Shaping the thrombotic cascade

Abstract

SUMMARY Venous thromboembolism is a common thrombotic disorder presenting complex clinical challenges, particularly regarding the optimal duration of anticoagulation therapy. Extending anticoagulation reduces the risk of recurrence but increases bleeding risk. Accurate prediction of recurrence risk could help guide decisions on whether to continue or stop anticoagulation. In Chapter 2, we investigated platelet and endothelial biomarkers and found that elevated levels of von Willebrand Factor during anticoagulation were associated with early recurrence, especially in men. These findings suggest VWF might be a useful biomarker for personalized risk prediction, enabling more tailored anticoagulation strategies if validated in further studies. Chapter 3 employed unbiased mass spectrometry to discover new biomarkers but did not find proteins that significantly differed between patients with and without recurrence, either during or after anticoagulation. Chapter 4 evaluated global coagulation assays focusing on thrombin generation and thrombin dynamics. We found that reduced endogenous thrombin potential (ETP) and reduced thrombin inhibition by antithrombin were linked to recurrence, particularly when excluding hormone-related VTE cases. Additionally, impaired thrombomodulin-mediated inhibition of thrombin generation was associated with recurrence risk. Chapter 5 shifted focus to patients with immune thrombocytopenia (ITP) , a patient group with heightened thrombotic risk. These patients showed elevated ETP, which increased further following eltrombopag treatment, highlighting a prothrombotic state. After establishing the importance of thrombin generation in predicting recurrence and assessing thrombotic state, Part II of the thesis examined thrombomodulin’s ability to modulate thrombin generation for diagnostic and therapeutic purposes, focusing on strategies to enhance or inhibit its activity. In Chapter 6, we developed liposomes conjugated with thrombomodulin (TM-liposomes) that more effectively inhibited thrombin generation in plasma and whole blood compared to soluble thrombomodulin. The lipid composition of these liposomes influenced their efficacy, with higher phosphatidylserine content enhancing protein C activation. These TM-liposomes may improve diagnostics by more closely mimicking the natural endothelial membrane environment and enabling better assessment of the activated protein C pathway in thrombin generation. Chapter 7 describes the design of a nanobody (VhH) that inhibits thrombin-activatable fibrinolysis inhibitor (TAFI) activation by thrombomodulin. We demonstrated that this VhH enhanced fibrinolysis in whole blood under flow conditions, offering a new strategy to enhance clot breakdown and possibly reduce thrombotic complications.