Diagnosis of von Willebrand disease

Summary. von Willebrand disease (vWD) is a bleeding disorder caused by quantitative or qualitative defects of von Willebrand factor (vWF). vWF is synthesized by endothelial cells and megakaryocytes and circulates in plasma as a multimeric high molecular weight glycoprotein. vWF plays a major role in the early phases of ostasis by promoting platelet-vessel wall and plateletplatelet interactions under high shear conditions. It is also the carrier of coagulation factor VIII (FVIII) in plasma. A deficiency of vWF results in impairment of both primary and secondary phases of ostasis. Therefore, patients with vWD manifest bleeding symptoms that are typical of defects of primary ostasis (mucocutaneous haemorrhages) but, in case of severe deficiency of vWF, there are also haemarthroses and haematomas, which are typical of those seen with coagulation defects. Several types and subtypes of vWD have been described with a high degree of heterogeneity. The diagnosis is based on measurements of plasma and platelet vWF, the ability of vWF to interact with its platelet receptor and the analysis of the multimeric composition of vWF. Due to the heterogeneity of vWF defects, a correct diagnosis of types and subtypes may be sometimes difficult but is very important for an appropriate treatment of patients with vWD.     

von Willebrand disease type IIC with different abnormalities of von Willebrand factor in the same sibship

A family with von Willebrand disease has been identified in which different members of the same sibship exhibit different abnormalities of von Willebrand factor (vWF). The two most severely affected sibs (bleeding time over 20 min) had abnormalities of vWF similar to those seen in type IIC. The smallest detectable multimer was increased and the triplet structure of individual multimers was replaced with a single band. The largest multimers could not be detected and there were relatively more small multimers than intermediate sized forms. vWF antigen (vWF:Ag) was decreased to 12.5-17% by electroimmunoassay (EIA) and to 3.2-5.5% by immunoradiometric assay (IRMA). In the less severely affected sibling (bleeding time 12.5 min) there was a similar relative increase in the smallest detectable multimer. However, the larger multimers were present and the relative concentration of large to small multimers was similar to normal. The triplet structure was altered in that the relative proportion of satellite bands to the central predominant band was decreased. vWF:Ag concentrations were moderately decreased (40-80% by EIA and 25-35% by IRMA). The father and grandfather showed a vWF multimeric pattern similar to the less severely affected sibling but there was no decrease in vWF:Ag concentration and their bleeding times were normal. These observations suggest that the interplay of several genetic factors is responsible for the expression of von Willebrand disease in this family.     

Clinical manifestations and complications of childbirth and replacement therapy in 385 Iranian patients with type 3 von Willebrand disease

Type 3 is the most severe form of von Willebrand disease (VWD) transmitted as an autosomal recessive trait. We collected data on the clinical manifestations of type 3 VWD by examining 385 patients from 300 Iranian kindreds, who were compared with 100 age-matched patients with severe haemophilia A. Joint and muscle bleeding was less frequent than in haemophiliacs, perhaps because factor VIII levels were in general higher (median value 4% vs. 1% or less). Mucosal tract haemorrhages such as epistaxis and menorrhagia were the most prevalent symptoms in VWD. Post-circumcision and oral cavity bleeding occurred frequently when prophylactic replacement therapy was not carried out or was inadequate. The course of pregnancy was usually uneventful, but increased bleeding occurred at parturition when affected women were treated with replacement therapy for less than 3-4 d. Ten of 385 (2.6%) of these multitransfused patients developed an alloantibody to VWF and 55% are chronically infected with the hepatitis C virus.     

Molecular Genetics of Type 2 von Willebrand Disease

Type 2 von Willebrand disease (VWD) is characterized by a wide heterogeneity of functional and structural defects. These abnormalities' cause either defective von Willebrand factor (VWF)-dependent platelet function in subtypes 2A, 2B, and 2M or defective VWF-factor VIII (FVIII) binding in subtype 2N. The diagnoses of types 2A, 2B, and 2M VWD may be guided by the observation of disproportionately low levels of ristocetin cofactor activity or collagen-binding capacity relative to VWF antigen. The abnormal platelet-dependent function is often associated with the absence of high molecular weight (HMW) multimers (type 2A, type 2B), but the HMW multimers may also be present (type 2M, some type 2B), and supranormal multimers may exist ("Vicenza" variant). The observation of a low FVIII-to-VWF:Ag ratio is a hallmark of type 2N VWD. in which the FVIII levels depend on the severity of the FVIII-binding defect. Today, the identification of mutations in particular domains of the pre-pro-VWF is helpful in classifying these variants and providing further insight into the structure-function relationship and the biosynthesis of VWF. Thus, mutations in the D2 domain, involved in the multimerization process, are found in patients with type 2A, formerly named IIC VWD. Mutations located in the D' domain or in the N terminus of the D3 domain define type 2N VWD. Mutations in the D3 domain characterize Vicenza and IIE patients. Mutations in the A1 domain may modify the binding of VWF multimers to platelets, either increasing (type 2B) or decreasing (type 2M, 2A/2M) the affinity of VWF for platelets. In type 2A VWD, molecular abnormalities identified in the A2 domain, which contains a specific proteolytic site, are associated with alterations in folding, impairing VWF secretion or increasing its susceptibility to proteolysis. Finally, a mutation localized in the carboxy-terminus CK domain, which is crucial for the dimerization of the VWF subunit, has been identified in a rare subtype 2A, formerly named IID.     

