Identification of an antigenic determinant of clavulanic acid responsible for IgE‐mediated reactions

Abstract

Background: Selective reactions to clavulanic acid (CLV) account for around 30% of immediate reactions after administration of amoxicillin-CLV. Currently, no immunoassay is available for detecting specific IgE to CLV, and its specific recognition in patients with immediate reactions has only been demonstrated by basophil activation testing, however with suboptimal sensitivity. The lack of knowledge regarding the structure of the drug that remains bound to proteins (antigenic determinant) is hampering the development of in vitro diagnostics. We aimed to identify the antigenic determinants of CLV as well as to evaluate their specific IgE recognition and potential role for diagnosis.Methods: Based on complex CLV degradation mechanisms, we hypothesized the formation of two antigenic determinants for CLV, AD-I (N-protein, 3-oxopropanamide) and AD-II (N-protein, 3-aminopropanamide), and designed different synthetic analogs to each one. IgE recognition of these structures was evaluated in basophils from patients with selective reactions to CLV and tolerant subjects. In parallel, the CLV fragments bound to proteins were identified by proteomic approaches.Results: Two synthetic analogs of AD-I were found to activate basophils from allergic patients. This determinant was also detected bound to lysines 195 and 475 of CLVtreated human serum albumin. One of these analogs was able to activate basophils in 59% of patients whereas CLV only in 41%. Combining both results led to an increase in basophil activation in 69% of patients, and only in 12% of controls.

Recent developments and highlights in drug hypersensitivity

“…An approach to increase in vitro test sensitivity is the inclusion of drug metabolites besides the native drug . This has been recently confirmed for AX‐CLV hypersensitivity, in which the use of one CLV synthetic determinant, together with CLV itself, improves significantly the sensitivity from 41% to 69% …”

“…Another study has suggested that patients could be immunized by benzylpenicillin‐haptenated peptides from proteins generated in dendritic cells during the processing and presentation to T cells . Moreover, CLV binds to Lys 195 and 475 residues of HSA, forming adducts able to activate basophils in 69% of CLV‐allergic patients . Regarding amoxicillin, haptenated intracellular proteins of B lymphocytes, such as EF‐2, HSP‐70, β‐actin and α‐enolase, have been identified.…”

“…119 This has been recently confirmed for AX-CLV hypersensitivity, in which the use of one CLV synthetic determinant, together with CLV itself, improves significantly the sensitivity from 41% to 69%. 120 The use of nanostructures bound to BLs has been evaluated in BAT, proving the importance of the carrier molecule and its tridimensional distribution for IgE recognition. Those nanostructures with higher diameter and number of determinants induced a higher level of basophil activation in allergic patients.…”

“…141 Moreover, CLV binds to Lys 195 and 475 residues of HSA, forming adducts able to activate basophils in 69% of CLV-allergic patients. 120…”

Biomarkers in the Diagnosis and Therapy of Allergic Diseases and Asthma

“…193 Interestingly, one antigenic determinant recognized by most patients with confirmed reactions to clavulanic acid has been recently identified. 196 Therefore, amoxicillin and any suspected BL must be included when performing STs. 192,195,197,198 For other drugs and for NIRs, the value of STs is very limited.…”