Low Von Willebrand Factor Research Articles

Von Willebrand factor in follicular fluid in women undergoing IVF/ICSI: a “ghost” associated with oocyte and embryo quality

Introduction: The purpose of the present study to determine the presence of Von Willebrand factor (vWF) in the follicular fluid of women undergoing IVF/ICSI and to investigate the association of its concentration with oocyte and embryo quality. Methods: We conducted a prospective observational study based in women undergoing in vitro fertilization treatment. vWF concentrations in follicular fluid samples collected during transvaginal oocyte retrieval were measured using an enzyme-linked immunosorbent assay (Elisa). They were also analyzed by the VIDAS® vWF method for confirmation of the results. The latter is an automated quantitative test for the measurement of vWF in human plasma, using the ELFA technique (enzyme-linked fluorescent assay). vWF concentrations were correlated to oocyte and embryo quality. Results: Contrary to previous reports, vWF was determined in follicular fluid. A higher percentage of good quality oocytes were found in the presence of lower vWF follicular fluid levels. Similarly, better quality embryos were derived from oocytes retrieved from follicles with lower vWF follicular fluid levels. Conclusions: Contrary to existing evidence to date, vWF was found in the follicular fluid of women undergoing IVF/ICSI treatment. In addition, the quality of oocytes and embryos was adversely related to vWF follicular fluid levels. In fact, a better outcome of IVF/ICSI treatment was suggested when the levels of vWF in follicular fluid were low. In this context, vWF follicular fluid levels could be used as a prognostic factor for oocyte and embryo quality.

Platelet activation and aggregation in different centrifugal-flow left ventricular assist devices

Left-ventricular assist devices (LVADs) improve outcomes in end-stage heart failure patients. Two centrifugal-flow LVAD systems are currently approved, HeartMate 3 (HM3) and Medtronic/Heartware HVAD (HVAD). Clinical findings suggest differences in thrombogenicity between both systems. We compared markers of platelet activation and aggregation between HM3 and HVAD. We prospectively included 59 LVAD patients (40 HM3, 19 HVAD). Platelet P-selectin expression, activated glycoprotein (GP) IIb/IIIa and monocyte-platelet aggregates (MPA) were assessed by flow-cytometry. Platelet aggregation was measured by light-transmission aggregometry (LTA) and multiple-electrode aggregometry (MEA). Von-Willebrand factor (VWF) antigen (VWF:Ag), VWF activity (VWF:Ac), and VWF multimer pattern analysis were determined. Soluble P-selectin (sP-selectin) was measured with an enzyme-linked immunoassay. P-selectin, GPIIb/IIIa and MPA levels in vivo and in response to arachidonic acid, adenosine diphosphate, and thrombin receptor activating peptide were similar between HM3 and HVAD (all p > .05). Likewise, agonist-inducible platelet aggregation by LTA and MEA did not differ between HM3 and HVAD (all p > .05). VWF:Ag levels and FVIII:C were similar between both systems (both p > .05), but patients with HVAD had significantly lower VWF:Ac (p = .011) and reduced large VWF multimers (p = .013). Finally, sP-selectin levels were similar in patients with HVAD and HM3 (p = .845). In conclusion, on-treatment platelet activation and aggregation are similar in HM3 and HVAD patients. Potential clinical implications of observed differences in VWF profiles between both LVAD systems need to be addressed in future clinical trials.

Management of elective procedures in low von Willebrand factor patients in the LoVIC study

BackgroundMost individuals with mild to moderate reductions in plasma von Willebrand factor (VWF) levels do not demonstrate increased bleeding. However, some patients with plasma VWF levels of 30–50 IU/dl do have a significant bleeding phenotype. Management of these “low VWF” patients, who may have significant bleeding scores >10, around times of elective procedures continues to pose a common clinical challenge because of a lack of evidence. ObjectiveTo investigate the safety and efficacy of different periprocedural management options for adult patients with low VWF. MethodsTreatment and outcomes were retrospectively reviewed for 160 invasive procedures performed in 60 patients with well characterized low VWF enrolled in the previously described Low Von Willebrand factor Ireland Cohort study. ResultsWe demonstrate that 1‐desamino‐8‐D‐arginine vasopressin is efficacious in preventing bleeding for both minor or major elective procedures in adult low VWF patients, even in those with significant bleeding histories. In addition, tranexamic acid alone is effective for low VWF patients undergoing nondental minor procedures. Importantly, age‐related increases in plasma VWF:antigen levels above 50 IU/dl were not necessarily associated with complete correction of bleeding phenotype. Procedure‐related bleeding complications were increased in low VWF patients who did not receive any hemostatic cover before their procedure. ConclusionElective procedures in adult patients with low VWF should be managed in liaison with a comprehensive care tertiary referral center so that personalized treatment plans may be implemented before all minor or major elective procedures.

