CHAPTER 135 VON WILLEBRAND DISEASE
CHAPTER 135 VON WILLEBRAND DISEASE
Definition and History
Etiology and Pathogenesis
The Von Willebrand Factor Gene and Cdna
Von Willebrand Factor Biosynthesis
The Function of Von Willebrand Factor
Molecular Genetics of Von Willebrand Disease
Von Willebrand Factor Antigen
Ristocetin Cofactor Activity
Ristocetin-Induced Platelet Agglutination (Ripa)
Dna Diagnosis of Von Willebrand Disease
Platelet-Type (Pseudo-) Von Willebrand Disease
Acquired Von Willebrand Disease
Therapy, Course, and Prognosis
Von Willebrand Factor Replacement Therapy
Other Nonreplacement Therapies
von Willebrand factor (vWF) is a central component of hemostasis, serving both as a carrier for factor VIII and as an adhesive link between platelets and the injured blood vessel wall. Abnormalities in vWF function result in von Willebrand disease (vWD), the most common inherited bleeding disorder in humans. The overall prevalence of vWD has been estimated to be as high as 1 percent of the general population, although the prevalence of clinically significant disease is probably closer to 1:1000. vWD is associated with either quantitative deficiency (type 1 and type 3) or qualitative abnormalities of vWF (type 2). The uncommon type 3 variant is the most severe form of vWD and is characterized by very low or undetectable levels of vWF, a severe bleeding diathesis, and a generally autosomal recessive pattern of inheritance. Type 1 vWD, the most common variant, is characterized by vWF that is normal in structure and function but decreased in quantity (in the range of 20 to 50 percent of normal). In type 2 vWD, the vWF is abnormal in structure and/or function. Type 2A vWD is associated with selective loss of the largest and most functionally active vWF multimers. Type 2A is further subdivided into group 1, due to mutations that interfere with biosynthesis and secretion, and group 2, in which the mutant vWF exhibits an increased sensitivity to proteolysis in plasma. Type 2N vWD is characterized by mutations within the factor VIII binding domain of vWF, leading to disproportionately decreased factor VIII and a disorder resembling mild hemophilia A, but with autosomal recessive inheritance. Type 2B vWD is due to mutations clustered within the vWF A1 domain, in a segment critical for binding to the platelet glycoprotein Ib (GPIb) receptor. These mutations produce a “gain of function” resulting in spontaneous vWF binding to platelets and clearance of the resulting platelet complexes, leading to thrombocytopenia and loss of the most active (large) vWF multimers. Type 1 vWD can often be effectively managed by treatment with DDAVP, which produces a two- to threefold increase in plasma vWF level. Response to DDAVP is generally poor in type 3 and most of the type 2 vWD variants. These disorders often require treatment with factor replacement in the form of plasma or selected factor VIII concentrates containing large quantities of intact vWF multimers.
Acronyms and abbreviations that appear in this chapter include: aPPT, activated partial thromboplastin time; DDAVP, 1-desamino-8-D-arginine vasopressin, or desmopressin; ER, endoplasmic reticulum; GPIb, glycoprotein Ib; HHT, hereditary hemorrhagic telangiectasia; PCR, polymerase chain reaction; RIPA, ristocetin-induced platelet aggregation; vWD, von Willebrand disease; vWF, von Willebrand factor.
DEFINITION AND HISTORY
In 1926, Eric von Willebrand described a bleeding disorder in 24 of 66 members of a family from the Åland Islands.1 Both sexes were afflicted, and the bleeding time was prolonged despite normal platelet counts and normal clot retraction. von Willebrand distinguished this condition from the other hemostatic diseases known at the time and recognized its genetic basis, calling the disorder “hereditary pseudohemophilia,” but incorrectly characterizing the inheritance as X-linked dominant. von Willebrand’s confusion about the inheritance pattern was probably due, at least in part, to the greater recognition of bleeding symptoms in women because of the hemostatic stresses of menstruation and parturition. The proband in the original family, Hjördis, was 5 years old at the time of von Willebrand’s initial evaluation and ultimately died at age 13 during her fourth menstrual cycle. Four of Hjördis’ sisters died between the ages of 2 and 4, and deaths in the family were also noted during childbirth.
An apparently similar disorder was independently reported in the United States by Minot and others in 1928. The original family in the Åland Islands was reexamined by von Willebrand and Jürgens in 1933, leading to the conclusion that the defect in this disorder was due to an impairment of platelet function. It was not until 1953 that Alexander and Goldstein demonstrated reduced levels of coagulation factor VIII in vWD patients, along with prolonged bleeding time. This observation was confirmed by others, including studies of the original von Willebrand pedigree by Nilsson and coworkers. In the late 1950s, the latter group demonstrated that a fraction of plasma referred to as “I-O” could correct the factor VIII deficiency and normalize the bleeding time, indicating that the defect in vWD was due to the deficiency of a plasma factor rather than an intrinsic platelet abnormality. Infusion of fraction I-O promptly increased the factor VIII level in a hemophilic patient, while in vWD the factor VIII level rose gradually, peaking at 5 to 8 h. Fraction I-O prepared from a hemophilia A patient was also shown to correct the defect in vWD, demonstrating that these disorders were due to deficiencies of distinct plasma factors (reviewed in ref.2 and 3).
It was not until 1971 that Zimmerman, Ratnoff, and Powell prepared the first antibodies against what was thought to be a highly purified form of factor VIII.4 This factor VIII–related antigen was found to be normal in hemophilia A patients but decreased in vWD. This puzzle was finally resolved with the demonstration that vWF and factor VIII are closely associated, with over 98 percent of the mass of the complex composed of vWF (see below). Thus, antibodies raised against this complex predominantly recognize vWF. The first direct assay of vWF function was based on the observation that the antibiotic ristocetin induced thrombocytopenia and the demonstration by Howard and Firkin5 that ristocetin-induced platelet aggregation was absent in some vWD patients. Weiss and coworkers6 used this observation to develop a quantitative assay for vWF function that remains a mainstay of laboratory evaluation for vWD to this day. In 1973, several groups succeeded in dissociating vWF from factor VIII procoagulant activity.7,8
Final proof that vWF and factor VIII are independent proteins encoded by distinct genes came with the cDNA cloning of the two molecules in 1984 and 1985.9,10,11,12,13 and 14 These discoveries also marked the beginning of the molecular genetic era for the study of vWF and factor VIII, leading to the identification of gene mutations in many patients with hemophilia and vWD as well as considerable insight into the structure and function of these related proteins.
Table 135-1 summarizes the current nomenclature and terminology for factor VIII and vWF. vWD is a heterogenous disorder with over 20 distinct variants described. The previous complex and confusing classification has recently been consolidated and simplified,15,16 as summarized in Table 135-2. Type 3 vWD is associated with very low or undetectable levels of vWF and severe bleeding. Type 1 vWD is characterized by concordant reductions in factor VIII activity, vWF antigen, and ristocetin cofactor activity, generally to the range of 20 to 50 percent of normal, in association with normal vWF multimer structure. Type 2 vWD is heterogeneous and further divided into four subtypes (2A, 2B, 2N, and 2M). Type 2A vWD is characterized by a disproportionately low level of ristocetin cofactor activity relative to vWF antigen and absence of large and intermediate-sized multimers. Type 2B vWD is also associated with reduced high-molecular-weight vWF multimers, but as the result of an abnormal vWF molecule with increased affinity for platelet GPIb. Abnormalities in vWF that result in decreased factor VIII binding to vWF have also been described (type 2N) and present as mild to moderate factor VIII deficiency. Many other subtypes have been reported, including platelet-type (pseudo-) vWD, which is actually an intrinsic platelet disorder due to mutations in GPIb (see Chap. 119). Finally, acquired forms of vWD also occur, generally due to autoantibody formation.
