Combined Use of Rapid Von Willebrand Factor (VWF) Activity,VWF-Propetide and Classical VWF Assays for Improved Diagnosisof Von Willebrand Disease Type 1, 2N and 2E Due to Mutations inthe D1, D2, D’, D3 and D4 Domains of the VWF Gene

Introduction: The Brno Antwerp and London VWD investigators used a complete set of Von Willebrand Factor (VWF) assays for the diagnosis and classification of Von Willebrand Disease (VWD) according to European Clinical Laboratory and Molecular (ECLM) criteria (Clinical Applied Thrombosis/ Hemostasis 2017; 23: 518). Aims: Thr Brno Antwerp London VWD investigators directly compared the rapid von Willebrand factor (VWF) assays VWF:GPIbM and VWF:GPIbR in Von Willebrand Disease (VWD) against the complete set of VWF assays using the ECLM criteria as the gold standard for VWD classification anno 2018. Between 2008 and 2018 we prospectively studied the Brno-Antwerp cohort with VWD type 1, 2N and 2 due to mutations in the D1, D2, D’and D3 domains of the vW gene. Methods: The complete set of rapid and classical VWF assays include Platelet Function Analyser Closure Time (PFA-CT) Von Willebrand Factor (VWF) Antigen (Ag), Ristocetine Cofactor activity (RCo), Collagen Binding (CB), Propeptide (pp), Ristocetine Induced Platelet Aggregation (RIPA), the rapid VWF activity assay VWF: GPIbM based on glycoprotein Ib (GPIb) binding to particles coated with G233V and M239V mutants in the absence of ristocetin, the rapid VWF: GPIbR assay in the presence of ristocetine, and the responses to DDAVP of FVIII: C and VWF parameters to pick up secretion and/or clearance defects of VWF. Results: The VWF: RCo/VWFAg, VWF: GPIbM/VWFAg and VWF: GPIbR/ VWF: Ag ratios are completely normal (above 0.7) in all variants of VWD type 1 and Low VWF. The VWF: RCo/VWF: Ag, GPIbR/VWF: Ag and GPIbM/VWF: Ag ratios vary around the cut off level of 0.70 in VWD due to multimerization defect in the D3 domain and therefore diagnosed as either type 1 E or type 2E VWD. Type 1 due to a heterozygous mutation in the D1 domain is featured by persistence of proVWF as the cause of VWF secretion/multimerization and FVIII binding defect mimicking VWD type 3 together with decreased values for VWFpp, VWFpp/Ag ratios. The majority of 22 different missense mutations in the D3 domain are of type 1 or 2 E multimerization defect usually associated with an additional secretion defect (increased FVIII: C/VWF: Ag ratio) and or clearance defect (increased VWFpp/Ag ratio). The majority of VWF mutations in the D4 and C1 to C6 are VWD type 1 SD with smeary (1sm) or normal (1m) multimers with no or a minor clearance defect. The heterozygous S2179F mutation in the D4 domain is featured by VWD type 1 Secretion and Clearance (SCD). The introduction of the rapid VWF:GPIbM or VWF:GPIbR assays as compared to the classical VWF: RCo assay did change VWD type 2 into type 1 in about 10 to 12%.