Criteria for low von Willebrand factor diagnosis and risk score to predict future bleeding

BackgroundImportant diagnostic and clinical aspects of moderately reduced von Willebrand factor (VWF) levels are still unknown. There is no clear evidence which cutoff value (0.50 vs 0.60 IU/ml) should be used to diagnose “low VWF.” Also, the incidence of bleeding after the diagnosis has been made, and risk factors for bleeding are unknown yet. ObjectivesTo investigate the incidence of postsurgical bleeding, postpartum hemorrhage (PPH), and traumatic and spontaneous bleeding after low VWF diagnosis, and to develop a risk score to predict future bleeding. MethodsWe performed a cohort study in patients with historically lowest VWF levels of 0.31 to 0.60 IU/ml. Clinical data of patients were retrospectively collected. ResultsWe included 439 patients with low VWF. During a follow‐up of 6.3 ± 3.7 years, 259 surgical procedures, 81 deliveries, and 109 spontaneous and traumatic bleeding episodes were reported. The incidence of postsurgical bleeding was 2.7%, whereas 10% of deliveries was complicated by PPH. Overall, 65 patients (14.8%) had bleeding requiring treatment, which was not different between patients with historically lowest VWF levels of 0.31–0.50 and 0.51–0.60 IU/ml (p = .154). Age <18 years, abnormal bleeding score at diagnosis, and being referred for bleeding symptoms at the time of diagnosis were independent risk factors for bleeding during follow‐up, and therefore included in the risk score. ConclusionsThe cutoff value of low VWF diagnosis should be set at 0.60 IU/ml. Furthermore, a risk score is developed to identify individuals with a high risk for bleeding after low VWF diagnosis.

Abstract P769: Remodeling of the Leptomeningeal Collaterals Following Stroke Involves Selective Upregulation of Von Willebrand Factor (VWF)

Introduction: The leptomeningeal collaterals (LMCs) are arterial anastomoses that interconnect the distal branches of the major cerebral arteries. They distribute blood flow from adequately to poorly perfused vascular territories when needed. In normal conditions, the LMCs experience little net flow. However, during ischemic stroke, the LMCs are recruited to provide rescue flow to the ischemic territory. During prolonged or permanent stroke, the LMCs can rapidly remodel to provide increased perfusion capacity. Von Willebrand factor (VWF) is a secreted protein expressed in cerebral endothelium and has recently been implicated in angiogenesis. We sought to determine if VWF expression was increased in remodeling LMCs in the post-stroke brain. Methods: C57BL6 mice were subjected to permanent distal middle cerebral artery occlusion (pdMCAO). At 3 days after pdMCAO, mice were perfused with fluorescent tomato lectin to label vascular endothelium. The cortical surface was “planed off” by vibratome to provide largely contiguous cortical surface vessel preparations containing distalmost arterioles (DMAs) and the connected LMCs. VWF immunofluorescence intensity was measured in LMCs and the connecting DMAs of hemispheres ipsilateral and contralateral to the stroke. Inner diameter of the LMCs and DMAs were measured with Image J. Results: At 3 days post-stroke, LMCs of the stroke hemisphere demonstrated significant outward remodeling. Mean inner diameter of the ipsilateral versus contralateral LMCs were 44.5 ± 5.9 um versus 28.2 ± 1.2 um, respectively (p&lt;0.02, 7-8 vessels from 3 mice). In contrast, the DMAs were unaltered in diameter, 35.2 ± 4.0 and 35.7 ± 3.1 um, respectively. VWF expression was significantly increased in the ipsilateral LMCs versus contralateral LMCs (2.8 fold, p&lt;0.05) and versus ipsilateral DMAs (3.7 fold, p&lt;0.02). The DMAs showed significantly lower VWF expression, which was unchanged between ipsilateral and contralateral hemispheres. Conclusions: We demonstrate that VWF is highly expressed in the remodeling LMCs (but not connecting DMAs) of the post-stroke brain. These findings suggest a novel and selective role for endothelial VWF in the LMC response to prolonged recruitment during ischemic stroke.

Toward Personalized Treatment for Patients with Low von Willebrand Factor and Quantitative von Willebrand Disease.

The biological mechanisms involved in the pathogenesis of type 2 and type 3 von Willebrand disease (VWD) have been studied extensively. In contrast, although accounting for the majority of VWD cases, the pathobiology underlying partial quantitative VWD has remained somewhat elusive. However, important insights have been attained following several recent cohort studies that have investigated mechanisms in patients with type 1 VWD and low von Willebrand factor (VWF), respectively. These studies have demonstrated that reduced plasma VWF levels may result from either (1) decreased VWF biosynthesis and/or secretion in endothelial cells and (2) pathological increased VWF clearance. In addition, it has become clear that some patients with only mild to moderate reductions in plasma VWF levels in the 30 to 50 IU/dL range may have significant bleeding phenotypes. Importantly in these low VWF patients, bleeding risk fails to correlate with plasma VWF levels and inheritance is typically independent of the VWF gene. Although plasma VWF levels may increase to > 50 IU/dL with progressive aging or pregnancy in these subjects, emerging data suggest that this apparent normalization in VWF levels does not necessarily equate to a complete correction in bleeding phenotype in patients with partial quantitative VWD. In this review, these recent advances in our understanding of quantitative VWD pathogenesis are discussed. Furthermore, the translational implications of these emerging findings are considered, particularly with respect to designing personalized treatment plans for VWD patients undergoing elective procedures.

