza

In vitro evaluation of IgE-mediated hypersensitivity reactions to quinolones

To cite this article: Aranda A, Mayorga C, Ariza A, Doña I, Rosado A, Blanca‐Lopez N, Andreu I, Torres MJ. In vitro evaluation of IgE‐mediated hypersensitivity reactions to quinolones. Allergy 2011; 66: 247–254.

Diagnostic evaluation of hypersensitivity reactions to beta‐lactam antibiotics in a large population of children

Hypersensitivity reactions to beta‐lactams (BLs) are often reported in children, with amoxicillin and, to a lesser extent, cephalosporins being the most frequent drugs involved. Although many of these children are considered to be allergic, a careful evaluation only confirms a low percentage.

Role of minor determinants of amoxicillin in the diagnosis of immediate allergic reactions to amoxicillin

To cite this article: Torres MJ, Ariza A, Fernández J, Moreno E, Laguna JJ, Montañez MI, Ruiz‐Sanchez AJ, Blanca M. Role of minor determinants of amoxicillin in the diagnosis of immediate allergic reactions to amoxicillin. Allergy 2010; 65: 590–596.

Protein haptenation by amoxicillin: High resolution mass spectrometry analysis and identification of target proteins in serum

Allergy towards wide spectrum antibiotics such as amoxicillin (AX) is a major health problem. Protein haptenation by covalent conjugation of AX is considered a key process for the allergic response. However, the nature of the proteins involved has not been completely elucidated. Human serum albumin (HSA) is the most abundant protein in plasma and is considered a major target for haptenation by drugs, including β-lactam antibiotics. Here we report a procedure for immunological detection of AX-protein adducts with antibodies recognizing the lateral chain of the AX molecule. With this approach we detected human serum proteins modified by AX in vitro and identified HSA, transferrin and immunoglobulins heavy and light chains as prominent AX-modified proteins. Since HSA was the major AX target, we characterized AX-HSA interaction using high resolution LTQ orbitrap MS. At 0.5mg/mL AX, we detected one main AX-HSA adduct involving residues Lys 190, 199 or 541, whereas higher AX concentrations elicited a more extensive modification. In molecular modeling studies Lys190 and Lys 199 were found the most reactive residues towards AX, with surrounding residues favoring adduct formation. These findings provide novel tools and insight for the study of protein haptenation and the mechanisms involved in AX-elicited allergic reactions.

Diagnosis of immediate hypersensitivity reactions to radiocontrast media

No consensus exists on the diagnostic approach for immediate hypersensitivity reactions (IHR) to radiocontrast media (RCM). We analyzed the diagnostic value of a skin test (ST), drug provocation test (DPT) and basophil activation test (BAT) in patients with symptoms compatible with IHR to RCM.

IgE‐mediated hypersensitivity reactions to methylprednisolone

To cite this article: Aranda A, Mayorga C, Ariza A, Doña I, Blanca‐Lopez N, Canto G, Blanca M, Torres MJ. IgE‐mediated hypersensitivity reactions to methylprednisolone. Allergy 2010; 65: 1376–1380.

Basophil Activation after Nonsteroidal Anti-inflammatory Drugs Stimulation in Patients with Immediate Hypersensitivity Reactions to These Drugs

Nonsteroidal anti‐inflammatory drugs (NSAIDs) are the drugs most frequently involved in allergic reactions of which two main types exist: IgE‐mediated and crossintolerance. The diagnosis of crossintolerance reactions is often based on the drug provocation test. The potential value of the basophil activation test (BAT) was evaluated using different basophil markers in the diagnosis of patients with crossintolerance to NSAIDs and cutaneous symptoms. We studied 46 patients with crossintolerance to NSAIDs and 45 tolerant controls. BAT was performed with acetyl salicylic acid, paracetamol, diclofenac, dipyrone, naproxen, and ibuprofen at four different concentrations using CD193 and CD203c as basophil markers and CD63 as activation marker. We compared BAT results using CD193+ or CD193+CD203c+ for basophil selection and found a significant increase in the stimulation index when using CD193+CD203c+ in both patients and controls (P = 0.004 and P = 0.017, respectively). Selection of living cells only produced an increase in basophil stimulation in patients for both CD193+ and CD193+CD203c+ (P < 0.001 for both), whereas in controls there was no change with CD193+ and a decrease with CD193+CD203c+ (P = 0.001). We found that CD193+CD203c+ increased the percentage of positive cases in patients and controls when compared with CD193+. When excluding dead cells, there was an increase of 21.7% in patients and 10% in controls. These results indicate that using CD193+CD203+, excluding dead cells, is the best approach for BAT although this test is not recommended for the diagnosis of patients with crossintolerance to NSAIDs owing to its low sensitivity and specificity.

