Von Willebrand Disease Patients Research Articles

Genetic variants, thrombocytopenia, and clinical phenotype of type 2B von Willebrand disease: a median 16-year follow-up study

BackgroundType 2B von Willebrand disease (VWD) is a bleeding disorder caused by gain-of-function variants in the VWF gene. The laboratory and clinical phenotype of type 2B VWD is heterogeneous. ObjectivesWe investigated associations between genotype and phenotype over a median of 16 years follow-up in a large cohort of well-characterized patients. MethodsWe included 64 genetically confirmed type 2B VWD patients from the national multicenter “Willebrand in the Netherlands” study and retrospectively collected clinical and laboratory data from electronic patient records. We analyzed associations between genotype and thrombocytopenia, bleeding phenotype, and events leading to endothelial activation and von Willebrand factor (VWF) secretion, including surgery, desmopressin administration, pregnancy, and delivery. ResultsThrombocytopenia manifested in 67.2% of patients, with varying occurrences between genetic variants (p.Arg1306Trp: 75.0%, p.Arg1308Cys: 58.3%). The most important determinant of thrombocytopenia was the p.Arg1306Trp VWF variant (odds ratio, 25.1). Platelet counts strongly varied over time and were continuously <150 × 109/L in 37.5% of patients with p.Arg1306Trp vs 8.3% in p.Arg1308Cys. In our analysis, endothelial activation was not an independent determinant (odds ratio, 1.3) for thrombocytopenia occurrence. No association was found between thrombocytopenia and cumulative bleeding scores or annual bleeding rates. Four women showed declining platelet counts in all full-term pregnancies (n = 8) during the third trimester with a sharp decrease in the week before delivery. Postpartum hemorrhage, defined as >500 mL estimated blood loss at delivery, occurred in 5 of 8 deliveries, despite prophylactic treatment with VWF concentrates. ConclusionThis study reveals a strong association between VWF variant p.Arg1306Trp and thrombocytopenia in type 2B VWD patients.

Higher Prevalence of Hepatitis B and C Infections among Indian Patients with Von Willebrand Disease

Background: The use of recombinant coagulation factors has reduced the incidence of hepatitis B and C infections in hemophilia patients. As recombinant von Willebrand factor is not easily available, patients with von Willebrand disease (VWD) may be at higher risk for acquiring hepatitis B and C infections. The prevalence of hepatitis B and C infections in the Indian population is ~0.95% and ~0.3%, respectively. Aims: This single tertiary center study aimed to assess the prevalence of hepatitis B and C infections and the profile of liver disease caused by these viruses among patients with VWD. Methodology: We retrospectively enrolled VWD patients treated in our center from January 2012 to December 2022 and analyzed the prevalence of hepatitis B and C infection and stage of liver disease. Results: Among 189 patients with VWD during the study period, 5 of 61 (8.1%) VWD patients screened were hepatitis B surface antigen positive (age: 34 [24–42] years; median (range); plasma VWF antigen: 10.1, [0–40] IU/dL). One patient had cholangiohepatoma at presentation, whereas none had chronic liver disease. Four of five patients received multiple bloodproduct transfusions before coming to our institution, with 10 (7–18) years interval between 1st transfusion and to detection of hepatitis B infection. Four of the 47 VWD patients (8.5%) screened were positive for hepatitis C virus antibody (48 [43–59] years, plasma VWF antigen: 20 [0–21.5] IU/dL). One patient had chronic liver disease. All four patients received multiple blood product transfusions before coming to our institution, with 15.5 (2–39) years interval between 1st transfusion and to the detection of hepatitis C infection. Conclusions: The prevalence of hepatitis B infection (8.1%) and hepatitis C infection (8.5%) was 8-fold and 28-fold higher, respectively, in VWD patients than the general population in India. VWD patients remain at high risk for acquiring transfusion-transmitted viral infections and appropriate interventions are needed to address this.

Epidemiological, Demographic, and Clinical Characteristics of Von Willebrand Disease Patients in Zahedan City, Iran: A Descriptive Study.

Background:Von Willebrand disease (VWD) is one of the most common coagulative diseases, so identifying the effective factors in preventing this complication is essential. The study aimed to evaluate the frequency of demographic and epidemiological findings in VWD patients referred to a hospital in Zahedan, Iran. Materials and Methods: This study was performed on 76 patients with VWD referred to Hazrat Ali-Asghar Hospital in Zahedan City, Sistan, and Baluchestan province. After obtaining consent from the patients, the demographic information and clinical symptoms of the disease were recorded. All statistical analyses were performed using SPSS 22.0 software. All descriptive data were expressed as mean ±SD and percent (%) depending on the continuous and dichotomous variables. A P-value ≤0.05 was considered significant statistically. Results:The present study results showed that the highest age group of VWD patients at the time of disease diagnosis was in the age group 1-5 years (47.3%), and most patients had type III VWD (80.3%). It was also found that 67.1% of patients had a positive family history and their parents' consanguineous marriage (77.6%). The most common complaints were epistaxis (88.15%), cutaneous bleeding (78.94%), and oral cavity bleeding (61.84%), respectively. Conclusion: Due to the high prevalence of VWD in consanguineous marriages and an increase in adverse complications and symptoms in VWD patients, proper diagnosis and screening at an early age, especially in people with family history, is essential. Efforts are needed to develop national registries and widely provide the required and available basic services for diagnosis and treatment.