Von Willebrand factor antigen levels are associated with burden of rare nonsynonymous variants in the VWF gene

Approximately 35% of patients with type 1 von Willebrand disease (VWD) do not have a known pathogenic variant in the von Willebrand factor (VWF) gene. We aimed to understand the impact of VWF coding variants on VWD risk and VWF antigen (VWF:Ag) levels, studying 527 patients with low VWF and VWD and 210 healthy controls. VWF sequencing was performed and VWF:Ag levels assayed. A combined annotation-dependent depletion (CADD) score >20 was used as a predicted pathogenicity measure. The number of rare nonsynonymous VWF variants significantly predicted VWF:Ag levels (P = 1.62 × 10−21). There was an association between average number of rare nonsynonymous VWF variants with VWD type 1 (P = 2.4 × 10−13) and low VWF (P = 1.6 × 10−27) compared with healthy subjects: type 1 subjects possessed on average >2 times as many rare variants as those with low VWF and 8 times as many as healthy subjects. The number of rare nonsynonymous variants significantly predicts VWF:Ag levels even after controlling for presence of a variant with a CADD score >20 or a known pathogenic variant in VWF (P = 2.7 × 10−14). The number of rare nonsynonymous variants in VWF as well as the presence of a variant with CADD >20 are both significantly associated with VWF levels. The association with rare nonsynonymous variants holds even when controlling for known pathogenic variants, suggesting that additional variants, in VWF or elsewhere, are associated with VWF:Ag levels. Patients with higher VWF:Ag levels with fewer rare nonsynonymous VWF gene variants could benefit from next-generation sequencing to find the cause of their bleeding.

Circulating neurotrophins and hemostatic risk factors of atherothrombotic cardiovascular disease at baseline and during sympathetic challenge: the SABPA study

Sympathetic activation may trigger acute coronary syndromes. We examined the relation between circulating neurotrophic factors and hemostatic risk factors of atherothrombotic cardiovascular disease at baseline and in response to acute mental stress to establish a brain–heart link. In 409 black and white South Africans, brain-derived neurotrophic factor (BDNF) and fibrinolytic measures were assessed at baseline. Glial cell-derived neurotrophic factor (GDNF), S100 calcium-binding protein (S100B), von Willebrand factor (VWF), fibrinogen and D-dimer were assessed at baseline and 10 min after the Stroop test. Neurotrophins were regressed on hemostatic measures adjusting for demographics, comorbidities, cardiometabolic factors and health behaviors. Higher baseline BDNF was associated with greater stress-induced increase in fibrinogen (p = 0.003) and lower D-dimer increase (p = 0.016). Higher baseline S100B was significantly associated with higher baseline VWF (p = 0.031) and lower fibrinogen increase (p = 0.048). Lower baseline GDNF was associated with higher baseline VWF (p = 0.035) but lower VWF increase (p = 0.001). Greater GDNF (p = 0.006) and S100B (p = 0.042) increases were associated with lower VWF increase. All associations showed small-to-moderate effect sizes. Neurotrophins and fibrinolytic factors showed no significant associations. The findings support the existence of a peripheral neurothrophin-hemostasis interaction of small-to-moderate clinical relevance. The implications for atherothrombotic cardiovascular disease need further exploration.

Increasing levels of von Willebrand factor and factor VIII with age in patients affected by von Willebrand disease

Unlabelled Box AbstractBackgroundIncreasing levels of von Willebrand factor (VWF) and factor VIII (FVIII:C) was associated with age in type 1 von Willebrand disease (VWD). ObjectivesTo evaluate VWF and FVIII:C increase with age in a large group of patients with VWD and low levels of VWF, in whom levels were repeatedly measured. MethodsClinical charts from all patients evaluated at the A. Bianchi Bonomi Center between 1970 and 2018 were reviewed and data on VWF and FVIII:C collected. Patients affected by type 3, severe type 1 and 2N VWD were excluded. The repeated measurements were evaluated by linear mixed‐effects models. A linear association between age and VWF/FVIII:C was shown after the age of 40 years in the linear mixed models and analyzed by calculating the regression slope coefficient (β). ResultsA total of 617 patients were included in the study (314 type 2, 112 type 1, 181 low VWF levels), with a median age at first measurement of 28 years (interquartile range 14/42) and a mean follow‐up of 16 years (standard deviation 11). VWF and FVIII:C increased with age in the whole group. The increase became linear after the age of 40 years (3.68 and 7.44 IU/dL per decade for VWF:activity and FVIII:C). In type 2, FVIII:C increased with age, whereas an increase of both VWF:activity and FVIII:C were shown in patients with type 1 VWD and low levels of VWF. ConclusionsA differential increase of VWF and FVIII:C with age was shown among in different ages and types of VWD.

