Biosynthetic and Functional Consequences of von Willebrand Disease Mutations

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DESCRIPTION (provided by applicant): Von Willebrand disease (VWD) is the most prevalent bleeding disorder in the world, affecting approximately 1% of the human population. It is divided into three general categories, types 1, 2 and 3. In type 2 VWD, most mutations result in replacement of a single amino acid in VWF. The functional consequences of these mutations are difficult to predict because of the complex nature of VWF synthesis and structure. The problem of type 2 VWD will be approached in this application with two specific aims. In Specific Aim 1, a variety of approaches are proposed to examine the effects of type 2B gain-of-function mutations on the disulfide bond structure of the VWF A1-A2-A3 region. These studies address the hypothesis that mutations that change the number of Cys residues or increase the negative charge in the region of the Cys1272-Cys1458 disulfide bond alter the disulfide-bond structure of the region. The disulfide-bond structure of 2B VWF mutants will be examined by mass spectrometry (MS) and correlated with functional alterations. In Specific Aim 2, plasma from a large number of well-characterized patients from a type 2 VWD kindred will be evaluated with a mathematical index of VWF quality and quantity that takes into account the percentage of mutant monomers incorporated into multimers, the relative content of multimers of different sizes, and the VWF quantity. This index will be correlated with VWF functional parameters and with the bleeding phenotype as quantified with a validated bleeding score questionnaire. These studies will be complemented with a longitudinal study of six patients with type 2B VWD (mutation R1308C) to compare the index with contemporaneously determined parameters of VWF function. It is expected that these studies will delineate both intra- and inter-individual variation in the VWF quality index. In aggregate, the proposed studies will shed light on the complex pathophysiology of type 2 VWD and have the potential to provide an accurate and quantitative assessment of bleeding risk in VWD patients.