TABLE 135-1 VON WILLEBRAND FACTOR AND FACTOR VIII TERMINOLOGY
TABLE 135-2 CLASSIFICATION OF VON WILLEBRAND DISEASE
ETIOLOGY AND PATHOGENESIS
vWF is synthesized exclusively in endothelial cells and megakaryocytes and performs two major functions in hemostasis. First, vWF serves as the initial critical bridge between circulating platelets and the injured blood vessel wall, accounting for the apparent defect in platelet function and prolonged bleeding time observed in vWD patients. The vWF monomer is assembled into higher-order multimers, a structure required for optimal adhesive function. Second, vWF serves as the carrier in plasma for factor VIII, ensuring its stability and localizing it to the initial platelet plug for participation in thrombin generation and fibrin clot formation (see Chap. 112). This tight, noncovalent interaction between vWF and factor VIII accounts for the copurification of these two molecules and the resulting initial confusion as to the origin of hemophilia and vWD. Factor VIII is encoded by the factor VIII gene on the X chromosome (see Chap. 112 and Chap. 123), while vWF is encoded by a distinct gene on human chromosome 12.
THE VON WILLEBRAND FACTOR GENE AND cDNA
The vWF cDNA was initially cloned from endothelial cells11,12,13 and 14 and the corresponding gene mapped to the short arm of chromosome 12 (12p13.3).11 The vWF mRNA is approximately 9.0 kb in length, encoding a primary translation product of 2813 amino acid residues with an estimated Mr of 310,000. Comparison of the primary peptide sequence obtained from plasma vWF17 with the vWF cDNA sequence established the pre-propolypeptide nature of vWF.18 Pre-propolypeptide vWF is composed of a 22–amino acid signal peptide, a 741–amino acid precursor polypeptide (propeptide) termed vWF antigen II, and the mature subunit.11,18,19,20 and 21 Cleavage of the 741–amino acid propeptide from the amino terminus produces the mature vWF subunit of 2050 amino acids (Fig. 135-1).
FIGURE 135-1 Schematic diagram of the human vWF gene, mRNA, and protein. The vWF gene and pseudogene are depicted at the top, with boxes representing exons and the solid black line introns. The vWF mRNA encoding the full prepro-vWF subunit is depicted in the middle as the bar and lettered boxes. The locations of signal peptide (sp) and propeptide (Pro) cleavage sites are indicated by arrowheads, and the lettered boxes denote regions of internally repeated sequence. The approximate localizations for known vWF functional domains within the mature vWF subunit are indicated at the bottom. Numbers underneath the domains refer to amino acid residues within the mature vWF subunit. The clusters of mutations responsible for type 2A, type 2B, and type 2N vWD are indicated. (aa, amino acids; chr, chromosome.) (Adapted from D Ginsburg and EJW Bowie222 with permission.)
Analysis of the vWF sequence identifies four distinct types of repeated domains: three A domains, three B domains, two C domains, and four D domains.19,22 The first pair of D domains is tandemly arranged in the vWF propeptide, followed by a partial and full D domain at the N terminus of the mature subunit. The final complete D domain is separated by a segment of more than 600 amino acids containing the triplicated A domains. The repeated domain structure of vWF suggests that the gene may have evolved via a complex series of partial duplications, although exon structure is not highly conserved between homologous domains.
Comparison of the vWF amino acid sequence to other proteins identifies a superfamily of related proteins that all share sequence similarity with the vWF A domains.23 The common theme among these potentially evolutionarily related genes is a role in extracellular matrix or adhesive function. Consistent with this notion, vWF functional domains for binding to the platelet receptor GPIb and specific ligands within the extracellular matrix have been localized to the vWF A repeats. A potential relationship between the vWF C domains and portions of thrombospondin and procollagen has also been proposed.24
The vWF gene spans approximately 180 kb and is divided into 52 exons.25 Exons range in size from 40 bases to 1.5 kb (exon 28). The latter exon is unusually large, encoding the entire Al and A2 domains and containing nearly all of the known type 2A and type 2B vWD mutations. The concentration of these defects within one exon has facilitated the identification of human mutations responsible for these vWD variants (see Molecular Genetics of von Willebrand Disease, below). A partial, nonfunctional duplication of the vWF gene, termed a pseudogene, has been localized to human chromosome 22.26 The pseudogene duplicates the middle portion of the vWF gene, from exons 23 to 34, and includes the intervening sequences. The pseudogene is approximately 97 percent identical in sequence to the authentic vWF gene, indicating that it is of fairly recent evolutionary origin.27
vWF is synthesized exclusively in megakaryocytes and endothelial cells and, as a result, has frequently been used as a specific histochemical marker to identify cells of endothelial cell origin. Although generally assumed to mark all endothelial cells, vWF is expressed at widely varying levels among endothelial cells, depending on the size and location of the associated blood vessel.28,29 A recent careful survey in the mouse identified wide differences in the level of vWF mRNA, with 5 to 50 times higher concentrations in the lung and brain, particularly in small vessels, than in comparable vessels in the liver and kidney. In general, the higher levels of vWF mRNA and antigen were found in the endothelial cells of large vessels rather than in microvessels and in venous rather than arterial endothelial cells.29
Specific DNA sequences within or near the proximal promoter of the vWF gene appear to be required for endothelium-specific gene expression,30 although it is likely that additional important regulatory elements exist outside of this region, perhaps at a great distance. A portion of the human vWF promoter from –487 to +246 has been shown to target vWF expression to blood vessels of the yolk sac and a subset of endothelial cells in the adult brain of the mouse.31 This heterogeneity in expression level among different endothelial cell subsets has only recently been appreciated.32
VON WILLEBRAND FACTOR BIOSYNTHESIS
The processing steps involved in the biosynthesis of vWF are similar in megakaryocytes33 and endothelial cells34,35 and 36 (reviewed in ref. 37). vWF is first synthesized as a large, precursor monomer polypeptide, depicted schematically in Fig. 135-1. vWF is unusually rich in cysteine, which accounts for 8.3 percent of its amino acid content. All cysteines in the mature vWF molecule are involved in disulfide bonds.38 Pro-vWF monomers are assembled into dimers through disulfide bonds at both C termini, and only dimers are exported from the endoplasmic reticulum (ER).38,39 and 40
Glycosylation begins in the ER, with 12 potential N-linked glycosylation sites present on the mature subunit and 3 on the propeptide. Extensive additional posttranslational modification of vWF occurs in the Golgi apparatus, including the addition of multiple O-linked carbohydrate structures, sulfation, and multimerization through the formation of disulfide bonds at the N termini of adjacent dimers. vWF is the only protein known to undergo extensive disulfide bond formation at this late stage, and this unique process appears to be catalyzed by a novel disulfide isomerase activity present within the vWF propeptide.41 Mutations at either of two specific cysteines within the propeptide that are thought to be critical for disulfide isomerase activity, or a shift in the spacing between them, results in loss of multimer formation.41 The multimerization process appears to require the slightly acidic environment of the distal Golgi.42 The vWF propeptide self-associates and may also serve to align vWF subunits for multimer assembly.43 However, the propeptide facilitates multimer assembly even when coexpressed as a separate molecule from the mature vWF monomer.44,45
Propeptide cleavage occurs late in vWF synthesis or just prior to secretion. Cleavage occurs adjacent to two basic amino acids, Lys-Arg at positions –2 and –1. An Arg at position –4 is also required for recognition by the intracellular protease responsible for propeptide cleavage.46 Multimerization and propeptide cleavage are not linked to each other. The multimers secreted by cultured endothelial cells contain both pro-vWF and mature subunits,36,47 and recombinant vWF with a point mutation inhibiting propeptide cleavage is still assembled into normal multimer structures.48 Although propeptide cleavage appears to occur primarily intracellularly, cleavage may also occur after secretion.