Correcting dominant‐negative von Willebrand disease

Correcting dominant‐negative von Willebrand disease

Von Willebrand factor in aortic or mitral valve stenosis and bleeding after heart valve surgery

ObjectivesLow von Willebrand factor (VWF) increases the risk of bleeding. The objective was to assess the influence of VWF on bleeding after valvular surgery. MethodsWe studied 82 consecutive patients in median age of 65.5 years with severe isolated aortic stenosis (AS, n = 62) or mitral stenosis (MS, n = 20), undergoing heart valve surgery in extracorporeal circulation. Preoperatively, we assessed VWF antigen (VWF:Ag) and activity (VWF:RCo), a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13 (ADAMTS13), and fibrinolysis inhibitors. ResultsCompared with AS, MS patients were more frequently female (80 vs. 55%, p = 0.045) with atrial fibrillation (AF) (80 vs. 8%, p < 0.0001), with no difference in age or comorbidities. Median postoperative drainage was 420 ml for AS, and 425 ml for MS (p = 0.37). Patients with AS had lower VWF:RCo (125.8 [88.5–160.8] vs. 188.0 [140.3–207.3] IU/dl, p = 0.003) and VWF:Ag (135.8 [112.0–171.2] vs. 191.7 [147.3–236.4] IU/dl, p = 0.01) than MS patients. Mean VWF:RCo/Ag ratio was 0.88 ± 0.17, with no intergroup differences. ADAMTS13 levels and activity were similar in both groups. In AS, both VWF:RCo and VWF:Ag correlated inversely with maximal (r = −0.39, p = 0.0003 and r = −0.39, p = 0.0004, respectively) and mean (r = −0.40, p = 0.0004 and r = −0.39, p = 0.0006, respectively) transvalvular pressure gradients. There was no difference in perioperative bleeding between patients following mitral and aortic valve surgery, and bleeding was not associated with VWF:Ag or VWF:RCo. ConclusionsIn severe AS, VWF levels and activity correlate inversely with transvalvular pressure gradients, and are lower than in severe degenerative MS, but do not affect blood loss after valvular surgery in extracorporeal circulation.

Diagnosis of von Willebrand disease in Western Mexico.

Von Willebrand disease (VWD) is the most common inherited bleeding disorder with a prevalence of 0.1%, characterised by quantitative or functional deficiency of von Willebrand factor (VWF). VWD diagnosis is based on symptomology, biochemical and genetic tests, but limited laboratory resources and VWD heterogeneity still generate an important subdiagnosis gap worldwide and in our country. To identify the type and subtype of VWD in a cohort of patients with a history of excessive bleeding in Western Mexico. This prospective cohort study from 2012 to 2019 included patients with mucocutaneous bleeding or abnormal laboratory tests. A standardised questionnaire and confirmatory tests were applied: FVIII:C, VWF activity, VWF antigen, and VWF multimeric analysis. Of the 297 patients recruited, 207 (69.7%) were excluded because their values exceeded 50% in VWF activity and VWF antigen. Of those 90 remaining, 54 (18.2%) had low VWF, and only 36 patients (12.1%) were diagnosed with VWD. Among them, 17 (47.2%) had quantitative deficiencies, of whom 14 were assigned as type 1 and 3 as type 3.The remaining 19 cases were diagnosed as type 2 (52.8%): type 2A and 2B were the most frequent with 6 and 7 cases respectively; 4 cases were possible type 2M and two suggestive of 2N, however, this was not confirmed. This study highlights the challenges of VWD diagnosis using a comprehensive panel of diagnostic tests which should extend to supplemental tests of VWF:CB, VWF:FVIIIB, and sequencing the VWD gene to confirm the results from the panel assays.

How we diagnose 2M von Willebrand disease (VWD): Use of a strategic algorithmic approach to distinguish 2M VWD from other VWD types.

von Willebrand disease (VWD) is the most common inherited bleeding disorder and caused by an absence, deficiency or defect in von Willebrand factor (VWF). VWD is currently classified into six different types: 1, 2A, 2B, 2N, 2M, 3. Notably, 2M VWD is more often misdiagnosed as 2A or type 1 VWD than properly identified as 2M VWD. To describe an algorithmic approach to better ensure appropriate identification of 2M VWD, and reduce its misdiagnosis, as supported by sequential laboratory testing. Comparative assessment of types 1, 2A, 2B and 2M VWD using various laboratory tests, including VWF antigen and several VWF activity assays, plus DDAVP challenge data, ristocetin-induced platelet agglutination (RIPA) data, multimer analysis and genetic testing. Types 1, 2A, 2B and 2M VWD give characteristic test patterns that can provisionally classify patients into particular VWD types. Notably, type 1 VWD shows low levels of VWF, but VWF functional concordance (VWF activity/Ag ratios >0.6), with both baseline assessment and post-DDAVP. Types 2A, 2B and 2M VWD show VWF functional discordance (low VWF activity/Ag ratio(s)) dependent on the defect, but type 2M separates from 2A/2B VWD based on specific test patterns, especially with collagen binding vs glycoprotein Ib binding assays. RIPA identifies 2B VWD. Multimers separate 2M from 2A/2B. We provide strategies to improve correct diagnosis of VWD, especially focussed on 2M VWD, and which can be used by most diagnostic haemostasis laboratories, reserving genetic analysis (if required) for confirmation.