Current Diagnosis of von Willebrand Disease in Italy: 3 Years Following the Release of the International Guidelines.

The American Society of Hematology-International Society on Thrombosis and Haemostasis-National Hemophilia Foundation-World Federation of Hemophilia 2021 International Guidelines (IGL) on von Willebrand disease (VWD) have pointed out many challenges, mainly in the diagnostic approach of VWD patients. To determine the impact of these IGL on the current clinical and laboratory diagnosis of Italian VWD patients, we have recently conducted a survey among 43 centers affiliated with the Italian Association of Hemophilia Centers (AICE). Directors and colleagues responsible for the management of VWD patients were invited to report in a detailed questionnaire how IGL recommendations about the assessment of the specific activities of von Willebrand Factor (VWF) could be applied at their local sites. Results from such a survey showed that bleeding assessment tools, VWF antigen, and factor VIII procoagulant are currently in use in all centers. The automated assays for platelet-dependent VWF activity with or without ristocetin described in IGL have been used since 2021 in 37/43 (86%) centers. Among other laboratory tests, VWF collagen binding, ristocetin-induced platelet agglutination, multimeric analysis, VWF propeptide, VWF:FVIII binding assay were available in 49, 63, 26, 7, and 28% of AICE, respectively. Analyses of VWF gene defects are available only at 3/43 (7%) centers. Desmopressin (DDAVP) infusion trials at diagnosis, with measurements of VWF activities at 1 and 4 hours post-DDAVP, is currently performed at 38/43 (88%) centers. Based on this information, a simplified clinical diagnosis using a few automated tests before and after DDAVP has been proposed. Such a diagnostic approach will be validated prospectively in a large cohort of Italian VWD patients.

Hemostatic management of von Willebrand disease during childbirth with a plasma-derived von Willebrand factor/factor VIII concentrate

BackgroundFemales with von Willebrand disease (VWD) do not show the same increases in von Willebrand factor and factor (F)VIII levels during pregnancy as females without VWD and are at higher risk of excessive bleeding associated with childbirth. Data on hemostatic management for childbirth in VWD patients are limited. ObjectivesTo evaluate the dosing, efficacy, and safety of plasma-derived von Willebrand factor/FVIII (wilate) for prevention of excessive bleeding associated with childbirth in females with any type of VWD. MethodsData for females with VWD who received wilate for hemostatic coverage for childbirth during 2 prospective clinical studies were analyzed. ResultsTen females with VWD and a mean age at enrolment of 29.6 years were treated with wilate to prevent excessive bleeding associated with childbirth. Two patients had type 1, 4 had type 2 (2 2A, 1 2B, and 1 2M), and 4 had type 3 VWD. Of the 10 deliveries, 5 were by cesarean section. Patients received a mean of 9.5 infusions of wilate over 6.8 exposure days, with a mean total dose of 234 IU/kg per delivery and 25 IU/kg per infusion. Hemostatic management for all deliveries was rated excellent or good, with no excessive bleeding during delivery and no postpartum bleeding during the period of wilate treatment in any patient. Two patients experienced 8 possible or probable treatment-related adverse events; all were mild or moderate and resolved. No thromboembolic events were observed. ConclusionThe results of this case series indicate that wilate provided effective hemostatic cover for childbirth in females with VWD during delivery and postpartum.

Genotyping R1336X and Eliminating the Pseudogene Amplification in Type 3 von Willebrand Disease Patients

Genotyping R1336X and Eliminating the Pseudogene Amplification in Type 3 von Willebrand Disease Patients

Diagnosis and treatment challenges in lower resource countries: State-of-the-art.

The 2022 World Federation of Haemophilia Annual Global Survey (AGS) reports that 454,690 patients with inherited bleeding disorders (IBD) have been identified globally. While this represents noteworthy progress, haemophilia epidemiology data indicate that 75% of people with inherited bleeding disorders living in low-income and low-to-middle-income countries have yet to be diagnosed. The AGS also revealed that 11 billion clotting factor units are available to treat haemophilia A and B globally. Due to a lack of finance, these treatments are unavailable to haemophilia in low-income countries with a consequence lack of access equity for haemophilia treatment in these communities. This sobering reality is not limited to haemophilia but applies to von Willebrand Disease (VWD). While VWD is the most prevalent IBD, only 103,844 people living with this condition have been diagnosed globally. Of the diagnosed patients, only a fraction live in low- or middle-income countries. Moreover, the majority of VWD patients are still treated sub-optimally without replacement therapies or prophylaxis, both of which are now accepted as global standards of care. In this state-of-the-art review, the authors reflect on three issues. First, the minimum elements required to diagnose haemophilia in a resource-constrained setting are identified. Second, this review points to the critical stakeholders and outlines their roles in removing access to haemophilia treatment barriers. Finally, the authors examine von Willebrand disease's ongoing diagnostic and treatment challenges and compare these to haemophilia. With the rapidly evolving novel therapies, the therapeutic landscape of all IBD will likely change for the better.

Targeting a higher plasma VWF level at time of delivery in pregnant individuals with von Willebrand disease: Outcomes at a single-institution cohort study.

