von Willebrand Disease: Diagnosis, Classification, and Treatment

J. Evan Sadler

David Lillicrap

von Willebrand disease (vWD) was discovered by Erik von Willebrand,¹^,² a Finnish internist who investigated a family living in Föglö, one of the Åland Islands in the Gulf of Bothnia, separating Sweden and Finland. The proband was a 5-year-old girl with a severe bleeding tendency. Four of her affected sisters died of hemorrhage before the age of 4 years, and the proband herself died at the age of 13 years with her 4th menstrual period.³ The parents and several other relatives of both sexes had mild bleeding symptoms, suggesting autosomal inheritance. The severity of bleeding was variable throughout this large pedigree, and some obligate heterozygous individuals were asymptomatic. The most common sites of bleeding were skin, uterus, and mucous membranes rather than deep tissues. Patients had a prolonged bleeding time, but had normal coagulation time, clot retraction, and platelet count. von Willebrand distinguished this condition from hemophilia and Glanzmann thrombasthenia but was unable to determine whether the defect lay in the blood, the vasculature, or the platelets.

Considerable progress has been made in understanding the pathophysiology of vWD. In the 1950s, patients with vWD were reported to have low levels of blood clotting factor VIII (FVIII), and transfusions with plasma fractions from healthy individuals or patients with hemophilia were shown to correct the bleeding tendency in vWD.⁴^,⁵^,⁶^,⁷^,⁸ These observations suggested that vWD was caused by abnormalities in a plasma protein, now known as von Willebrand factor (vWF). In 1972, vWF was purified,⁹ and genetic variants of vWD were identified and correlated with structural differences in vWF.¹⁰ The development of multimer gel electrophoresis¹¹^,¹²^,¹³ uncovered additional heterogeneity in vWD. In 1985, the structure of vWF was determined by protein sequencing and cDNA cloning.¹⁴^,¹⁵^,¹⁶^,¹⁷ This was followed almost immediately by the characterization of mutations that cause severe vWD.¹⁸^,¹⁹ Today, hundreds of mutations in many subtypes of vWD have been described.²⁰^,²¹ An online database of vWD mutations is maintained by the International Society on Thrombosis and Haemostasis at the University of Sheffield and is accessible at http://www.vwf.group.shel.ac.uk/index.html.

We now understand many aspects of vWF biosynthesis, structure, and function, as discussed in Chapter 13. vWF is a multimeric blood protein that performs two major roles in hemostasis; it mediates the adhesion of platelets to sites of vascular injury and it is a carrier protein for FVIII. These activities require the assembly of vWF into large multimers and interactions with several ligands. Inherited defects in vWF may interfere with biosynthetic processing, or disrupt specific ligand-binding sites, thereby causing bleeding by impairing either platelet adhesion or blood clotting. Depending on the disease mechanism, vWF mutations may cause autosomal dominant or recessive vWD. This knowledge provides a framework to understand the pathogenesis and classify the many variants of vWD.

DIAGNOSIS AND CLASSIFICATION OF VON WILLEBRAND DISEASE

Three major categories of vWD are distinguished: partial quantitative deficiency (type 1), qualitative deficiency (type 2), and total deficiency (type 3) (see Table 52.1).²²^,²³ Qualitative type 2 vWD is divided further into four variants (2A, 2B, 2M, and 2N) on the basis of details of the clinical phenotype. These six categories correspond to distinct pathophysiologic mechanisms and they correlate with distinct clinical features and therapeutic requirements, as discussed in a subsequent section. The laboratory features of vWD subtypes are summarized in Table 52.2.