vWF is secreted from endothelial cells via both constitutive and regulated pathways.37 vWF is stored in tubular structures within the a granule of platelets and within the Weibel-Palade body in endothelial cells.49,50,51 and 52 Weibel-Palade bodies are derived from the Golgi apparatus and are found in most endothelial cells, though the number varies considerably. Although a number of other hemostatic proteins are also stored in the platelet a granule, the Weibel-Palade body appears to be relatively specific for vWF and its propeptide.53,54 The transmembrane glycoprotein P-selectin is also found in the membranes of both the a granule and the Weibel-Palade body.55 The only other known component of the Weibel-Palade body is CD63, a lysosomal protein also found on activated platelets.56,57
Regulated secretion of vWF from its storage site in the Weibel-Palade body is triggered by a number of secretagogues, including thrombin,58 fibrin,59 histamine,60 and the C5b-9 complement complex.61 While the vasopressin analog desmopressin acetate (DDAVP) causes marked release of vWF in vivo, it has no direct effect on endothelial cells in culture,62 suggesting that its effects are controlled by a secondary mediator. Constitutive secretion of vWF occurs evenly at the apical and basolateral surface, while regulated secretion from the Weibel-Palade body is highly polarized in the basolateral direction.63 While constitutively secreted multimers are of relatively small size, the multimers stored within the Weibel-Palade body are the largest, most biologically potent form.54,64 The vWF stored in platelet a-granules is also enriched for large multimers.65 The N-terminal D domains appear to be required for vWF storage, with deletion of any of the individual domains resulting in constituent secretion.66,67 It also appears that cleavage of the vWF prosequence is required for efficient formation of storage granules.68
The concentration of vWF in plasma is approximately 10 µg/ml, with approximately 15 percent of circulating vWF localized to the platelet compartment.69 Bone marrow transplants between normal and vWD pigs demonstrate that platelet vWF is derived entirely from synthesis within the marrow and does not contribute to the normal plasma vWF pool.70,71 and 72 These studies also demonstrate that both the plasma and the platelet vWF pools are required for full hemostasis, although the plasma pool appears to be more critical.
Plasma vWF appears to be further processed through cleavage by a specific protease in the circulation, resulting in reduction in the size of the largest multimers.73,74 The major proteolytic cleavage site has been mapped to the peptide bond between Tyr842 and Met843 in the vWF A2 domain,75 and recombinant vWF missing the A2 domain is resistant to proteolysis.76 vWF carrying a subgroup of type 2A vWD mutations exhibits increased susceptibility to cleavage by this protease,77 and this is the proposed mechanism for the selective loss of large vWF multimers in this group of patients (see Molecular Genetics of von Willebrand Disease, below). Recently, this specific vWF processing protease activity has been proposed to play a central role in the pathophysiology of chronic relapsing thrombotic thrombocytopenic purpura (see Chap. 117).78,79 and 80
THE FUNCTION OF VON WILLEBRAND FACTOR
vWF is a large multivalent adhesive protein that plays an important role in platelet attachment to subendothelial surfaces, platelet spreading, and platelet-platelet interactions, that is, aggregation at sites of vessel injury. vWF also stabilizes factor VIII. The interaction of vWF and factor VIII is important for the protection of factor VIII from inactivation or degradation. Factor VIII bound to vWF may localize to cells and/or sites where it can more readily participate in the promotion of blood coagulation and/or thrombus formation.
vWF is required for the adhesion of platelets to the subendothelium, particularly at moderate to high shear force. vWF performs this bridging function by binding to two platelet receptors, GPIb and GPIIb/IIIa, as well as to specific ligands within the exposed subendothelium at sites of vascular injury (reviewed in ref. 81,82 and 83). Binding of vWF to its platelet receptors generally does not occur in the circulation under normal conditions. However, the interaction of vWF with its ligands in the vessel wall, combined with high shear stress conditions, facilitates vWF binding to platelet GPIb and subsequent platelet adhesion and activation. Activation of platelets leads to the exposure of the GPIIb/IIIa complex, an integrin receptor that can bind to fibrinogen and other ligands, including vWF, to form the platelet-platelet bridges required for thrombus growth. Platelet adhesion to vWF immobilized at a site of injury appears to be a two-step process, with the initial tethering of the rapidly moving platelet dependent on the vWF/GPIb interaction and subsequent firm adhesion occurring through GPIIb/IIIa after platelet activation.84,85
VON WILLEBRAND FACTOR BINDING TO THE VESSEL WALL
vWF binds to several different types of collagens, including types I through VI. Two distinct binding domains for the fibrillar collagens, types I and III, have been localized to specific segments within the vWF A1 and A3 repeats (see Fig. 135-1),86,87 and a potential third domain has been identified in the propeptide.88 Studies of recombinant vWF suggest that the A3 collagen-binding domain may be the most important.89,90 The physiologic relevance of vWF interactions with fibrillar collagens has been questioned, since vWF still binds to extracellular matrix depleted of these molecules by treatment with collagenase.91 vWF has also been shown to bind to the nonfibrillar collagen type VI, which is resistant to collagenase92 and colocalizes with vWF in the subendothelium.93 Type VI collagen supports the binding of vWF under high shear through cooperative interactions between binding domains within the vWF A1 and A3 repeat.94 Although vWF binding has also been demonstrated in a number of other potential components of the subendothelium, including glycosaminoglycans95,96 and sulfatides,97 the biologic significance of these interactions remains to be demonstrated.
VON WILLEBRAND FACTOR BINDING TO PLATELETS
vWF interacts with a receptor complex on the surface of platelets composed of the disulfide-linked GPIba and GPIbb chains noncovalently associated with GPIX and GPV. The binding site for vWF is within a 293–amino acid segment at the N terminus of GPIb and requires sulfation of several key tyrosine residues for optimal binding.98 The GPIb binding domain within vWF has been mapped to the A1 segment, within the disulfide loop formed between the cysteine residues at 509 and 695 (see Fig. 135-1).99,100 Ristocetin binds to both vWF and platelets, but the mechanism by which it enhances the vWF/GPIb interaction is still poorly understood.101,102 The snake venom botrocetin appears to induce GPIb binding through a different alteration in the vWF A1 domain and is also used to study this interaction.103 Scanning mutagenesis studies of recombinant vWF have characterized a number of amino acid residues within the vWF A1 domain that are critical for binding to GPIb and for interaction with botrocetin.104 Several mutations were also identified that increase platelet binding, an effect similar to that of mutations associated with type 2B vWD (see Molecular Genetics of von Willebrand Disease, below). These natural and synthetic mutations cluster in a small area on the surface of the vWF A1 domain structure, as revealed by x-ray crystallographic studies.105 The structure of the A1 domain closely resembles that of other previously studied A domains, including the vWF A3 domain.106,107
The structure of the vWF A1 domain and the mutations that cause enhanced GPIb binding, together with the observations that these mutations enhance binding to similar extents and are not additive when inserted into the same molecule, suggest an allosteric “on-off” model for vWF function.83,108 In this model, the vWF GPIb binding domain is in an “off” configuration when at rest in the circulation. Binding to collagen (or another ligand in the vessel wall) or interaction with soluble modulators such as ristocetin or botrocetin induces a switch to the “on” conformation, resulting in platelet binding. In an alternative model, it is the immobilization of vWF on a surface, together with high shear force, that facilitates the multivalent interaction of vWF with the platelet surface, rather than a specific conformational switch within the vWF A1 domain.82,109
The Arg-Gly-Asp-Ser (RGDS) sequence at amino acids 1744–1747 of the mature vWF subunit serves as the binding site within vWF for GPIIb/IIIa. The latter complex, also known as aIIb/b3, is a member of the integrin family of cell surface receptors. GPIIb/IIIa undergoes a conformational change to a high-affinity ligand-binding state following platelet activation and, in addition to vWF, can bind a number of other adhesive proteins, including fibrinogen. Although vWF is present in blood at much lower concentrations than is fibrinogen, evidence suggests that vWF may be a critical ligand under flow conditions.84,85 An RGD sequence is also present in the vWF propeptide (vWF antigen II), although its functional significance is unknown.
