PB0198 - A novel plasminogen activator anti-fibrin-uPA increases the sensitivity of the plasmin generation assay to regulators of fibrinolysis

Sunday, June 23, 2024

13:45 – 14:45 ICT

Room: Exhibition Hall

Presenting Author(s)

Ilaria De Simone, PhD

PostDoc
Synapse Research Institute
Maastricht, Limburg, Netherlands

Co-Author(s)

KP

Kieran Pocknell

PhD student
Aberdeen Cardiovascular and Diabetes Centre, School of Medicine, Medical Sciences and Nutrition, Institute of Medical Sciences, University of Aberdeen, Aberdeen, UK
Aberdeen, Scotland, United Kingdom
ST

Samia Tufaha

student
Synapse Research Institute
Maastricht, Limburg, Netherlands
Joke Konings, PhD

Senior Scientist
Synapse Research Institute
Maastricht, Limburg, Netherlands
Mark Roest

PhD, Clinical Director
Synapse Research Institute
Maastricht, Limburg, Netherlands
Alisa S. Wolberg, PhD

Professor
University of North Carolina
Chapel Hill, North Carolina, United States
Claire S. Whyte, PhD

PhD, Advanced Research Fellow
Aberdeen Cardiovascular & Diabetes Centre, Institute of Medical Sciences, School of Medicine, Medical Sciences & Nutrition, University of Aberdeen
Aberdeen, Scotland, United Kingdom
Bd

Bas de Laat

General Director
Synapse Research Institute
Maastricht, Limburg, Netherlands
Nicola J. Mutch, MSc Hons, PhD

Professor
Aberdeen Cardiovascular & Diabetes Centre, Institute of Medical Sciences, School of Medicine, Medical Sciences & Nutrition, University of Aberdeen
Aberdeen, Scotland, United Kingdom
Dana Huskens, PhD

PhD, Senior Research Project Leader
Synapse Research Institute
Maastricht, Limburg, Netherlands

Background: Pharmacological modifiers of fibrinolysis are used to modulate bleeding and thrombosis. There is a need for a sensitive plasmin generation (PG) assay, to study the regulatory mechanisms in fibrinolysis and to identify abnormal fibrinolytic states, and monitor antifibrinolytic medication.

Aims: To establish reference values of PG parameters and study different regulators of fibrinolysis, using a novel calibrated, automated PG assay.

Methods: A calibrated, automated method was used to assess PG in the plasma of 120 healthy individuals, normal pooled plasma (NPP), or factor deficient plasma, with a plasmin-specific substrate. Trigger solution consisted of phospholipids, tissue factor (TF) or Russell's viper venom-factor X activator (RVV-X), together with tissue plasminogen activator (tPA) or the novel urokinase plasminogen activator anti-fibrin-uPA (aFn-uPA).

Results: Reference values for PG parameters were established in plasma from healthy individuals. Median plasmin peaks (% normalized to NPP) induced with TF/tPA, TF/aFn-uPA, RVV-X/tPA and RVV-X/aFn-uPA, were 97.9(85.9-110.0), 99.8(91.0-114.6), 96.1(86.8-107.4), 97.7(88.5-108.3), respectively. Plasmin was not generated in plasminogen or fibrinogen deficient plasma unless supplemented with plasminogen or fibrinogen, respectively. In NPP, plasminogen activator inhibitor 1 (PAI-1), thrombomodulin (TM), alpha 2-antiplasmin (α2-AP) and tranexamic acid (TXA) reduced PG. Interestingly, the PG assay became sensitive to PAI-1 when induced with aFn-uPA, an effect abolished using an anti-PAI-1 antibody (Figure). Furthermore, the inhibitory effect of TM in PG, was counteracted using carboxypeptidase inhibitor (CPI), an inhibitor specific to thrombin-activatable fibrinolysis inhibitor (TAFI), confirming a TAFI-dependent role of TM in PG.

Conclusion(s): The calibrated, automated PG assay is a specific and efficient tool to assess modulators of fibrinolysis, and has potential as a clinical tool to identify abnormal fibrinolytic profiles in patients, and monitor antifibrinolytic drugs, such as TXA. This assay is particularly effective when aFn-uPA is used as trigger for fibrinolysis.