von Willebrand Disease Type 1

vWD type 1 includes partial quantitative deficiency of vWF and is commonly inherited as an autosomal dominant trait. vWD type 1 accounts for more than 70% of all vWD, although the prevalence of vWD type 1 is not known precisely. Symptomatic vWD that leads to medical treatment affects approximately 35 to 100 per million individuals, which is comparable to the prevalence of hemophilia A.²⁴ Screening programs often identify many more individuals with low vWF and mild symptoms and suggest that vWD could have a prevalence of approximately 1%,²⁵ but very few patients identified by population screening subsequently have significant bleeding.²⁶ Finally, screening patients attending the offices of primary care physicians has indicated a prevalence of symptomatic vWD of approximately 1 in 1,000.⁴⁵ vWD type 1 includes variants previously designated as type IA²⁷; type I-1, I-2, and I-3²⁸; type I-platelet normal and type I-platelet low²⁹; and vWD Vicenza. vWD Vicenza was formerly classified as vWD type 2M, but the consensus now favors classifying it as vWD type 1. Most studies indicate that vWF Vicenza has a normal vWF function/antigen ratio (vWF:RCo/vWF:Ag),³⁰^,³¹ but some suggest the protein is dysfunctional.³²

Many factors make vWD type 1 difficult to diagnose with confidence. Mild bleeding symptoms are sufficiently common that coincidental association with low vWF levels also is frequent, even when patients have a positive family history of bleeding.³³ Furthermore, bleeding symptoms are not specific for vWD, and apparent transmitters of low vWF or a bleeding phenotype may themselves be phenotypically normal. Therefore, the criteria used for diagnosis are important determinants of the apparent prevalence of vWD type 1.

Table 52.1 Classification of vWD

Type	Description
1	Partial quantitative deficiency of vWF
2	Qualitative vWF defects
2A	Decreased vWF-dependent platelet adhesion and a selective deficiency of high molecular weight vWF multimers
2B	Increased affinity for platelet GPIb
2M	Decreased vWF-dependent platelet adhesion without a selective deficiency of high molecular weight vWF multimers
2N	Markedly decreased binding affinity for FVIII
3	Virtually complete deficiency of vWF
vWF, von Willebrand factor.
From Sadler JE, Budde U, Eikenboom JC, et al. Update on the pathophysiology and classification of von Willebrand disease: a report of the Subcommittee on von Willebrand factor. J Thromb Haemost 2006;4(10):2103-2114.

Bleeding Symptoms

Few large studies of symptoms in vWD have been published, and they rarely distinguish among subtypes of vWD. For example, the extensive compilations of Silwer³⁴ and of Buchanan and Leavell³⁵ include patients with all variants of vWD. Because other variants are relatively uncommon, these surveys remain useful guides to the clinical features of vWD type 1. The most common symptoms (see Table 52.3) are epistaxis, skin bruises, hematomas, prolonged bleeding from trivial wounds, oral cavity bleeding, and excessive menstrual bleeding. Gastrointestinal bleeding appears to be relatively rare, but may be serious when it occurs.

Table 52.2 Selected laboratory features of vWD subtypes

vWD	FVIII	vWF:Ag	vWF:RCo or vWF:CB	RIPA	Multimer Structure
Type 1	Decreased	Decreased	Decreased	Decreased	Normal in plasma and platelets
Type 2A	Decreased or normal	Decreased or normal	Decreased relative to vWF:Ag	Decreased relative to vWF:Ag	Large and intermediate multimers absent from plasma; variable in platelets
Type 2B	Decreased or normal	Decreased or normal	Decreased or normal	Increased at low ristocetin concentration	Large and intermediate multimers absent from plasma; normal in platelets
Type 2M	Decreased or normal	Decreased or normal	Decreased relative to vWF:Ag	Decreased relative to vWF:Ag	Large and intermediate multimers present in plasma and platelets
Type 2N	Moderately decreased (<20 IU/dL)	Normal	Normal	Normal	Normal in plasma and platelets
Type 3	Moderately decreased (<20 IU/dL)	Absent or trace	Absent	Absent	None or trace in plasma or platelets
vWD, von Willebrand disease; vWF, von Willebrand factor; RIPA, ristocetin-induced platelet aggregation.

