Defects Of Von Willebrand Factor Research Articles

Recombinant vs plasma-derived von Willebrand factor to prevent postpartum hemorrhage in von Willebrand disease

Abstractvon Willebrand disease (VWD) is a congenital bleeding disorder characterized by deficient or defective von Willebrand factor (VWF). Among women with VWD, postpartum hemorrhage (PPH) is common. Treatment options at delivery include plasma-derived VWF (pdVWF) and recombinant VWF (rVWF). However, limited data are available regarding their efficacy. We conducted a retrospective observational study comparing PPH in women with VWD treated at the Hemophilia Center of Western Pennsylvania between 1 February 2017 and 31 January 2018 with either rVWF or pdVWF. We compared postpartum outcomes, including PPH frequency and estimated blood loss (EBL) at delivery. There were a total of 12 deliveries, 7 vaginal and 5 cesarean. At delivery and for 3 days postpartum, 6 women received 80 IU/kg of rVWF and 6 received 80 IU/kg of pdVWF, based on prepregnancy weight, insurance, and/or patient choice. Treatment groups had similar demographics, including median age (32.0 vs 27.0 years; P = .075), bleeding scores (3.0 vs 3.5; P = .734), and prepregnancy body mass index (29.0 vs 29.2 kg/m2; P = .691). PPH occurred in 3 (25.0%) of 12 deliveries, with no difference by treatment group (2 of 6 rVWF vs 1 of 6 pdVWF; P = 1.000) and no difference in EBL by treatment group (685 vs 462 mL; P = .384) or delivery type (vaginal, P = .722 vs cesarean, P = .531). In summary, PPH occurred in one-fourth of the deliveries in women with VWD, despite a higher dose (80 IU/kg) of rVWF or pdVWF. Future trials are needed to develop and assess novel strategies to prevent PPH in VWD.

The Effect of Age on von Willebrand Factor and Bleeding Symptoms in von Willebrand Disease.

von Willebrand disease (VWD) is a quantitative or qualitative defect in von Willebrand factor (VWF) resulting in mucocutaneous bleeding symptoms and hemorrhage following hemostatic challenges, such as trauma or surgery. VWD-specific therapy, DDAVP (1-desamino-8-D-arginine vasopressin) and VWF concentrates, is necessary periprocedurally to ensure adequate hemostasis. The aging VWD patient may complicate this matter. The plasma concentration of many coagulation proteins, including VWF, increases with age. While it has been established that VWF levels increase with age in a healthy population, emerging research demonstrates this occurs in certain subtypes of VWD, too. Thus, the management of periprocedural VWD-specific therapy in the aging VWD patient is problematic when VWF levels increase over time to normal, and hematologists are left with uncertainty regarding whether or not periprocedural VWD-specific therapy is still necessary. In this article, we will review the current state of the literature regarding the effect of age on VWF levels in the healthy population and VWD while exploring possible etiologies for this phenomenon. Further, we will detail how this affects bleeding symptoms and highlight what research remains to be done to optimize care in this patient population.

Navigating the Myriad of von Willebrand Factor Assays.

von Willebrand factor (VWF) represents a large and complex adhesive plasma protein whose main function is to provide a bridge between blood platelets and damaged endothelium, and thus facilitate primary hemostasis. VWF also binds to FVIII, preventing early proteolysis, and delivers this cargo to sites of vascular injury, thereby promoting clot formation and secondary hemostasis. An absence, deficiency, or defect in VWF can lead to a bleeding diathesis called von Willebrand disease (VWD), considered the most common inherited bleeding disorder. Contemporary laboratory assays used in VWD diagnosis/exclusion comprise a myriad of assays that identify the quantity (level) of VWF, as well as the multitude of VWF activities. These may use the following test abbreviations: VWF:Ag, VWF:RCo, VWF:CB, VWF:GPIbR, VWF:GPIbM, VWF:FVIIB, VWF:Ab. The current review explains what these assays are, as well as their place in VWD diagnostics.

