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Abstract: TH-PO652

Proteolytic Rewiring and Signaling through the Human Complement System

Session Information

Category: Glomerular Diseases

  • 1402 Glomerular Diseases: Clinical, Outcomes, and Therapeutics

Authors

  • Rinschen, Markus M., Aarhus Universitet, Aarhus, Midtjylland, Denmark
  • Lassé, Moritz, Universitatsklinikum Hamburg-Eppendorf, Hamburg, Hamburg, Germany
  • Thiel, Steffen, Aarhus Universitet, Aarhus, Midtjylland, Denmark
  • Hoxha, Elion, Universitatsklinikum Hamburg-Eppendorf, Hamburg, Hamburg, Germany
  • Melderis, Simon, Universitatsklinikum Hamburg-Eppendorf, Hamburg, Hamburg, Germany
  • Saleem, Moin A., University of Bristol Bristol Renal, Bristol, United Kingdom
  • Demir, Fatih, Aarhus Universitet, Aarhus, Midtjylland, Denmark
Background

Proteolysis is substantially involved during the manifestation of damage in the kidney in multiple diseases and is pivotal for exacerbation and resolution of inflammatory processes. The human complement system is a prime example for a clinically relevant proteolytic cascade. Systematic lupus erythematosus (SLE) is a severe chronic, inflammatory disease with renal complications, which affects mostly women (>80%).

Methods

We developed a high-throughput, automated analytical workflow for enrichment of N-termini from human EDTA-plasma through negative selection with the mass spectrometry-based “High-sensitivity Undecanal N-TERmini enrichment” (HUNTER) method. We mapped the enriched protease cleavage sites in a variety of clinically relevant populations, including complement inhibitor-treated individuals, SLE and lupus nephritis patients (cross-sectional and longitudinal). Furthermore, in vitro assays for selected proteases and specific activity assays for the human complement system were performed. In addition, we performed serum N-termini enrichment from stable-isotope labeled mice sera to identify proteolytic fragments, which have a lower turnover rate.

Results

We identified > 11 500 protease-generated N-termini in human plasma from more than 150 patients. Most of those N-termini did not correspond to already known fragments and a considerable fraction displayed a substantial alteration in SLE. Additionally, we could identify specific substrates for four central proteases of the human complement system and link this information to our identified N-termini from the human SLE cohorts. We also discovered GFR and proteinuria-associated cleavage products. Some of the identified protein N-termini were clearly linked to renal function, but also to the global activity of the human complement system and could pave the way for potential interventions in the future.

Conclusion

We could identify proteolytic processing events and determine functions of proteolytic modulation in SLE, with implications for lupus nephritis. The data suggest an unanticipated complexity of proteolytic processing throughout the human complement system.

Funding

  • Private Foundation Support