Epistaxis may be severe and prolonged. Easy bruising is common in vWD. Prolonged bleeding after minor trauma to skin or mucous membranes is characteristic of vWD. Severe hemorrhage after major surgery may continue for several weeks. Prolonged oral bleeding is common from gums, or with biting of the tongue or lips. Heavy bleeding is also common after tooth extraction or other oral surgery such as tonsillectomy and adenoidectomy. Menorrhagia is a typical presenting symptom in women with vWD, but is also frequent in otherwise healthy women.³⁴

Coexistent conditions or drug interactions can strongly influence the severity of symptoms in vWD. Bleeding may be exacerbated by the use of aspirin or corticosteroids and decreased by the use of oral contraceptives. Treatment with sodium valproate for epilepsy can lower the level of vWF and induce bleeding.³⁶ The risk of hemorrhage may be increased by liver disease, uremia, defects in connective tissue such as Ehlers-Danlos syndrome,³⁷ hypothyroidism,³⁸ and focal lesions such as gastric or duodenal ulcers, or gastrointestinal angiodysplasia.³⁹

Standardized questionnaires have been developed to describe and quantitate bleeding symptoms.⁴⁰^,⁴¹ These assessment tools are being refined and validated for prospective use with pediatric and adult patients to diagnose vWD.⁴²^,⁴³^,⁴⁴^,⁴⁵ These tools may help direct the intensity of laboratory investigation and also facilitate communication of the bleeding severity between health care professionals.

Laboratory Testing

The goal of laboratory testing in vWD type 1 is to exclude other causes of bleeding, document vWF deficiency, and exclude qualitative vWF defects. As a complete laboratory evaluation for vWD is quite elaborate, the efficiency of testing can be increased by using initial screening tests for vWD. vWF levels vary with physiologic
stress, and where possible, tests should not be performed in proximity to hemorrhagic events, pregnancy, acute infection, or strenuous exercise. Optimal testing strategies have not yet been validated experimentally, and the approach described here is an attempt to synthesize various expert opinions.

Table 52.3 Frequency (%) of bleeding symptoms in vWD

		Buchanan and Leavell^a
Symptoms	Swedish Series^b (264 Cases)	Men (95 Cases)	Women (104 Cases)	Normal^b (500 Cases)
Nose bleeding	62.5	73	64.5	4.6
Menorrhagia	60^c	—	51	25.3^c
Bleeding after tooth extraction	51.5	28.5	38	4.8
Ecchymoses and hematomas	49.2	—	—	11.8
Ecchymoses	—	55.7	67	—
Hematomas	—	8.5	7.5	—
Bleeding from minor cuts or abrasions	36	37	33.5	0.2
Gingival bleeding	34.8	33	36	7.4
Postoperative bleeding	28	19	20	1.4
Bleeding at delivery	23.3^c	—	10.5	19.5^c
Gastrointestinal bleeding	14	24.5	9	0.6
Traumatic oral and lip bleeding	11.7	—	—	0.6
Petechiae	11.5	16	18.5	1.2
Joint bleeding	8.3	12.5	5	0
Hematuria	6.8	7.5	2	0.6
Ovarian bleeding	6.8^c	—	—	—
Bleeding after tonsillectomy	6.1	—	—	—
Bleeding during shedding of teeth	4.9	—	—	—
Bleeding at abortion	3.8^c	—	—	—
Intramuscular, deep subcutaneous, or submucosal bleeding	2.7	—	—	—
^aData of Buchanan JC, Leavell BS. Pseudohemophilia: report of 13 new cases and statistical review of previously reported cases. Ann Intern Med 1956;44:241-256. ^bData of Silwer J. von Willebrand’s disease in Sweden. Acta Paediatr Scand Suppl 1973;238:1-159. ^cFor women older than 15 years.

