Acquired von Willebrand Syndrome and Desmopressin Resistance During Venovenous Extracorporeal Membrane Oxygenation in Patients With COVID-19: A Prospective Observational Study.

OBJECTIVES: Although COVID-19 is associated with high von Willebrand factor (vWF) parameters promoting thrombosis, venovenous extracorporeal membrane oxygenation (vvECMO) is associated with the development of acquired von Willebrand syndrome (AVWS) promoting bleeding. This study was designed to assess both the incidence and severity of AVWS in COVID-19 patients undergoing vvECMO, and the benefit of comprehensive vWF analyses. DESIGN: Prospective observational study. SETTING: ICU at a tertiary-care center. PATIENTS: Twenty-seven consecutive COVID-19 patients with acute respiratory distress syndrome (ARDS) requiring vvECMO. MEASUREMENTS AND MAIN RESULTS: Comprehensive vWF analyses (including sodium dodecyl-sulfate polyacrylamide gel electrophoresis) were performed before, during, and after vvECMO. In a subgroup of 12 patients with AVWS, effectiveness of treatment with desmopressin was assessed. The patients' mean age was 53 years (range, 23-73), 70% were male, and all had various comorbidities. Following markedly elevated vwf antigen (vWF: Ag; mean, 546% ( sd , 282]), vWF collagen binding capacity (mean, 469% [ sd , 271]), vWF activity (vWF:A; mean, 383% [ sd , 132]), and factor VIII activity (mean, 302% [ sd , 106]), and only borderline decreases in high-molecular-weight (HMW) vWF multimers before vvECMO, all of these variables decreased and HMW vWF multimers became undetectable within hours following initiation of vvECMO. All variables fully recovered within 3-38 hours after discontinuation of vvECMO. During vvECMO, decreases in the vWF:A/vWF:Ag ratio correlated with absent HMW vWF multimers. Desmopressin did not affect vWF parameters. CONCLUSIONS: In patients with COVID-19-associated ARDS, AVWS developed soon after initiation of vvECMO. The vWF:A/vWF:Ag ratio was a suitable screening test for AVWS. As desmopressin was ineffective, bleeding during vvECMO-associated AVWS should preferably be treated with concentrates containing vWF.

Emicizumab in children: bleeding episodes and outcome before and after transition to Emicizumab.

PURPOSE: Real-world data and study data regarding therapy with Emicizumab in pediatric cohorts with haemophilia A is scarce. Especially, data on previously untreated pediatric patients (PUPs) and minimally treated patients (MTPs) are missing. METHODS: Thirteen pediatric patients with haemophilia A and treatment with Emicizumab were retrospectively evaluated for Annual Bleeding Rates (ABR) pre-and post-Emicizumab treatment. Safety data and data on management of minor surgery as well as laboratory results were collected. Additionally, we describe the clinical features of two PUPs and one MTP that are included in our cohort. RESULTS: Median age at initiation of Emicizumab was 5.3 (range: 0.26-17.5) years, three patients were younger than one year at initiation of treatment with Emicizumab. Median follow-up time on Emicizumab was 23.8 (range: 0.7-40) months. Total ABR (p = 0.009) as well as spontaneous (p = 0.018), traumatic (p = 0.018), and joint (p = 0.027) ABR reduced significantly post-Emicizumab transition. Safety profile was favourable as only one local site reaction occurred; no cessation of treatment was necessary. Surgery was successfully performed in three patients receiving rFVlla pre- and post-surgery. Emicizumab trough levels showed a median of 43.2 µg/ml (range: 23.9-56.8) after three doses of 3 mg/kg and 51.9 µg/ml (range: 30.4-75) at first follow-up with 1.5 mg/kg. CONCLUSION: Emicizumab is safe and efficient in pediatric patients with and without inhibitors. More data on larger multicenter cohorts and especially on PUPs/MTPs are still needed.

Hypercoagulopathy, acquired coagulation disorders and anticoagulation before, during and after extracorporeal membrane oxygenation in COVID-19: a case series.

