Type 2M Research Articles

Management of von Willebrand Disease for Orthognathic Surgery: Evidenced-Based Recommendations and Scoping Review of the Literature

BackgroundThe general management of von Willebrand Disease has evolved over the past several decades. Advances in blood product replacement manufacturing have led to changes in perioperative management of the bleeding diathesis in its various subtypes. Surgical management follows changes in medical management, but unique perioperative considerations exist depending on the type of surgery. Orthognathic surgery is considered a major surgery that can have life-threatening complications in patients with von Willebrand Disease (vWD) and updates in evidenced-based management are presented. ObjectivesThe objectives of this report are to review available literature regarding management of patients with von Willebrand Disease and other bleeding disorder undergoing orthognathic surgery and to provide a case report regarding the management of a patient vWD type 2M undergoing orthognathic surgery. Eligibility CriteriaArticles included for review were studies addressing patients with bleeding disorders undergoing orthognathic surgery. Sources of EvidencePubMed and Ovid were queried using the terms “orthognathic surgery OR jaw deformity” AND “von Willebrand OR bleeding disorders OR hemophilia.” Additional references relevant to the review were obtained from the reference lists of available articles ultimately deemed appropriate for inclusion in the review. MethodsA recent example case of a patient with type 2M vWD undergoing orthognathic surgery and the patient’s perioperative management is presented for consideration. A scoping review of the literature was undertaken for evaluation of available literature using the Preferred Reporting Items for Systematic Reviews and Meta-Analyses extension for Scoping Review (PRISMA-ScR). ResultsInsufficient evidence exists for significant analysis of perioperative management strategies previously reported compared to the currently utilized management strategy for the case presented. A review of the available literature with reference to updated literature regarding management of patients with vWD undergoing surgical procedures other than orthognathic surgery yields primarily case reports, two randomized controlled trials, and one systematic review. The results indicate that an evolution in the general management of patients with vWD undergoing surgery can be applied to patients undergoing orthognathic surgery and that the addition of hemostatic agents such as tranexamic acid may be considered with a low level of evidence. ConclusionsFor patients undergoing orthognathic surgery, the use of cryoprecipitate in large quantities has been replaced with the use of Humate-P infusion and will likely be replaced with the use of recombinant von Willebrand factor (VWF). The timing of infusion of adjuncts preoperatively for patients with vWD can potentially be extended to the previous day, and the addition of medications to aid in hemostasis such as tranexamic acid may be considered. Prophylactic use of additional VWF adjuncts should be considered in the postoperative setting, and close monitoring is prudent. The use of newer agents may allow for the expansion of craniomaxillofacial surgery procedures that can be safely considered in patients with vWD.

Von Willebrand disease.

von Willebrand disease (VWD) is the most common inherited bleeding disorder. The disorder is characterized by excessive mucocutaneous bleeding. The most common bleeding manifestations of this condition include nosebleeds, bruising, bleeding from minor wounds, menorrhagia or postpartum bleeding in women as well as bleeding after surgery. Other less frequent symptoms include gastrointestinal bleeding, haematomas or haemarthroses. VWD pathophysiology is complex and results from defects in von Willebrand factor (VWF) glycoprotein. Quantitative deficiencies are responsible for type 1 VWD with a partial decrease of VWF and type 3 with the complete absence of VWF. Qualitative abnormalities cause type 2 VWD, being further divided into types 2A, 2B, 2M and 2N. Although common, VWD is at risk of misdiagnosis, overdiagnosis and underdiagnosis owing to several factors, including complex diagnosis, variability of bleeding symptoms, presence of external variables (blood groups and other physiological modifiers such as exercise, thyroid hormones, oestrogens, and ageing), and lack of disease awareness among non-specialist health-care providers. Establishing the correct VWD diagnosis requires an array of specialized phenotypic assays and/or molecular genetic testing of the VWF gene. The management of bleeding includes increasing endogenous VWF levels with desmopressin or infusion of exogenous VWF concentrates (plasma-derived or recombinant). Fibrinolytic inhibitors, topical haemostatic agents and hormonal therapies are used as effective adjunctive measures.