Initial tests for any hemostatic disorder include a complete blood count with platelet count, prothrombin time (PT), and activated partial thromboplastin time (aPTT). Additional tests specific for vWD include ristocetin cofactor activity (vWF:RCo), collagen-binding activity (vWF:CB), vWF antigen (vWF:Ag), and FVIII level.

vWF:RCo is a useful initial assay for vWF concentration and platelet-binding function,²⁵ but the test is difficult to standardize and is not very precise. Therefore, vWF:Ag may be used to confirm low values for vWF:RCo. Like the vWF:RCo test, vWF:CB also depends on vWF concentration but measures the interaction of vWF with immobilized collagen. vWF:CB tests are not as widely available as vWF:RCo, but are useful for a comprehensive evaluation of vWF functional defects.⁴⁶^,⁴⁷

Discrepancies between vWF:RCo and vWF:Ag, or between vWF:CB and vWF:Ag, suggest a qualitative vWF defect that should be investigated along with other tests, as discussed in the section, von Willebrand Disease Type 2. However, some common vWF sequence variations affect the vWF:RCo assay. For example, African Americans have relatively decreased vWF:RCo, and this variation is explained by differences in the frequency of the vWF Asp1472His polymorphism. vWF 1472His has normal hemostatic function but decreased vWF:RCo compared to vWF 1472Asp. vWF 1472His occurs in 19% of whites and 62% of African Americans, which accounts for the observed differences in vWF:RCo/vWF:Ag between these populations and can lead to the overdiagnosis of vWD type 2M.⁴⁸ The vWF mutation Pro1467Ser also profoundly decreases vWF:RCo/vWF:Ag (<0.1) with no impairment of hemostasis.⁴⁹ Conversely, the polymorphism Ala1381Thr modestly increases vWF:RCo.⁵⁰ Mutations associated with vWD type 2B Malmö/New York may also belong to this category.

vWF levels may be normal intermittently in vWD, and abnormal results should be confirmed.⁵¹ For example, the PT, aPTT, FVIII, and vWF tests may be repeated after an interval of 2 weeks. If the results are inconsistent, they may be performed again. Unfortunately, such repeated testing is impractical or not feasible for many patients. With the limitations discussed in the subsequent text, vWD is excluded by arriving at repeated normal values of FVIII and concordant vWF:RCo, vWF:CB, and vWF:Ag on serial testing. If these initial tests are abnormal, other tests are indicated.

An abnormal aPTT is evaluated in concert with the FVIII level. An FVIII that is disproportionately lower than the vWF:Ag would suggest either hemophilia A, a defect that impairs vWF ability to bind FVIII (vWF:FVIIIB) as occurs in vWD type 2N, or another cause of FVIII deficiency such as an antibody against FVIII. If the FVIII is normal, then other causes of a prolonged aPTT should be considered.

Low values for vWF by any test can be hard to evaluate because the range of normal values for plasma vWF concentration is very broad. vWF levels also depend on ABO blood type (see Table 52.4), and the mean vWF:Ag level is approximately 25% lower for individuals of blood type O compared to other blood types.⁵² Some additional variation has been attributed to the secretor locus that specifies some oligosaccharide antigens of the Lewis blood group system.⁵³^,⁵⁴ However, most of the variation in vWF levels is due to unknown factors other than either vWD or blood type. The problem is illustrated in FIGURE 52.1, using data for blood type O. For this population of normal type O blood donors, the mean level of vWF:Ag was 86 IU/dL, with a range (±2 SD) of 41 to 179 IU/dL.⁵² Therefore, blood type O alone can cause vWF levels low enough to suggest the diagnosis of vWD. By halving the values for this normal population, the distribution for individuals of blood type O and only one functional vWF allele can be estimated to have a mean vWF:Ag of 43 IU/dL (see FIGURE 52.1), but there is considerable overlap with the normal distribution. Because individuals heterozygous for vWD mutations constitute much <1% of the population,²⁴ moderately low vWF levels cannot reliably identify patients with vWD type 1. Whether blood type-adjusted means should be used routinely remains controversial, but it seems increasingly likely that the ABO blood type is one of perhaps many genetic modifiers that influence vWF plasma levels. In these initial tests, comparison of functional assays with the level of vWF:Ag must not suggest a qualitative abnormality of vWF, as discussed in the section on vWD type 2. If available, ristocetin-induced platelet aggregation (RIPA) should not indicate abnormal sensitivity to ristocetin, and the plasma vWF multimer distribution should have a normal complement of large multimers.