Coexistence of Von Willebrand disease and gastrointestinal stromal tumor (G.I.S.T): Case report of a rare and challenge association

Von Willebrand disease (VWD) is the most common inherited bleeding disorder and is caused by a quantitative (type 1 and 3) or qualitative (type 2) defect of Von Willebrand factor (VWF). Bleeding from the gastrointestinal (GI) tract is not uncommon in VWD and is usually associated with angiodysplasia. We report herein on the management of a patient affected by VWD2B with severe GI bleeding secondary to gastrointestinal stromal tumor (GIST) complicated by deep vein thrombosis (DVT). The current case demonstrated that the hemostatic balance, in RBDs under specific circumstances, can range from a tendency toward a hemorrhagic to normal or prothrombotic state. In these patients, a close collaboration between hematologists and surgeons can guarantee appropriate management in high-risk clinical scenarios.

Increased von Willebrand factor levels in polycythemia vera and phenotypic differences with essential thrombocythemia

BackgroundAcquired von Willebrand factor (VWF) deficiency was described in Philadelphia‐negative myeloproliferative neoplasms, especially in essential thrombocythemia (ET). VWF phenotype in contemporary patients with polycythemia vera (PV) remains less explored. ObjectivesTo characterize the VWF phenotype in PV and to compare VWF phenotype in PV with matched healthy subjects and ET patients. Patients/MethodsWe studied 48 PV patients, treated according to current recommendations (hematocrit ≤ 45%, on low‐dose aspirin prophylaxis); 48 healthy and 41 subjects with ET, all sex, age, and blood group matched. We measured VWF antigen, activity, multimeric pattern, ADAMTS‐13, and factor VIII (FVIII) antigen. ResultsIn patients with PV, VWF antigen and activity were significantly higher than in healthy subjects (antigen: 119[96‐137] vs 93[79‐107] IU/dL; activity: 114[95‐128] vs 90[79‐107] IU/dL, respectively, medians and interquartile, P < 0.01), with normal multimeric distribution. ADAMTS‐13 levels were similar between patients with PV and healthy subjects. FVIII levels were higher in PV than in healthy subjects (141[119‐169] versus 98[88‐123] IU/dL, respectively, P < 0.01). By multivariable analysis, JAK2‐p.V617F allelic burden, erythrocyte count, and male sex significantly predicted VWF antigen and activity levels. As compared to patients with ET, patients with PV showed similar VWF antigen levels but approximately 40% higher activity (79[49‐104] vs 112[93‐125] IU/dL, respectively, P < 0.01). ConclusionsPatients with PV show increased VWF and FVIII levels, predicted by JAK2‐p.V617F burden and erythrocyte count. At variance with ET, acquired VWF defect was not observed in PV. High VWF/FVIII levels may sustain the thrombotic diathesis of PV and may be investigated as biomarkers for risk stratification.

Patients with aortic stenosis have von Willebrand factor abnormalities and increased proliferation of endothelial colony forming cells

BackgroundPatients with aortic stenosis (AS) can experience bleeding complications including gastrointestinal bleeding from angiodysplastic lesions due to acquired von Willebrand syndrome. Studies have pointed to a role for von Willebrand factor (VWF) in angiogenesis. ObjectiveThe objective of this study was to assess VWF defects in AS patients over time and the impact on angiogenesis using patient‐derived endothelial colony‐forming cells (ECFCs). Patients/MethodsPlasma sample collection and ECFC isolations were performed before valve replacement surgery, 3 to 5 days after, and 6 months after surgery. Plasma VWF antigen, activity, propeptide, collagen binding, multimers, factor VIII coagulant activity, and ADAMTS13 activity (a disintegrin‐like and metalloprotease with thrombospondin type 1 motifs 13) were determined. ECFCs were assessed for VWF and angiopoietin‐2 (Ang‐2) storage and secretion, cell proliferation, and tubule formation in Matrigel. Results and ConclusionsAortic stenosis patients exhibited quantitative and qualitative abnormalities of VWF including significantly increased VWF antigen, activity, and propeptide levels following surgery (P < .01). Increased high molecular weight VWF multimers were observed at all time points and in particular 3 to 5 days after surgery (mean = 14% ± 6%) relative to before (mean = 10% ± 4%), suggesting increased proteolysis by ADAMTS13 pre‐operatively in a shear‐dependent manner. ECFCs from patients with aortic stenosis were more proliferative than controls (P < .05) and had increased retention of Ang‐2 (P < .05) suggesting epigenetic modification of the cells. Overall, there are hemostatic changes in AS patients that are present before valve replacement surgery and these persist long after surgery has occurred. These findings have implications for the current clinical management of AS patients.