ADAMTS13 activity, high VWF and FVIII levels in the pathogenesis of deep vein thrombosis

BackgroundDeep vein thrombosis (DVT) is a common multi-factorial disease with a partially understood aetiology. Although the roles of high factor (F)VIII and von Willebrand factor (VWF) levels are recognized, that of ADAMTS13 is still unclear. AimTo assess the association between ADAMTS13 activity levels, VWF antigen (VWF:Ag) and FVIII coagulant activity (FVIII:C) levels and DVT. Materials and methods365 Italian DVT patients and 292 age- and sex-matched controls were considered. Plasma ADAMTS13 activity was measured using FRETS-VWF73 assay. VWF:Ag and FVIII:C were measured using immunoassay and one-stage clotting assay (ACL TOP analyzer), respectively. Quartile analyses were performed to evaluate the individual association between ADAMTS13 activity, VWF:Ag, FVIII:C and DVT. The combined effect of high VWF levels (> 4th quartile) and low ADAMTS13 levels (< 1st quartile) was evaluated using binary variables. All models were age- and sex-adjusted. Estimated risks were reported as Odds ratio (OR) with 95% confidence intervals (CI). ResultsADAMTS13 activity was lower in DVT patients (94% vs. 98% of controls). Patients with an ADAMTS13 activity <1st quartile (86%) showed a 1.6-fold increased risk of DVT (95%CI, 1.05–2.55). The combination of low ADAMTS13 activity and high VWF:Ag levels was associated with a 15–fold increased risk (95%CI, 7.80–33.80). VWF:Ag and FVIII:C were associated to DVT with a dose-response relationship. ConclusionsADAMTS13 activity < 86% was associated with a moderate risk of DVT. The co-presence of low ADAMTS13 activity and high VWF levels resulted in a strong synergistic effect on DVT risk. The association of VWF:Ag and FVIII:C with DVT was confirmed.

Low Von Willebrand Factor: Cut-Off Value for Diagnosis and Risk Score to Predict Bleeding Incidence

Introduction Although large studies have recently provided valuable insights on the diagnosis, bleeding phenotype, and treatment outcomes of VWD patients, these aspects remain poorly understood in individuals with low VWF. Firstly, there is no clear evidence which cut-off value should be used to diagnose low VWF. Although 0.50 IU/mL is the most recommended cut-off value, some centers use the lower limit of normal (0.60 IU/mL). Secondly, the incidence of post-surgical bleeding, postpartum hemorrhage (PPH) and traumatic- or spontaneous bleeding after diagnosis of low VWF are still unknown. Lastly, it is hard to predict which individuals with low VWF have an increased bleeding risk. Therefore, we investigated the bleeding phenotype of individuals with historically lowest VWF levels of 0.31-0.50 IU/mL and 0.51-0.60 IU/mL, and the incidence of post-surgical bleeding, PPH and traumatic- and spontaneous bleeding after their initial diagnosis of "low VWF". Methods We performed a retrospective cohort study from January 2007 to November 2019 at the Erasmus MC, University Medical Center Rotterdam. All patients evaluated for the presence of a bleeding disorder with VWF antigen (VWF:Ag) and/or VWF activity (VWF:Act) and/or VWF collagen binding (VWF:CB) levels between 0.31-0.60 IU/mL, were included. Patients with VWF:Ag and/or VWF:Act and/or VWF:CB ≤0.30 IU/mL, acquired VWD and those with a concomitant bleeding disorder were excluded. For each individual we collected data from electronic patient files on baseline characteristics, reason for referral, family history of bleeding disorders, ISTH-BAT and laboratory measurements at diagnosis. Retrospective follow-up started from initial date of low VWF diagnosis through November 2019, during which we collected data on surgical procedures, pregnancies, and incidence of spontaneous- and traumatic bleeding. Results We included 439 patients; 269 patients with historically lowest VWF levels 0.31-0.50 IU/mL and 170 patients 0.51-0.60 IU/mL. Mean age at diagnosis was 28.8 ±17.7 years. Most patients were female (74.3%) and had blood group O (76.4%, Table 1). The bleeding score (BS) was similar in patients with historically lowest VWF levels of 0.31-0.50 IU/mL (3.7 ±3.0) and 0.51-0.60 IU/mL (4.0 ±2.9, p=0.209, Table 1). During the mean follow-up period of 6.3 ±3.7 years, 259 surgical procedures were performed in 146 patients, 81 deliveries in 56 women, and 109 spontaneous- or traumatic bleedings in 71 patients. The incidence of post-surgical bleeding was 7 (2.7%) during follow-up, whereas 8 deliveries (10%) were complicated by PPH. Overall, 65 out of 439 patients (14.8%) had a bleeding episode requiring treatment during follow-up, resulting in an incidence of bleeding requiring treatment of 0.5 ±1.9 per patient per decade. No difference was found in the incidence of bleeding requiring treatment between patients with historically lowest VWF levels of 0.31-0.50 IU/mL and 0.51-0.60 IU/mL (Figure 2A, p=0.154). We found that referral for a personal bleeding diathesis, a younger age at diagnosis and an abnormal BS at diagnosis were strong and independent risk factors for bleeding requiring treatment during follow-up, respectively HR=2.32 (95%CI: 1.16-4.63), HR=1.18 (95%CI: 1.01-1.38) and HR=1.77 (95%CI: 1.04-3.01). These risk factors were combined to develop a risk score to identify low VWF patients with an increased risk for bleeding requiring treatment (Figure 2B). The risk score performed excellent to differentiate in bleeding requiring treatment between low risk, intermediate risk and high risk patients (p&amp;lt;0.001, Figure 2C). The number of patients with bleeding requiring treatment was 8/126 (6.3%) in patients with low risk, 18/143 (12.6%) in intermediate risk and 39/170 (22.9%) in high risk patients (p&amp;lt;0.001). Likewise, the incidence of bleeding requiring treatment per patient per decade was 0.22 ±1.08 in low risk, 0.28 ±1.25 in intermediate risk and 0.87 ±2.61 in high risk patients (p=0.004, Figure 2D). Conclusion To conclude, there is no difference in the bleeding phenotype of individuals with historically lowest VWF levels of 0.31-0.50 IU/mL and 0.51-0.60 IU/mL. Therefore, the cut-off value to diagnose low VWF should be set at 0.60 IU/mL. Furthermore, the risk score developed in the current study may assist to identify low VWF patients with low, intermediate and high risk for future bleeding. Disclosures Atiq: SOBI: Other: travel grant; CSL Behring: Research Funding. Kruip:Boehringer Ingelheim: Research Funding; Pfizer: Research Funding; Bayer: Research Funding; Daiichi Sankyo: Research Funding; SOBI: Research Funding; Bayer: Speakers Bureau. Cnossen:Takeda: Research Funding; Shire: Research Funding; Baxter: Research Funding; Bayer: Research Funding; Sobi: Research Funding; CSL behring: Research Funding; Nordic Pharma: Research Funding; Novo Nordisk: Research Funding; Pfizer: Research Funding. Leebeek:CSL Behring: Research Funding; Shire/Takeda: Research Funding; Uniqure: Consultancy; Shire/Takeda: Consultancy; Novo Nordisk: Consultancy; SOBI: Other: Travel grant; Roche: Other: DSMB member for a study.