Table 52.4 Influence of ABO blood groups on vWF:Ag in volunteer blood donors

		vWF:Ag (Percentage of Pooled Normal Plasma Standard)
ABO Type	n	Geometric Mean IU/dL	Geometric Mean ± 2 SD IU/dL
O	456	86	41-179
A	340	121	55-267
B	196	134	65-275
AB	109	141	73-271
The groups were statistically different from each other as follows: O versus A, B, and AB, P < 0.01; A versus AB, P < 0.01; B versus A, P < 0.05.
vWF, von Willebrand factor.
Modified from Gill JC, Endres-Brooks J, Bauer PJ, et al. The effect of ABO blood group on the diagnosis of von Willebrand disease. Blood 1987;69:1691-1695. The modified table includes vWF:Ag values converted to WHO international units per deciliter (IU/dL).²³

FIGURE 52.1 Distribution of vWF:Ag levels in normal blood donors of blood type O and hypothetical distribution of vWF:Ag levels in individuals of blood type O who are heterozygous for vWF null alleles. The reference normal value of 100 IU/dL is based on the World Health Organization standard pooled plasma. (Adapted from tabulated data of Gill JC, Endres-Brooks J, Bauer PJ, et al. The effect of ABO blood group on the diagnosis of von Willebrand disease. Blood 1987;69:1691-1695.)

The template bleeding time has been used to diagnose vWD, but its value for this purpose is difficult to demonstrate. A prolonged bleeding time is not specific for vWD, and does not predict bleeding at surgery.⁵⁵^,⁵⁶^,⁵⁷ The bleeding time is subject to even greater normal variation than vWF levels, has poor reproducibility, and often is normal in vWD.⁵¹ The test can distinguish populations that are defined independently, but generally it is not useful when applied to individuals. The bleeding time may be helpful for assessing the efficacy of therapeutic interventions in selected patients with vWD, but it does not appear to have sufficient sensitivity or reproducibility to contribute strongly to the diagnosis or exclusion of vWD. With all of these considerations, performance of the bleeding time has now been discontinued in many institutions.

A platelet function analyzer (PFA)-100 has been developed to test platelet plug formation in vitro by forcing a sample of citrated whole blood at high shear through a porous membrane coated with collagen plus adenosine 5′-diphosphate or collagen plus epinephrine. Normal platelets become activated upon contact with the membrane and form occlusive aggregates. The time required for blood flow to stop is termed the closure time, which is prolonged by defects in vWF or platelet function. The PFA-100 has been proposed as a sensitive and reproducible alternative to the bleeding time, but lack of specificity remains a concern and the clinical utility of PFA-100 testing remains uncertain.⁵⁸^,⁵⁹^,⁶⁰ As previously reported for the bleeding
time,⁵⁵^,⁵⁶^,⁵⁷ a prolonged PFA-100 closure time was not found to predict increased surgical bleeding.⁶¹

Other vWF tests may be performed in specialized diagnostic or research laboratories. Approximately 15% of vWF in the blood is located within the α-granules of platelets and is secreted upon platelet activation.⁶² Platelet vWF can be assayed by methods similar to those used for plasma vWF, and the results can help identify some variants of vWD type 1.²⁹ Botrocetin is a snake venom protein that binds vWF.⁶³^,⁶⁴ Like ristocetin, botrocetin induces high-affinity binding to platelet glycoprotein (GP) Ibα. Tests of botrocetin-induced platelet aggregation or “botrocetin cofactor activity” have been developed that are analogous to vWF assays employing ristocetin, although botrocetin and ristocetin affect vWF function by distinct mechanisms.⁶⁵ A variant of the vWF:RCo assay has been described that replaces platelets with recombinant, gain-of-function GPIbα immobilized on microtiter plates. This enzyme-linked immunosorbent assay style test is very precise and sensitive compared to conventional vWF:RCo assays⁶⁶ but is not yet available commercially.