THE INTERACTION OF VON WILLEBRAND FACTOR WITH FACTOR VIII
The noncovalent interaction between factor VIII and vWF is required for the stability of factor VIII in the circulation, as is evident from the factor VIII levels of less than 10 percent that are observed in most severe vWD patients. Although each vWF subunit appears to carry a binding site for factor VIII, the stoichiometry for the vWF/FVIII complex found in normal plasma is approximately 1 to 2 factor VIII molecules per 100 vWF monomers.110 Factor VIII bound to vWF is also protected from proteolytic degradation by activated protein C (reviewed in ref. 83 and 111).
The factor VIII binding domain within vWF has been localized to the first 272 N-terminal amino acids of the mature subunit,112 with antibody studies suggesting a particularly critical role for amino acids 78–96.113,114 The mutations identified in patients with type 2N vWD, in which vWF binding to factor VIII is specifically affected (see Molecular Genetics of von Willebrand Disease, below), are all clustered in this region, including the most common type 2N mutation, at Arg91.115 It is noteworthy that the same amino acid substitution at Arg89 is a common polymorphism that does not affect factor VIII binding.116 The corresponding binding site for vWF on factor VIII includes an acidic region at the N terminus of the light chain (residues 1669–1689)117 and requires sulfation of Tyr1680 for optimal binding.118 Thrombin cleavage after Arg1689 activates and releases factor VIII from vWF. Thus, vWF may serve to efficiently deliver factor VIII to the sites of clot formation, where it can complex with factor IXa on the platelet surface.
MOLECULAR GENETICS OF VON WILLEBRAND DISEASE
vWD is an extremely heterogenous and complex disorder, with over 20 distinct subtypes reported (reviewed in ref. 83,119 and 120). A large number of mutations within the vWF gene have now been identified (Fig. 135-2). A partial list is maintained by a consortium of vWD investigators and can be accessed through the internet at http://mmg2.im.med.umich.edu/vWF.121,122 These findings form the basis for the simplified classification of vWD outlined in Table 135-215,16 and used throughout this chapter. Types 1 and 3 vWD are defined as pure quantitative deficiencies of vWF that are either partial (type 1) or complete (type 3). Type 2 vWD is characterized by qualitative abnormalities of vWF structure and/or function. The quantity of vWF found in type 2 vWD may be normal, but it is usually mildly to moderately decreased (see Table 135-2).
FIGURE 135-2 vWD mutations. The location of all point mutations and small, in-frame insertions and/or deletions associated with vWD, as reported to the vWD database (http://mmg2.im.med.umich.edu/vWF), are depicted within the vWF coding sequence. Shown below are the relative positions of all 52 vWF gene exons. (Adapted from WC Nichols and D Ginsburg119 with permission.)
TYPE 3 VON WILLEBRAND DISEASE
Patients with type 3 vWD have very low or undetectable levels of plasma and platelet vWF antigen and ristocetin cofactor activity and generally present early in life with severe bleeding.123 Factor VIII coagulant activity is markedly reduced but usually detectable at levels of 3 to 10 percent of normal. Type 3 vWD appears to be inherited as an autosomal recessive trait in most families, but parents of affected individuals may have mildly reduced vWF levels and are occasionally given the diagnosis of mild type 1 vWD.
Southern blot analysis has identified gross gene deletion as the molecular mechanism for type 3 vWD in only a small subset of families26,124,125 and 126; however, large deletions may confer an increased risk for the development of alloantibodies against vWF.26,126 A similar correlation has been reported for hemophilia B (see Chap. 123). Comparative analysis of vWF genomic DNA and platelet vWF mRNA has identified nondeletion defects resulting in complete loss of vWF mRNA expression as a molecular mechanism in some patients with type 3 vWD.127,128 A number of nonsense and frameshift mutations that would be predicted to result in loss of vWF protein expression or in expression of a markedly truncated or disrupted protein have been identified in some type 3 vWD families (see Fig. 135-2).119,121,129,130 A frameshift mutation in exon 18 appears to be a particularly common cause of type 3 vWD in the Swedish population and has been shown to be the defect responsible for vWD in the original Åland Island pedigree.131,132 This mutation results in a stable mRNA encoding a truncated protein that is rapidly degraded in the cell.133 This mutation also appears to be common among type 3 vWD patients in Germany134 but not in the United States.133
TYPE 1 VON WILLEBRAND DISEASE
Type 1 is the most common form, accounting for approximately 70 percent of vWD patients. Type 1 vWD is generally autosomal dominant in inheritance and is associated with coordinate reductions in factor VIII, ristocetin cofactor activity, and vWF antigen with maintenance of the full complement of multimers (Fig. 135-3). Subgroups within type 1 vWD have been proposed based on the relative levels of vWF present in the plasma and platelet pools.135,136,137 and 138
FIGURE 135-3 Agarose gel electrophoresis of plasma vWF. vWF multimers from plasma of patients with various subtypes of vWD are shown. The brackets to the left encompass three individual multimer subunits, including the main band and its associate satellite bands. N indicates normal control lanes. Lanes 5 through 7 are rare variants of type 2A vWD. The former designations for these variants are indicated in parentheses below the lanes (IIC through E). (Adapted from SD Berkowitz et al223 with permission.)