Utility of the platelet function analyser (PFA-100/200) for exclusion or detection of von Willebrand disease: A study 22 years in the making

Introductionvon Willebrand disease (VWD) is the most common inherited bleeding disorder and may alternatively arise as an acquired condition (AVWS). These represent deficiency and/or defects in von Willebrand factor (VWF). Closure times (CTs) obtained from the platelet function analyser (PFA) are highly sensitive to both VWD/AVWS. The current study has evaluated the utility of the PFA-100/-200 to exclude or detect laboratory identified VWD. Materials and methodsAn evaluation of the success or otherwise of prospective PFA testing to help exclude or detect VWD using data from a locally maintained database. This database contains patient information, results of PFA testing, results of VWF testing, platelet count and hematocrit, and additional information, for a current total of 3678 entries representing over 2 decades of testing. ResultsAccording to selection criteria, a total of 142 samples were identified as derived from patients with VWD. All but one of these were also identified to have abnormal PFA CTs. Additional data sets of patients with ‘low VWF’ (n = 137), or borderline normal VWF (n = 163) were also identified, as well as patients with thrombocytopenia and/or low hematocrit (n = 487). In these cohorts, PFA CTs were often (but not always) abnormal. There was a strong association between VWF test parameter values and PFA CTs. Additional study samples comprised cases with normal VWF parameters but prolonged CTs (n = 594), as well as cases with normal VWF and normal CTs (n = 1292), permitting calculation of sensitivity of abnormal PFA for VWD (99.3%), as well as specificity (68.5%), negative predictive value (99.9%) and positive predictive value (19.2%), for a prevalence of 7.0%. Comparatively, normal PFA CTs were better able to exclude VWD than normal test results for individual VWF parameters. ConclusionThis study reports on an evaluation of PFA CTs for identification or exclusion of VWD. In our dataset, representing over 22 years of experience, normal PFA CTs were able to predict absence of VWD with higher sensitivity than individual VWF test results.

Von Willebrand factor (VWF) as a determinant of silent cerebral microbleeds during TAVR in a prospective MRI cohort

Cerebral microbleeds have been observed in “asymptomatic” elderly patients undergoing systematic MRIs and might be related with cognitive decline. A defect in Von Willebrand factor (VWF) after Trans Aortic-valve Replacement (TAVR) enhanced bleeding risk and possibly contributes to vascular malformations. To define the occurrence of “acute” microbleeds during TAVR and the clinical and procedural factors associated with this occurrence including a post-procedural defect in VWF. A prospective cohort of patients with aortic stenosis referred for TAVR was included. A standardized neurological assessment, a cerebral magnetic resonance imaging (MRI) and a biological analysis including quantification of VWF multimers and PFA-CADP were performed before and after TAVR. Eighty-four patients completed the imaging protocol. On pre-procedural MRI, 22 patients (26%, 95%CI = 17–37%) had at least one microbleeds. After TAVR, new microbleeds were observed in 19 (23%, 95%CI = 14–33%) patients. The occurrence of new microbleeds was independent of the presence microbleeds at baseline. A lower HMW-multimer-ratio as measured immediately at the end of the procedure was associated with the occurrence of new microbleed detected on the MRI performed 3 days after the procedure (0.90 ± 0.14 vs. 1.06 ± 0.27, P = 0.009). The same observation was made with a higher CT-ADP (165 ± 72 vs. 127 ± 57, P = 0.02). A prolonged procedure (RR = 1.21 [1.01–1.170] for every 5 min of fluoroscopy time, P = 0.04) and post-procedural acquired-VWF-defect (RR = 1.42 [1.08–1.89] for every lower “0.1 unit” of HMW-multimer-ratio, P = 0.004) were the only factors associated with the occurrence of new post-procedural microbleed(s). We report for the first time a high incidence (23%, 95%CI = 14–33%) of new post-procedural cerebral microbleeds occurring during TAVR. Procedural management and persistence of acquired-VWF defect could play a major role in the occurrence of those new microbleeds.

