Abstract: TH-PO0661
Mapping Conformational Epitopes in Phospholipase A2 Receptor 1 (PLA2R1), the Major Autoantigen of Membranous Nephropathy
Session Information
- Glomerular Diseases: Immunopathogenesis and Targeted Therapeutics
November 06, 2025 | Location: Exhibit Hall, Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Glomerular Diseases
- 1401 Glomerular Diseases: Mechanisms, including Podocyte Biology
Authors
- Abdel Sater, Alice, Institut de Pharmacologie Moleculaire et Cellulaire, Valbonne, Provence-Alpes-Côte d'Azur, France
- Dolla, Guillaume, Institut de Pharmacologie Moleculaire et Cellulaire, Valbonne, Provence-Alpes-Côte d'Azur, France
- Justino, Joana, Institut de Pharmacologie Moleculaire et Cellulaire, Valbonne, Provence-Alpes-Côte d'Azur, France
- Brglez, Vesna, Institut de Pharmacologie Moleculaire et Cellulaire, Valbonne, Provence-Alpes-Côte d'Azur, France
- Wu, Tsai-yi, Chang Gung University College of Medicine, Taoyuan, Taoyuan City, Taiwan
- Tu, Kun-Hua, Chang Gung University College of Medicine, Taoyuan, Taoyuan City, Taiwan
- Tomas, Nicola M., Universitatsklinikum Hamburg-Eppendorf, Hamburg, HH, Germany
- Seitz-Polski, Barbara, Centre Hospitalier Universitaire de Nice, Nice, Provence-Alpes-Côte d'Azur, France
- Ronco, Pierre M., Hopital Tenon, Paris, Île-de-France, France
- Lambeau, Gerard J., Institut de Pharmacologie Moleculaire et Cellulaire, Valbonne, Provence-Alpes-Côte d'Azur, France
Background
Membranous nephropathy (MN) is caused by autoantibodies targeting PLA2R1 in 55% of patients. We previously demonstrated that anti-PLA2R1 autoantibodies target conformational epitopes in up to five PLA2R1 domains, with immunodominant epitopes in the cysteine-rich (CR) and C-type lectin domain 1 (C1) domains, classifying patients in two groups: iCR (60%) and iC1 (40%), with different clinical outcomes and responses to treatment.
Methods
To precisely map the conformational epitope(s) within the CR domain, we generated 184 single-point mutants (alanine scanning and other point mutations) and 69 chimeras between CR and homologous domains. All constructs were validated for expression as folded proteins in HEK293 cells by western blot and ELISA. The reactivities of mutants and chimeras were tested by ELISA against mouse monoclonal anti-PLA2R1 antibodies as controls and polyclonal sera from nine patients (iCR/iC1) from France and Taiwan, as well as six CR-specific recombinant monoclonal antibodies (mAbs) cloned from three Taiwanese patients. In parallel, complexes between soluble PLA2R1 and the CR domain with single-chain variable fragments (scFvs) and fragment antibodies (Fabs) from patients’ antibodies were designed for structural studies using X-ray crystallography and cryo-EM.
Results
Using our epitope mapping strategy, we identified a single immunodominant epitope region in CR targeted by all patients. In silico modeling with AlphaFold3 of CR-Antibody complexes independently validated our results obtained by mutagenesis. The epitope region consists of two distinct but adjacent epitopes and differs from the epitope region previously reported. To definitely validate our findings, we are solving the structures of CR-antibody complexes using X-ray crystallography and Cryo-EM experiments.
Conclusion
These insights into the immunodominant CR domain contribute to a better understanding of the molecular pathophysiology of MN, including disease etiology, and pave the way for better diagnosis, prognosis, and the development of targeted therapeutic strategies.
Funding
- Government Support – Non-U.S.