Prediagnostic Appearance of Thrombospondin Type-1 Domain 7A Autoantibodies in Membranous Nephropathy

Abstract

Background: Pathogenic autoantibodies against thrombospondin type-1 domain 7A (THSD7A) are present in approximately 3% of patients with membranous nephropathy (MN). Compared to PLA2R antibodies, less is known about THSD7A autoantibodies (AB) due to the relative rarity and lack of a commercially, available quantitative immunoassay. Methods: Here we describe the development and validation of a highly quantitative, luciferase immunoprecipitation systems (LIPS) assay for detecting THSD7A AB and used it to study dominant THSD7A epitopes, disease associations and monitoring disease activity. The Department of Defense Serum Repository (DODSR) was then used to analyze THSD7A AB in 371 longitudinal serum samples collected before clinical diagnosis of MN from 110 PLA2R-negative MN subjects. Results: LIPS analysis demonstrated that a near full-length THSD7A (amino acids 1-1656) detected robust autoantibody levels in all known seropositive MN patients with 100% sensitivity and specificity compared to ELISA and/or Western blotting. The majority of THSD7A seropositive subjects in our pilot cohort had evidence of coexisting autoimmunity or cancer. Moreover, three THSD7A seropositive cases undergoing immunosuppressive therapy showed longitudinal autoantibody levels that tracked clinical status. Additional epitope analysis of two smaller protein THSD7A fragments spanning amino acids 1-416 and 1-671 demonstrated lower sensitivity of 32% and 44%, respectively. In the DODSR cohort, THSD7A seropositivity was detected in 4.5% of PLA2R negative MN cases. In one "primary" and in one secondary MN-associated with cancer, THSD7A AB were detectable less than one month prior to biopsy-proven diagnosis. In addition, 3 cases of lupus membranous nephropathy had detectable THSD7A AB years before hypoalbuminemia and biopsy-proven diagnosis. Conclusions: While further studies are needed to explore the significance of THSD7A AB in lupus membranous nephropathy, this study describes a novel, highly sensitive LIPS immunoassay for detecting THSD7A AB and adds to the existing literature on THSD7A-associated MN.

Serologic Studies in Membranous Nephropathy: Novel Strategies and Strengthened Associations

“…Burbelo et al identified the presence of prediagnostic antibodies through screening of a large biorepository of blood samples donated by veterans with the Department of Defense serum repository. 1 The presence of circulating antibodies before transplantation in patients developing early disease recurrence provides further evidence of anti-THSD7A and PLA2R antibodies preceding disease manifestations. [7][8][9] Antibody titers for THSD7A and PLA2R correlate with disease activity, increasing with worsened proteinuria and falling with disease remission.…”

“…As such, the entire extracellular domain of THSD7A was used as a source of antigen with adequate expression and high sensitivity for antibody detection. 1 LIPS technology has been used for detection of antibodies of multiple autoimmune and infectious diseases S9-12 . It also provides the capability for multiplexing for testing panels of autoantibodies, such as antinuclear antibodies specificities S13 .…”

“…In this issue, Burbelo et al characterize the temporality of anti-THSD7A antibodies with disease activity and demonstrate evidence of seropositivity before disease onset in their original research article "Prediagnostic Appearance of THSD7A Autoantibodies in Membranous Nephropathy." 1 The authors also use a highly sensitive and novel technique of serologic detection through use of the luciferase immunoprecipitation system (LIPS), which may hold promise for the development of future serologic assays in MN and other autoimmune diseases.…”

“…Although it is well known that anti-PLA2R antibodies correlate with disease activity and disease onset, this is less established for circulating antibodies against minor MN autoantigens because of a lower frequency of positivity in disease, including thrombospondin type-1 domain-containing 7A (THSD7A). In this issue, Burbelo et al characterize the temporality of anti-THSD7A antibodies with disease activity and demonstrate evidence of seropositivity before disease onset in their original research article “ Prediagnostic Appearance of THSD7A Autoantibodies in Membranous Nephropathy .” 1 The authors also use a highly sensitive and novel technique of serologic detection through use of the luciferase immunoprecipitation system (LIPS), which may hold promise for the development of future serologic assays in MN and other autoimmune diseases.…”

The expanding spectrum and utility of antigens in membranous nephropathy

“…In primary MN, circulating autoantibodies traverse the GBM and bind endogenous podocyte antigens, resulting in subepithelial deposits. However, patients may have circulating antibodies for years prior to clinical presentation [73,74 ▪▪ ]. The dynamics underlying the sudden loss of immune tolerance is unclear but may be informed by factors such as titers of anti-PLA2R antibodies, the ability of antibodies to reach the subepithelial space, the PLA2R epitope specificity, and the ability to activate complement.…”