The Von Willebrand Factor Antigen Plasma Concentration: a Monitoring Marker in the Treatment of Aortic and Mitral Valve Diseases

Von Willebrand disease is a commonly inherited bleeding disorder caused by defects of von Willebrand factor (vWF). In the most common valve diseases, aortic valve stenosis (AVS) and mitral valve regurgitation (MVR), a bleeding tendency has been described in a number of patients. This has been associated to a high turbulence of blood flow through the compromised valve, promoting degradation of vWF with loss of high-molecular-weight multimers of vWF (HMWM), leading to an acquired von Willebrand syndrome (AvWS). We analysed three groups of patients, one affected by AVS, treated with transcatheter aortic valve implantation (TAVI), the second group of patients affected by MVR, treated with Mitraclip® mitral valve repair. The third group was represented by patients also affected by AVS, but not eligible for TAVI and treated with standard surgery. A fourth group of patients that underwent percutaneous coronary intervention (PCI) with stenting was used as a control. Our results demonstrated that the level of vWF measured as antigen concentration (vWF:Ag) increases in all cohorts of patients after treatment, while in control PCI patients, no modification of vWF:Ag has been registered. Western blot analysis showed only a quantitative loss of vWF in the pre-treatment time, but without significant HMWM modification. The monitoring of the vWF:Ag concentration, but not the quality of HMWM, can indicate the status of blood flow in the treated patients, thus introducing the possibility of using the vWF antigen detection in monitoring the status of replaced or repaired valves.

Disease progression and defects in primary hemostasis as major cause of bleeding in multiple myeloma

In patients with multiple myeloma (MM), unexpected bleeding complications remain a major issue. Since routine coagulation parameters are often inconspicuous, diagnosis and treatment of the underlying coagulation disorders are challenging. In our single-center observational study, we analyzed 164 patients with MM for coagulation disorders and bleeding complications. Prolonged closure times (CTs), measured by PFA-100, were the most common, abnormal coagulation test, found in 66% of bleeding patients vs 5% in non-bleeding, followed by qualitative defects of von Willebrand factor (VWF:CB/VWF:Ag ratios), found in 34% vs 1% in the non-bleeding group. Increased serum free light chains (SFLC) and SFLC ratios were significantly associated with prolonged CTs and acquired von Willebrand syndrome (AVWS). Prolonged CTs and AVWS were associated with disease progression, determined by dynamics of SFLC ratios (P<.001), serum creatinine level (P=.013), Beta-2 microglobulin (P=.03), LDH (P=.016), and bone marrow infiltration (P<.001). Of note, response to myeloma therapy was frequently correlated with normalization of coagulation parameters. Bleeding complications in MM are predominantly caused by defects in primary hemostasis and associated with disease progression. In a peri-interventional workup, determination of CTs and VWF:CB/VWF:Ag ratios are of significant importance to assess bleeding risk.

Von Willebrand Disease Minimize Menorrhagia (VWDMin) Trial

Von Willebrand Disease Minimize Menorrhagia (VWDMin) Trial

Utility of repeat testing in the evaluation for von Willebrand disease in pediatric patients

BackgroundVon Willebrand disease (VWD) is the most common inherited bleeding disorder and is caused by quantitative and qualitative defects in von Willebrand factor (VWF). The laboratory diagnosis of VWD in pediatric patients is complicated by VWF interassay and intra‐assay variability, stress‐induced elevations in VWF levels, and a lack of significant bleeding history with which to correlate test results. ObjectiveGuidelines recommend repeat testing in patients with a high suspicion of VWD and unclear laboratory assay results; however, no studies have evaluated the utility of repeat VWF testing in pediatric patients. MethodsThis retrospective single‐center cohort study aimed to determine clinical variables associated with requiring more than one test to diagnose VWD and to establish a cutoff VWF value above which further testing is not informative. ResultsOf 811 patients evaluated for a suspected bleeding disorder, 22.2% were diagnosed with VWD, with ~70% diagnosed on the first test. Patients with VWD were younger (5.8 vs. 8.5 years, P = .002) and more likely to have a family history of VWD (38% vs. 22%, P < .001) than those without VWD. Univariate analysis failed to identify any clinical variables that correlated with needing multiple tests for a VWD diagnosis. A cutoff of 100 IU/dL for VWF antigen or activity on the first test yielded negative predictive values >95%. ConclusionsWe demonstrate that the majority of pediatric patients had diagnostic VWF values on the first set of testing. Pediatric patients without a family history of VWD and VWF levels >100 IU/dL may not need further testing to rule out the diagnosis of VWD.

