Novel VWF Sequence Variations Identified in Normal Controls and Index Cases Enrolled in the TS Zimmerman Program for the Molecular and Clinical Biology of VWD (ZPMCB-VWD).

Von Willebrand disease (VWD) is caused by quantitative (types 1 and 3) and qualitative (type 2) defects in von Willebrand factor (VWF). The TS Zimmerman Program for the Molecular and Clinical Biology of VWD is a multinational Program Project established to further the study of VWD in the United States and to contrast these studies with the studies initiated previously in the EU and Canada. As one of the components of this study we sought further insight into the clinical expression and penetrance of established types of VWD by performing full gene DNA sequence analysis in VWD patients and normal controls. This report is an interim report of the first 50 index cases and 113 normal individuals recruited into this study. Twenty four of these index cases were found to have known mutations, four of which had a second new mutation, and 11 cases had 1 or 2 new mutations. In cases where mutations were identified, 46% of the identified mutations were new mutations that have not been reported in the Sheffield VWF Mutation Database. In 15 patients, no mutations were identified in the coding region, although analysis of the non-coding regions is still in progress. Five of the mutations were deletions, insertions, or nonsense mutations that have clear functional consequences. The other 12 mutations were missense mutations. Since VWF polymorphisms are not well characterized in all exons, we have also completed studies of the first 113 normal control individuals in our study. These are individuals without a bleeding history and in whom full VWF laboratory testing and VWF sequencing was also undertaken. Since some estimates in the EU and Canadian studies have determined the prevalence of VWF mutations varies by the severity of type 1 VWD patients, we wanted to determine the frequency of VWF variation in a normal population and determine if sequence variations correlate with VWF levels. There were three linked common polymorphims identified in normal African Americans that are discussed elsewhere and are not included in this present analysis. We found 19 new sequence variations in the normal control group of which three (2900G>A, 6554G>A, 7997C>T) were found individually in 4–6% of the normal control samples. In addition, in 12 normal control samples we identified 6 sequence variations that were previously reported as VWF mutations. Four were reported as type 1 mutations (2220 G>A, 3686T>G, 3692A>C, 6859C>T) and two as type 2N mutations (2451T>A, 2771G>A). The 2220G>A and 2451T>A mutations were seen in 6 normal controls (5%) and 5 of these 6 normal controls had both mutations. In another normal control, both 3686T>G and 3692A>C were identified. Although the reported prevalence of VWD is 1% or greater, the frequency of these mutations in our normal controls is higher than expected (as high as 5%). In our normal control group, the mean VWF:Ag concentration in the patients with polymorphisms/mutations did not differ from the normal control group as a whole and did not cluster on the lower end of the normal range. Thus, the data on our normal individuals suggest that VWF gene variation is considerable and that many mutations and polymorphisms remain to be identified. Differentiation of those that affect the diagnosis of VWD and/or hemorrhagic risk continues to be difficult.