Antigenic determinants of asthma-associated allergens for design of immunotherapy

Conducting structural and functional analysis of antigenic determinants on the major group 1 house dust mite allergens Der p 1 (from Dermatophagoides pteronyssinus) and Der f 1 (from D. farinae) to define specificity and cross-reactivity associated with IgE antibody recognition. Using x-ray crystal structures of allergen–antibody complexes and site-directed mutagenesis, the team examined how particular amino acid residues within epitopes contribute to binding by three murine monoclonal antibodies (mAbs) that interfere with IgE binding: the Der p 1-specific mAbs 5H8 and 10B9, and the cross-reactive mAb 4C1. Structural superposition of Der p 1–antibody complexes (PDB codes 3RVW, 4PP1, 4PP2) defined spatial relationships among these epitopes and identified overlapping regions between 4C1 and 10B9.

The investigators substituted selected residues in epitopes for mAbs 5H8 and 4C1 and observed impaired antibody binding to Der p 1, demonstrating the functional importance of those residues for mAb recognition. Conversely, they engineered an epitope for the Der p 1-specific mAb 10B9 into Der f 1 by introducing one to three amino acid changes; these mutations were sufficient to confer binding by mAb 10B9 to Der f 1. Analysis showed that the residues enabling 10B9 binding form hydrogen bonds with complementarity-determining regions (CDRs) of the antibody other than heavy chain CDR3 (H CDR3), revealing an exception to the commonly observed dominant role of H CDR3 in antigen recognition.

Functional assays extended the structural observations to human IgE antibody recognition. Inhibition assays using pooled plasma from mite-allergic patients demonstrated that multiple-epitope mutants of Der p 1 reduced IgE binding compared with wild-type protein. Specifically, triple and quadruple mutants targeting mAb 4C1 and mAb 5H8 epitopes, respectively, decreased IgE binding, while the Der f 1 triple mutant showed increased IgE binding relative to wild-type Der f 1. These findings identify critical amino acids required for both monoclonal antibody and IgE recognition of group 1 mite allergens.

Collectively, the study maps key antigenic determinants on Der p 1 and Der f 1 and clarifies molecular bases for specificity and cross-reactivity among antibodies targeting these allergens. By pinpointing residues that govern antibody interactions and demonstrating that limited targeted mutations can alter antibody binding patterns, the work provides actionable information for engineering allergen variants with reduced IgE binding. Such engineered mutants could serve as candidate molecules for safer, more effective immunotherapy approaches by maintaining structural integrity while diminishing IgE reactivity. Chruszcz and collaborators thereby advance understanding of allergen–antibody interactions at atomic resolution and offer a structural roadmap to guide the rational design of modified allergens for therapeutic use.