Fluoroquinolone Photodegradation Influences Specific Basophil Activation

Fluoroquinolones (FQs) are photoreactive drugs, but it is not known whether laboratory light exposure can influence the induction of photoproducts and modify in vitro test results. The basophil activation test (BAT) has proven to be useful for evaluating immunoglobulin E (IgE)-mediated hypersensitivity to FQs, with a higher percentage of positive responders with ciprofloxacin (CIP) than with moxifloxacin (MOX). We studied the effect of laboratory light on CIP and MOX degradation, and drug-protein conjugate formation, and its influence on the BAT for evaluating IgE-mediated hypersensitivity to FQs. The results showed an important decrease in fluorescence emission intensity under light compared to dark conditions for MOX, and that BAT positivity was lower in light (17.9%) than in dark (35.7%) conditions. No changes were found for CIP in either fluorescence emission intensity or BAT results (46.4% in both conditions). We can conclude that light exposure is a critical factor in BAT results when photolabile drugs like MOX are used. Therefore, light is important when interpreting in vitro results.

Synthetic Approach to Gain Insight into Antigenic Determinants of Cephalosporins: In Vitro Studies of Chemical Structure−IgE Molecular Recognition Relationships

Cephalosporins, after penicillins, are the most widely used antibacterial agents in infectious diseases and the cause of adverse immune reactions in the world. Whether a patient with a suspected allergy to a β-lactam can safely take a cephalosporin is often a matter of debate. However, there are no tests with enough sensitivity to detect allergy to cephalosporins. Understanding the way in which the drug metabolizes after protein conjugation is important if we are to make advances in the diagnosis of clinical allergy. Structural studies of cephalosporin-protein adducts have never been addressed successfully and are difficult to investigate. Our approach to determine the requirements involved in antigenic determinant structures consisted of designing and synthesizing a proposed skeleton that remains linked to the carrier protein after chemical degradation in cephalosporin conjugated to carrier proteins. In this study, a series of proposed epitopes were efficiently synthesized following a versatile methodology, involving the condensation of the R(1) acyl side chains of native cephalosporins, with the nuclear fragment structures (derived from amino acids or other aminofunctionalized molecules). The final well-defined structures 1-4 (a-f), representing a fragment from the proposed cephalosporin-Lys(protein) adduct intermediate, consist of closely related low-molecular-weight molecules, differing only in the functional group at C-3 and the R(1) side chains. They were assessed with sera from patients allergic to cephalosporins to study structure-IgE molecular recognition relationships. These IgE showed an enhanced recognition to proposed new skeleton epitopes with adequate functionality at C-3, with the specifities mainly related to the R(1) acyl side chain. Thus, this study led us to refine the model haptenic structures of cephalosporins and gain insight into the chemical mechanism of epitope formation.

Hypersensitivity to fluoroquinolones: The expression of basophil activation markers depends on the clinical entity and the culprit fluoroquinolone.

AbstractAlthough fluoroquinolones (FQs) are generally well-tolerated antibiotics, increasing numbers of hypersensitivity reactions have been reported. These can be evaluated in vitro by basophil activation tests (BATs); however, sensitivity is not optimal. Many factors could influence sensitivity such as basophil activation markers. The objective of this study was to evaluate the influence of 2 different activations markers, CD63 and CD203c, on the sensitivity of BAT to FQ. We studied 17 patients with immediate allergic reactions to FQ. BAT was performed with moxifloxacin and ciprofloxacin using CD193 (CCR3) for basophil selection and CD203c or CD63 as activation markers. Stimulation with ciprofloxacin induced a significantly higher expression of CD63 in ciprofloxacin-allergic patients compared to moxifloxacin-allergic patients (P = 0.002). In patients allergic to moxifloxacin with anaphylactic shock, we have observed an increase in the percentage of cells that upregulate CD203c, whereas patients with anaphylaxis preferentially upregulate CD63. The best sensitivity–specificity was obtained using a cutoff of 3 and the culprit FQ, using CD203c for moxifloxacin-allergic patients (sensitivity = 36.4%; specificity = 94.4%), and CD63 for ciprofloxacin-allergic patients (sensitivity = 83.3%; specificity = 88.9%). A negative correlation was found between the upregulation of CD63 and CD203c and the time interval between the reaction occurrence and the performance of the test (Spearman r = −0.446; P < 0.001 for CD63 and Spearman r = −0.386; P < 0.001 for CD203c). The performance of BAT for FQ allergy must be optimized for each drug, taking into account possible differences in the stimulation mechanism that leads to the upregulation of different activation markers.