Von Willebrand Disease: From In Vivo to In Vitro Disease Models.

Von Willebrand factor (VWF) plays an essential role in primary hemostasis and is exclusively synthesized and stored in endothelial cells and megakaryocytes. Upon vascular injury, VWF is released into the circulation where this multimeric protein is required for platelet adhesion. Defects of VWF lead to the most common inherited bleeding disorder von Willebrand disease (VWD). Three different types of VWD exist, presenting with varying degrees of bleeding tendencies.The pathophysiology of VWD can be investigated by examining the synthesis, storage and secretion in VWF producing cells. These cells can either be primary VWF producing cells or transfected heterologous cell models. For many years transfected heterologous cells have been used successfully to elucidate many aspects of VWF synthesis. However, those cells do not fully reflect the characteristics of primary cells. Obtaining primary endothelial cells or megakaryocytes with a VWD phenotype, requires invasive procedures, such as vessel collection or a bone marrow biopsy. A more recent and promising development is the isolation of endothelial colony forming cells (ECFCs) from peripheral blood as a true-to-nature cell model. Alternatively, various animal models are available but limiting, therefore, new approaches are needed to study VWD and other bleeding disorders. A potential versatile source of endothelial cells and megakaryocytes could be induced pluripotent stem cells (iPSCs).This review gives an overview of models that are available to study VWD and VWF and will discuss novel approaches that can be considered to improve the understanding of the structural and functional mechanisms underlying this disease.

The role of genetics in the pathogenesis and diagnosis of type 1 Von Willebrand disease.

Von Willebrand disease (VWD) is a common bleeding disorder, but diagnosis of VWD is challenging, particularly with type 1 VWD. Although most clinicians use specific tests of von Willebrand factor (VWF) activity to classify patients with VWD, genetic testing for VWF defects is another potential method of diagnosis. Studies of patients with type 1 VWD report consistently that many, but not all, study participants have VWF gene defects. Certain populations, including those with VWF levels less than 30 IU/dl and those with clearance defects, are more likely to have a VWF sequence variant. In addition, a number of loci outside the VWF gene have been shown to affect VWF levels, including ABO, CLEC4M, STXBP5, and STAB2. Genetic defects in VWF are common, but not all defects lead to disease. Type 1 VWD in particular does not always have an associated VWF sequence variant. New data stemming from genome-wide association studies on modifier genes suggest that the etiology of type 1 VWD is multifactorial.

Molecular Genetics of von Willebrand Disease in Korean Patients: Novel Variants and Limited Diagnostic Utility of Multiplex Ligation-Dependent Probe Amplification Analyses.

Backgroundvon Willebrand disease (VWD), characterized by quantitative or qualitative defects of von Willebrand factor (VWF), is the most common inheritable bleeding disorder. Data regarding the genetic background of VWD in Korean patients is limited. To our knowledge, this is the first comprehensive molecular genetic investigation of Korean patients with VWD.MethodsTwenty-two unrelated patients with VWD were recruited from August 2014 to December 2017 (age range 28 months–64 years; male:female ratio 1.2:1). Fifteen patients had type 1, six had type 2, and one had type 3 VWD. Blood samples were collected for coagulation analyses and molecular genetic analyses from each patient. Direct sequencing of all exons, flanking intronic sequences, and the promoter of VWF was performed. In patients without sequence variants, multiplex ligation-dependent probe amplification (MLPA) was performed to detect dosage variants. We adapted the American College of Medical Genetics and Genomics guidelines for variant interpretation and considered variants of uncertain significance, likely pathogenic variants, and pathogenic variants as putative disease-causing variants.ResultsVWF variants were identified in 15 patients (68%): 14 patients with a single heterozygous variant and one patient with two heterozygous variants. The variants consisted of 13 missense variants, one small insertion, and one splicing variant. Four variants were novel: p.S764Efs*16, p.C889R, p.C1130Y, and p.W2193C. MLPA analysis in seven patients without reportable variants revealed no dosage variants.ConclusionsThis study revealed the spectrum of VWF variants, including novel ones, and limited diagnostic utility of MLPA analyses in Korean patients with VWD.