Type 2M/2A von Willebrand disease: a shared phenotype between type 2M and 2A

AbstractFour variants have been continuously subjected to debate and received different von Willebrand disease (VWD) classifications: p.R1315L, p.R1315C, p.R1374H, and p.R1374C. We chose to comprehensively investigate these variants with full set of VWD tests, protein-modeling predictions and applying structural biology. Patients with p.R1315L, p.R1315C, p.R1374H, and p.R1374C were included. A group with type 2A and 2M was included to better understand similarities and differences. Patients were investigated for phenotypic assays and underlying disease mechanisms. We applied deep protein modeling predictions and structural biology to elucidate the causative effects of variants. Forty-three patients with these variants and 70 with 2A (n = 35) or 2M (n = 35) were studied. Patients with p.R1315L, p.R1374H, or p.R1374C showed a common phenotype between 2M and 2A using von Willebrand factor (VWF):GPIbR/VWF:Ag and VWF:CB/VWF:Ag ratios and VWF multimeric profile, whereas p.R1315C represented a type 2M phenotype. There was an overall reduced VWF synthesis or secretion in 2M and cases with p.R1315L, p.R1374H, and p.R1374C, but not in 2A. Reduced VWF survival was observed in most 2A (77%), 2M (80%), and all 40 cases with p.R1315L, p.R1374H, and p.R1374C. These were the only variants that fall at the interface between the A1-A2 domains. p.R1315L/C mutants induce more compactness and internal mobility, whereas p.R1374H/C display a more extended overall geometry. We propose a new classification of type 2M/2A for p.R1315L, p.R1374H, and p.R1374C because they share a common phenotype with 2M and 2A. Our structural analysis shows the unique location of these variants on the A1-A2 domains and their distinctive effect on VWF.

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), <1 in type 2A patients (p.Arg1374His), to >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.

PB0850 Management of Type 2M von Willebrand Disease in Lower Limb Surgery

PB0850 Management of Type 2M von Willebrand Disease in Lower Limb Surgery

Formation of Heterobimetallic Complexes by Addition of d10-Metal Ions to [(Me3P)xM(2-C6F4PPh2)2] (x = 1, 2; M = Ni and Pt): A Synthetic and Computational Study of Metallophilic Interactions.

Treatment of the bis(chelate) complexes trans-[M(κ2-2-C6F4PPh2)2] (trans-1M; M = Ni, Pt) and cis-[Pt(κ2-2-C6F4PPh2)2] (cis-1Pt) with equimolar amounts or excess of PMe3 solution gave complexes of the type [(Me3P)xM(2-C6F4PPh2)2] (x = 2: 2Ma, 2Mb x = 1: 3Ma, 3Mb; M = Ni, Pt). The reactivity of complexes of the type 2M and 3M toward monovalent coinage metal ions (M' = Cu, Ag, Au) was investigated next to the reaction of 1M toward [AuCl(PMe3)]. Four different complex types [(Me3P)2M(μ-2-C6F4PPh2)2M'Cl] (5MM'; M = Ni, Pt; M' = Cu, Ag, Au), [(Me3P)M(κ2-2-C6F4PPh2)(μ-2-C6F4PPh2)M'Cl]x (x = 1: 6MM'; M = Pt; M' = Cu, Au; x = 2: 6PtAg), head-to-tail-[(Me3P)ClM(μ-2-C6F4PPh2)2M'] (7MM'; M = Ni, Pt; M' = Au), and head-to-head-[(Me3P)ClM(μ-2-C6F4PPh2)2M'] (8MM'; M = Ni, Pt; M' = Cu, Ag, Au) were observed. Single-crystal X-ray analyses of complexes 5-8 revealed short metal-metal separations (2.7124(3)-3.3287(7) Å), suggestive of attractive metal-metal interactions. Quantum chemical calculations (atoms in molecules (AIM), electron localization function (ELF), non-covalent interaction (NCI), and natural bond orbital (NBO)) gave theoretical support that the interaction characteristics reach from a pure attractive non-covalent to an electron-shared (covalent) character.