Guidelines on the management of pregnant individuals with von Willebrand disease (VWD) at the time of delivery recommend that von Willebrand factor (VWF) and factor VIII:C (FVIII:C) levels be ≥50% to prevent postpartum haemorrhage (PPH). Yet, high PPH rates persist despite these levels or with prophylactic factor replacement therapy to achieve these levels. The current practice at our centre has been to target peak plasma VWF and FVIII:C levels of ≥100IU/dL at time of delivery. The objective of this study was to describe obstetric outcomes in pregnant individuals with VWD who were managed at our centre. Demographics and outcomes on pregnant individuals with VWD who delivered between January 2015 and April 2023 were collected. Forty-seven singleton deliveries (among 41 individuals) resulting in 46 live births and one foetal death were included. Twenty-one individuals had at least one prior birth by the start date of this study, of which 11 (52.4%) self-reported a history of PPH. Early PPH occurred in 12.8% (6/47) of deliveries. Two individuals required blood transfusion, of which one also had an unplanned hysterectomy and transfer to ICU. There were no thrombotic events reported. The strategy of targeting higher peak plasma VWF and FVIII:C levels (≥100IU/dL) at the time of delivery may be effective in reducing the risk of delivery-associated bleeding complications in VWD patients. Yet, the rate of early PPH remains unsatisfactory compared to the non-VWD population.

Progress in von Willebrand Disease Treatment: Evolution towards Newer Therapies.

von Willebrand disease (VWD) is a very heterogenous disease, resulting in different phenotypes and different degrees of bleeding severity. Established therapies (i.e., desmopressin, antifibrinolytic agents, hormone therapy for heavy menstrual bleeding, and von Willebrand factor [VWF] concentrates) may work in some subtypes, but not in all patients. In recent years, progress has been made in improving the diagnosis of VWD subtypes, allowing for more specific therapy. The impact of VWD on women's daily lives has also come to the fore in recent years, with hormone therapy, tranexamic acid, or recombinant VWF as treatment options. New treatment approaches, including the replacement of lacking factor VIII (FVIII) function, may work in those subgroups affected by severe FVIII deficiency. Reducing the clearance of VWF is an alternative treatment pathway; for example, rondaptivon pegol is a VWFA1 domain-binding aptamer which not only improves plasma VWF/FVIII levels, but also corrects platelet counts in thrombocytopenic type 2B VWD patients. These approaches are currently in clinical development, which will be the focus of this review. In addition, half-life extension methods are also important for the improvement of patients' quality of life. Targeting specific mutations may further lead to personalized treatments in the future. Finally, a few randomized controlled trials, although relatively small, have been published in recent years, aiming to achieve a higher level of evidence in future guidelines.

Haemostaseological Changes of VWF and FVIII during Pregnancy and the Oestrus Cycle in a Porcine Model of Von Willebrand Disease

Pregnancy and the oestrus cycle are challenging for female patients suffering from von Willebrand disease (VWD). Therefore, our study aimed to investigate the changes in von Willebrand factor (VWF) and factor VIII (FVIII) during pregnancy and the oestrus cycle in our porcine model of von Willebrand disease compared with the wild-type. Plasma analyses regarding primary hemostasis, secondary hemostasis, and VWF multimers, as well as immunohistochemistry analyses of VWF in the uterus and ovary, were performed. For levels of VWF and FVIII activities, significant elevations were seen in the last trimester. Primary hemostasis improved towards the end of pregnancy. In the oestrus cycle, significantly lower VWF values can be seen in the immunohistochemistry of the ovaries during the oestrus, while values were highest in the metoestrus. VWF multimer patterns in pigs were similar to the ones in human VWD patients. In summary, the course of VWF and FVIII during pregnancy and the oestrus cycle in porcine VWD were investigated for the first time. The porcine model seems to be suitable for haemostaseological studies on VWD. This provides an advantage for investigating reproduction-related bleeding and understanding the underlying mechanisms of post-partum hemorrhage or miscarriage in women with VWD.

Von Willebrand factor/factor VIII concentrate (Wilate) prophylaxis in children and adults with von Willebrand disease

AbstractLong-term prophylaxis with a von Willebrand factor (VWF) concentrate is recommended in patients with von Willebrand disease (VWD) who have a history of severe and frequent bleeds. However, data from prospective studies are scarce. WIL-31, a prospective, noncontrolled, international phase 3 trial, investigated the efficacy and safety of Wilate prophylaxis in severe patients with VWD. Male and female patients 6 years or older with VWD types 1, 2 (except 2N), or 3 who had completed a prospective, 6-month, on-demand, run-in study (WIL-29) were eligible to receive Wilate prophylaxis for 12 months. At baseline, patients (n = 33) had a median age of 18 years. Six (18%) patients had severe type 1, 5 (15%) had type 2, and 22 (67%) had type 3 VWD. The primary end point of a >50% reduction in mean total annualized bleeding rate (TABR) with Wilate prophylaxis vs prior on-demand treatment was met; mean TABR during prophylaxis was 5.2, representing an 84.4% reduction. The bleeding reduction was consistent across age, sex, and VWD types. The mean spontaneous ABR was 3.2, representing an 86.9% reduction vs on-demand treatment. During prophylaxis, 10 (30.3%) patients had 0 bleeding events and 15 (45.5%) patients had 0 spontaneous bleeding events. Of 173 BEs, 84.4% were minor and 69.9% treated. No serious adverse events related to study treatment and no thrombotic events were recorded. Overall, WIL-31 showed that Wilate prophylaxis was efficacious and well-tolerated in pediatric and adult patients with VWD of all types. The WIL-29 and WIL-31 trials were registered at www.ClinicalTrials.gov as #NCT04053699 and #NCT04052698, respectively.