Healthcare Utilization and Health Related Quality of Life in Persons with Von Willebrand Disease

Healthcare Utilization and Health Related Quality of Life in Persons with Von Willebrand Disease

Genotype Analysis of Adolescents with Heavy Menstrual Bleeding and Low Von Willebrand Activity - Report of a Multi-Center Study

Introduction: Low von Willebrand factor (VWF) activity is prevalent in adolescents with heavy menstrual bleeding (HMB). There is a need to better genetically characterize these patients and improve our understanding of the pathophysiology of their bleeding risk. Methods: One of the main objectives of this multi-center, single arm, observational cohort study was to genotype adolescent females with HMB and low VWF (≥ 30 and ≤ 50 IU/dL) by means of whole exome sequencing (WES), to identify variants throughout the exome that may modulate risk for bleeding. Post-menarchal females &amp;lt; 21 years with HMB (defined as PBAC score &amp;gt;100) and low VWF were eligible for the study. All patients were enrolled by participating centers where blood samples were collected. Exome data for 86 cases and 900 unrelated pediatric (&amp;lt;21 years) controls taken from a genetic study on Chiari 1 Malformation (CM1) were aligned and sorted, and variants called using the Sentieon software package. Annotation including allele frequencies, function, amino acid change and Clinvar rating among others was obtained using ANNOVAR. Variants were retained if there was a genotype call rate of &amp;gt;0.8, GQ&amp;gt;20, AB between 0.3-0.7 and min DP&amp;gt;8. Case/control analyses included common and rare SNP associations, gene burden analysis of both rare nonsynonymous variants and ClinVar 'pathogenic' variants, and gene-set burden analyses. Variants were considered rare if they had a minor allele frequency of &amp;lt;1% in the Genome Aggregation Database (gnomAD). Count differences between cases/controls were determined by Fisher's Exact test. Results: Of the 113 subjects enrolled, 86 had sufficient blood samples collected for WES. The median age was 16.2 years (range: 11.5-19.6). 36% of cases showed variants in VWF vs. 26% of controls (p=0.14). After multiple test correction, WES revealed a significant common variant association in FERMT2 with an LD block with a frequency of 24% in cases and 6% in controls (p=7.5x10-7). Rare variant analysis showed a significant association with an intronic variant in ABCA13 (p=1.6x10-7). Among the gene burden analysis using rare nonsynonymous variants, 2 genes of interest passed multiple test correction: IL12B and DTNBP1. When using ClinVar 'pathogenic' variants as the input for the gene burden analysis, 4 genes of interest passed multiple test correction: HBM, MYLK, RUNX1 and CD36. Gene-set burden analysis revealed 5 significant pathways of interest, including platelet degranulation, platelet alpha granule lumen and erythrocyte differentiation. We then focused a subset of known risk genes and compared the number of missense, nonsense and ClinVar 'pathogenic' variants between cases and controls. GP6, and MTHFR had significantly more missense variants in controls vs. cases (p=0.003 and 0.01, respectively), and F13B approached significance, with more missense variants in controls than cases (p=0.07). Conclusion: We found VWF variants in 36% of subjects, in accordance with previous reports. We found novel SNP associations with variants in FERMT2 and ABCA13.FERMT2 encodes for the integrin, Kindlin 2, which has been shown to be critical for supporting vascular integrity. Several other relevant genes passed multiple test correction in gene burden analyses, including genes known to cause Hermansky-Pudlak syndrome (DTNBP1), familial platelet disorder (RUNX1), platelet glycoprotein IV deficiency (CD36) and aortic aneurysm (MYLK). This association with platelet disorders was strengthened by our gene-set burden results, which implicate several platelet-related pathways. These data suggest that while a subset of HMB patients may have their bleeding explained by variants in VWF, there is a role for other hemostasis, platelet biology and vascular integrity related gene variants which can contribute to the variation in bleeding severity in adolescents with low-VWF related HMB. Study supported by an investigator-initiated research grant from Shire US Inc., now part of Takeda Disclosures O'Brien: Bristol Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees. Mullins:Takeda, Bayer: Other: Advisory Board. Sidonio:Takeda: Research Funding. Ragni:Sangamo: Consultancy, Research Funding; Takeda: Research Funding; Bioverativ: Consultancy, Research Funding; Spark: Consultancy, Research Funding; BioMarin: Consultancy, Research Funding; Alnylam/Sanofi, ATHN, BioMarin, Bioverativ, Sangamo, Spark: Research Funding; Alnylam/Sanofi, BioMarin, Bioverativ, Spark: Consultancy; Alnylam Pharmaceuticals Inc., Baxalta/Takeda, BioMarin, Bioverativ, and Spark Therapeutics: Membership on an entity's Board of Directors or advisory committees; American Thrombosis Hemostasis Network: Other: Committee work; Baxalta/Takeda, CSL Behring, Genentech, a member of the Roche Group, OPKO Biologics, and Vascular Medicine Institute: Research Funding. Kulkarni:Sanofi/ Bioverativ, Bayer, Biomarin, Shire/Takeda, Novo Nordisk, Freeline: Other: clinical trial research grants ; Bioverativ/Sanofi, BPL, Genentech, Kedrion, Novo Nordisk, Octapharma, Pfizer, Takeda, Catalyst Bioscience Bayer: Membership on an entity's Board of Directors or advisory committees. Srivaths:Shire US Inc., now part of Takeda: Research Funding.