Genetic determinants of enhanced von Willebrand factor clearance from plasma

BackgroundEnhanced von Willebrand factor (VWF) clearance from plasma is associated with von Willebrand disease (VWD). However, the genetic background of this disease mechanism is not well defined. ObjectiveTo determine VWF variants that are associated with reduced VWF survival. MethodsTwo hundred fifty-four patients with VWD (type 1 = 50 and type 2 = 204) were investigated, and the results were compared with 120 healthy controls. The patients were comprehensively characterized for phenotypic and genetic features. The ratio of VWF propeptide (VWFpp)/VWF antigen (VWFpp ratio) was used to establish in each patient the VWF clearance state. ResultsOut of 92 variants associated with type 1 (7 were novel) and type 2 VWD, 19 had a VWFpp ratio ranging from 1.7 to 2.2, 24 had a VWFpp ratio between 2.3 and 2.9, and 24 variants had a ratio of ≥3. The VWFpp median ratio in healthy controls was 0.98 (0.55-1.6) so that a cut-off value of >1.6 was considered an indicator of accelerated VWF clearance from plasma. An enhanced VWF clearance was observed in 34% of type 1 cases, 100% of type 1 Vicenza cases, 81% of 2A cases, 77% of 2B cases, 88% of 2M cases, and 36% of 2N cases. ConclusionsAn accelerated VWF clearance was found in most patients with type 2A, 2B, and 2M VWD, with a lower proportion of type 1 and 2N. Sixty-seven different variants alone or in combination with other variants were associated with an increased VWFpp ratio. The variants with the highest VWFpp ratio were mostly located in the D3-A1 VWF domains.

A nanobody against the VWF A3 domain detects ADAMTS13-induced proteolysis in congenital and acquired VWD

Abstractvon Willebrand factor (VWF) is a multimeric protein, the size of which is regulated via ADAMTS13-mediated proteolysis within the A2 domain. We aimed to isolate nanobodies distinguishing between proteolyzed and non-proteolyzed VWF, leading to the identification of a nanobody (designated KB-VWF-D3.1) targeting the A3 domain, the epitope of which overlaps the collagen-binding site. Although KB-VWF-D3.1 binds with similar efficiency to dimeric and multimeric derivatives of VWF, binding to VWF was lost upon proteolysis by ADAMTS13, suggesting that proteolysis in the A2 domain modulates exposure of its epitope in the A3 domain. We therefore used KB-VWF-D3.1 to monitor VWF degradation in plasma samples. Spiking experiments showed that a loss of 10% intact VWF could be detected using this nanobody. By comparing plasma from volunteers to that from congenital von Willebrand disease (VWD) patients, intact-VWF levels were significantly reduced for all VWD types, and most severely in VWD type 2A–group 2, in which mutations promote ADAMTS13-mediated proteolysis. Unexpectedly, we also observed increased proteolysis in some patients with VWD type 1 and VWD type 2M. A significant correlation (r = 0.51, P < .0001) between the relative amount of high–molecular weight multimers and levels of intact VWF was observed. Reduced levels of intact VWF were further found in plasmas from patients with severe aortic stenosis and patients receiving mechanical circulatory support. KB-VWF-D3.1 is thus a nanobody that detects changes in the exposure of its epitope within the collagen-binding site of the A3 domain. In view of its unique characteristics, it has the potential to be used as a diagnostic tool to investigate whether a loss of larger multimers is due to ADAMTS13-mediated proteolysis.

Type 2M VWD Laboratory Defects More Consistent Than Bleeding Symptoms across Family Members