Characterization and Quantification of Von Willebrand Factor Proteoforms in Von Willebrand Disease Patients and Healthy Controls: Insights from the Win-Study

Background: Von Willebrand disease (VWD) is a commonly inherited bleeding disorder which is characterized by either low (type 1) or absent (type 3) levels of Von Willebrand factor (VWF) or by the occurrence of dysfunctional VWF proteoforms (type 2) in plasma. The complexity of diagnosing and managing VWD arises from limited understanding of the diverse VWF proteoforms present in both healthy individuals as well as in VWD patients. Objective: The aim of this study was to investigate the plasma proteome and to characterize VWF proteoforms in Von Willebrand disease (VWD) patients, that were enrolled in the Willebrand in the Netherlands (WiN) 1 study, compared with healthy controls. Method: We selected type 1, type 2A, type 2M, type 2N and type 3 patients from the WiN cohort for which extensive laboratory- and genotype data were available. Plasma from 64 VWD patients and ten healthy controls was analyzed using a combination of discovery- and targeted mass spectrometry-based approaches. Results: Both discovery-based as well as targeted quantitative proteomics analysis revealed significant variation in VWF levels across the selected VWD patient group, which was in line with reported clinical laboratory data. Elevated VWF-propeptide/VWF ratios were observed in plasma samples from a range of VWD patients, with notable increases observed in VWD type 2A, B, and M, indicating accelerated clearance of these VWF proteoforms. Discovery-based proteomics revealed a strong VWF-F8 relationship in all VWD subtypes and healthy controls, except for type 2N VWD. We were able to identify peptides corresponding with common polymorphisms ( e.g. p.Thr798Ala, p.Gln852Arg, p.Thr1381Ala), for which the observed peptide distribution was in line with the minor allele frequency reported by the Broad institute. In addition, we identified peptides corresponding with 16 pathogenic VWF variants (e.g. p.Arg1306Trp, p. Phe1293Leu, p.Cys1190Arg, p.Arg1374His or p.Tyr1584Cys) of which p.Cys1190Arg wild-type and variant peptide aligned with the zygosity at DNA level (Figure X). Based on spike-in with stable isotope-labeled peptides, the variant-to-wildtype ratios varied among VWD patients with distinct pathogenic variants, ranging from 0.5-2.5 in type 2M (p.Phe1293Leu) and type 1 patients (p.Tyr1584Cys), &amp;lt;1 in type 2A patients (p.Arg1374His), to &amp;gt;8 in type 2B patients (p.Arg1306Trp), suggesting distinct stoichiometry of VWF variants among these patients (Figure 2). Conclusion: Our study highlights the feasibility of VWF proteoform identification and quantification from plasma and suggest a potential role in unravelling their contribution to the biological variations and clinical diversity observed in VWD. 1 De Wee et al., Determinants of bleeding phenotype in adult patients with moderate or severe von Willebrand disease. Thromb Haemost. 2012 Oct;108(4):683-92.

Efficacy of Regular Prophylaxis with a Plasma-Derived Von Willebrand Factor/Factor VIII Concentrate in Reducing Nose Bleeds in Children and Adults with Von Willebrand Disease

Background: The use of a von Willebrand factor (VWF) concentrate for long-term prophylaxis is recommended for individuals with von Willebrand disease (VWD) who have a history of frequent and severe bleeds, regardless of their age or the type of VWD. Bleeding from the nose (epistaxis) is one of the most common types of bleeding in VWD patients. The WIL-31 study demonstrated the efficacy of prophylaxis with a plasma-derived VWF/factor VIII (pdVWF/FVIII) concentrate containing VWF and FVIII in a 1:1 activity ratio (wilate ®) in adults and children with VWD of all types. The primary endpoint was met, showing a decrease of 84% in the mean total annualized bleeding rate (ABR) compared with prior on-demand treatment. Aims: To investigate the efficacy of regular prophylaxis with a pdVWF/FVIII concentrate in reducing nose bleeds in people with VWD, compared with previous on-demand treatment. Methods: WIL-31 (NCT04052698) was a prospective, non-controlled, international, multicenter phase 3 trial that enrolled male/female patients, aged ≥6 years with VWD type 1 (VWF:RCo &amp;lt;30 IU/dL), type 2 (except 2N) or type 3. Prior to entering the WIL-31 study, all patients had received on-demand treatment with a VWF-containing product during a 6-month, prospective, observational, run-in study (WIL-29); patients who experienced at least 6 bleeding episodes, excluding menstrual bleeds, of which ≥2 were treated with a VWF-containing product, were eligible to enter WIL-31. Patients in WIL-31 received regular prophylaxis with pdVWF/FVIII 2-3 times per week at a dose of 20-40 IU/kg for 12 months. In this post-hoc analysis, nose bleeds during WIL-29/-31 were described, and the mean ABRs were compared. Results: The study population included 33 patients with a median (range) age of 18 (7-61) years; 9 (27.3%) patients were 6-11 years old, 6 (18.2%) were 12-16 years old, and 18 (54.5%) were ≥17 years old. All VWD types were represented: type 1 (n=6), type 2 (n=5); and type 3 (n=22). There were 173 breakthrough bleeds in total during WIL-31, of which nose bleeds were the most common (89/173; 51.4%). Nose bleeds occurred in 16 of 33 patients during prophylaxis vs 26 of 33 during on-demand treatment. Of the 89 nose bleeds, 17 were traumatic, 69 spontaneous, and 3 due to allergic reaction (not product related). The mean number of spontaneous nose bleeds per patient during prophylaxis was 2.1, 2.3, 1.2, and 2.2, for the overall population, and the 6-11, 12-16, and ≥17 years age groups, respectively. Eight nose bleeds (9.0%) were major (all in the same patient). The mean nose ABR was reduced by 76% during prophylaxis vs on-demand treatment. The reduction was consistent across age groups (74-77%, Figure 1) but not across VWD types, with mean nose ABR reductions of 48, 89, and 80% for VWD type 1, type 2, and type 3 patients, respectively. During prophylaxis, a higher number of VWD type 3 patients, compared with type 1 and type 2 patients, experienced nose bleeds (12 vs 3 vs 1). The mean duration of nose bleeds decreased during prophylaxis vs on-demand for 12-17 years old, ≥17 years old, type 2 and type 3 VWD patients, but increased for 6-11 years old and severe type 1 patients. No relationship was observed between the number of nose bleeds and patient age (R 2=0.082). The majority (80.9%; 72/89) of nose bleeds were treated with pdVWF/FVIII with 86.1% (62/72) requiring only a single infusion; two required concomitant treatment with tranexamic acid. All treated nose bleeds were rated as “successfully treated”. Conclusion: The data indicate thatprophylaxis with pdVWF/FVIII was effective at reducing nose bleeds in patients with VWD type 2 and 3, and across all age groups.