Platelet Function Analyzer-100 Shows Poor Predictive Value for Children with Low VWF and Mild Platelet Function Defects

Background: The Platelet Function Analyser (PFA)-100 (Siemens) has proven to be a useful screening tool for primary hemostasis. Studies have demonstrated a pooled sensitivity around 90% for all individuals with Von Willebrand Disease (VWD), however the sensitivity for Type 1 is notably poorer than types 2 and 3. Our institution utilizes the PFA-100 as part of a panel of laboratory studies called the "bleeding screen"; this panel is often used as first line screening for inherited bleeding disorders across the hospital system. As the relative frequency of Type 1 VWD and low VWF (defined as Von Willebrand factor (VWF) levels 30-50%) greatly exceed the frequency of individuals with Type 2 and Type 3, when used as a general screen, the PFA-100 likely has reduced sensitivity than documented in the literature. Methods: To evaluate the utility of the PFA-100 as a screening test for VWD and platelet function defects, we retrospectively examined the ordering practices of PFA-100 in relation to the Von Willebrand panel (VWF:antigen, VWF:ristocetin cofactor, Factor VIII), platelet aggregation studies and platelet electron microscopy. We compared results of the testing with diagnostic codes to characterize the test characteristics of the PFA-100 for the diagnosis of VWD or platelet function defects in a pediatric population. We included all patients tested with the PFA-100 from 2017 to 2018 at any Children's Healthcare of Atlanta facility. Exclusion criteria include age under 30 months (reference range established for those over 30 months), hematocrit &amp;lt;35% or platelets &amp;lt;150,000/uL at time of testing due to impact on PFA-100 results. Demographics, laboratory results, bleeding symptoms and diagnoses were collected via chart review. Bleeding disorders were determined by ICD-9 and ICD-10 codes and confirmed via chart review. Descriptive statistics were calculated with frequency and median (interquartile range); comparative statistics calculated with t-tests, chi-square, Fisher's Exact, Mann-Whitney and Spearman rank tests. Test characteristics for PFA-100 and VWD were determined by correlation between PFA-100 and VWD studies sent concurrently; a ristocetin co-factor &amp;lt;50% was considered positive for VWD in the comparative analysis. Test characteristics determined for platelet function testing were dependent on results of platelet aggregation studies and/or platelet electron microscopy. Results: There were 609 children who met inclusion criteria. Median age was 12.2 years (7.3-15.6). The majority were female (62.4%), white (58.5%) and non-Hispanic (77.0%). The most common reason for testing was epistaxis (28.1%), followed by heavy menstrual bleeding (24.8%) and bruising (18.4%). Almost 30% had no bleeding symptoms documented. The majority of individuals also had a VWD profile sent (91.5%), with 418 having the PFA-100 and VWD Profile sent concurrently. The majority of PFA-100 tests were normal (70.6%). Only 91 (14.9%) had additional platelet evaluation with platelet aggregation studies or electron microscopy. Overall, 170 individuals (27.9%) were found to have a bleeding disorder. VWD was diagnosed in 146 individuals (24.0%), a platelet function defect 14 (2.3%) and 11 individuals had other diagnoses including Factor VII deficiency, Hemophilia A and Hemophilia B. The sensitivity of the PFA-100 for VWD was 60.4% (95%CI 46.9-73.6%) and the sensitivity was 60.0% (26.2-87.4%) for platelet function defects. For those with an abnormal PFA, the results of a VWD profile did not determine the likelihood of a provider sending additional testing with platelet aggregation studies or platelet microscopy. Of the 104 individuals with an abnormal PFA and normal VWD studies, only 31 had platelet aggregation testing. There were no differences in demographics or reported symptoms between the two groups. With normal VWD studies, individuals with an abnormal PFA were about 3 times more likely to have platelet aggregation studies sent (p&amp;lt;0.0001). Conclusion: The PFA-100 has poor sensitivity for VWD and mild platelet function defects in pediatric patients. Most individuals screened with a PFA-100 also had a complete VWD panel sent concurrently indicating that this is used as a screen for platelet function defects in our hospital. Given the poor sensitivity for platelet function defects, advanced platelet function testing should be sent based on clinical concern regardless of PFA-100 results when VWD testing is normal. Disclosures Brown: National Hemophilia Foundation, Takeda Clinical Fellowship: Research Funding. Woods:Takeda: Membership on an entity's Board of Directors or advisory committees.