Background Von Willebrand disease (VWD) is an inherited bleeding disorder characterized by a defect or decrease of von Willebrand factor (VWF). Type 2 VWD results from qualitative defects in VWF. Type 2M VWD is defined by defective platelet binding to GPIb and/or collagen. While the inheritance of type 2M VWD is autosomal dominant, the extent of symptoms across family members has not been well characterized. Methods Subjects included index cases with type 2M VWD and all enrolled family members from a large US study of VWD (Zimmerman Program). Index cases were enrolled due to a prior diagnosis of VWD. All available family members were enrolled if willing. VWF testing performed included VWF antigen (VWF:Ag), VWF platelet binding by either VWF ristocetin cofactor activity (VWF:RCo) or VWF:GPIbM, collagen binding with types I, III or IV collagen (VWF:CB1, VWF:CB3, VWF:CB4) and VWF multimer distribution. Bleeding scores were calculated using the ISTH bleeding assessment tool (BAT). DNA sequencing was performed for the full length VWF gene. Statistics were calculated using GraphPad Prism with a p value of <0.05 considered significant. Family members were defined as affected if they had VWF activity levels consistent with type 2M VWD and presence of genetic variant if identified. Results 24 index cases, 51 affected family members and 40 unaffected family members were available for analysis. The mean VWF:RCo/VWF:Ag ratio for index cases was 0.5 and the mean VWF:CB/VWF:Ag ratio was 1.05. The VWF:RCo/VWF:Ag ratio was similar in affected family members (0.44, p=NS) but higher as expected in unaffected members (0.89, p<0.001 for index case vs. unaffected, p<0.001 for affected vs. unaffected). In contrast, there was no difference in VWF:CB/VWF:Ag ratios between index cases and affected family members (0.94, p=NS). Data for VWF:GPIbM, collagens I and IV were available only on index cases, and showed low VWF:GPIbM/VWF:Ag (mean 0.79), higher VWF:CBI/VWF:Ag (mean 1.31) and lower VWF:CBIV/VWF:Ag (mean 0.85). There was a negative correlation between VWF:GPIbM and VWF:CBIV, with subjects having either low VWF:GPIbM and normal VWF:CBIV or low VWF:CBIV and normal VWF:GPIbM. Similar results were seen for VWF:CB1 although only 1 subject was diagnosed with type 2M VWD and low VWF:CB1. Median bleeding scores were highest in index cases (7.5), intermediate in affected family members (3) and normal in unaffected family members (1). There was a significant difference between index cases and affected family members (<0.015), and between index cases and unaffected family members (p<0.001). One subject had a genetic variant in the VWF A3 domain, one in the VWF D3 domain and the remainder of index cases had a defect in the VWF A1 domain. Conclusions These findings suggest that decreases in platelet and collagen binding activity increase bleeding risk in families with type 2M VWD. Index cases had higher bleeding scores as compared to affected family members, suggesting that diagnosis of family members may require more attention. Precise understanding of these defects may assist in better diagnosis and treatment of von Willebrand disease.

A case report of type 2M Von Willebrand disease with unique mutations

Abstract Von Willebrand disease (vWD) is the most common inherited bleeding disorder. It results from quantitative or qualitative defects of the von Willebrand factor (vWF) protein, and is divided into three types, namely type 1, type 2(2A, 2B, 2M, and 2N), and type 3 vWD. Here we report a case of 55-year-old female who presented with recurrent gastrointestinal bleeding due to angiodysplasia. Her past medical history is significant for vWD (type unknown) with hemoptysis, epistaxis and menorrhagia requiring Humate-P replacement multiple times. She has one sister with mild vWD of unknown type and 4 other siblings with no bleeding history. Coagulation testing shows normal PT and aPTT. Von Willebrand panel shows vWF antigen of 33%, VWF ristocetin cofactor activity of 15%, and Factor VIII of 87%. The ratio (VWF: RCo/VWF: Ag) is low (0.45) and the multimer pattern is normal. This is consistent with type 2M vWD. VWD gene sequencing shows heterozygous variants of c.4241T&amp;gt;G (p.Val1414Gly) and c.5227G&amp;gt;A (p.Val1743Met). Of note, one variant (c.4241T&amp;gt;G/p.Val1414Gly) has been reported in two individuals with type 2A but not in type 2M vWD. This may truly represent different types of vWD caused by same mutation or could be due to variation in testing and/or result interpretation among laboratories. The other variant c.5227G&amp;gt;A(p.Val1743Met) in A3 domain has never been reported with uncertain significance. Further investigation including vWF panel testing and genetic analysis in family members would help characterize those mutations.

Thermodynamic stabilization of von Willebrand factor A1 domain induces protein loss of function.