Clinical efficacy of highly purified, doubly virus-inactivated factor VIII/von Willebrand factor concentrate (Fanhdi®) in the treatment of von Willebrand disease: a retrospective clinical study

The goal of therapy in patients with von Willebrand disease (vWD) is to correct the dual defect of primary haemostasis and intrinsic coagulation reflected by low levels of von Willebrand factor (vWF) and factor VIII coagulant activity (FVIII:C). Factor VIII/von Willebrand factor (FVIII/vWF) concentrates are currently the treatment of choice in vWD patients unresponsive to desmopressin (DDAVP). However, only few studies on their clinical use are available so far. The main objective of this study was to retrospectively evaluate the clinical efficacy of a highly purified, doubly virus-inactivated FVIII/vWF concentrate with a high content of FVIII/vWF (Fanhdi). Twenty-two patients with congenital vWD have been treated from 1999 to 2001 at eight specialized centres belonging to the Italian Association of Hemophilia Centers (AICE). Ten males and 12 females, median age 28.5 years, range 5-70 years) had type 3 vWD (six cases), DDAVP-unresponsive type 1 (nine cases) and type 2B (seven cases). The study drug was given to stop or prevent 12 bleeding episodes or to prevent excessive bleeding during 14 surgical or invasive procedures. Overall, replacement therapy with the concentrate showed an excellent to good clinical efficacy in 92% of bleeding episodes and in 93% of surgical procedures. No adverse events occurred during 1,601 infusions, accounting for a total of 304,500 IU of FVIII:C administered. These results confirm the efficacy and safety of this concentrate in the management of bleeding episodes and in the prevention of excessive bleeding during major and minor surgery.     

Two novel mutations (Pro864His, Val867Glu) causing type 2A von Willebrand disease and affecting a single restriction site in exon 28

We detected two transversions in two unrelated Italian patients with type 2A von Willebrand disease (VWD): a C to A at nucleotide 8821 and a T to A at nucleotide 8830, resulting in the missense mutations Pro864His and Val867Glu respectively. Both mutations were in the heterozygous form and abolished the BstXI restriction site in exon 28 of the VWF gene. In both mutations plasma VWF multimer pattern improved by antiproteases. Moreover, DDAVP normalized plasma VWF multimers in the Pro864His patient, especially when protease inhibitors were present. These new mutations appear to be of the 2A VWD subtype due to the increased susceptibility to proteases.     

Evidence-based recommendations on the treatment of von Willebrand disease in Italy

Background

von Willebrand disease (VWD) is the most common hereditary bleeding disorder affecting both males and females. It arises from quantitative or qualitative defects of von Willebrand factor (VWF) and causes bleeding of mucous membranes and soft tissues. The aim of treatment is to correct the dual defect of haemostasis caused by the abnormal/reduced VWF and the concomitant deficiency of factor VIII (FVIII).

Material and methods

This document contains evidence-based recommendations for the management of VWD compiled by AICE (the Italian Association of Haemophilia Centres). All the evidence supporting these recommendations are based on non-randomised comparative studies or case series, because randomised controlled clinical trials or meta-analyses are not available for this disease.

Results and conclusions

Desmopressin (DDAVP) is the treatment of choice for patients with type 1 VWD with FVIII and VWF levels of 10 U/dL or more, while VWF/FVIII concentrates are indicated for those who are unresponsive or insufficiently responsive to DDAVP (severe type 1, type 2 and 3 VWD). VWF concentrates devoid of FVIII, not yet licensed in Italy, may be considered for short-term prophylaxis in elective surgery or for long-term secondary prophylaxis.     

When von Willebrand disease comes into age – a matter of change?