The Importance and Complications of Sequencing of Von Willebrand Gene in Von Willebrand Disease

Abstract Genetic testing in patients with von Willebrand disease completes phenotypic testing with an aim to confirm the von Willebrand factor defect at a molecular level. Structure of the VWF gene was described 30 years ago; since then a large number of mutations leading to VWD have been described in this gene. Thanks to describing these mechanisms it is possible to understand the pathogenesis of the most common congenital bleeding disorder. In the Slovak Republic genetic testing is still not a routine part of VWD diagnostics. The National Center of Hemostasis and Thrombosis in Martin is the first department in Slovakia which has begun genetic testing of patients with VWD. Sequencing of the VWF gene has many limitations which are referred in more details within this article. Therefore, we decided to use the methods of new generation sequencing in combination with Sanger sequencing. We believe that soon we will have the first results which will help us to identify the possible cause of VWD in these patients.

Analysis of von Willebrand Disease in the South Moravian Population (Czech Republic): Results from the BRNO-VWD Study.

von Willebrand disease (VWD) is an inherited bleeding disorder caused by a quantitative (type 1 and 3) or qualitative (type 2) defect of von Willebrand factor (VWF). The heterogeneity of laboratory phenotyping makes diagnosing difficult. A cross-sectional, family-based VWD study in a collaboration between University Hospital Brno (Czech Republic) and Antwerp University Hospital (Belgium) to improve the understanding of laboratory phenotype/genotype correlation. A total of 205 patients with suspected VWD were identified from historical records. Complete laboratory analysis was established using all available VWD assays including VWF multimers and genetic analysis. Based on the current International Society of Thrombosis and Haemostasis (ISTH) - Scientific and Standardization Committee VWD classification and type 2A sub-division into 2A/IIA, IID, IIC and IIE, the majority was characterized as a type 1 VWD, followed by type 2. Proposed laboratory phenotypes were confirmed by their multimeric pattern within 98% of this cohort. All type 2, 3 and 75% of type 1 VWD patients were confirmed by underlying causative mutations. Forty-six different causal mutations (117 not previously described in the literature) could be identified. Fifty per cent of all cases was represented by eight individual mutations, mainly p.Pro812ArgfsX31. Thirteen patients had a large heterozygous gene alteration. Although an extensive panel of tests was used, VWD classification and (sub)typing remains difficult and fluid. This study provides a cross-sectional overview of the VWD population in the Czech Republic and provides important data to the ISTH/European Association for Haemophilia and Allied Disorders VWD mutation database in linking causal mutations with unique VWD (sub)types. It also identifies new, as not previously described in the literature, causal mutations.

Rethinking internal quality control and external quality assessment for laboratory diagnostics of von Willebrand disease

von Willebrand disease (VWD) is recognised as the most common congenital bleeding disorder, and an acquired form named von Willebrand syndrome (AVWS) may arise in a variety of disease states. Thus, laboratory testing for VWD and AVWS represents a key diagnostic activity of hematology facilities performing advanced hemostasis diagnostics. VWD and AVWS arise due to deficiency and/or defect(s) in the adhesive plasma protein called von Willebrand factor (VWF). VWF represents a key element of primary hemostasis, and also contributes to secondary hemostasis. Thus, deficiency and/or defects in VWF, either congenital or acquired, leads to a bleeding diathesis in affected individuals. A wide variety of laboratory tests may be performed to investigate, diagnose or exclude VWD/AVWS, including screening assays, more complex functional assays, and also molecular analysis. Thus, VWD testing now incorporates a variety of processes, and potential methodologies. Internal quality control (IQC) and external quality assessment (EQA) is critical to ensure the quality of laboratory testing, but is made difficult in VWD diagnostics by the nature and variety of the tests available, and the heterogeneity of VWD/AVWS. The current review looks at laboratory testing for VWD/AVWS from the perspective of IQC and EQA, and considers some novel approaches to ensure the accuracy and quality of this activity in the future diagnostics landscape.