Criteria for Diagnosis of von Willebrand Disease Type 1

No matter what criteria are used, the extreme range of normal vWF levels complicates the diagnosis of vWD type 1, and a satisfactory general answer to this dilemma has not yet been devised. Because of this diagnostic uncertainty, the risk and benefit of specifically diagnosing vWD type 1 should be balanced for each patient.

The diagnosis of vWD type 1 is easy to make when the patient has significant mucocutaneous bleeding, the vWF level is clearly low (e.g., <30 IU/dL), and the family history is strongly positive with high penetrance. Such families usually show clear coinheritance of low vWF with bleeding symptoms, and usually, mutations in the vWF gene can be found.²⁹^,⁶⁷^,⁶⁸^,⁶⁹^,⁷⁰^,⁷¹^,⁷²^,⁷³^,⁷⁴ For these patients, a diagnosis of vWD type 1 seems appropriate because they often can benefit from changes in lifestyle, or from specific treatment to prevent or control bleeding. Also, genetic counseling is straightforward when the inheritance of the disorder can be predicted with confidence. Unfortunately, this combination of features is relatively rare.

More commonly, patients have a medical history of only menorrhagia or other infrequent mild bleeding, their vWF levels are borderline low (e.g., 30 to 50 IU/dL), and their family members do not consistently have either low vWF or bleeding symptoms. In such cases, low vWF level and bleeding are likely to associate coincidentally rather than causally.³³ Family studies usually show that bleeding symptoms and low vWF levels are not coinherited, and linkage to the vWF gene cannot be demonstrated.²⁶^,⁴¹^,⁷⁰^,⁷¹^,⁷²^,⁷³^,⁷⁴^,⁷⁵^,⁷⁶ vWF levels that are slightly more than 2 SD below the population mean confer a relatively small relative risk of menorrhagia or other bleeding³³^,⁷⁷^,⁷⁸ and do not correlate with increased bleeding at surgery.⁷⁹ To label these patients as vWD type 1 has little clinical utility. It may, in fact, harm them by affecting their relationship with employers and insurance providers, by causing unnecessary changes in lifestyle, and by inducing unjustified fear of transmitting a genetic disease. A modestly low vWF level may be treated instead, as a biomarker for modestly increased bleeding risk. Empiric therapy to raise vWF may be considered for these patients with low vWF if they should bleed.

Molecular Defects in von Willebrand Disease Type 1

Recent studies have indicated that vWD type 1 is associated with candidate mutations at the vWF locus in approximately 65% of cases.⁷¹^,⁷³^,⁷⁴ There is significant allelic heterogeneity, with >100 different mutations being documented to date. Missense mutations predominate, with the Tyr1584Cys substitution being the most frequent variant reported, showing a prevalence of 5% to 15% in different populations.⁷¹^,⁷³^,⁷⁴ This mutation is associated with a mild phenotype (vWF:Ag -40 IU/dL in heterozygotes), usually accompanies blood type O, and has a penetrance of approximately 70%.⁸⁰

vWD type 1 can occasionally be caused by frameshifts, nonsense mutations, or deletions that are similar or identical to mutations found in patients with vWD type 3.⁸¹^,⁸²^,⁸³ As first shown by Erik von Willebrand, some heterozygous members of such families may not have bleeding symptoms, so that possession of a single such mutant allele does not consistently cause bleeding symptoms. Variation in symptoms may be due to chance coinheritance of a second mutated vWF allele. For example, in two families in the Netherlands, compound heterozygosity for vWD type 1 and vWD type 2N was found in patients with substantial bleeding, whereas relatives with either single mutant allele were asymptomatic.⁸⁴

