Sample conditions determine the ability of thrombin generation parameters to identify bleeding phenotype in FXI deficiency.

Individuals with Factor XI (FXI) deficiency have a variable bleeding tendency that does not correlate with FXI:C levels or genotype. Comparing a range of sample conditions, we tested whether the thrombin generation assay (TGA) could discriminate between control subjects (n = 50) and FXI-deficient individuals (n = 97), and between those with bleeding tendency (n = 50) and without (n = 24). The comparison used platelet-rich plasma (PRP) and platelet-poor plasma (PPP), either with or without corn trypsin inhibitor (CTI) to prevent contact activation, over a range of tissue factor (TF) concentrations. When contact activation was inhibited and platelets were absent, FXI:C levels did not correlate with thrombin generation parameters, and control and FXI-deficient individuals were not distinguished. In all other sample types, the best discrimination was obtained using TF 0.5 pM and assay measures: endogenous thrombin potential (ETP) and peak height. We showed that although a number of conditions could distinguish differences between the groups tested, TGA measured in PRP with CTI best differentiated between bleeders and nonbleeders. These measures provided high sensitivity and specificity (peak height receiver operating characteristic [ROC] area under the curve [AUC] = 0.9362; P < .0001) (ETP ROC AUC = 0.9362; P < .0001). We conclude that by using sample conditions directed to test specific pathways of FXI activation, the TGA can identify bleeding phenotype in FXI deficiency.

A novel deletion mutation is recurrent in von Willebrand disease types 1 and 3.

Direct sequencing of VWF genomic DNA in 21 patients with type 3 von Willebrand disease (VWD) failed to reveal a causative homozygous or compound heterozygous VWF genotype in 5 cases. Subsequent analysis of VWF mRNA led to the discovery of a deletion (c.221-977_532 + 7059del [p.Asp75_Gly178del]) of VWF in 7 of 12 white type 3 VWD patients from 6 unrelated families. This deletion of VWF exons 4 and 5 was absent in 9 patients of Asian origin. We developed a genomic DNA-based assay for the deletion, which also revealed its presence in 2 of 34 type 1 VWD families, segregating with VWD in an autosomal dominant fashion. The deletion was associated with a specific VWF haplotype, indicating a possible founder origin. Expression studies indicated markedly decreased secretion and defective multimerization of the mutant VWF protein. Further studies have found the mutation in additional type 1 VWD patients and in a family expressing both type 3 and type 1 VWD. The c.221-977_532 + 7059del mutation represents a previously unreported cause of both types 1 and 3 VWD. Screening for this mutation in other type 1 and type 3 VWD patient populations is required to elucidate further its overall contribution to VWD arising from quantitative deficiencies of VWF.

Inherited and de novo von Willebrand disease 'Vicenza' in UK families with the R1205H mutation: diagnostic pitfalls and new insights.

von Willebrand disease (VWD) caused by the R1205H mutation has distinct and reproducible clinical and laboratory features. This report describes the phenotypic and molecular investigation of seven kindreds with VWD Vicenza R1205H. All affected individuals have historically been diagnosed with moderate to severe type 1 VWD. Amongst all families with highly penetrant type 1 VWD investigated at our centre, heterozygosity for the R1205H mutation was found to be the most common underlying molecular defect. A severe laboratory phenotype associated with a bleeding history that was milder than expected was commonly observed, consistent with previous published case reports; however, abnormal ultralarge high molecular weight multimers were not detected in resting plasma samples. We also provide evidence that the R1205H mutation may arise de novo--evidence that a common genetic origin for this mutation is unlikely.

The prevalence of the cysteine1584 variant of von Willebrand factor is increased in type 1 von Willebrand disease: co-segregation with increased susceptibility to ADAMTS13 proteolysis but not clinical phenotype.

The molecular pathogenesis of type 1 von Willebrand disease (VWD) is uncertain in most patients. We examined 30 type 1 VWD families in the UK Haemophilia Centre Doctors' Organization study. Heterozygosity for Y/C1584 was present in eight of 30 (27%) families and 19 of 76 (25%) individuals with type 1 VWD recruited into the study. Eighteen (95%) of these 19 individuals were blood group O. C1584 did not co-segregate with VWD in four families, and co-segregated in one family; the results were equivocal in three families. In all families increased susceptibility of von Willebrand factor (VWF) to a disintegrin and metalloprotease with thrombospondin motifs (ADAMTS) 13 proteolysis co-segregated with C1584 in affected and unaffected individuals. These data show that C1584, associated with blood group O, is prevalent among patients with type 1 VWD but not necessarily causative of disease and should not be used in isolation to diagnose VWD. Increased susceptibility of C1584 VWF to ADAMTS13 proteolysis may be physiologically significant and increase an individual's risk of bleeding and presenting with VWD.