The population of elderly patients with von Willebrand Disease (VWD) is growing because of the improvement of medical care. The increase in comorbidities and their treatment as well as standard preventive measures like antiplatelet or anticoagulation therapy constitutes a challenge in the overall treatment of patients with coagulation disorders. Because of the lack of literature on older patients with VWD, we discuss different aspects related to ageing in those patients. Plasma levels of von Willebrand factor and factor VIII, bleeding symptoms, treatment requirements and comorbidities are possibly changing with age. Development of an evidence-based approach to the management of ageing VWD patients is therefore necessary.     

The molecular characterization of von Willebrand disease: good in parts

Since the cloning of the gene that encodes von Willebrand factor (VWF), 27 years ago, significant progress has been made in our understanding of the molecular basis of the most common inherited bleeding disorder, von Willebrand disease (VWD). The molecular pathology of this condition represents a range of genetic mechanisms, some of which are now very well characterized, and others that are still under investigation. In general, our knowledge of the molecular basis of type 2 and 3 VWD is now well advanced, and in some instances this information is being used to enhance clinical management. In contrast, our understanding of the molecular pathogenesis of the most common form of VWD, type 1 disease, is still at an early stage, with preliminary evidence that this phenotype involves a complex interplay between environmental factors and the influence of genetic variability both within and outside of the VWF locus.     

von Willebrand disease: still an intriguing disorder in the era of molecular medicine

von Willebrand disease (vWD) is a single-locus disorder resulting from a deficiency of von Willebrand factor (vWF): a multimeric multifunctional protein involved in platelet adhesion and platelet-to-platelet cohesion in high shear stress vessels, and in protecting from proteolysis and directing circulating factor VIII (FVIII) to the site of injury. vWD is the most frequent bleeding disorder, with an estimated prevalence in the general population of 1%. Almost all these cases are represented by a partial quantitative deficiency of von Willebrand factor (vWF) (type 1 vWD). Type 1 is transmitted as an autosomal dominant trait with an extremely variable penetrance and expressivity. A consensus figure for the prevalence of cases with significant bleeding symptoms, requiring some form of treatment, is approximately 100 cases million-1. Among these cases, more than 70-80% are represented by type 1 and respond to deamino-D-arginine vasopressin (DDAVP; desmopressin) administration. The remaining cases are represented by type 2 vWD (qualitative), some of which require substitutive treatment. Only 3-5 cases million-1 result from a total deficiency of vWF in plasma and platelets because of the recessive inheritance of two defective alleles. These cases may have severe bleeding episodes and may require frequent substitutive treatment. The molecular basis of type 2 (missense mutation in the functional domains of the vWF subunit) and type 3 (nonsense or large deletions) is quite well understood. On the contrary the molecular basis for most type 1 cases remains largely unknown, and many genetic factors (e.g. ABO blood group) and environmental or circumstantial factors (e.g. age, stress, drugs, pregnancy, and inflammation) are superimposed on to the genetic background determined by the vWF gene to produce a continuous spectrum from normality to mild type 1 cases. It is extremely difficult to make a clear distinction between mild type 1 cases and normal people because of the wide 'normal' range of laboratory measurements (e.g. length of bleeding time, and levels of vWF and FVIII) and of bleeding symptoms. Molecular testing is useless in this situation, and only good history-taking and repeated laboratory testing of vWF-related measurements in the propositus and his/her family members can help in clinical diagnosis, albeit imprecisely. This difficult task is the main focus of this review which is aimed at alerting the physician toward a balanced approach that should take into consideration both the risk of over- and under-diagnosis of this frequent disorder and the unavoidable production of a number of false positive and false negative cases.     

Von Willebrand factor testing ratios in the diagnosis and subtyping of von Willebrand disease

Von Willebrand disease (VWD) is a common bleeding disorder of platelet adhesion with six currently recognized subtypes. Laboratory diagnosis consists of an initial test panel including antigen, activity and factor VIII measurements, sometimes followed by further specialized testing. VWF activity/antigen testing ratios help to differentiate type 1 and type 2 disease, which is important for selection of proper therapy. Recommended ratio cutoffs differ by guideline, ranging from 0.5 to 0.7, with 0.7 commonly recommended. The ratio cutoff used affects the sensitivity and specificity for type 2 diagnosis. Variability in VWD due to underlying mutations and patient factors, as well as variability in VWF tests, impact the accuracy of ratios for VWD subtyping. This review discusses the use of activity/antigen ratios in the diagnosis and subtyping of VWD with a focus on technical aspects of the tests.     