vWD type 1 sometimes is inherited as an autosomal dominant trait with very high penetrance, and this strongly dominant phenotype is usually associated with missense mutations.⁷⁰^,⁷¹^,⁷²^,⁷³^,⁷⁴ Many of these missense mutations cause intracellular retention of mutant vWF subunits.⁸⁵ In one family, a Cys1149Arg missense mutation was identified that caused the retention of mutant subunits in the endoplasmic reticulum and also inhibited the secretion of normal vWF subunits, possibly by causing the intracellular retention of heterodimers between mutant and normal subunits.⁸⁶^,⁸⁷ Another missense mutation at a cysteine residue, Cys1130Arg, was found in two other families with a similar dominant phenotype, although the effect on vWF biosynthesis was not reported.⁸⁶ Two independent mutations that cause the heterozygous deletion of vWF exons 4 to 5 were shown to dominantly inhibit the secretion of normal vWF.⁸⁸ This dominant mechanism provides a plausible biochemical explanation for the transmission of exceptionally low vWF levels and bleeding symptoms with high penetrance.

Quantitative vWF deficiency also can be caused by mutations that lead to the increased clearance of vWF from the blood, provided that the clearance mechanism does not exhibit a strong preference for large multimers.⁸⁹^,⁹⁰^,⁹¹ For example, the mutation Arg1205His causes vWD Vicenza, which is characterized by vWF:Ag < 15 IU/dL and plasma vWF multimers that are sometimes even larger than those found in normal plasma. vWF Vicenza appears to be cleared from the blood approximately fivefold more rapidly than normal vWF.³¹ It has been proposed that these accelerated clearance variants, which comprise approximately 15% of type 1 vWD cases, be subclassified as vWD type 1c.⁸⁹

Other mutations resulting in vWD type 1 may be located in vWF introns or in distant regulatory elements. Furthermore, the results of a large genome-wide association study meta-analysis indicate that several genes involved in storage organelle formation and protein clearance functions are strongly associated with plasma vWF levels.⁹² Variation at these novel loci may act as primary causative mutations or genetic modifiers in vWD type 1.

von Willebrand Disease Type 2

For vWF to perform its hemostatic functions, it must be assembled into large multimers, and functional binding sites must be constructed for FVIII, for platelet receptors, and for ligands in
connective tissue. Mutations in vWD type 2 can disrupt any of these properties, and vWD type 2 accounts for between 7% and 30% of all vWD in various reports. Such qualitative abnormalities of vWF often are associated with discrepancies between the level of vWF:Ag and functional assays such as vWF:RCo, vWF:CB, or vWF:FVIIIB. Four subtypes are currently recognized (Tables 52.1 and 52.2). The prevalence of the subtypes appears to be increasing as awareness of them increases, although the distribution of subtypes differs considerably among treatment centers. A survey in France found that patients with vWD type 2 were divided as 30% type 2A, 28% type 2B, 8% type 2M (or unclassified), and 34% type 2N.⁹³ A similar study in Germany found 74% type 2A, 10% type 2B, 13% type 2M, and 3.5% type 2N.⁹⁴

Discrimination between qualitatively normal vWF and abnormal vWF can be difficult by laboratory testing, especially because vWF:RCo assays become somewhat unreliable when the vWF:Ag level is <20 IU/dL. A common strategy is to compare values obtained in functional assays (vWF:RCo, vWF:CB, vWF:FVIIIB) with vWF:Ag; a ratio <0.6⁹⁵ or <0.7⁹⁶^,⁹⁷ is inferred to be abnormal, and a ratio larger than the selected value is interpreted as normal. This approach seems reasonable, although the sensitivity and specificity of a particular ratio would depend on the performance characteristics of the individual assays. The discriminant power of vWF multimer analysis is also a significant determinant of vWD type 2 diagnosis.

von Willebrand Disease Type 2A

A disproportionately low vWF:RCo or vWF:CB, relative to vWF:Ag, may reflect decreased affinity of vWF for platelets or collagen, respectively. The most common cause of this defect is the absence of large vWF multimers (see FIGURE 52.2), which is characteristic of vWD type 2A. This subtype can be diagnosed by the combination of markedly reduced vWF:RCo or vWF:CB and compatible multimer gel analysis (FIGURE 52.2). vWD type 2A appears to account for most of vWD type 2.⁹⁸ vWD type 2A includes variants previously designated as type IIA¹²; type IIA-1, IIA-2, and IIA-3²⁸; type I-platelet discordant²⁹; type IB²⁷; type IIC⁹⁹; type IID¹⁰⁰; type IIE¹⁰¹; type IIF¹⁰²; type IIG¹⁰³; type IIH¹⁰⁴; and type II-I.¹⁰⁵