BACKGROUND: Thromboembolism and bleeding contribute to Coronavirus disease 2019 (COVID-19)'s morbidity and mortality and are also frequent complications of venovenous extracorporeal membrane oxygenation (vvECMO). As the interaction of the underlying pathologies caused by vvECMO in COVID-19 is barely understood, we designed this study to better differentiate coagulation disorders in COVID-19 patients before, during and after vvECMO-support. METHODS: Observational case series, six consecutive patients with Coronavirus acute respiratory distress syndrome supported with vvECMO treated in the anaesthesiologic ICU in a third level University ECMO-centre. We measured routine coagulation parameters and assessed coagulation factors. We also conducted advanced von Willebrand factor (VWF) multimer analysis, platelet aggregometry and immunological screening. RESULTS: We identified various phases of coagulation disorders: Initially, intensely activated coagulation with highly increased VWF and factor VIII activity in acute COVID-19, then severe acquired von Willebrand syndrome and platelet dysfunction during vvECMO leading to spontaneous bleeding and finally, hypercoagulopathy after vvECMO explantation. Five of six patients developed immunological abnormalities enhancing coagulation. CONCLUSIONS: Coronavirus-induced coagulopathy and bleeding disorders during vvECMO cannot be discriminated via 'routine' coagulation tests. Precise and specific analyses followed by the appropriate treatment of coagulation disorders may help us develop tailored therapeutic concepts to better manage the phases described above.

Novel Likely Pathogenic Variant in the A3 Domain of von Willebrand Factor Leading to a Collagen-Binding Defect.

Von Willebrand disease (VWD) is the most prevalent congenital bleeding disorder. Diagnosis and classification of VWD is complex due to its heterogeneity regarding clinical manifestations and molecular genetic analysis. Genetic investigations became an inherent part of diagnosis and help distinguish different types/subtypes of VWD. Although many variants have been listed being causative for VWD, the genetic etiology remains undefined in a lot of patients. We report about two siblings with severely reduced values for von Willebrand factor collagen-binding activity (VWF:CB). Genetic analysis using panel sequencing identified a heterozygous non-synonymous single nucleotide variant in exon 30. At the protein level, the alteration (p.Ser1731Leu) is located in the A3 collagen-binding domain. The amino acid position is already known to be important for collagen binding because p.Ser1731Thr has been reported to affect the VWF:CB.

Successful Secondary Endovascular Intervention in Pediatric Patients with Venous Thromboembolic Events.

BACKGROUND: In the past, pediatric patients with venous thromboembolic events (VTE) were treated with low-molecular-weight heparin (LMWH) which was successful in around 70% of the cases. However, anticoagulation alone might not restore patency in all patients, and advanced therapeutic options to prevent postthrombotic syndrome are needed. During recent years, endovascular interventions have become a treatment option for pediatric patients with persistent thrombotic occlusion, not only in life- or limb-threatening VTE. METHODS: We evaluated 12 consecutive patients (11-17 years) with newly diagnosed VTE being treated at our department during the last 4 years (2017-2020). In case follow-up examination showed persistent venoocclusion under anticoagulation, patients received secondary interventional therapy like recanalization, percutaneous transluminal angioplasty with or without catheter-directed thrombolysis, and stenting. Patients with no clinical signs of venoocclusion or regredient thrombosis in imaging examination received anticoagulation alone. RESULTS: Six of 12 (50%) patients underwent catheter intervention. Median time from diagnosis to intervention was 4 months (0-12 months). Reintervention was necessary in one (8%) case and complete recanalization failed in one (8%) case. There were no major bleeding events or other major postinterventional complications, no acute or late local recurrence, and all patients reported clinical improvement after the procedure. CONCLUSION: If endovascular intervention is used in teenage patients with persistent symptomatic VTE, reduction of postthrombotic symptoms is possible, even if intervention is performed secondary to failure of anticoagulation. Multidisciplinary treatment decisions can be based on the clinical course and follow-up imaging.

Laboratory Findings, Medical Imaging, and Clinical Outcome in Children with Cerebral Sinus Venous Thrombosis.

