Anouk Donners

69 Comparing FVIII concentration and activity INTRODUCTION Haemophilia A is a hereditary bleeding disorder resulting from a deficiency or dysfunction of endogenous coagulation factor VIII (FVIII) with a prevalence of 1:5,000 male live births [1-3]. The International Society of Thrombosis and Haemostasis classifies the severity of haemophilia A based on the endogenous FVIII activity as severe (<1 IU/dL), moderate (1–5 IU/dL), or mild (>5–40 IU/dL), all three with a specific phenotype [4]. Patients with severe haemophilia (approximately 40% of haemophilia patients) have spontaneous or provoked bleeds in soft tissue and joints, causing arthropathy, impaired quality of life, and higher risks of intracranial haemorrhage or early death. Patients with moderate haemophilia, in contrast, are less affected but suffer from, for example, prolonged bleeding or easy bruising. Patients with mild haemophilia only experience bleeding problems during and after major trauma or surgery [5-7]. The standard of care in the developed regions with access to costly FVIII products, preferably entails an intravenous substitution of exogenous FVIII products based on disease severity and bleeding phenotype. Typically, severe patients receive regular prophylactic infusions with FVIII, and mild or moderate patients are treated in case of bleeding only (on-demand). The dose is often based on an individualised pharmacokinetic profile of a patient’s FVIII activity. To minimise bleeding risk and to prevent bleedings, many protocols aim at maintaining minimum trough levels of FVIII activity (>1 IU/dL) in patients with severe haemophilia [8]. The FVIII activity (>1 IU/dL) is currently used as a biomarker to assess disease severity and for monitoring treatment with FVIII products which is dependent on an accurate and precise quantification. The FVIII activity can be measured in clinical laboratories with the one-stage clotting assay (OSA) and/or the chromogenic assay (CSA). The OSA is based on the activated partial thromboplastin time (aPTT), making it easily automated, simple, fast, and inexpensive compared to CSA. The CSA is perceived to be more complex and technically challenging as a consequence of the two-stage principle with factor X activation and an additional chromogenic substrate step [9]. For diagnosing, it is recommended to perform multiple OSA measurements, to combine both OSA and CSA to ascertain the absence of discrepancies, or to evaluate the mutation profile of a patient [10]. Unfortunately, FVIII activity measuring has several limitations. Not only can these limitations result in misclassification of disease severity leading to under- or overestimation of the bleeding phenotype in specific subgroups, but also can result in suboptimal treatment monitoring of patients receiving FVIII replacement products [11, 12]. Both OSA and CSA are hampered by interference of different drugs (e.g., heparin, direct oral anticoagulants) and endogenous inhibitors such as lupus anticoagulant. Results from the assays are also affected by inter-laboratory variability, caused by the use of a wide variety of instruments, reagents, standards, and dilution algorithms [14, 15]. 4