The von Willebrand disease (vWD) is the most common hereditary bleeding disorder caused by defects of the von Willebrand Factor (vWF), a large extracellular protein in charge of adhering platelets to sites of vascular lesions. vWF performs this essential homeostatic task via specific protein-protein interactions between the vWF A1 domain and the platelet receptor, the glycoprotein Ib alpha (GPIBα). The two naturally occurring vWF A1 domain mutations G1324A and G1324S, near the GPIBα binding site, induce a dramatic decrease in platelet adhesion, resulting in a bleeding disorder classified as type 2M vWD. However, the reason for the drastic phenotypic response induced by these two supposedly minor modifications remains unclear. We addressed this question using a combination of equilibrium-molecular dynamics (MD) and nonequilibrium MD-based free energy simulations. Our data confirms that both mutations maintain the highly stable Rossmann fold of the vWF A1 domain. G1324A and G1324S mutations hardly changed the per-residue flexibility of the A1 domain but induced a global conformational change affecting the region near the binding site to GPIBα. Furthermore, we observed two significant changes in the vWF A1 domain upon mutation, the global redistribution of the internal mechanical stress and the increased thermodynamic stability of the A1 domain. These observations are consistent with previously reported mutations increasing the melting temperature. Overall, our results support the idea of thermodynamic conformational restriction of A1-before the binding to GPIBα-as a crucial factor determining the loss-of-function of the G1324A(S) vWD mutants.

Generalised idiopathic polymyositis mimicking masticatory myositis in a dog

AbstractMasticatory myositis (MM) and generalised idiopathic polymyositis (gIPM) are the most common immune‐mediated myositis in dogs. In humans, the diagnosis and classification of myositis have continuously evolved over the past few decades with the combination of histopathological and immunohistochemical evaluation alongside the identification of myositis‐specific autoantibodies (MSAs). In dogs, however, MSAs have only been described in MM and breeds such as Newfoundland and Boxer. We report a case of a dog that presented with sub‐acute pain when chewing and exophthalmos, followed by chronic progressive masticatory muscle atrophy and enophthalmos. Clinical signs and electromyography were primarily consistent with MM, but serological autoantibodies against type 2M myofibres were negative. Creatine kinase activity, histological and immunohistochemical examination of the temporal and triceps brachii muscles and negative infectious disease tests were consistent with gIPM. This atypical case highlights the requirement for multiple muscle biopsies and appeals for extended immunohistochemical studies in canine myositis.

Postpartum Hemorrhage in Women with von Willebrand Disease: Consider Other Etiologies

ObjectiveHigher rates of postpartum hemorrhage (PPH) have been reported for women with von Willebrand disease (VWD). Comprehensive multidisciplinary care reduces these rates; thus PPH may not be secondary to VWD. MethodsWe conducted a retrospective review for the period of 2009–2018, including all VWD pregnancies at 2 tertiary care academic hospitals to determine rates, etiology, and timing of PPH. ResultsA total of 63 women with 80 pregnancies were included. Three women had twin pregnancies. Sixty-six pregnancies (82.5%) involved type 1 VWD; 4 (5.0%), type 2 (unclear subtype); 3 (3.8%) type 2A; 3 (3.8%) type 2B; and 2 (2.5%), type 2M. Median age of patients was 32.9 years (range 19–43 y). Most patients were blood type O (65%), and 33 of 80 pregnancies (41.3%) were nulliparous. The mean bleeding assessment score was 8 (range 0–16). Thirty-seven pregnancies (46.3%) received prophylactic hemostatic treatment prior to delivery. Seventy-four percent of pregnancies were delivered vaginally, and 88% received epidural anaesthesia. The majority of pregnancies (78.8%) had von Willebrand factor (VWF) levels assessed during the third trimester, with most (71.3%) achieving VWF levels above 1.00 IU/mL. Four pregnancies (5.2%) were complicated by primary PPH; uterine atony in 2 and placenta previa in 1. Delayed postpartum bleeding occurred in 5 pregnancies (6.3%). ConclusionMultidisciplinary care of pregnancies with VWD improves outcomes. Rates of primary and delayed PPH in this study are lower than previously described and are similar to those of women without VWD. In women with VWD, uterine etiologies for primary PPH need to be considered, in a manner similar to the assessment of women without VWD, to ensure hemostasis is achieved.