Type 1 vWD is generally assumed to simply represent the heterozygous form of type 3 vWD. However, the majority of heterozygous carriers of vWF gene deletions, as well as carriers of vWF mRNA expression defects,26,124,125,127,128 are asymptomatic and have normal vWF laboratory values, consistent with an autosomal recessive pattern of inheritance for type 3 vWD. Nonetheless, in some families with nonsense or frameshift mutations, heterozygotes with apparent type 1 vWD have been identified, indicating that some or all type 1 vWD may be due to such defects within the vWF gene (see Fig. 135-2). Mutations that give rise to defective vWF subunits that interfere in a dominant negative way with the normal allele may be particularly likely to cause symptomatic vWD in the heterozygote.130 Mutations have been identified at several cysteine residues in the vWF D3 domain of patients with moderately severe type 1 vWD. vWF carrying one of these mutations was shown to be retained in the ER and appeared to exert a dominant negative effect on the normal vWF allele.139
To date, all mutation studies and genetic linkage analysis of type 1 vWD have been consistent with defects within the vWF gene (reviewed in ref. 130). However, the possibility that a subset of type 1 vWD is due to defects in genes outside of vWF (locus heterogeneity) must still be considered. Given the complex biosynthesis and processing of vWF, defects at a number of other loci could be expected to result in quantitative vWF abnormalities. However, no such example has yet been reported. It is interesting to note that a mouse model for type 1 vWD associated with an up to twentyfold reduction in plasma vWF is due to an unusual mutation in a glycosyltransferase gene, leading to aberrant posttranslation processing of vWF and accelerated clearance from plasma.140 A similar mechanism may explain the modifying effect of the ABO blood group glycosyltransferases on plasma vWF level.141
TYPE 2A VON WILLEBRAND DISEASE
Type 2A is the most common qualitative variant of vWD and is generally associated with autosomal dominant inheritance and selective loss of the large and intermediate vWF multimers from plasma (see Fig. 135-2). A 176-kDa proteolytic fragment present in normal individuals is markedly increased in quantity in many type 2A vWD patients. This fragment is due to proteolytic cleavage of the peptide bond between Tyr842 and Met843.75,142 Based on this observation, initial DNA sequence analysis in patients centered on vWF exon 28, in the region encoding this segment of the vWF protein, leading to the identification of the first point mutations responsible for vWD.143 Since that time, a large number of mutations have been identified, accounting for the majority of type 2A vWD patients.119 Most of these mutations are clustered within a 134–amino acid segment of the vWF A2 domain (between Gly742 and Glu875; see Fig. 135-3), and the most common, Arg834Trp, appears to account for about one-third of type 2A vWD patients.119,121
Expression of recombinant vWF containing type 2A vWD mutations has identified two distinct molecular mechanisms for the loss of large vWF multimers characteristic of this disorder.144 In the first subset, classified as group 1, the type 2A vWD mutation results in a defect in intracellular transport, with retention of mutant vWF in the ER. In the second subset, or group 2, mutant vWF is normally processed and secreted in vitro, and thus loss of multimers in vivo presumably occurs due to increased susceptibility to proteolysis in plasma.75,144,145,146 and 147 As noted above, the protease that appears to be responsible for this cleavage has been identified.73,74,77
The multimer structure of platelet vWF correlates well with this subclassification. Group 1 patients show loss of large vWF multimers within platelets due to defective synthesis, while group 2 patients have normal vWF multimers within the protected environment of the a granule.144 These observations confirm the earlier subclassification of type 2A vWD based on platelet multimers.135 Subclassification into group 1 or 2 might be expected to predict response to desmopressin therapy, although this remains to be demonstrated.
In addition to the major class of type 2A vWD described above, a number of rare variants previously classified as types IIC–H, type IB, and “platelet discordant” are now included in the new, more general type 2A category. Most of these rare variants were distinguished on the basis of subtle differences in the multimer pattern (see Fig. 135-3; reviewed in ref. 120). The IIC variant is usually inherited as an autosomal recessive trait and is associated with loss of large multimers and a prominent dimer band. Several mutations have been identified in the vWF propeptide of these patients,148,149 presumably interfering with multimer assembly. A mutation at the C terminus of vWF, interfering with dimer formation, was recently described in a patient with the IID variant.150 Most of the other reported variants of type 2A vWD are quite rare, often limited to single case reports.
TYPE 2B VON WILLEBRAND DISEASE
Type 2B vWD is usually inherited as an autosomal dominant disorder and is characterized by thrombocytopenia and loss of large vWF multimers. The plasma vWF in type 2B vWD binds to normal platelets in the presence of lower concentrations of ristocetin than does normal vWF and often binds spontaneously. Accelerated clearance of the resulting complexes between platelets and the large, most adhesive forms of vWF accounts for the thrombocytopenia and the characteristic multimer pattern (see Fig. 135-3).
The peculiar functional abnormality characteristic of type 2B vWD suggested a molecular defect within the GPIb binding domain of vWF. For this reason, initial DNA sequence analysis focused on the corresponding portion of vWF exon 28.151,152 Nearly all of these mutations are located within the vWF A1 domain, at one surface of the recently described crystallographic structure.105 The four most common mutations are clustered within a 35–amino acid stretch between Arg543 and Arg578 (see Fig. 135-2); together, these account for more than 80 percent of type 2B vWD patients.121 Functional analysis of mutant recombinant vWF108,153,154,155 and 156 confirms that these single–amino acid substitutions are sufficient to account for increased GPIb binding and the resulting characteristic type 2B vWD phenotype.
Three families have been described that exhibit enhanced vWF binding to GPIb but a normal distribution of vWF multimers. These variants, previously referred to as type I New York, type I Malmö, and type I Sydney, are now all designated as type 2B vWD. Type I New York and type I Malmö have now been shown to be due to the same mutation, Pro503Leu. This mutation is located within the cluster of type 2B mutations in the vWF A1 domain and results in a similar increase in platelet GPIb binding.157
TYPE 2N VON WILLEBRAND DISEASE
As described in Chap. 123, hemophilia A results from defects in the factor VIII gene and is inherited in an X-linked recessive manner. Rare families have been reported in which the inheritance of hemophilia appears to be autosomal, based on the occurrence of affected females or direct transmission from an affected father.158,159 Several cases of an apparent autosomal recessive decrease in factor VIII have been shown to be due to decreased binding of factor VIII by vWF.160,161 and 162 This disorder has also been referred to as vWD Normandy, after the province of origin of the first patient. DNA sequence analysis has identified a total of 11 mutations associated with this disorder, all located at the vWF N terminus (see Fig. 135-2).115,120 One of these mutations, Arg91Gln, appears to be particularly common and may contribute to variability in the severity of type 1 vWD in some cases.163
TYPE 2M VON WILLEBRAND DISEASE
This category is reserved for rare vWD variants in which a defect in vWF platelet-dependent function leads to significant bleeding but vWF multimer structure is not affected (although some have subtle multimer abnormalities). Most of these variants were previously classified as type I. The variant previously referred to as type B is associated with absent ristocetin cofactor activity but normal platelet binding with other agonists. This variant has been shown to be due to a mutation in the A1 domain (Gly561Ser).164 Mutations have also been identified in a number of other families with normal vWF multimers and disproportionately decreased ristocetin cofactor activity.120,165 Several families have been described with a vWD variant (vWD Vicenza) characterized by larger than normal vWF multimers.166 Although the mutation responsible for this disorder has not been identified, genetic linkage analysis indicates that the defect lies within the vWF gene.167
Type 1, the most common form of vWD, is generally transmitted as an autosomal dominant disorder and accounts for approximately 70 percent of clinically significant vWD. However, disease expressivity is variable, and penetrance is incomplete.130 Laboratory values and clinical symptoms can vary considerably, even within the same individual, and establishing a definite diagnosis of vWD is often difficult. In two large families with type 1 vWD, only 65 percent of individuals with both an affected parent and an affected descendent had significant clinical symptoms.168 For comparison, 23 percent of the unrelated spouses of the patients, who presumably did not have a bleeding disorder, were judged to have a positive bleeding history.
A number of factors are known to modify vWF levels, including ABO blood group, Lewis antigen, estrogens, thyroid hormone, age, and stress.169,170 ABO blood group is the best characterized of these factors. Mean vWF antigen levels for type O individuals are approximately 75 percent and for type AB individuals 123 percent when compared to a pool of normal donor plasmas. Thus, it may be difficult to differentiate between a low-normal value and mild type 1 vWD in blood group O individuals.169 The variable expressivity and incomplete penetrance of type 1 vWD has complicated the determination of accurate incidence figures for vWD. The prevalence of type 1 vWD has been estimated to be as high as 1 percent and as low as 3 to 4 per 100,000.171,172
In general, the type 2 variants are more uniformly penetrant. Type 2A and type 2B vWD account for the vast majority of patients with qualitative vWF abnormalities. No accurate incidence figures are available for these subtypes, but the type 2 variants are generally felt to comprise 20 to 30 percent of all vWD diagnoses. The type 2 variants are generally autosomal dominant in inheritance, although rare cases of apparent recessive inheritance have been reported.