The Impacts of Genetic Polymorphisms on the Von Willebrand Factor Level in Type 1 Von Willebrand Disease

Background: von Willebrand disease (VWD) is the most common inherited bleeding disease, which results from deficiency or defect of von Willebrand factor (VWF). In type 1 VWD, there are only 53-81% patient whose mutations of VWF gene can be identified. Some studies reported that variants outside the VWF gene may also contribute to the VWF level. There are 7 genetic variants of single nucleotide polymorphism ( SNP ) reported to be associated with the variation of VWF levels, including CLEC4M, STXBP5, SCARA5, ABO, VWF, STAB2 and UFM1. The aims of this study were to explore the frequency of SNPs of the 7 genes among type 1 VWD patients and the effects of the 7 genetic variants on the VWF levels in type 1 VWD patients. Patients and methods: There were 62 patients ( the ratio of M/F: 1 ) with type 1 VWD enrolled in this study, including 35 (56.5%) patients with found mutations of VWF gene. Each patient's bleeding symptoms were assessed by the MCMDM-1 VWD bleeding score. The results of VWF levels: VWF:Ag and VWF activity ( VWF:ACL or VWF:RCo ) and FVIII: C at the VWD diagnosis were retrospectively collected, as shown in Table 1. Each enrolled patient's genomic DNA was extracted and the SNPs of the 7 gene were targeted by PCR amplification followed by direct sequencing. Results: There was high correlations between VWF:Ag and VWF activity ( r = 0.73, p &amp;lt; 0.0001 ) and between VWF:Ag and FVIII:C ( r = 0.60, p &amp;lt; 0.0001 ), respectively. The patients without variant ( A/A) of UFM1 has significantly lower median VWF:Ag of 33% than 44.7% of patients with heterozygous variant ( A/C ) type ( p = 0.033 ). Patients without variant (A/A ) of STXBP gene had both significantly lower median value of VWF:Ag ( 31.9% vs 46.1%, p = 0.017 ) and VWF activity (26.55% vs 30.4%, p = 0.0467 ) than those patients with heterozygous variant ( A/G ). Four alleles substitution of SNP of UFM1( C&amp;gt;A ) and STXBP genes ( G&amp;gt;A ) variant has remarkably lowest median VWF antigen level of 27.4%, which was significantly lower than 43.05% of the three alleles substitution ( p = 0.0014 ) and the 48.6% of the two alleles substitution ( p = 0.0279 ), as shown in Figure 1. Conclusions: Our study demonstrates that both C to A variant of UFM1 gene and G to A variant of STXBP5 gene were associated with lower VWF: Ag level than those without variant, respectively. The combination of four allelic variant of UFM1 and STXBP5 genes had the most reducing effect on VWF: Ag level.

Efficacy of Regular Prophylaxis with a Plasma-Derived Von Willebrand Factor/Factor VIII Concentrate in Reducing Heavy Menstrual Bleeding in Females with Von Willebrand Disease