Desmopressin response depends on the presence and type of genetic variants in patients with type 1 and type 2 von Willebrand disease

AbstractPatients with type 1 and type 2 von Willebrand disease (VWD) can be treated with desmopressin. Although a previous study has shown that the location of the causative VWF gene variant is associated with desmopressin response in type 1 VWD, the association between variants in the VWF gene and desmopressin response is not yet fully understood. Our primary aim was to compare desmopressin response in type 1 VWD patients with and without a VWF gene variant. Secondly, we investigated whether desmopressin response depends on specific VWF gene variants in type 1 and type 2 VWD. We included 250 patients from the Willebrand in the Netherlands study: 72 type 1 without a VWF gene variant, 108 type 1 with a variant, 45 type 2A, 16 type 2M, and 9 type 2N patients. VWF gene was analyzed with ion semiconductor sequencing and Multiplex Ligation-dependent Probe Amplification. Complete response to desmopressin was observed in all type 1 VWD patients without a variant, 64.3% of type 1 patients with a variant, and 31.3% of type 2 patients (P < .001). Despite a large interindividual variability in desmopressin response, patients with the same variant had comparable desmopressin responses. For instance, in 6 type 1 patients with exon 4 to 5 deletion, mean VWF activity at 1 hour after desmopressin was 0.81 IU/mL, with a coefficient of variation of 22.9%. In conclusion, all type 1 VWD patients without a VWF gene variant respond to desmopressin. In type 1 and type 2 VWD patients with a VWF variant, desmopressin response highly depends on the VWF gene variants.

Von Willebrand disease type 2M: Correlation between genotype and phenotype

BackgroundAn appropriate clinical diagnosis of von Willebrand disease (VWD) can be challenging because of a variable bleeding pattern and laboratory phenotype. Genotyping is a powerful diagnostic tool and may have an essential role in the diagnostic field of VWD. ObjectivesTo unravel the clinical and laboratory heterogeneity of genetically confirmed VWD type 2M patients and to investigate their relationship. MethodsPatients with a confirmed VWD type 2M genetic variant in the A1 or A3 domain of von Willebrand factor (VWF) and normal or only slightly aberrant VWF multimers were selected from all subjects genotyped at the Radboud university medical center because of a high suspicion of VWD. Bleeding scores and laboratory results were analyzed. ResultsFifty patients had a clinically relevant genetic variant in the A1 domain. Median bleeding score was 5. Compared with the nationwide Willebrand in the Netherlands study type 2 cohort, bleeding after surgery or delivery was reported more frequently and mucocutaneous bleedings less frequently. Median VWF activity/VWF antigen (VWF:Act/VWF:Ag) ratio was 0.32, whereas VWF collagen binding activity/VWF antigen (VWF:CB/VWF:Ag) ratio was 0.80. Variants in the A3 domain were only found in two patients with low to normal VWF:Act/VWF:Ag ratios (0.45, 1.03) and low VWF:CB/VWF:Ag ratios (0.45, 0.63). ConclusionGenetically confirmed VWD type 2M patients have a relatively mild clinical phenotype, except for bleeding after surgery and delivery. Laboratory phenotype is variable and depends on the underlying genetic variant. Addition of genotyping to the current phenotypic characterization may improve diagnosis and classification of VWD.

Quantitative Analysis of Von Willebrand Factor (VWF) Multimers in Von Willebrand Disease (VWD) Patients Recruited through the Zimmerman Program for the Molecular and Clinical Biology of VWD

Quantitative Analysis of Von Willebrand Factor (VWF) Multimers in Von Willebrand Disease (VWD) Patients Recruited through the Zimmerman Program for the Molecular and Clinical Biology of VWD

A New Frontier Has Emerged That Enables the Reliable Diagnosis and Subtyping of Von Willebrand Disease (vWD) Clinical Patients in 48 Hours

