Abstract: TH-OR003
Detection and Characterization of Serum Anti-EXT1/2 Antibodies in Exostosin-Associated Membranous Nephropathy
Session Information
- Advances in Renal Pathology and Laboratory Medicine
October 22, 2026 | Location: Room 605, Convention Center
Abstract Time: 04:50 PM - 05:00 PM
Category: Pathology and Lab Medicine
- 1700 Pathology and Lab Medicine
Authors
- Arivett, Brock A., Meharry Medical College School of Medicine, Nashville, Tennessee, United States
- Caza, Tiffany, Arkana Laboratories, Little Rock, Arkansas, United States
- Storey, Aaron J., Arkana Laboratories, Little Rock, Arkansas, United States
- Chapla, Digantkumar G., University of Georgia, Athens, Georgia, United States
- Larsen, Christopher Patrick, Arkana Laboratories, Little Rock, Arkansas, United States
- Moremen, Kelley, University of Georgia, Athens, Georgia, United States
- Borza, Dorin-Bogdan, Meharry Medical College School of Medicine, Nashville, Tennessee, United States
Background
The EXT1/EXT2 complex is the third most common antigen identified in subepithelial immune deposits from patients with membranous nephropathy (MN). However, whether EXT1 or EXT2 are true autoantigens or mere disease biomarkers is still controversial because no assays to date have successfully detected circulating anti-EXT antibodies. Our goals were to determine whether anti-EXT1/2 autoantibodies actually exist and, if so, to develop a quantitative immunoassay for measuring their serum levels.
Methods
Immune complexes were recovered from residual frozen biopsy tissue of MN biopsies by protein G immunoprecipitation, followed by data-independent analysis-mass spectrometry for determination of antigen type. Serum samples were available from 8 patients with a biopsy diagnosis of MN that were EXT1-positive by immunostaining. Three preparations of recombinant EXT1/EXT2 protein with different affinity tags were evaluated as solid-phase antigens in an ELISA immunoassay. The binding of antibodies from patient and control sera to EXT1/EXT2-coated wells was assayed using HRP-conjugated anti-human IgG followed by colorimetric detection. IgG1-IgG4 subclasses were similarly measured.
Results
EXT1 and EXT2 peptides were specifically detected in immune complexes recovered from kidney biopsies of EXT-positive MN patients and were absent within PLA2R-, THSD7A-, or NELL1-MN controls. The EXT1/EXT2 antigen showing the highest immunoreactivity in ELISA demonstrated good discrimination between EXT-positive MN cases and controls (area under the ROC curve = 0.93; 95% CI 0.81-1.00, P = 0.0011). At the optimal cutoff, anti-EXT1/2 IgG antibodies were detected in 7/8 sera from EXT-associated MN patients (87.5% assay sensitivity) with 92.3% specificity. Pre-treatment of the EXT1/EXT2 antigen with denaturing agents prior to coating significantly reduced IgG binding, suggesting conformation-dependent epitopes. Among the seven sera found to contain anti-EXT1/2 IgG, the predominant subclass was IgG1 in three, IgG4 in three, and one was co-dominant for IgG1/IgG3/IgG4.
Conclusion
Our results show that anti-EXT1/2 IgG antibodies are present in not only glomerular immune complexes but also sera from patients with EXT-associated MN. The novel ELISA immunoassay affords rapid, high-throughput, quantitative measurement of serum anti-EXT1/2 IgG, which could be used to monitor disease activity and response to treatment.
Funding
- Other NIH Support