INTRODUCTION: Cerebral sinus venous thrombosis (CSVT) is a rare disease, especially in children. Therefore, thrombophilia markers, risk factors, treatment strategy, and MRI, as well as clinical outcome need further investigation to support future diagnostic and therapeutic guidelines for children. METHODS: We retrospectively identified all children with CSVT treated in our center between January 1, 2000, and December 31, 2015. Risk factors and laboratory findings were investigated. Furthermore, outcome and treatment satisfaction were evaluated using magnetic resonance imaging (MRI) analyses and a modified questionnaire. RESULTS: All 43 patients, who agreed to participate, were treated with therapeutic levels of heparin; 86% of children had an increased risk for thromboembolic events upon onset of CSVT (acute disease: 58.1%, perinatal risk factors: 9.3%, medical intervention/immobility: 14%, chronic disease: 16.3%). Thrombophilia markers showed positive results (e.g., reduced values for protein C/S, factor-V-Leiden mutation) in 58% of children at the time of CSVT diagnosis but dropping to 20.9% over the course of the disease. Forty-two of 43 patients received MRI follow-ups and the outcome showed complete recanalization in 69% of the patients and partial recanalization in 31%. At the onset of CSVT, 88% of patients reported restrictions in everyday life due to CSVT; at follow-up this percentage declined to 18%. Satisfaction with the outcome among parents/patients according to the questionnaire was high with 1.7 (German school grades from 1 to 6). CONCLUSIONS: All 42 children with MRI follow-up demonstrated complete or partial recanalization under anticoagulation. This positive result underlines the need for future studies on anticoagulation to optimize therapy regimens of pediatric CSVT.

Prepartum Anemia and Risk of Postpartum Hemorrhage: A Meta-Analysis and Brief Review.

Postpartum hemorrhage (PPH) is responsible for 30% to 50% of maternal deaths. There is conflicting evidence if prepartum anemia facilitates PPH. A comprehensive analysis of studies describing their relation is missing. An extensive database search was conducted applying the terms "anemia" OR "hemoglobin" AND "postpartum hemorrhage." We used a random-effects meta-analysis model to estimate an overall odds ratio (OR) for PPH and prepartum anemia, separating studies that were conformant and non-conformant with the World Health Organization (WHO) definitions for anemia. The search yielded 2519 studies, and 46 were appropriate for analysis. The meta-analyses of WHO-conformant (n = 22) and non-conformant (n = 24) studies showed that the risk of PPH was increased when anemia was present. The ORs were 1.45 (CL: 1.23-1.71) for WHO-conformant studies, 2.88 (CL: 1.38-6.02) for studies applying lower thresholds for anemia, and 3.28 (CL: 2.08-5.19) for undefined anemia thresholds. PPH risk appeared to increase with lower anemia thresholds. Prepartum anemia is associated with an increased risk of PPH, an observation that is important regarding improved anemia correction strategies such as iron supplementation.

Acquired von Willebrand Syndrome in Children.

Acquired von Willebrand syndrome (AVWS) is a rare bleeding disorder caused by various underlying diseases or conditions and should be distinguished from the inherited type of von Willebrand disease. AVWS is associated with underlying diseases such as cardiovascular, autoimmune, malignant, proliferative disorders, or with mechanical circulatory support (MCS). AVWS was first reported in 1968 and most case reports describe AVWS in adults. However, AVWS can appear in pediatric patients occasionally as well. Because bleeding complications are rare in everyday life, AVWS may be underdiagnosed in pediatric patients. Therefore, the diagnosis should be suspected in a pediatric patient who is known for one of these underlying diseases or conditions and who presents with an onset of bleeding symptoms, especially before the child will undergo an invasive procedure. Here, we present an overview of the diagnostic analyses regarding AVWS and of the underlying diseases or conditions in which AVWS should be considered. Importantly, the patient's history should be investigated for bleeding symptoms (mucocutaneous or postoperative bleeding). As no single routine coagulation test can reliably confirm or exclude AVWS, the diagnosis may be challenging. Laboratory investigations should include analysis of von Willebrand factor (VWF):antigen, VWF:collagen-binding capacity, VWF:activity, and VWF multimeric analyses. For treatment, tranexamic acid, 1-desamino-8-D-arginine vasopressin, and VWF-containing concentrate can be used. AVWS disappears after the underlying disease has been successfully treated or the MCS has been explanted.

Comprehensive Analyses of Coagulation Parameters in Patients with Vascular Anomalies.