Remarkably, classical von Willebrand disease (vWD) laboratory work-ups have been shown to require repeat laboratory testing 1 to 20 times to confirm diagnosis [1]. Furthermore, most diagnostic laboratory algorithms terminate with molecular confirmation [2, 3]. Here we present the method validation of a diagnostic laboratory approach that allows for the simultaneous functional and genetic diagnosis and subtyping von Willebrand disease (vWD) and pseudo- or platelet-type vWD within 48 hours. The clinical sensitivity of this test is approximately 85% for subtypes 2A, 2B, 2N, 2M and 3 vWD [4].The method validation is compliant with current Clinical Laboratory Improvement Act (CLIA) and applicable current Good Laboratory Practices (cGLP, 21 CFH Part 58) guidelines and therefore is suitable for clinical utilization and clinical trial utilization. The method validation protocol included the creation of reference intervals, reportable ranges, intra-run precision, inter-run precision, accuracy, lower limit of quantification (LLQ), linearity, analytical and clinical sensitivity assessment, analytical and clinical specificity assessment, interfering substances, stability and longitudinal quality control performance [Table 1.].Historically, no one test has been able to diagnose vWD. Classical laboratory work-ups include aPTT, factor VIII (8) activity, Ristocetin cofactor activity, vWF antigen, ratio calculations, multimeric composition and others. Each of these tests is influenced by numerous factors, such as inflammation, stress, infection, hormone replacement therapy, age, acute phase response, menstrual cycle, pregnancy exercise, ABO blood type, lupus anticoagulant and other factors, making traditional diagnosis a challenging moving target [3, 5, 6].The methodology used in this study employed simultaneous classical vWD laboratory testing and rapid next generation sequencing (NGS) optimized for 48-hour result availability [Table 2]. Furthermore, normal healthy and vWD 2N subtype samples can clearly be distinguished in this testing approach [Table 3].vWD occurs with bleeding symptoms at rate of 1 in 1,000 in the general population [6]. In 10% of those cases the bleeding is severe [3]. This prevalence makes vWD the most common congenital bleeding disorder known [6]. Acquire vWD (1-5% of cases) can be assessed by contrasting functional von Willebrand testing with genetic testing in a specialized coagulation, platelet and genetics laboratory. Common symptoms of vWD include menorrhagia, prolonged bleeding or bruising from minor trauma, recurrent nose bloods, excessive bleeding at childbirth and excessive bleeding associated with surgery.In conclusion, the standard clinical approach of diagnosing vWD can take months to years and require one or more referrals to specialized clinicians to establish diagnosis and subtyping. The data presented in this study proposes that an alternative method of rapidly characterizing suspected vWD patients simultaneously with both functional and genetic tests can revolutionize speed of diagnosis and treatment.vWD Subtypes: (each can have severe clinical presentations)Type 1 = 60%Type 2 = 30%Type 2A = 15%Type 2B = 5%Type 2M < 5%Type 2N <5% (clinically similar to mild hemophilia A)Type 3 = 5-10%Pseudo-vWD or Platelet-type vWD = rare (defect in GP1BA gene)Acquired vWD = 1-5%

Self-Reported Reproductive Health Experiences in Women with Von Willebrand Disease: A Qualitative Interview-Based Study

Background: Women with von Willebrand disease (VWD) may experience multiple reproductive health challenges, particularly related to menstruation and pregnancy. Patient-reported experiences in this setting have not been well characterized.Aim: To elicit patient-reported menstrual health experiences in women with VWD.Methods: We conducted a physician-administered telephone survey of women seen at Mayo Clinic for workup and/or management of VWD. Patients were asked to discuss their diagnosis of VWD and questioned about menstrual health, contraception, and pregnancy and asked to describe how their VWD had impacted their reproductive health experiences. Quantitative data were analyzed using R (version 3.2.0). Qualitative comments were analyzed by frequency of response (specific adjectives) and general thematic elements.Results: Of 130 women diagnosed with VWD, 82 (63%) agreed to participate, including 58 (71%) with type 1, 9 (11%) with type 2A, 4 (5%) with type 2B, 8 (10%) with type 2M, 1 (1%) with type 2N, and 3 (2%) with type 3. 89% of women self-reported a history of heavy menstrual bleeding; for 23 (28%) this was their first bleeding event. 28 women (34%) ultimately underwent hysterectomy for heavy menstrual bleeding. Women used several forms of contraception, most commonly estrogen-containing oral contraceptives (61, 74%) and intrauterine device (9, 11%). 65 women (79%) reported at least one pregnancy. There were 200 total pregnancies (median 3 per woman, range 1-10). 142 pregnancies (71%) resulted in live term birth, 47 (24%) in spontaneous abortion (at a median of 12 weeks), 9 (4%) in a live preterm birth, and 2 (1%) in therapeutic abortion. Patients' self-reported comments are described in the Table, with responses in bold indicating the same themes were used by more than one patient. Women used primarily negative adjectives to describe heavy menstrual bleeding and reported a significant negative impact on their academic, home, and sexual lives. Despite this, women often reported that they “just dealt with it” and “thought it was normal.” Regarding pregnancy and VWD, women again used primarily negative adjectives to describe their experiences. Many described pregnancy-associated bleeding events (generally postpartum hemorrhage). Overall, women reported that it was often “uncomfortable” to discuss their reproductive concerns with their medical providers (including hematologists). One woman reported that she “wished [she] had been more confident speaking with [her] doctor” and another that “it is important to be your own advocate.”Conclusions: VWD is a relatively common bleeding condition which appears to significantly impact women's reproductive health. In this qualitative interview study, women often used negative adjectives to describe their reproductive experiences as related to VWD. Additionally, many were uncomfortable discussing their reproductive health with their providers. Physicians (particularly hematologists and obstetrician-gynecologists) should be trained to recognize the impact of VWD on reproductive health and include reproductive health as part of their essential health maintenance conversations with their patients. More compassionate and informational patient-physician conversations may help to optimize reproductive health experiences and outcomes in women with VWD. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Type 2A and 2M von Willebrand Disease: Differences in Phenotypic Parameters According to the Affected Domain by Disease-Causing Variants and Assessment of Pathophysiological Mechanisms.