Estimates of prevalence for severe (type 3) vWD range from 0.5 to 5.3 per 1,000,000.173,174 and 175 Although this variant is frequently defined as autosomal recessive in inheritance, this is not a consistent finding. As described above, one or both parents of a severe vWD patient are frequently clinically asymptomatic and often have entirely normal laboratory test results, but many families have also been reported in which one or both parents appear to be affected with classic type 1 vWD. Thus, in some families, severe vWD may represent the homozygous form of type 1 vWD. In this model, the apparent recessive inheritance in a subset of families could simply be the result of the incomplete penetrance of type 1 vWD. Alternatively, there may be a fundamental difference in the molecular mechanisms responsible for type 1 and type 3 vWD.130
Mucocutaneous bleeding is the most common symptom in patients with type 1 vWD.168 It is important to note that over 20 percent of normal individuals may give a positive bleeding history.176 This observation, together with the limited sensitivity and specificity of the currently available laboratory tests (see below), makes the diagnosis of mild vWD quite difficult and probably contributes to the wide range of prevalence figures for type 1 vWD currently in the literature.
Epistaxis occurs in approximately 60 percent of type 1 vWD patients, 40 percent have easy bruising and hematomas, 35 percent have menorrhagia, and 35 percent have gingival bleeding. Gastrointestinal bleeding occurs in approximately 10 percent of patients.177 An apparent association between hereditary hemorrhagic telangiectasia (HHT) and vWD had been reported in several families. Genetic defects causing HHT have been localized to chromosomes 3, 9, and 12 (see Chap. 123), and thus most cases are unlikely to be linked to the vWF gene on chromosome 12. However, since inheriting vWD is likely to increase the severity of bleeding from HHT, the diagnosis is more likely to be made in patients inheriting both defects.178 Mucocutaneous bleeding is common after trauma, with about 50 percent of patients reporting bleeding after dental extraction, about 35 percent after trauma or wounds, 25 percent postpartum, and 20 percent postoperatively. Spontaneous atraumatic hemarthroses occur almost exclusively in patients with type 3 vWD. Hemarthroses in patients with moderate disease are extremely rare and are generally only encountered after major trauma.
Patients with type 3 vWD can suffer from severe clinical bleeding and experience hemarthroses and muscle hematomas, as in severe hemophilia A (see Chap. 123). The bleeding time is very prolonged. After infusion of vWF-containing plasma fractions, some of these patients develop anti-vWF antibodies that neutralize vWF. Development of antibodies has been correlated with the presence of gene deletions.26,126
The bleeding symptoms can be quite variable among patients within the same family and even in the same patient over time. An individual may experience postpartum bleeding with one pregnancy but not with others, and clinical symptoms in mildly to moderately affected type 1 individuals often ameliorate by the second or third decade of life. Aside from an infrequent type 3 patient, death from bleeding rarely occurs in vWD.
Thrombocytopenia is a common feature of type 2B vWD and is not seen in any other form of vWD. Most patients only experience thrombocytopenia at times of increased vWF production or secretion, such as during physical effort, in pregnancy, in newborn infants, postoperatively, or if an infection develops. The platelet count rarely drops sufficiently to contribute to clinical bleeding.179,180 Infants with type 2B vWD may present with neonatal thrombocytopenia, which could be confused with neonatal sepsis or congenital thrombocytopenia.
Patients who are homozygous or compound heterozygous for type 2N vWD generally have normal levels of vWF antigen and ristocetin cofactor activity and normal vWF platelet adhesive function. However, factor VIII levels are moderately decreased, resulting in a mild to moderate hemophilia-like phenotype.115 However, in contrast to patients with classic hemophilia A (factor VIII deficiency), these patients do not respond to infusion of purified factor VIII and should be treated with vWF-containing concentrates. Heterozygotes for this disorder may have mildly decreased factor VIII levels but are generally asymptomatic. Although type 2N vWD appears to be considerably less common than classic hemophilia A, it should be considered in the differential diagnosis of factor VIII deficiency, particularly if any features suggest an autosomal pattern of inheritance. Although the factor VIII level rarely drops below 5 percent, at least one type 2N vWD mutation has been associated with factor VIII levels as low as 1 percent, when coinherited with a type 3 vWD allele.181 The latter observation suggests that a diagnosis of type 2N vWD should also be considered in patients with marked reductions of factor VIII.
In the initial laboratory evaluation of patients suspected by history of having vWD, the following tests are routinely performed: assay of factor VIII activity, vWF antigen (vWF:Ag), and ristocetin cofactor activity. In a large epidemiologic study, the ristocetin cofactor assay was found to be more sensitive than the vWF:Ag for the diagnosis of type 1 vWD.182 Other tests that are commonly used include the bleeding time, ristocetin-induced platelet aggregation (RIPA), and vWF multimer analysis. As noted above, results of these tests can all be normal in some patients with type 1 vWD. In addition, the wide range of normal and the considerable overlap with the levels observed in type 1 vWD make borderline levels difficult to interpret. A variety of concurrent diseases and drugs may modify the results of individual tests, including aspirin or other nonsteroidal anti-inflammatory drugs, which often prolong the bleeding time. Many conditions, such as pregnancy, time of the menstrual cycle, hypo- or hyperthyroidism, uremia, recent exercise, liver disease, infection, diabetes, estrogen therapy, or myeloproliferative syndromes, affect the factor VIII activity, vWF:Ag, and ristocetin cofactor activity levels. These values can be regarded as acute-phase reactants, and many minor illnesses can increase their levels to normal. Even controlling for many of these factors, the coefficients of variation of repeated vWF:Ag and ristocetin cofactor assays in a single person are quite large.183 For this reason, repeated measurements are usually necessary, and the diagnosis of vWD or its exclusion should not be based on a single set of laboratory values unless they are well below or well above the limits of normal.
The bleeding time has long been used as a standard screening test for vWD and other abnormalities of platelet function.184 However, results can vary considerably with the experience of the operator and a variety of other factors, and its value as a screening test has been questioned. There is now a general consensus that the bleeding time should not be used for routine patient screening in the preoperative setting.185,186 and 187 While the bleeding time should also probably not be used as a routine screening test for vWD, it may still be of value in selected patients when taken together with the clinical history and the results of other laboratory tests. It may also be useful as a means of monitoring therapy in some settings.
Factor VIII levels in vWD patients are generally coordinately decreased along with plasma vWF. Levels in type 3 vWD generally range from 3 to 10 percent. In contrast, the levels in type 1 and the type 2 vWD variants (other than 2N) are variable and usually only mildly or moderately decreased. The factor VIII level in type 2N vWD is more severely decreased, but rarely to less than 5 percent. The activated partial thromboplastin time (aPTT) can be prolonged in vWD, although only as a reflection of the reduced factor VIII level.
VON WILLEBRAND FACTOR ANTIGEN
Plasma vWF:Ag is usually quantitated by electroimmunoassay, radioimmunoassay, or an ELISA technique. In type 1 vWD, the vWF:Ag assay usually parallels the ristocetin cofactor activity, but it has lower specificity and sensitivity than the ristocetin cofactor assay. In patients with type 2A vWD, the vWF:Ag is usually low but can be normal.183
RISTOCETIN COFACTOR ACTIVITY
The standard measure of vWF activity quantitates the ability of plasma vWF to agglutinate platelets in the presence of ristocetin,188 also referred to as the ristocetin cofactor assay. Normal platelets washed free of plasma vWF are used either as fresh platelets or after formaldehyde fixation. This assay appears to be the most sensitive and specific single test for the detection of vWD.182 While it is generally decreased coordinately with vWF:Ag and factor VIII in type 1 vWD patients, ristocetin cofactor activity is usually disproportionately decreased in the type 2A variants, due to the greater dependence of the latter assay on the larger vWF multimers.