BACKGROUND: Vascular anomalies comprise a diverse group of rare diseases with altered blood flow and are often associated with coagulation disorders. The most common example is a localized intravascular coagulopathy in venous malformations leading to elevated D-dimers. In severe cases, this may progress to a disseminated intravascular coagulopathy with subsequent consumption of fibrinogen and thrombocytes predisposing to serious bleeding. A separate coagulopathy is the Kasabach-Merritt phenomenon in kaposiform hemangioendothelioma characterized by platelet trapping leading to thrombocytopenia and eventually consumptive coagulopathy. Our previous work showed impaired von Willebrand factor and platelet aggregometry due to abnormal blood flow, i.e., in ventricular assist devices or extracorporeal membrane oxygenation. With altered blood flow also present in vascular anomalies, we hypothesized that, in particular, the von Willebrand factor parameters and the platelet function may be similarly impacted. METHODS: We prospectively recruited 73 patients with different vascular anomaly entities and analyzed their coagulation parameters. RESULTS: Acquired von Willebrand syndrome was observed in both of our patients with Kasabach-Merritt phenomenon. In six out of nine patients with complex lymphatic anomalies, both the vWF antigen and activity were upregulated. Platelet aggregometry was impaired in both patients with Kasabach-Merritt phenomenon and in seven out of eight patients with an arteriovenous malformation. CONCLUSIONS: The analysis of coagulation parameters in our patients with vascular anomalies advanced our understanding of the underlying pathophysiologies of the observed coagulopathies. This may lead to new treatment options for the, in part, life-threatening bleeding risks in these patients in the future.

Novel GNE Gene Variants Associated with Severe Congenital Thrombocytopenia and Platelet Sialylation Defect.

The GNE gene encodes an enzyme that initiates and regulates the biosynthesis of N-acetylneuraminic acid, a precursor of sialic acids. GNE mutations are classically associated with Nonaka myopathy and sialuria, following an autosomal recessive and autosomal dominant inheritance pattern. Reports show that single GNE variants cause severe thrombocytopenia without muscle weakness. Using panel sequencing, we identified two novel compound heterozygous variants in GNE in a young girl with life-threatening bleedings, severe congenital thrombocytopenia, and a platelet secretion defect. Both variants are located in the nucleotide-binding site of the N-acetylmannosamin kinase domain of GNE. Lectin array showed decreased α-2,3-sialylation on platelets, consistent with loss of sialic acid synthesis and indicative of rapid platelet clearance. Hematopoietic stem cell transplantation (HSCT) normalized platelet counts. This is the first report of an HSCT in a patient with an inherited GNE defect leading to normal platelet counts.

A Novel GATA1 Variant in the C-Terminal Zinc Finger Compared with the Platelet Phenotype of Patients with A Likely Pathogenic Variant in the N-Terminal Zinc Finger.

The GATA1 transcription factor is essential for normal erythropoiesis and megakaryocytic differentiation. Germline GATA1 pathogenic variants in the N-terminal zinc finger (N-ZF) are typically associated with X-linked thrombocytopenia, platelet dysfunction, and dyserythropoietic anemia. A few variants in the C-terminal ZF (C-ZF) domain are described with normal platelet count but altered platelet function as the main characteristic. Independently performed molecular genetic analysis identified a novel hemizygous variant (c.865C>T, p.H289Y) in the C-ZF region of GATA1 in a German patient and in a Spanish patient. We characterized the bleeding and platelet phenotype of these patients and compared these findings with the parameters of two German siblings carrying the likely pathogenic variant p.D218N in the GATA1 N-ZF domain. The main difference was profound thrombocytopenia in the brothers carrying the p.D218N variant compared to a normal platelet count in patients carrying the p.H289Y variant; only the Spanish patient occasionally developed mild thrombocytopenia. A functional platelet defect affecting αIIbß3 integrin activation and α-granule secretion was present in all patients. Additionally, mild anemia, anisocytosis, and poikilocytosis were observed in the patients with the C-ZF variant. Our data support the concept that GATA1 variants located in the different ZF regions can lead to clinically diverse manifestations.

Pathogenic Aspects of Inherited Platelet Disorders.

