3 [4]. It spans approximately 178 kb and contains 52 exons that range in size from 1.3 kb (exon 28) to 40 bp (exon 50) [5]. Analysis of the VWF gene is complicated by at least two other factors in addition to size: (1) there is a partial pseudogene on chromosome 22 with 97% sequence homology to exons 23–34 that necessitates the use of carefully selected gene-specific PCR amplification primers for this region [6] and (2) the VWF locus is highly polymorphic (to date >150 polymorphisms have been reported) (http://www.vwf.group.shef.ac.uk). This makes direct VWF sequencing the methodology of choice for genetic analysis, given that mutation screening Ku 0059436 approaches such as conformation sensitive gel electrophoresis and denaturing

high performance liquid chromatography will be complicated LY2606368 nmr by the frequent sequence variants. The role of genetic testing for each of the current VWD subtypes (Types 1, 2A, 2B, 2M, 2N and 3 VWD), established by the International Society on Thrombosis and Haemostasis, will be reviewed below [7]. (1) Type 1 VWD, a partial deficiency of qualitatively normal VWF, represents the most common form of the disease and is the most problematic in terms of its diagnosis. The genetic

basis of Type 1 VWD has been the focus of much recent investigation and three large multicentre trials have reported consistent results on ∼300 families [11–13]. Mutations (predominantly missense) were identified in ∼65% of index cases and were found more frequently, and with higher penetrance, in cases with medchemexpress lower VWF levels. The most frequently reported genetic variation (10–20% of index cases) identified in all studies was a missense mutation resulting in an amino acid substitution of tyrosine to cysteine at codon 1584 (Y1584C) [14]. Importantly however, some Type 1 VWD patients had no obvious VWF mutation identified and in these (often milder)

cases, the genetic determinants are likely to be more complex and could involve other genetic loci. These studies have therefore confirmed prior suspicions that the genetic basis of this condition is highly variable. This genetic complexity precludes the use of molecular genetic testing as a complementary diagnostic aid in the majority of Type 1 VWD cases at the present time. Type 2A VWD accounts for ∼10% of all VWD cases and is characterized by the loss of high and intermediate molecular weight multimers. Type 2A VWD has been associated with more than 50 different missense mutations that result in two types of pathogenetic mechanisms: either aberrant VWF dimer or multimer biosynthesis (group I mutations) or the synthesis of a protein with enhanced susceptibility to A disintegrin-like and metalloprotease with thrombospondin type 1 results (ADAMTS13)-mediated proteolysis (group II mutations) [15,16]. In addition to providing further insights into VWF structure/function, genetic testing for Type 2A VWD can be employed when phenotypic uncertainty exists.