Background: Candidate von Willebrand factor (VWF) mutations were identified in 70% of index cases in the European study 'Molecular and Clinical Markers for the Diagnosis and Management of type 1 von Willebrand Disease'. The majority of these were missense mutations.
Objectives: To assess whether 14 representative missense mutations are the cause of the phenotype observed in the patients and to examine their mode of pathogenicity.
Methods: Transfection experiments were performed with full-length wild-type or mutant VWF cDNA for these 14 missense mutations. VWF antigen levels were measured, and VWF multimer analysis was performed on secreted and intracellular VWF.
Results: For seven of the missense mutations (G160W, N166I, L2207P, C2257S, C2304Y, G2441C, and C2477Y), we found marked intracellular retention and impaired secretion of VWF, major loss of high molecular weight multimers in transfections of mutant constructs alone, and virtually normal multimers in cotransfections with wild-type VWF, establishing the pathogenicity of these mutations. Four of the mutations (R2287W, R2464C, G2518S, and Q2520P) were established as being very probably causative, on the basis of a mild reduction in the secreted VWF or on characteristic faster-running multimeric bands. For three candidate changes (G19R, P2063S, and R2313H), the transfection results were indistinguishable from wild-type recombinant VWF and we could not prove these changes to be pathogenic. Other mechanisms not explored using this in vitro expression system may be responsible for pathogenicity.
Conclusions: The pathogenic nature of 11 of 14 candidate missense mutations identified in patients with type 1 VWD was confirmed. Intracellular retention of mutant VWF is the predominant responsible mechanism.