A serologically assessed neo-epitope biomarker of cellular fibronectin degradation is related to pulmonary fibrosis

Annika Hummersgaard Hansen*, Helene Wallem Breisnes, Thomas Skovhus Prior, Ole Hilberg, Daniel Guldager Kring Rasmussen, Federica Genovese, Marie Vestergaard Lukassen, Birte Svensson, Lasse Løcke Langholm, Tina Manon-Jensen, Morten Asser Karsdal, Diana Julie Leeming, Elisabeth Bendstrup, Jannie Marie Bülow Sand

*Corresponding author for this work

Research output: Contribution to journalJournal articleResearchpeer-review

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Abstract

Background: Idiopathic pulmonary fibrosis (IPF) is characterized by excessive extracellular matrix (ECM) remodeling, herein ECM degradation. Fibronectin (FN) is an important component of the ECM that is produced by multiple cell types, including fibroblasts. Extra domain B (EDB) is specific for a cellular FN isoform which is found in the ECM. We sought to develop a non-invasive test to investigate whether matrix metalloproteinase 8 (MMP-8) degradation of EDB in cellular FN results in a specific protein fragment that can be assessed serologically and if levels relate to pulmonary fibrosis.

Method: Cellular FN was cleaved in vitro by MMP-8 and a protein fragment was identified by mass spectrometry. A monoclonal antibody (mAb) was generated, targeting a neo-epitope originating from EDB in cellular FN. Utilizing this mAb, a neo-epitope specific enzyme-linked immunosorbent assay (FN-EDB) was developed and technically validated. Serum FN-EDB was assessed in an IPF cohort (n = 98), registered at clinicaltrials.gov (NCT02818712), and in healthy controls (n = 35).

Results: The FN-EDB assay had high specificity for the MMP-8 degraded neo-epitope and was technically robust. FN-EDB serum levels were not influenced by age, sex, ethnicity, or BMI. Moreover, FN-EDB serum levels were significantly higher in IPF patients (median 31.38 [IQR 25.79–46.84] ng/mL) as compared to healthy controls (median 28.05 [IQR 21.58–33.88] ng/mL, p = 0.023).

Conclusion: We developed the neo-epitope specific FN-EDB assay, a competitive ELISA, as a tool for serological assessment of MMP-8 mediated degradation of EDB in cellular FN. This study indicates that degradation of EDB in cellular FN is elevated in IPF and warrants further investigation.
Original languageEnglish
Article number110599
JournalClinical Biochemistry
Volume118
Number of pages8
ISSN0009-9120
DOIs
Publication statusPublished - 2023

Keywords

  • Fibronectin
  • EDB
  • Neo-epitope
  • Extracellular matrix
  • Idiopathic pulmonary fibrosis
  • Biomarker
  • Serological biomarker
  • Non-invasive test

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