Inherited platelet disorders (IPDs) constitute a large heterogeneous group of rare bleeding disorders. These are classified into: (1) quantitative defects, (2) qualitative disorders, or (3) altered platelet production rate disorders or increased platelet turnover. Classically, IPD diagnostic is based on clinical phenotype characterization, comprehensive laboratory analyses (platelet function analysis), and, in former times, candidate gene sequencing. Today, molecular genetic analysis is performed using next-generation sequencing, mostly by targeting enrichment of a gene panel or by whole-exome sequencing. Still, the biochemical and molecular genetic characterization of patients with congenital thrombocytopathias/thrombocytopenia is essential, since postoperative or posttraumatic bleeding often occurs due to undiagnosed platelet defects. Depending upon the kind of surgery or trauma, this bleeding may be life-threatening, e.g., after tonsillectomy or in brain surgery. Undiagnosed platelet defects may lead to additional surgery, hysterectomy, pulmonary bleeding, and even resuscitation. In addition, these increased bleeding symptoms can lead to wound healing problems. Only specialized laboratories can perform the special platelet function analyses (aggregometry, flow cytometry, or immunofluorescent microscopy of the platelets); therefore, many IPDs are still undetected.

Type 2B von Willebrand Disease: Early Manifestation as Neonatal Thrombocytopenia.

Here, we report about a preterm female newborn with a prolonged course of severe thrombocytopenia and hematomas. The family history was positive for von Willebrand disease type 2B (VWD 2B). Diagnosis of VWD 2B was identified analyzing von Willebrand factor (VWF) parameters (VWF:antigen, VWF:activity, VWF multimer analyses) and performing light transmission aggregometry (with half concentration of ristocetin). In addition, the diagnosis was confirmed by molecular genetic analysis: identification of a disease-causing missense mutation (Val1316Met) in the VWF gene associated with a severe course of VWD 2B, which had been previously reported. Treatment with a VWF-containing plasma concentrate was initiated. Because the combination of prematurity and very low platelet count is often associated with intracranial bleeding, at the beginning platelet concentrates were transfused. Fortunately, the patient did not develop serious bleeding episodes. Interestingly, the patient had a mutation in the VWF gene, which had been described to be associated with aggravation of thrombocytopenia especially in stressful situations. Therefore, we replaced venous blood withdrawals by capillary blood samplings when possible and, consequently, we observed an increase of the platelet count after this change in management. At the age of 2 months, the patient was discharged after stabilization of the platelet count without any bleeding signs and without a need of long-term medication.

Hermansky-Pudlak Syndrome: Identification of Novel Variants in the Genes HPS3, HPS5, and DTNBP1 (HPS-7).

Hermansky-Pudlak syndrome (HPS), a rare heterogeneous autosomal recessive disorder, is characterized by oculocutaneous albinism (OCA) and a bleeding diathesis due to a defect regarding melanosomes and platelet delta (δ)-granule secretion. Interestingly, patients with HPS type 2 (HPS-2) or HPS type 10 (HPS-10) present additionally with an immunological defect. We investigated three patients (IP1, IP2, and IP3) who suffer from a bleeding diathesis. Platelet aggregometry showed impaired platelet function and flow cytometry revealed a severely reduced platelet CD63 expression hinting to either a defect of platelet delta granule secretion or a decreased number of delta granules in these patients. However, only IP3 presents with an apparent OCA. We performed panel sequencing and identified a homozygous deletion of exon 6 in DTNBP1 for IP3. Western analysis confirmed the absence of the encoded protein dysbindin confirming the diagnosis of HPS-7. Interestingly, this patient reported additionally recurrent bacterial infections. Analysis of lymphocyte cytotoxicity showed a slightly reduced NK-degranulation previously documented in a more severe form in patients with HPS-2 or HPS-10. IP1 is carrier of two compound heterozygous variants in the HPS3 gene (c.65C > G and c.1193G > A). A homozygous variant in HPS5 (c.760G > T) was identified in IP2. The novel missense variants were classified as VUS (variant of uncertain significance) according to ACMG guidelines. For IP1 with the compound heterozygous variants in HPS3 a specialized ophthalmological examination showed ocular albinism. HPS3 and HPS5 encode subunits of the BLOC-2 complex and patients with HPS-3 or HPS-5 are known to present with variable/mild hypopigmentation.

Case Report: Hepatic Adenoma in a Child With a Congenital Extrahepatic Portosystemic Shunt.