A specific assay of FVIII binding to vWF has been developed and is used to confirm the diagnosis of type 2N vWD.189 Although this assay is widely used in European hemostasis laboratories, its availability in the United States is currently limited to a few specialized reference laboratories.
A number of other assays for vWF activity have been proposed, including measurement of platelet agglutination induced by botrocetin and other snake venom proteins,190 assays based on collagen binding,191 and a new functional assay that measures platelet binding under high shear.192 While the latter device shows some promise, none of these assays is currently available in the routine clinical laboratory.
RISTOCETIN-INDUCED PLATELET AGGLUTINATION (RIPA)
The addition of ristocetin to normal platelet-rich plasma causes platelet clumping. This activity is generally reduced in most vWD patients. Hyperresponsiveness to ristocetin-induced platelet agglutination results either from a type 2B vWD mutation or an intrinsic defect in the platelet (platelet-type or pseudo-vWD). In these disorders, patient platelet-rich plasma agglutinates spontaneously or at ristocetin concentrations of only 0.2 to 0.7 mg/ml. At these concentrations, normal platelet-rich plasma does not agglutinate. Type 2B and platelet-type vWD can be distinguished by RIPA experiments performed with separated patient platelets or plasma mixed with the corresponding component from a normal individual.
Analysis of plasma vWF multimers is critical for the proper diagnosis and subclassification of vWD (see Fig. 135-3). This is generally accomplished by agarose gel electrophoresis of plasma vWF to separate vWF multimers on the basis of molecular size, with the largest multimers migrating more slowly than the intermediate or smaller multimers. The multimers may be visualized by autoradiography after incubation with 125I-monospecific anti-human vWF antibody or by nonradioactive immunologic techniques. The normal multimeric distribution is an orderly ladder of major protein bands of increasing molecular weight, going from the smallest to the largest vWF multimers (see Fig. 135-3). Each normal multimer has a fine structure consisting of one major component and two to four satellite bands.193 Type 2B and most of the type 2A variants were initially distinguished from each other on the basis of subtle variations in the satellite band pattern.
Given the mild clinical phenotype of most patients with the common variants of vWD, prenatal diagnosis for the purpose of deciding on terminating the pregnancy is rarely performed. However, type 3 vWD patients often have a profound bleeding disorder, similar to or more severe than classic hemophilia, and so some families may request prenatal diagnosis. In those cases of vWD in which the precise mutation is known, DNA diagnosis can be performed rapidly and accurately by PCR from amniotic fluid or chorionic villus biopsies.194 In those cases where the mutation is unknown, diagnosis can still be attempted by genetic linkage analysis using the large panel of known polymorphisms within the vWF gene.122 One of these polymorphisms, a TCTA tetranucleotide repeat of variable length in intron 40, is particularly useful, with over 100 known polymorphic alleles. Several cases of successful prenatal diagnosis have been reported.194,195 and 196 Although all cases of vWD analyzed to date appear to be linked to the vWF gene, the possibility of locus heterogeneity (i.e., a similar phenotype due to a mutation in a gene other than vWF) should be considered.130
DNA DIAGNOSIS OF VON WILLEBRAND DISEASE
With advances in understanding the molecular genetics of vWD, it is now possible to precisely diagnose and subclassify many variants of vWD on the basis of specific DNA mutations identified in the research laboratory. Unfortunately, DNA testing for vWD is not currently available in the clinical setting. As molecular testing is gradually introduced into the clinical laboratory, DNA diagnosis should be particularly straightforward for type 2B vWD, where a panel of four mutations detects over 80 percent of patients. Similar panels of mutations should be able to correctly identify the defect in the majority of type 2A and type 2N vWD. The analysis of type 3 and type 1 vWD will be more complex, since the currently known mutations account only for a small subset of these patients, except in selected populations.131
PLATELET-TYPE (PSEUDO-) VON WILLEBRAND DISEASE
Platelet-type (pseudo-) vWD is a platelet defect that phenotypically mimics vWD (see Chap. 119).197 The plasma vWD lacks the largest multimers, RIPA is enhanced at low concentrations of ristocetin, and thrombocytopenia of variable degree is often present. Clinically, these patients have primarily mucocutaneous bleeding. Molecular analysis has identified mutations within the GPIba chain as the molecular basis for pseudo-vWD. These mutations are located within the segment of GPIb thought to encode the vWF binding domain and appear to induce the conformational change complementary to that produced in the corresponding fragment of vWF by type 2B vWD mutations.197
The specialized RIPA test should be performed at low ristocetin concentrations to distinguish type 2B and platelet type vWD from type 2A vWD. Purified plasma vWF or cryoprecipitate causes platelet aggregation when added to platelet-rich plasma from patients with platelet-type vWD, distinguishing this disorder from type 2B vWD. In addition, type 2B vWD plasma transfers the enhanced RIPA to normal platelets, whereas plasma from patients with platelet-type vWD interacts normally with control platelets.
ACQUIRED VON WILLEBRAND DISEASE
Acquired vWD usually presents as a late-onset bleeding diathesis in a patient with no prior bleeding history and a negative family history of bleeding. Decreased levels of factor VIII, vWF:Ag, and ristocetin cofactor activity are common, and the bleeding time is usually prolonged. Acquired vWD is usually associated with another underlying disorder and has been reported to occur in patients with myeloproliferative disorders,198 hypothyroidism,199 benign or malignant B-cell disorders,200 several solid tumors (particularly Wilm’s tumor),201 or certain cardiac or vascular defects,202 or in association with several drugs, including ciprofloxacin and valproic acid.203,204
A variety of B-cell disorders have been associated with the development of anti-vWF autoantibodies. In most cases the acquired vWD appears to be due to rapid clearance of vWF induced by the circulating inhibitor, although these antibodies may also interfere with vWF function. Hypothyroidism results in decreased vWF synthesis,199 and, in some cases of malignancy, the acquired vWD is thought to be due to selective adsorption of vWF to the tumor cells. In acquired vWD associated with valvular heart disease or certain drugs, vWF may be lost by accelerated destruction or proteolysis.203,204
The vWF multimers in acquired vWD usually exhibit a type 2A pattern, with relative depletion of the large multimer forms. Distinguishing acquired vWD from genetic vWD can be difficult, since testing for the associated autoantibodies is generally not available in the clinical setting. The diagnosis often rests on the late onset of the disease, the absence of a family history, and the identification of an associated underlying disorder.
Management of acquired vWD is generally aimed at treating the underlying disorder. vWF levels and bleeding symptoms often improve with successful treatment of hypothyroidism or an associated malignancy. Refractory patients have been treated with corticosteroids, plasma exchange, intravenous gamma globulin, DDAVP, and vWF-containing factor VIII concentrates.204
THERAPY, COURSE, AND PROGNOSIS
The choice of treatment in any given patient depends upon the type and severity of vWD, the clinical setting, and the type of hemostatic challenge that must be met. A previous history of trauma or surgery and the success of previous treatment are important parameters to include in assessing the risk of bleeding. In general, the goals of therapy are to normalize the factor VIII activity and the bleeding time.