A novel mutation (Leu817Pro) causing type 2A von Willebrand disease

We studied a patient affected by von Willebrand disease type 2A who experienced several mild bleeding episodes and was characterized by markedly reduced haemostatic parameters. In the exon 28 of von Willebrand factor (vWF) gene a T to C transition at nucleotide 8680, resulting in the missense mutation Leu817Pro, was found in the heterozygous form in the patient and in two affected relatives.
As suggested by the presence in platelets of a complete spectrum of vWF multimers as well as by the increased vWF antigen levels and improved haemostasis after DDAVP treatment, the mutation is compatible with normal multimerization, and could be responsible for a reduced stability or an impaired physiological secretion of vWF.     

Optimizing therapy with factor VIII/von Willebrand factor concentrates in von Willebrand disease

In von Willebrand disease, the main goals of treatment are to correct the dual defect of haemostasis caused by a reduced or abnormal von Willebrand factor (vWF), i.e. the prolonged bleeding time (BT) and the deficiency of factor VIII coagulant activity (FVIII:C). The synthetic vasopressin analogue, desmopressin (DDAVP), has reduced the need for transfusions in most of the mild forms of von Willebrand disease but DDAVP is ineffective in type 3 and in other severe cases of types 1 and 2 von Willebrand disease. For many years cryoprecipitate has been the mainstay of replacement therapy but, after the introduction of virucidal methods, concentrates containing FVIII/vWF have been considered much safer than cryoprecipitate and proposed in von Willebrand disease management. FVIII/vWF concentrates have been produced and tested by many authors but there is only one report describing four virus-inactivated FVIII/vWF concentrates evaluated in a cross-over randomized trial. According to these in vitro and pharmacokinetic data, the following information can be derived: (a) no FVIII/vWF concentrate had an intact multimeric structure similar to that of normal plasma or of cryoprecipitate; (b) all FVIII/vWF concentrates were equally effective in attaining normal and sustained levels of FVIII:C postinfusion, although peak levels were more delayed in the concentrate devoid of FVIII:C; (c) no FVIII/vWF concentrate consistently normalized the BT in a sustained fashion. On the other hand, clinical haemostasis can be achieved in the management of bleeding episodes and of surgery for most of von Willebrand disease cases regardless of whether the BT is corrected; in the few rare cases with mucosal bleeding not controlled by FVIII/vWF concentrates, infusion of DDAVP or platelet concentrates can be administered in addition.     

Multimeric pattern discrepancy between platelet and plasma von Willebrand factor in type IIC von Willebrand disease

von Willebrand factor (vWF) from platelet lysate and plasma, collected in the presence of protease inhibitors, was studied in two patients with type IIC von Willebrand disease (vWD). Platelet and plasma vWF showed the smallest multimer increased, but the latter had a repeating single band whereas the former had a repeating "doublet." This platelet-plasma discrepancy observed for the first time in these patients suggests that the repeating "doublet" or single band described in other type IIC patients represent minor subgroups of type IIC vWD.     

A novel mutation (Leu817Pro) causing type 2A von Willebrand disease

We studied a patient affected by von Willebrand disease type 2A who experienced several mild bleeding episodes and was characterized by markedly reduced haemostatic parameters. In the exon 28 of von Willebrand factor (vWF) gene a T to C transition at nucleotide 8680, resulting in the missense mutation Leu817Pro, was found in the heterozygous form in the patient and in two affected relatives. As suggested by the presence in platelets of a complete spectrum of vWF multimers as well as by the increased vWF antigen levels and improved haemostasis after DDAVP treatment, the mutation is compatible with normal multimerization, and could be responsible for a reduced stability or an impaired physiological secretion of vWF.     

New developments in von Willebrand disease

Von Willebrand disease (VWD) constitutes the most common inherited human bleeding disorder. It is associated with a mucocutaneous bleeding phenotype that can significantly impact upon quality of life. Despite its prevalence and associated morbidity, the diagnosis and subclassification of VWD continue to pose significant clinical challenges. This is in part attributable to the fact that plasma von Willebrand factor (VWF) levels vary over a wide range in the normal population, together with the multiple different physiological functions played by VWF in vivo. Over recent years, substantial progress has been achieved in elucidating the biological roles of VWF. Significant advances have also been made into defining the pathophysiological mechanisms underpinning both quantitative and qualitative VWD. In particular, several new laboratory assays have been developed that enable more precise assessment of specific aspects of VWF activity. In the present review, we discuss these recent developments in the field of VWD diagnosis, and consider how these advances can impact upon clinical diagnostic algorithms for use in routine clinical practice. In addition, we review some important recent advances pertaining to the various treatment options available for managing patients with VWD.