FIGURE 52.2 Multimer patterns in vWD. Samples of plasma from a normal individual (N) and from patients with vWD type 1, type 2A (two unrelated patients), type 2B, and type 3 were electrophoresed through a 1.3% agarose/sodium dodecyl sulfate (SDS) gel and von Willebrand factor (vWF) was detected by Western blotting. (Reprinted from Sadler JE. von Willebrand disease. In: Bloom AL, Forbes CD, Thomas DP, et al., eds. Haemostasis and thrombosis, 3rd ed. Edinburgh, UK: Churchill-Livingstone, 1994:843-857, with permission.)

vWD type 2A usually is a dominant disorder and is caused in approximately 80% of cases by single amino acid substitutions in or near domain A2 (residues 1,480 to 1,672) of the mature vWF subunit²⁰ (see FIGURE 52.3). These mutations lead to the absence of large vWF multimers either by impairing intracellular multimer assembly or by promoting intravascular proteolysis of vWF.¹⁰⁶ Some mutations may cause vWD type 2A by a combination of these mechanisms. Defects in multimer assembly produce an absence of large multimers in both plasma and platelet vWF. In contrast, increased susceptibility to proteolysis selectively depletes the large multimers of plasma vWF because
platelet vWF is relatively protected from proteolysis. The plasma metalloprotease, a disintegrin and metalloprotease with thrombospondin type 1 motifs member 13 (ADAMTS13), cleaves the Tyr1605-Met1606 bond within domain A2 of vWF,¹⁰⁷^,¹⁰⁸ and this cleavage accounts for the major proteolytic fragments of the vWF subunit that are found in plasma vWF.¹⁰⁹ Mutations that increase the susceptibility of vWF to ADAMTS13 cause the loss of large, hemostatically effective multimers from plasma and result in bleeding. Conversely, deficiency of ADAMTS13 causes thrombotic thrombocytopenic purpura associated with unusually large vWF multimers, as discussed in Chapter 99.

FIGURE 52.3 Mutations in vWD type 2. Amino acid residues are numbered by codon number 1 to 2,813 for prepro-vWF. The signal peptide contains residues 1 to 22, the propeptide contains residues 23 to 763, and the mature subunit contains residues 764 to 2,813. The locations of conserved structural domains (A, B, C, D, and CK) are indicated. Intersubunit disulfide bonds (not shown) connect CK to CK domains and D3 to D3 domains. The locations of binding sites are indicated for FVIII, platelet glycoprotein (GP)Ib, collagen, and platelet α_IIbβ₃. The metalloprotease ADAMTS13 cleaves a Tyr-Met bond in domain A2 (arrow). Circles mark the positions of mutations that cause the vWD phenotypes noted at the left. For vWD type 2A, the labeled brackets show the location of mutations that correspond to variants with characteristic multimer patterns discussed in the text. These include variants with increased sensitivity to proteolysis (IIA) or defective multimer assembly due to mutations in the propeptide (IIC), the D3 domain (IIE), or the CK domain (IID). References to the individual mutations can be found in the database of vWD mutations²⁰ maintained by the International Society on Thrombosis and Haemostasis at the University of Sheffield and accessible at http://www.shef.ac.uk/vwfvwf.group.shel.ac.uk/index.html. (Adapted from White GC II, Sadler JE. Von Willebrand disease: clinical aspects and therapy. In: Hoffman R, Benz EJ, Shattil SJ, eds. Hematology: basic principles and practice, 4th ed. Philadelphia, PA: Elsevier Churchill Livingstone, 2005:2121-2136, with permission.)

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