Distribution of Pathogenic Variants in Von Willebrand Disease Type 2A in the American Population: Results from a National Clinical Reference Laboratory

Introductionvon Willebrand disease (VWD) is a common inherited bleeding disorder with a reported incidence ranging from 0.01% to 1%. VWD is classified as quantitative (types 1 and 3) and qualitative (type 2) defects in von Willebrand factor (VWF). Classical VWD type 2A, the most common qualitative defect of VWD, is characterized by decreased VWF-dependent platelet adhesion and a selective deficiency of high molecular weight VWF multimers. The majority of type 2A VWD is inherited in an autosomal dominant fashion. This subtype is caused by missense variants acting through a variety of mechanisms including defects in multimer assembly or an intrinsically increased sensitivity to cleavage by ADAMTS-13. The majority of pathogenic mutations are located in exon 28, predominantly affecting the A2 domain. Missense mutations have also been reported in the D3 and A1 domains (exons 22 and 25-28). Less common missense mutations include those responsible for dimerization (CK domain, exon 52) and multimerization defects (D2 domain, exons 11-17). Our objective was to determine the genomic distribution of mutations underlying type 2A VWD in patients referred to a USA-based clinical reference laboratory.MethodsIn a clinical reference laboratory, BloodCenter of Wisconsin, VWF gene sequence analysis results from January 2009 to July 2018 were reviewed. Molecular testing was performed by polymerase chain reaction (PCR) amplification and bi-directional Sanger sequencing or capture-based enrichment and next-generation sequencing (NGS). Complete coding region and splice junction of VWF gene was compared to the reference sequence (NM_00552.3). Variant interpretation was performed according to American College of Medical Genetics (ACMG) standards and guidelines by utilizing genome databases, population frequency data, variant specific literature search and predictive computational tools. Variant interpretation was reviewed by a team comprising of genetic counselors, molecular experts and experts in coagulation disorders correlating these findings with the patient's clinical phenotype provided.ResultsPathogenic variants were identified in 46 cases received. Of these, 56.5 % (26 of 46) were in exon 28 of the VWF gene.(Figure 1) 21.7% of mutations (10 out of 46) were identified in exon 26, 4.3% were identified in exon 25 (2 of 46) and 6.5% (3 out of 46) were identified in exon 52 of VWF gene. The remaining 10.7% were identified in exons 6,11,12,22 and 27 (1 each). The distribution of these mutations according to VWF domains was also studied. 39% of these mutations were found in the A2 domain, 30.4% were identified in the D3 domain, and 13% were found in the A1 domain. A small number of these mutations were identified in the D1 domain (1/46), D2 domain (2/46) and CK domain (2/46). Phenotypic information was available for most patients evaluated.ConclusionsIn a predominantly Caucasian population with others of mixed ethnicities, the distribution of VWD type 2A mutations was studied in patients evaluated in a clinical reference laboratory. Most mutations were identified in VWF Exon 28 and in the A2 domain. Next generation sequencing platform enables sequencing of large amounts of DNA in parallel thereby allowing all VWF exons to be evaluated at one time. However, in smaller laboratories, without access to a NGS platform, initial evaluation of the VWF Exon 28 for evaluation of VWD type 2A variants is a cost effective measure. Subsequently, sequencing of other less implicated exons may be performed utilizing a reflexive algorithm. Genetic testing can be very useful in diagnosis of type 2 variants, where mutations in specific regions correspond to the defect in VWF function. [Display omitted] DisclosuresFriedman:Shire: Membership on an entity's Board of Directors or advisory committees; CSL Behring: Membership on an entity's Board of Directors or advisory committees.