Epinephrine, insulin, and vasopressin given to normal volunteers induce short-lived increases in factor VIII coagulant activity and vWF levels. Desmopressin (1-desamino-8-D-arginine vasopressin, DDAVP) is an analog of antidiuretic hormone and was originally produced for the treatment of diabetes insipidus. When DDAVP is administered to healthy subjects, it causes sustained increases of factor VIII and ristocetin cofactor activity for approximately 4 h.205 DDAVP also releases tissue plasminogen activator and plasminogen activator inhibitor, presumably from endothelial cells. Patients with type 1 vWD treated with DDAVP release unusually high-molecular-weight vWF multimers into the circulation for 1 to 3 h after the infusion.205,206 Therapy with DDAVP increases the factor VIII activity, vWF:Ag, and ristocetin cofactor activity to two to five times the basal level and, in many instances, corrects the bleeding time of type 1 vWD patients.
DDAVP has become a mainstay for the treatment of mild hemophilia and vWD.207 It is regularly used in the setting of mild to moderate bleeding and for prophylaxis of patients undergoing surgical procedures. DDAVP is most commonly administered at a dose of 0.3 µg/kg, with an upper limit of 20 µg. Common side effects are mild cutaneous vasodilatation resulting in a feeling of heat, facial flushing, tingling, and headaches. The potential for dilutional hyponatremia, especially in elderly and very young patients, requires appropriate attention to fluid restriction, since it may result in seizures. There have been isolated reports of acute arterial thrombosis associated with administration of DDAVP, but the risk appears to be very low when judged against the total number of patients treated.
An intranasal form of DDAVP is also available and appears to be similar in efficacy to intravenous administration,208,209 although the response may be more variable. Patients receiving DDAVP at closely spaced intervals of less than 24 to 48 h can develop tachyphylaxis. However, in one study, 22 type 1 vWD patients showed a departure of less than 20 percent from the mean factor VIII peak level calculated from two separate infusions. In addition, the consistency of response in one patient reliably predicted the future response of that patient and other affected family members.210 For patients requiring repeated infusions of DDAVP, the factor VIII activity and vWF responses may not be of the same magnitude as after the first infusion. Although this decay in response has considerable individual variability, after one infusion of DDAVP per day for 4 days it was found that the responses on days 2 to 4 were reduced approximately 30 percent compared to day 1.208,209,210 and 211
Approximately 80 percent of type 1 vWD patients have excellent responses to DDAVP. In patients for whom DDAVP is potentially the treatment of choice, a test dose should be given (with measurements of before and after vWF and factor VIII levels) in advance of the first required course of treatment to ensure an adequate therapeutic response. For patients with type 1 vWD who are undergoing surgical procedures, DDAVP can be administered 1 h before surgery and approximately every 12 h thereafter. The response of factor VIII and ristocetin cofactor activity should be monitored when DDAVP is administered at frequent intervals. vWF-containing factor VIII concentrates and/or cryoprecipitate should be available for transfusion as backup.
Approximately 20 to 25 percent of patients with vWD do not respond adequately to DDAVP. This includes many type 2 vWD patients and nearly all patients with type 3 vWD. The response to DDAVP of patients with type 2A vWD is variable. Although most patients respond only transiently, some patients exhibit complete hemostatic correction after DDAVP infusion.212,213 The differences in DDAVP efficacy among type 2A patients may correspond to the type of mutation, with better responses predicted in patients with group 2 mutations, although this hypothesis remains to be tested.
Many experts consider DDAVP to be contraindicated in the treatment of type 2B vWD, as the high-molecular-weight vWF released from storage sites has an increased affinity for binding to GPIb and might be expected to induce spontaneous platelet aggregation and worsening thrombocytopenia.214 However, there are two reports of DDAVP used successfully in type 2B vWD patients, with an associated shortening or correction of the bleeding time and variable thrombocytopenia.215,216
VON WILLEBRAND FACTOR REPLACEMENT THERAPY
It is important to determine the response to DDAVP for each individual in order to avoid the unnecessary use of plasma products. For type 3 vWD patients and other patients unresponsive to DDAVP, the use of selected virus-inactivated, vWF-containing factor VIII concentrates is generally safe and effective.217 Cryoprecipitate has been successfully used in the past, but since it is not currently treated to inactivate viruses, it is less desirable. Solvent-detergent-treated plasma is available, and cryoprecipitate prepared from such plasma may be an appropriate choice. It is important to note that most standard factor VIII concentrates are not effective for vWD, presumably because the vWF is either removed or undergoes degradation during processing. Only preparations that contain large quantities of vWF with well-preserved multimer structure are suitable for use in vWD patients. A recent study reported the analysis of 11 different factor VIII concentrates.217 Humate P and VHP are currently the two most frequently used concentrates, but only the former is available in the United States. Both of these concentrates have been shown to contain large vWF multimers resembling those found in normal plasma.218,219
Replacement therapy is largely empiric. In instances of serious bleeding or major surgical interventions, treatment may have to be repeated at least once a day. Although in general there is a correlation between normal hemostasis and correction of the bleeding time and factor VIII activity, this does not occur in all cases. In patients who have concomitant thrombocytopenia associated with or in addition to vWD, it may be necessary to transfuse platelets in addition to factor VIII concentrates. It is recommended that patients be treated for 7 to 10 days after major surgical procedures and for approximately 3 to 5 days after minor surgical procedures. Since postpartum hemorrhage can occur for up to a month or more after delivery, therapy may need to be prolonged in certain patients with severe disease. Sufficient therapy should be given to ensure normalization of factor VIII activity and shortening or correction of the bleeding time. If clinical bleeding continues, additional replacement therapy must be given and searches undertaken for other hemostatic defects. An occasional type 3 vWD patient will develop an alloantibody against the infused vWF, severely complicating replacement therapy.220 The development of such a vWF inhibitor appears to be more common among type 3 vWD patients with large gene deletions.26,126 A variety of approaches to the management of vWD inhibitors have been tried, including immunosuppression, similar to the treatment of factor VIII inhibitors in hemophilia A (see Chap. 123).
OTHER NONREPLACEMENT THERAPIES
Estrogens or oral contraceptives have been used empirically in treating menorrhagia. In addition to their effects on the ovaries and uterus, estrogens also tend to increase plasma vWF levels. Patients with vWD frequently normalize their levels of factor VIII, vWF:Ag, and ristocetin cofactor activity during pregnancy. The mechanism of action of estrogens may be related in part to the increased production of vWF through a direct effect on endothelial cells.221 In pregnant patients with type 1 vWD, the factor VIII and ristocetin cofactor activities usually rise above 50 percent. These patients usually do not require any specific therapy at the time of parturition. In contrast, individuals who have 30 percent or less factor VIII or variant forms of vWD are more likely to require prophylactic therapy before delivery. Postpartum hemorrhage in all forms of vWD may occur as long as 1 month postpartum. Some patients are treated with plasma products prophylactically. Postpartum hemorrhage within the first few days after parturition may be related to the relatively rapid return to prepregnancy levels of factor VIII and vWF activities.
Fibrinolytic inhibitors such as e-aminocaproic acid have been used effectively in some vWD patients. Fibrinolytic inhibitors have been suggested as an adjunct to DDAVP infusion, given the potential for enhanced fibrinolysis as a result of the release of tissue plasminogen activator along with vWF. However, fibrinolytic inhibitors are not generally used in this setting and are generally restricted to prophylactic treatment for dental procedures or empiric treatment of chronic menorrhagia or recurrent epistaxis.
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Ernest Beutler, Marshall A. Lichtman, Barry S. Coller, Thomas J. Kipps, and Uri Seligsohn