Background: Heavy menstrual bleeding (HMB) is the dominant symptom affecting ~80-90% of women with VWD and is a common cause of outpatient visits and hospitalizations. Long-term prophylaxis with a von Willebrand factor (VWF) concentrate is recommended in von Willebrand disease (VWD) patients with a history of severe, frequent bleeds, and for women with VWD who experience HMB. A recent prospective study found that recombinant VWF was not superior to tranexamic acid at reducing HMB in women with VWD. The WIL-31 study demonstrated the efficacy of prophylaxis with a plasma-derived VWF/factor VIII (pdVWF/FVIII) concentrate containing VWF and FVIII in a 1:1 activity ratio (wilate ®) in adults and children with VWD of all types. The primary endpoint was met, showing a decrease of 84% in the mean total annualized bleeding rate (ABR) compared with prior on-demand treatment. The impact of pdVWF/FVIII prophylaxis on HMB was evaluated as a secondary endpoint of the WIL-31 study. Aims: To investigate the efficacy of regular prophylaxis with a pdVWF/FVIII concentrate in reducing the incidence of HMB in females with VWD who had previously been treated on demand. Methods: WIL-31 (NCT04052698) was a prospective, non-controlled, international, multicenter phase 3 trial that enrolled male/female patients, aged ≥6 years old with VWD type 1 (VWF:RCo &amp;lt;30 IU/dL), type 2 (except 2N) or type 3. Prior to entering the WIL-31 study, all patients had received on-demand treatment with a VWF-containing product during a 6-month, prospective, observational, run-in study (WIL-29). Patients in WIL-31 received regular pdVWF/FVIII prophylaxis 2-3 times per week at a dose of 20-40 IU/kg for 12 months. Exploratory analyses included comparison of heavy menstrual ABR (HMABR). HMB was defined as any menstrual bleed that impeded the ability to perform daily activities (such as work, housework, exercise, or social activities) during menstrual periods. Criteria for HMB included changing pads more frequently than hourly, menstrual bleeding lasting 7 or more days, or the presence of clots &amp;gt;1 cm combined with a history of flooding or a Pictorial Blood Assessment Chart (PBAC) score ≥185. PBAC scores were reviewed from patient diaries at each study visit. Results: Of the 33 patients included in the overall population, 14 (42%) were female and half (n=7; 13-43 years old) were of childbearing age. In these females, all VWD types were represented: type 1 (n=2), type 2 (n=1); and type 3 (n=4). Six (86%) females were on hormonal birth control during WIL-31. Prophylaxis with pdVWF/FVIII reduced the mean HMABR by 64% compared with on-demand treatment (3.0 vs 8.3 in WIL-31 and WIL-29, respectively). During on-demand treatment, 86% (6/7) of females experienced ≥1 HMB episode, which was reduced to 29% (2/7) during prophylaxis with four patients experiencing no HMB (Figure 1). During on-demand treatment, 6 females experienced 32 HMB episodes; of these, 7 (22%) required further treatment with pdVWF/FVIII and 2 (6%) required concomitant medication. Under prophylaxis, two patients (aged ≥17 years), experienced 12 HMBEs; of these, only one (8%) impeded daily activity and none required additional pdVWF/FVIII treatment, hospitalization, or concomitant medication. Mean (SD) intra-individual median PBAC scores (N=4) decreased by 43% from 227 (91.7) during on-demand treatment to 131 (59.9) with prophylaxis, a 96-point reduction. Conclusion: Prophylaxis with pdVWF/FVIII was efficacious in reducing HMB compared with on-demand treatment in 7 female patients with VWD. None of the cases of HMB during prophylaxis was severe enough to require additional treatment. Among women with VWD, HMB remains a major burden and further research is required.

Effect of Factor VIII and Von Willebrand Factor Activities on Number of Bleeding Events in Previously Treated Patients with Von Willebrand Disease on Regular Prophylaxis with a Plasma-Derived Von Willebrand Factor/Factor VIII Concentrate

Background: von Willebrand disease (VWD) is characterized by a deficiency or reduction of activity in von Willebrand factor (VWF). As VWF has a role in stabilizing coagulation factor VIII (FVIII), VWD patients often suffer from secondary FVIII deficiency. Current guidelines recommend that VWD patients with a history of severe and frequent bleeds should use long-term prophylaxis with factor replacement therapy. The WIL-31 study demonstrated the efficacy of prophylaxis with a plasma-derived VWF/factor VIII concentrate containing VWF and FVIII in a 1:1 activity ratio (pdVWF/FVIII; wilate ®) in adults and children with VWD. The primary endpoint was met, showing a decrease of 84% in the mean total annualized bleeding rate (ABR). As elevated levels of FVIII are associated with an increased risk for thrombosis, it is important to determine whether repeated dosing with pdVWF/FVIII leads to accumulation of FVIII over time. Aims: To investigate the activity levels of VWF and FVIII in patients with VWD receiving regular prophylaxis with pdVWF/FVIII. Methods: WIL-31 (NCT04052698) was a prospective, non-controlled, international, multicenter phase 3 trial that enrolled male/female patients, aged ≥6 years old with VWD type 1 (VWF:RCo &amp;lt;30 IU/dL), type 2 (except 2N) or type 3. Prior to entering the WIL-31 study, all patients had received on-demand treatment with a pdVWF/FVIII concentrate during a 6-month, prospective, observational, run-in study (WIL-29). Patients in WIL-31 received regular pdVWF/FVIII prophylaxis 2-3 times per week at a dose of 20-40 IU/kg for 12 months. VWF activity (VWF:RCo) and FVIII activity (measured by the chromogenic [Chr] and one-stage [OS] assays) were performed at baseline and throughout the study. Full pharmacokinetic (PK) analyses were performed in all patients aged 6-16 years, a single dose of 60 ± 10 IU/kg pdVWF/FVIII was administered. Samples were collected at up to 1 hour prior to injection and then at multiple time points post-injection (1-72 hours). Safety, including the occurrence of thrombotic events, was monitored throughout the study. Results: The overall study population included 33 patients, with a median age of 18 years consisting of 9 (27.3%) patients aged 6-11 years, 6 (18.2%) patients aged 12-16 years, and 18 (54.5%) patients aged ≥17 years. Six (18.2%) patients had severe type 1 VWD, 5 (15.2%) patients had type 2, and 22 (66.7%) patients had type 3. Baseline FVIII and VWF activities in WIL-31 were similar regardless of age group, and as expected were lower in patients with type 3 VWD compared with severe type 1 and type 2 (Figure 1). There was a negative correlation of baseline FVIII and VWF activity with the number of spontaneous bleeding episodes (BEs) during prophylaxis. Patients who had no spontaneous BEs during prophylaxis had higher baseline FVIII and VWF levels compared with those who had spontaneous BEs. Mean peak and trough FVIII and VWF activity levels remained consistent during the study and there was no accumulation of either factor over the 12-month prophylaxis period (Figure 2). Full PK assessments were performed according to protocol in 13 patients aged 6-16 years. The mean ± standard deviation in vivo half-life was 16.1±6.7, 16.2±7.7, and 9.8±7.7 hours, for FVIII Chr, FVIII OS, and VWF:RCo, respectively. No thrombotic events were observed. Conclusion: No accumulation of FVIII or VWF in the plasma was reported in 33 patients who received regular prophylaxis with pdVWF/FVIII for 12 months, regardless of VWD type and age.