Iron Deficient Anemia May Obscure a Diagnosis of Low VWF, VWD or Mild Hemophilia a in Biological Females with Heavy Menstrual Bleeding

INTRODUCTION Heavy menstrual bleeding (HMB) is one of the most common disease manifestations of biological females with inherited bleeding disorders and is often accompanied by iron deficient anemia (IDA). The most common bleeding disorders identified in biological females are von Willebrand Disease (VWD)/low von Willebrand Factor levels (low VWF) and heterozygosity for factor VIII genetic defects. Several pre-analytical variables including elevated adrenergic stress and estrogen-states can make it challenging to accurately diagnose these conditions as these variables are known to influence VWF and factor VIII activity levels. We sought to investigate whether an iron depleted state impacts the results of a laboratory assessment for VWD/low VWF or mild hemophilia A. METHODS Subjects were recruited from the Spots and Dots clinic (female-identifying bleeding disorder clinic) through the University of Colorado's Hemophilia and Thrombosis Center. Eligible subjects included biological females of any age who had at least two sets of laboratory assessments for von Willebrand levels (VWF antigen, VWF activity, and factor VIII activity level) and markers for total body iron stores (complete blood count and ferritin). Age at time of first testing, iron prescriptions, and underlying bleeding disorder diagnoses were also recorded. IDA was defined as a ferritin less than 20ng/mL with accompanying anemia (Hb &amp;lt;14.3 g/dL). Descriptive statistics and Fisher's exact tests were used to evaluate the association of factor VIII/VWF activity levels and iron stores. RESULTS Thirty-seven subjects were included in the final analysis. The average age at time of first testing was 21.5 years (range 9-50 years). IDA was present in 22 patients at the time of first assessment and in 20 patients at follow up. Serum ferritin improved overall in 23 patients between assessments. Oral iron was prescribed in 20 patients and intravenous iron was prescribed in three. An improvement in serum ferritin predicted a decrease in factor VIII activity (p = 0.0092) on follow-up assessment but did not predict a significant decrease in VWF activity levels (p = 0.0819) (Figure 1). Retesting factor VIII and VWF activity levels after iron repletion led to a diagnosis of low VWF in 4 cases (10.8%) and VWD in 2 cases (5.4%) that would have otherwise been classified as normal without retesting. CONCLUSIONS IDA is frequent in women with HMB and may be insufficiently managed with oral iron therapy in adolescents and those with an underlying bleeding disorder. The fact that many patients remained iron deficient at the time of follow up testing may have blunted the results of this analysis. Despite this limitation, this study still suggests that iron deficient anemia may be a significant enough biological stressor to increase factor VIII and possibly VWF activity levels to a point that may obscure an underlying diagnosis of VWD, low VWF, or hemophilia A. Further investigations are warranted to confirm if universal retesting of VWF and factor VIII activity levels are diagnostically imperative after correction of IDA. Such a recommendation may have a significant impact on this patient population as discovery of an underlying bleeding disorder diagnosis can not only provide an explanation for a patient's HMB but also can identifying individuals who may be at increased risk for bleeding at other sites or during invasive procedures. Disclosures No relevant conflicts of interest to declare.

Semiautomatic VWF Multimer Densitometric Analysis: Validation of the Clinical Accuracy and Clinical Implications in Von Willebrand Disease

Semiautomatic VWF Multimer Densitometric Analysis: Validation of the Clinical Accuracy and Clinical Implications in Von Willebrand Disease