Two novel mutations, Q1053H and C1060R, located in the D3 domain of von Willebrand factor, are responsible for decreased FVIII‐binding capacity

Abstract

Summary. In type 2N von Willebrand disease (VWD), vonWillebrand factor (VWF) is characterized by a markedly decreased affinity for Factor VIII (FVIII), and the mutations responsible are essentially located in the D¢ domain of VWF. We report the identification, in seven unrelated French families, of two novel type 2N VWD mutations, Q1053H and C1060R (Gln290His and Cys297Arg in mature VWF sequence), in exon 24 of the VWF gene. These missense mutations have been identified in the heterozygous, homozygous or hemizygous states. Using site-directed mutagenesis and transient expression in COS-7 cells, we showed that both mutations, although located in the D3 domain of VWF, outside the tryptic fragment containing the FVIII domain, dramatically decrease the binding of VWF to FVIII. In contrast, the R924Q substitution, which was identified in a patient who was heterozygous for C1060R, was shown to be a polymorphism.

The mutational spectrum of type 1 von Willebrand disease: results from a Canadian cohort study

“…15,23,32 Additionally, the R924Q change is controversial. Although this variation was originally reported as a polymorphism, 33 it is currently listed on the ISTH SSC VWF Database (http://www.vwf.group.shef.ac.uk/index.html) 24 as a type 2N VWD mutation and is scored as a benign variant by both of the in silico protein algorithms used in this study. Similarly, the R1315C missense mutation is listed in the VWF Mutation Database 24 as either a type 2M or unclassified mutation.…”

Analysis of intracellular storage and regulated secretion of 3 von Willebrand disease–causing variants of von Willebrand factor

“…Consistent with this, it has been shown that rVWF-Cys1149Arg and rVWF-Cys1060Arg, which are involved in intrachain bonds, do not prevent formation of low and intermediate molecular weight VWF multimers. 17,32 To be able to fully assess the functioning of VWF, we believe that analysis of its ability to form WPBs, using expression of VWF in a heterologous system, should be part of a comprehensive study. The system should be such that expression of WT rVWF leads to formation of elongated organelles with internal striations that recruit appropriate membrane proteins and that respond to secretagogue.…”

“…Following electrophoresis, VWF was detected in the gel by means of rabbit antihuman VWF (Dako, Glostrup, Denmark) and visualized by means of alkaline phosphatase-conjugated swine antirabbit immunoglobulins (Dako, Denmark) and the AP Conjugate Substrate kit (Biorad, Hertfordshire, United Kingdom). 17 …”

Type 2N von Willebrand disease: Characterization and diagnostic difficulties

“…12 All the abovementioned mutations are located in the D' domain, but others have been reported in exon 17 13 or D3 domains. [14][15][16] Type 2N VWD patients are homozygotes or compound heterozygotes for different type 2N…”