Impact of Anticoagulant Function By the Addition of Heparinoid in Vitro in the Plasmas from Patients with Hemophilia and Von Willebrand Disease

Background: Heparinoid (Hirudoid ®) is a local anticoagulant preparation as skin application that relieves pain and inflammation with addition to promoting healing in superficial thrombophlebitis and bruising. Heparinoid is mainly composed of chondroitin sulfate and has high moisturizing ability due to the large number of hydrophilic group. Thus, it is used as topical treatments for sebum deficiency in Japan. Since heparinoid has a structure similar to heparin, the package insert states the contraindication for hemorrhagic diseases such as hemophilia and von Willebrand disease (VWD) in Japan. Hemophilia A and B (HA/HB) are inherited bleeding disorders caused by a defect in factor (F) VIII and FIX coagulant protein, respectively. Based on its activity levels, the clinical severity is classified as severe (&amp;lt;1 IU/dL), moderate (1-5 IU/dL), and mild (&amp;gt;5-40 IU/dL). VWD is the common congenital bleeding disorders and is caused by a deficiency of von Willebrand factor (VWF). VWD is classified into partial deficiency (type 1), qualitative deficiency (type 2), and complete quantitative deficiency (type 3) of VWF, based on the VWF antigen (VWF:Ag) and ristocetin cofactor activity (VWF:RCo) level. In the clinical practice, there are cases of hemorrhagic patients using heparinoid at the discretion of their families. However, the influence of heparinoid on coagulant function in human subjects remains to be investigated, and it is unclear if the patients with hemophilia and VWD can use heparinoid safely. Aim: To examine the impact of anticoagulant function of heparinoid in vitro in the plasmas from patients with hemophilia and VWD. Methods: The maximum blood concentration from absorbed heparinoid was estimated to be 2.5 µg/mL, based on the previous report, when heparinoid was applied to the skin as stated in package insert. Heparinoid (0-6.5 µg/mL) or unfractionated heparin (0-0.6 IU/mL) were added to normal plasma (NP), followed by measuring activated partial thromboplastin time (aPTT) by one-stage clotting assay. A modified clot waveform analysis (CWA) using an aPTT reagent were performed on the CS-2400™ (Sysmex), followed by measuring clotting time (CT) and adjusted maximum coagulation velocity (Ad|min1|). Heparinoid was also added to the patients' plasmas with HA, HB, or VWD, followed by CWA assessment. The CT and Ad|min1| of 20 healthy individuals were 31.1±0.1 sec and 6.90±0.05 (median±SEM). Results: aPTT and CT values in NP by the spike of heparinoid were prolonged and Ad|min1| was decreased dose-dependently. The addition of 2.5 µg/mL heparinoid showed that aPTT (34.3 sec; 1.2-fold of control (no heparinoid)), CT (36.3 sec; 1.2-fold of control), and Ad|min1| (9.5; 0.96-fold of control) were comparable to the effect of ~0.2 IU/mL heparin, suggesting that heparinoid exhibited slight anticoagulant effect for NP. To examine the effect of heparinoid on HA/HB plasmas, recombinant (r) FVIII or rFIX (0-50 IU/dL) were added to FVIII- or FIX-deficient plasmas (3 lots each) together with heparinoid (0-6.3 µg/mL), respectively. The addition of heparinoid had prolonged the CT and decreased the Ad|min1| at each concentration of FVIII and FIX dose-dependently. The addition of 2.5 µg/mL heparinoid showed modest anticoagulant effect for HA and HB at each concentration of FVIII and FIX (CT; 1.3- and 1.5-fold of control, Ad|min1|; 0.82- and 0.87-fold of control). Next, the effect of heparinoid on the plasma from one VWD patient each with type 1 (VWF:Ag/VWF:RCo 28 IU/dL/26 IU/dL), type 2 (20 IU/dL/9 IU/dL), and type 3 (&amp;lt;2 IU/dL/&amp;lt;3 IU/dL) was assessed. The addition of 2.5 µg/mL heparinoid showed slight anticoagulant effect for each type of VWD (CT; ~1.3-fold of control, Ad|min1|; ~0.91-fold of control). Conclusion: The presence of heparinoid demonstrated mild anticoagulant effect in vitro in the plasmas of patients with hemophilia and VWD. It might be possible to apply the heparinoid-contained product as skin treatment for these patients.