Recombinant VWF for Delivery in Women with Von Willebrand Disease

Background: Von Willebrand disease (VWD) is the single most common congenital bleeding disorder, affecting 1% of the population and characterized by deficient and/or defective von Willebrand factor (VWF). Among women with VWD, reproductive tract bleeding is common, with up to 80% developing postpartum hemorrhage (PPH), defined as 24-hr blood loss ≥500 cc after vaginal delivery, or ≥1,000 cc after cesarean delivery. Despite current guidelines suggesting 50 IU/kg plasma-derived VWF (pdVWF) at delivery if VWF:RCo is less than 0.50 IU/ml at the 8thmonth of gestation, PPH occurs even when that level has normalized, and, moreover, despite factor replacement, VWF levels are lower and blood loss greater at delivery than in controls. These data suggest VWF levels should be higher at delivery to prevent PPH. As blood volume increases 1.5-fold during pregnancy, and blood loss at delivery is 1.4-fold greater than controls, we have proposed a 1.5-fold higher dosing, ~80 IU/kg, to prevent PPH. Yet, experience with these agents at delivery is limited in women with VWD.Methods: This was an observational study to compare postpartum blood loss in women with VWD, cared for at the Hemophilia Center of Western Pennsylvania, between February 1, 2017 and January 31, 2018, treated with recombinant von Willebrand factor (rVWF, Vonvendi®) or plasma-derived von Willebrand factor (pdVWF, Humate®). Medical records of all women with VWD undergoing delivery during the 12-month period were reviewed. The study was approved by the University of Pittsburgh IRB as an exempt study PRO18010198. Mean, median, and standard deviation or frequency (percentage) were determined for clinical variables including age, race, VWD type, bleeding score, bleeding history, body surface area, estimated blood loss, and comorbidity. Continuous variables were compared by student's t test, and discrete data were analyzed by chi-square or Fisher's exact test. A p-value of < 0.05 was considered statistically significant.Results:A totalof 12 women with VWD, including 11 with type 1 and 1 with type 2B VWD, underwent delivery, 7 vaginal and 5 cesarean section, under coverage of clotting factor during the 12-month study period. These included 6 who received rVWF 80 IU/kg and 6 who received pdVWF 80 IU/kg at delivery and for up to 3 days postpartum. Treatment dose was based on pre-pregnancy weight and determined by insurance and patient choice. Demographic data were comparable between rVWF and pdVWF treatment groups, including age, 31.7 vs 26.3years (p=0.067); race, 5/6 vs 4/6 Caucasian (p=0.409); and blood type, 6/6 vs 4/6 type O (p=0.227). Bleeding history was similar between rVWF and pdVWF groups as were pre-pregnancy bleeding scores, 3.2 vs. 3.0(p=0.867). Co-morbidities were not different by treatment group, including anemia, p=.0227, diabetes, p=0.500, hypertension, p=0.500, obesity, p=1.00, and smoking, p=0.500. PPH occurred in 58.3% overall, with no difference in PPH by treatment group, overall, 5 of 6 (83.3%) vs 2 of 4 (50.0%) (p=0.114), and no difference by type delivery, vaginal (p=0.343) or cesarean delivery (p=0.400). Estimated blood loss by group in vaginal deliveries was 433vs 381 cc (p=0.703), and 1173 vs 950 cc (p=0.473) in cesarean. At delivery, groups had comparable hemoglobin, 11.6 vs 11.7, gm/dl, p=0.900, and platelet count, 183 vs 219 x106/ml (p=0.606), and none required transfusion. Length of stay was 3.8 vs 2.8 days (p=0.156). Comparing groups, VWF:RCo levels increased over 2.0-fold during pregnancy from baseline, 0.43 vs 0.48 (p=0.666), to 8thmonth, 0.85 vs 1.25 (p=0.235); VWF:Ag increased 2.5-fold from baseline, 0.53 vs 0.66 (p=0.895) to 8thmonth, 1.32 vs 1.66 (p=0.434); and VIII:C increased over 2.2-fold from baseline, 0.69 vs 0.86 (p=0.245); to 8thmonth, 1.53 vs 1.97 (p=0.321). BMI increased up to 1.3-fold from baseline, 27.7 vs 28.8 (p=0.882); to delivery, 35.9 vs 35.2 (p=0.882).Conclusion:Postpartum blood loss was similar in rVWF and pdVWF treatment groups, whether vaginal or cesarean delivery. Dosing at 80 IU/kg appears safe, but over half developed PPH. Future trials are needed to evaluate blood-volume based VWF dosing to prevent PPH. [Display omitted] DisclosuresRagni:Alnylam: Honoraria, Research Funding; Bayer: Consultancy, Provision of Drug for Investigator Initiated Trial; Biomarin: Consultancy, Honoraria, Research Funding; Bioverativ: Consultancy, Research Funding; CSL Behring: Research Funding; MOGAM: Consultancy; NovoNordisk: Research Funding; Opko Biologics: Research Funding; Sangamo: Research Funding; SPARK: Consultancy, Research Funding; Shire: Other: Provision of Drug for Investigator Initiated Trial; Institute for Clinical & Economic Review: Consultancy.