Congenital extrahepatic portosystemic shunts (CEPS), previously also described as Abernethy malformations, are rare malformations in which the extrahepatic portal system directly communicates with the vena cava inferior, thereby bypassing the liver. A hypoplastic portal vein (PV) exists in most cases. CEPS have been associated with the development of liver nodules, ranging from mostly focal nodular hyperplasia (FNH) to hepatic adenoma (HA) and even hepatocellular carcinoma (HCC). Tumor development in CEPS may be due to changes in perfusion pressures, oxygen supply or endocrine imbalances. It is important to rule out CEPS in children with liver tumors, because resection could impede future shunt occlusion procedures, and benign masses may regress after shunt occlusion. Here, we review the case of a 9-years-old male with CEPS and hepatic nuclear Factor 1-alpha (HNF-1-alpha) inactivated HA to raise awareness of this condition and review histopathological changes in the liver of CEPS.

The Multiple Object Test as a performance-based tool to assess the decline of ADL function in Parkinson's disease.

INTRODUCTION: As cognitive-driven worsening of activities of the daily living (ADL) in Parkinson's disease (PD) is the core feature of PD dementia (PDD), there is great need for sensitive quantitative assessment. Aim of our study was the evaluation of cognitive-driven worsening of ADL by the performance-based Multiple Object Test (MOT), offering an essential clinical advantage as it is quick and easy to apply in a clinical context even on severely impaired patients. METHODS: 73 PD patients were assessed longitudinally over a period of 37 (6-49) months. According to their neuropsychological profile the sample was divided into two groups: PD patients with (n = 34, PD-CI) and without cognitive impairment (n = 39, PD-noCI). The MOT comprises five routine tasks (e.g. to make coffee) quick and easy to apply. Quantitative (total error number, processing time) and qualitative parameters (error type) were analyzed using non-parametric test statistic (e.g.Wilcoxon signed-rank test, binary logistic regression). RESULTS: Median number of total errors (p = 0.001), processing time (p<0.001), perplexity (p = 0.035), and omission errors (p<0.001) increased significantly from baseline to follow-up in the total sample. Worsening of MOT performance was correlated to cognitive decline in the attention/ executive function and visuo-constructive domain. PD-CI showed an increase in omission errors (p = 0.027) compared to PD-noCI over time. This increase in omission errors between visits was further identified as a risk marker for PDD conversion. CONCLUSION: The MOT, especially frequency of omission errors, is a promising tool to rate PD patients objectively and might help to identify patients with a high risk for having mild cognitive impairment or dementia.

The Multiple Object Test as a Performance Based Tool to Assess Cognitive Driven Activity of Daily Living Function in Parkinson's Disease.

BACKGROUND: There is need for multidimensional quantitative assessment of cognitive driven activities of daily living (ADL) functions in Parkinson's disease (PD). OBJECTIVE: To determine whether there is an ADL profile related to cognitive impairment in PD assessed by the Multiple Object Test (MOT). We assumed MOT performance to be lower in PD patients versus controls and in PD patients with more severe cognitive impairment. METHODS: 50 PD patients with no cognitive impairment (PD-NC), 54 patients with PD-mild cognitive impairment (PD-MCI), 29 with Parkinson's disease dementia (PDD), and 40 healthy controls (HC) were investigated. Besides comprehensive cognitive testing, the MOT, a performance based test consisting of five routine tasks (e.g., preparing a cup of coffee), was applied. Quantitative (total errors and time) and qualitative (error type) MOT parameters were analyzed. RESULTS: Total time and number of MOT errors was increased in PD patients compared to controls (p < 0.001). These parameters also differentiated PDD patients from other cognitive groups (p < 0.05). No control subject had ≥ 4 errors in the MOT, but 30% of PD patients, especially PDD, scored above this cut-off. Omission (p < 0.001) and mislocation (p < 0.03) errors were more prominent in PDD than other cognitive groups. Perplexity errors did not differ between PD-MCI and PDD but between PD-NC and PDD (p = 0.01). MOT parameters discriminating between cognitive groups correlated mainly with lower test performance in psychomotor speed and executive function. CONCLUSION: Performance based testing is promising to identify quantitative and qualitative ADL aspects differentiating between different cognitive groups which might be helpful for an early detection of PDD.