Population-based prevalence and mutational landscape of von Willebrand disease using large-scale genetic databases

Von Willebrand disease (VWD) is a common bleeding disorder caused by mutations in the von Willebrand factor gene (VWF). The true global prevalence of VWD has not been accurately established. We estimated the worldwide and within-population prevalence of inherited VWD by analyzing exome and genome data of 141,456 individuals gathered by the genome Aggregation Database (gnomAD). We also extended our data deepening by mining the main databases containing VWF variants i.e., the Leiden Open Variation Database (LOVD) and the Human Gene Mutation Database (HGMD) with the goal to explore the global mutational spectrum of VWD. A total of 4,313 VWF variants were identified in the gnomAD population, of which 505 were predicted to be pathogenic or already reported to be associated with VWD. Among the 282,912 alleles analyzed, 31,785 were affected by the aforementioned variants. The global prevalence of dominant VWD in 1000 individuals was established to be 74 for type 1, 3 for 2A, 3 for 2B and 6 for 2M. The global prevalences for recessive VWD forms (type 2N and type 3) were 0.31 and 0.7 in 1000 individuals, respectively. This comprehensive analysis provided a global mutational landscape of VWF by means of 927 already reported variants in the HGMD and LOVD datasets and 287 novel pathogenic variants identified in the gnomAD. Our results reveal that there is a considerably higher than expected prevalence of putative disease alleles and variants associated with VWD and suggest that a large number of VWD patients are undiagnosed.

Characterization of copy-number variants in a large cohort of patients with von Willebrand disease reveals a relationship between disrupted regions and disease type

BackgroundGenetic analysis for von Willebrand disease (VWD) commonly utilizes DNA sequencing to identify variants in the von Willebrand factor (VWF) gene; however, this technique cannot always detect copy-number variants (CNVs). Additional mapping of CNVs in patients with VWD is needed. ObjectivesThis study aimed to characterize CNVs in a large sample of VWF mutation-negative VWD patients. MethodsTo determine the role of CNVs in VWD, a VWF high-resolution comparative genomic hybridization array was custom-designed to avoid multiple sequence variations, repeated sequences, and the VWF pseudogene. This was performed on 204 mutation-negative subjects for whom clinical variables were also available. ResultsAmong the 204 patients, 7 unique CNVs were found, with a total of 24 CNVs (12%). Of the 7 unique CNVs, 1 was novel, 1 was found in a VWF database, and 5 were previously reported. All patients with type 1C VWD and a CNV had the same exon 33 and 34 in-frame deletion. Certain clinical variables were also significantly different between those with and without CNVs. ConclusionThe in-frame deletion in patients with type 1C VWD exactly matches the D4N module of the D4 domain, a region where mutations and deletions are known to affect clearance. We observed significantly higher VWF–to–ristocetin cofactor levels in patients with type 1C VWD and a CNV than in patients without a CNV, suggesting a relationship between CNVs and the increased clearance observed in patients with type 1C VWD. Glycoprotein IbM activity was significantly lower in patients with type 1 VWD and a CNV than in patients without a CNV, suggesting that platelet binding is more affected by CNVs than single base pair mutations. This work elucidates some of the underlying genetic mechanisms of CNVs in these patients.

The necessity of repeat testing for von Willebrand disease in adult patients with mild to moderate bleeding disorders

BackgroundIn patients with mild-to-moderate bleeding disorders (MBD), von Willebrand disease (VWD) is diagnosed at von Willebrand factor (VWF) levels ≤50 IU/dL. Although VWF levels are unstable, repeated testing for VWD diagnosis is not necessarily advised in recent guidelines. ObjectivesTo analyze the relevance of repeated VWF testing to diagnose VWD in patients with MBD. MethodsData of 277 patients with MBD from the Vienna Bleeding Biobank with at least 2 separate assessments of VWF antigen (VWF:Ag) and activity (VWF:Act) were analyzed. ResultsIn repeated VWF measurements, 36 patients (13.0%) had “changing” VWF levels (≤/>50 IU/dL), 27 (9.7%) had persistent levels ≤50 IU/dL (“pathologic”), and 214 (77.3%) had levels >50 IU/dL (“normal”). Of the 36 changing patients, 22 (61%) were diagnosed with VWD at baseline, whereas the others only met VWD diagnostic criteria at repeated measurements. Using logistic regression, we estimated a probability of change of 26.4% (95% CI, 12.5-47.4) at baseline VWF levels of 30 IU/dL, 50.8% (95% CI, 35.6-65.8) at 50 IU/dL, 18.8% (95% CI, 12.3-27.6) at 60 IU/dL, and 1.2% (95% CI, 0.3-4.9) at 80 IU/dL. Baseline VWF was a strong predictor for changing status (Χ2 = 49.9; P < .001), while age, sex, Vicenza score, and blood type O had limited added value (Χ2 = 5.1; P = .278). Baseline VWF:Ag or VWF:Act cutoffs of 80 IU/dL had negative predictive values of 98.1% and 99.1% for changing status, respectively. ConclusionOur data emphasize an overlap between patients with VWD and MBD with bleeding disorder of unknown cause and underline the need for repeated VWF testing, especially in patients with VWF levels <80 IU/dL.