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Identification of a major epitope recognized by PLA2R autoantibodies in primary membranous nephropathy.

ABSTRACT: Phospholipase A2 receptor 1 (PLA2R) is a target autoantigen in 70% of patients with idiopathic membranous nephropathy. We describe the location of a major epitope in the N-terminal cysteine-rich ricin domain of PLA2R that is recognized by 90% of human anti-PLA2R autoantibodies. The epitope was sensitive to reduction and SDS denaturation in the isolated ricin domain and the larger fragment containing the ricin, fibronectin type II, first and second C-type lectin domains (CTLD). However, in nondenaturing conditions the epitope was protected against reduction in larger fragments, including the full-length extracellular region of PLA2R. To determine the composition of the epitope, we isolated immunoreactive tryptic fragments by Western blotting and analyzed them by mass spectrometry. The identified peptides were tested as inhibitors of autoantibody binding to PLA2R by surface plasmon resonance. Two peptides from the ricin domain showed strong inhibition, with a longer sequence covering both peptides (31-mer) producing 85% inhibition of autoantibody binding to PLA2R. Anti-PLA2R antibody directly bound this 31-mer peptide under nondenaturing conditions and binding was sensitive to reduction. Analysis of PLA2R and the PLA2R-anti-PLA2R complex using electron microscopy and homology-based representations allowed us to generate a structural model of this major epitope and its antibody binding site, which is independent of pH-induced conformational change in PLA2R. Identification of this major PLA2R epitope will enable further therapeutic advances for patients with idiopathic membranous nephropathy, including antibody inhibition therapy and immunoadsorption of circulating autoantibodies.

SUBMITTER: Fresquet M

PROVIDER: S-EPMC4310666 | biostudies-literature | 2015 Feb

REPOSITORIES: biostudies-literature

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Identification of a major epitope recognized by PLA2R autoantibodies in primary membranous nephropathy.

Fresquet Maryline M Jowitt Thomas A TA Gummadova Jennet J Collins Richard R O'Cualain Ronan R McKenzie Edward A EA Lennon Rachel R Brenchley Paul E PE

Journal of the American Society of Nephrology : JASN 20141006 2

Phospholipase A2 receptor 1 (PLA2R) is a target autoantigen in 70% of patients with idiopathic membranous nephropathy. We describe the location of a major epitope in the N-terminal cysteine-rich ricin domain of PLA2R that is recognized by 90% of human anti-PLA2R autoantibodies. The epitope was sensitive to reduction and SDS denaturation in the isolated ricin domain and the larger fragment containing the ricin, fibronectin type II, first and second C-type lectin domains (CTLD). However, in nonden ...[more]

PMID: 25288605

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Project description:BackgroundM-type phospholipase A2 receptor (PLA2R) is known as a major antigen on podocytes, which is involved with the pathogenesis of idiopathic membranous nephropathy (iMN). Many studies have shown that serum anti-PLA2R autoantibodies (sPLA2R) are prevalent in patients with iMN but are rarely detected in secondary membranous nephropathy (SMN) or other glomerulonephritis. The anti-PLA2R is considered as a promising serum biomarker in iMN but reports about its diagnostic value are variable and inconsistent.ObjectiveTo evaluate the diagnostic test accuracy (DTA) of anti-PLA2R and glomerular PLA2R antigen (gPLA2R) for diagnosing iMN.MethodMEDLINE, EMBASE, WEB OF SCIENCE, and COCHRANE LIBRARY were searched from 2009 January to February 2018. Heterogeneity was evaluated by Q test and I2. Source of heterogeneity was explored by subgroup analysis and meta-regression. Meta-analysis was executed and reported according to the Preferred Reporting Items for Systematic Reviews and Meta-analyses statement.ResultsTotally, 35 studies were retrieved under the pre-set study eligibility criteria. Twenty-eight studies were included to evaluate the DTA of anti-PLA2R for differentiating iMN from non-iMN. They indicated a pooled sensitivity of 65% (63-67%), specificity of 97% (97-98%), positive likelihood ratio of 15.65 (9.95-24.62), and negative likelihood ratio of 0.37 (0.32-0.42) with a diagnostic OR (sDOR) of 50.41 (31.56 to 80.52) and AUC of 0.9393. No threshold effect was detected. The heterogeneity analysis for sDOR showed that I2 = 50.3% and Cochran-Q = 54.29, df = 27 (p = 0.0014). Heterogeneity was significant. Meta-regression revealed that sample size might be the potential source of heterogeneity. Subgroup analysis demonstrated that method type and ratio of patients with nephrotic-range proteinuria at baseline might be the source of heterogeneity. Sixteen studies reported the diagnostic value of glomerular PLA2R antigen for differentiating iMN from non-iMN. The pooled sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, sDOR, and AUC were 79% (76-81%), 90% (88-92%), 8.17 (5.60-11.93), 0.25 (0.19-0.33), 39.37 (22.18-60.13), and 0.9278. Heterogeneity analysis showed that Cochran-Q = 35.36; df = 15 (p = 0.002), and I2 for sDOR was 57.6%.ConclusionsPLA2R and gPLA2R demonstrated a good diagnostic accuracy in differentiating iMN and non-iMN.

Project description:Primary membranous nephropathy (PMN) is a renal-specific autoimmune disease caused by circulating autoantibodies that target glomerular podocyte antigens (PLA2R/THSD7A). However, very little is known on the molecular mechanisms controlling B cell response in this nephropathy. The present study was aimed at correlating the serum levels of B cell activators BAFF/BLyS and APRIL with the presence of anti-PLA2R antibodies in PMN patients and with long-term clinical outcome. To this aim, 51 patients with anti-PLA2R-positive biopsy-proven PMN and nephrotic range proteinuria (>3.5 g/24 hours) were enrolled between January 2009 and December 2015 and treated with conventional 6-month immunosuppressive therapy. After 6 months, 29 patients (56.9%) cleared circulating anti-PLA2R, while in remaining 22 (43.1%), they persisted. Intriguingly, in the first group, baseline serum levels of BAFF/BLyS and APRIL were significantly lower than those in the second one. Moreover, after 6 months of immunosuppressive therapy, an overall reduction in both cytokine serum levels was observed. However, in PMN patients with anti-PLA2R clearance, this reduction was more prominent, as compared with those with anti-PLA2R persistence. When related to clinical outcome, lower baseline BAFF/BLyS (<6.05 ng/mL) and APRIL (<4.20 ng/mL) serum levels were associated with significantly higher probability to achieve complete or partial remission after 24-month follow-up. After dividing the entire study cohort into three groups depending on both cytokine baseline serum levels, patients with both BAFF/BLyS and APRIL below the cut-off showed a significantly higher rate of complete or partial remission as compared with patients with only one cytokine above the cut-off, while the composite endpoint was achieved in a very low rate of patients with both cytokines above the cut-off. Taken together, these results provide new insights into the role of BAFF/BLyS and APRIL in both the pathogenesis of anti-PLA2R-positive PMN and the response to immunosuppressive therapy.

Dataset Information

Identification of a major epitope recognized by PLA2R autoantibodies in primary membranous nephropathy.

Publications

Identification of a major epitope recognized by PLA2R autoantibodies in primary membranous nephropathy.

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