The number of patients receiving anticoagulant treatment with vitamin K antagonists (VKA) is still increasing and a substantial number of these patients experience major bleeding complications as a consequence of this therapy, 1–3% yearly [1,2]. It is of utmost importance to prevent bleeding complications from anticoagulant treatment. Identification of risk factors for bleeding complications will help to improve the safety of VKA treatment.

It is well known that the intensity of VKA treatment is the major determinant of the risk of bleeding in VKA-treated patients. However, other factors also contribute to the risk and play a significant role [1,3]. In the search for novel risk factors associated with bleeding in VKA-treated patients, the FACTORS (FACTors in ORal anticoagulation Safety) study was initiated [3]. The FACTORS study is a case–control study including patients on VKA treatment that have (cases) or have not (controls) experienced major bleeding. The definition of major bleeding was: bleeding leading to death, hospitalization or a decrease of more than 1.25 mm of hemoglobin; intracranial bleeding; and muscle, joint, or intraocular bleeding. The indications for VKA treatment of the patients participating in the FACTORS study were atrial fibrillation, venous thromboembolism, postoperative prophylaxis, mechanical heart valve, vascular surgery, and ischemic heart disease. In the study cases and controls were recruited from the Thrombosis Services in Leiden and Amsterdam and they were matched for the indication of VKA treatment, age, gender, intensity and type of VKA (acenocoumarol or phenprocoumon). The FACTORS study was previously described in detail [3]. It has been shown that high levels of soluble thrombomodulin (s-TM) are associated with an increased risk of bleeding [4]. However, the risk was only linked to the highest quartile of s-TM in the FACTORS study [3]. Thus additional, possibly novel genetic risk factors may play a role in the bleeding in patients treated with VKA.

A candidate genetic risk factor is the von Willebrand factor (VWF) Y1584C polymorphism. This polymorphism was shown to be linked to increased susceptibility of VWF to proteolysis by ADAMTS-13 [5]. The VWF Y1584C polymorphism was found at a frequency of 1% (heterozygotes) in two different cohorts of normal individuals (2/200 [5] and 1/100 [6]). A much higher frequency of 14% of this polymorphism was found in type 1 von Willebrand disease (VWD) patients in a Canadian study [6]. However, the polymorphism did not always cosegregate with VWD and some of the heterozygous carriers were unaffected and showed normal VWF antigen (VWF:Ag) levels and VWF multimer patterns [5,6]. Even in the presence of normal VWF:Ag levels this polymorphism may affect the formation of the platelet plug through increased in vivo proteolysis of VWF multimers under shear stress, as suggested by Bowen et al. [5]. Hence, the effect of this polymorphism on VWF multimer survival in vivo and primary hemostasis may be substantial. Consequently, it may modulate the risk for bleeding and may be considered a potential general risk factor for bleeding, particularly in patients already hemostatically challenged by VKA treatment.

To investigate whether VWF Y1584C is indeed associated with bleeding, we screened 327 patients from the FACTORS study for the presence of the VWF Y1584C polymorphism; 110 of these were patients with bleeding episodes during VKA treatment (cases) and 217 were patients without bleeding episodes during VKA treatment (controls). The screening was performed on genomic DNA by amplification of exon 28 by polymerase chain reaction (PCR) with a forward primer specific for the VWF gene, 5′-TG GTT CTG GAT GTG GCG TTC (complementary to nts 24/1015–1034), and a reverse primer, 5′-CAA GGC CAT GCC AGC CCT CG (complementary to nts 24/1622-1641, numbering according to Mancuso [7]). The use of a gene-specific primer prevents inadvertent coamplification of the VWF pseudogene. Subsequently, the 627-bp PCR product was digested with the restriction enzyme Acc65I. Homozygosity for the common allele would yield two fragments of 269 and 358 bp, whereas heterozygosity would yield three fragments of 269, 358 and 627 bp, due to the loss of the Acc65I site in the 1584C allele.

In the population of 327 individuals, two patients that were included because of bleeding (cases) were heterozygous for the polymorphism (Table 1). The first patient was an 81-year-old male using VKA for atrial fibrillation and who had experienced gastrointestinal bleeding. His VWF:Ag level was 196%. The second patient was a 68-year-old female who received VKA for atrial fibrillation and ischemic stroke. She experienced muscle hemorrhage and her VWF:Ag level was 155%. No heterozygotes were observed in the control group of 217 individuals that did not experience major bleeding.

Table 1.   Distribution of the VWF Y1584C polymorphism in cases and controls
VWFCase bleeding +Control bleeding –OR*95% CI
  1. *Odds ratio. †Reference category. ‡According to [8].


In conclusion, we present some evidence that the VWF Y1584C polymorphism may be associated with bleeding in VKA-treated patients. However, because of the rarity of this polymorphism in our population, a much larger group of patients and controls would be required to assess reliably the bleeding risk associated with this polymorphism. In the two patients tested positive for the Y1584C polymorphism VWF levels were high. Although the polymorphism was previously found to be associated with type 1 VWD [5], it can not be considered a causative VWD mutation. The contribution to bleeding risk in other patient categories than those treated with VKA remains to be determined.


  1. Top of page
  2. Acknowledgements
  3. References

This work was financially supported by grants from the Netherlands Organization for Scientific Research [(NWO/ZonMW) no. 906-26-209 to J.C.J.E.], the Van den Tol Foundation (to J.C.J.E.), Zorgonderzoek Nederland (no. 2100.0009 to P.H.R.) and the Netherlands Heart Foundation (no. 99.165 to P.H.R.). We thank Vincent van Marion (LUMC, Leiden, the Netherlands) for technical support.


  1. Top of page
  2. Acknowledgements
  3. References
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