Type 2A and 2M von Willebrand disease (VWD) broadly show similar phenotypic parameters, but involve different pathophysiological mechanisms. This report presents the clinical and laboratory profiles of type 2A and type 2M patients genotypically diagnosed at one large center. Higher bleeding score values and a higher incidence of major bleeding episodes were observed in type 2A compared with type 2M, potentially reflective of the absence of large and intermediate von Willebrand factor (VWF) multimers in 2A. In type 2A, most of disease-causing variants (DCVs) appeared to be responsible for increased VWF clearance and DCV clustered in the VWF-A1 domain resulted in more severe clinical profiles. In type 2M, DCV in the VWF-A1 domain showed different laboratory patterns, related to either reduced synthesis or shortened VWF survival, and DCV in the VWF-A2 domain showed patterns related mainly to shortened survival. VWF-type 1 collagen binding/Ag (C1B/Ag) showed different patterns according to DCV location: in type 2A VWD, C1B/Ag was much lower when DCVs were located in the VWF-A2 domain. In type 2M with DCV in the VWF-A1domain, C1B/Ag was normal, but with DCV in the VWF-A2 domain, C1B/Ag was low. The higher frequency of major bleeding in VWD 2M patients with DCV in the VWF-A2 domain than that with DCV in the VWF-A1 domain could be a summative effect of abnormal C1B/Ag, on top of the reduced VWF-GPIb binding. In silico modeling suggests that DCV impairing the VWF-A2 domain somehow modulates collagen binding to the VWF-A3 domain. Concomitant normal FVIII:C/Ag and VWFpp/Ag, mainly in type 2M VWD, suggest that other nonidentified pathophysiological mechanisms, neither related to synthesis/retention nor survival of VWF, would be responsible for the presenting phenotype.

Surgical stabilisation of hiatal hernia and gastroesophageal reflux associated with idiopathic inflammatory polymyopathy in a Wire Fox Terrier

A one-and-a-half-year-old male Wire Fox Terrier weighing 3 kg presented with continuous drooling and vomiting. Its body condition score was 3/9, and severe atrophy of the temporal/masticatory muscle, trismus, and enophthalmos was observed on physical examination. The radiographic examination and fluoroscopic oesophagography revealed a type 1 hiatal hernia with gastroesophageal reflux. The serology tests revealed increased muscle enzyme activities. The antibody tests for acetylcholine receptor, type 2M fibre, and infectious agents were negative. A conventional surgical treatment was performed, and a thickened, stretched, and flaccid diaphragmatic muscle and an extended inelastic oesophageal hiatus were observed. On the histological examination of the diaphragmatic muscle, a diffuse histiocytic myositis was confirmed. Although the postoperative gastroesophageal reflux totally disappeared, the oesophageal motility and clinical signs did not improve significantly. Medical treatment with immunosuppressive agents was attempted and was effective in alleviating the clinical signs and abnormal oesophageal motility. The health condition of the dog was adequately maintained in the 12-month monitoring period. These findings suggest that, although the medical and surgical treatment have different therapeutic effects, they should be considered simultaneously for the management of a hiatal hernia associated with polymyopathy in dogs.