Correcting dominant‐negative von Willebrand disease

2021 ◽  
Vol 19 (1) ◽  
pp. 55-57
Author(s):  
Ellie Karampini ◽  
James S. O’Donnell
Keyword(s):  
Von Willebrand Disease ◽  
Dominant Negative ◽  
Von Willebrand

scholarly journals In vivo modulation of a dominant‐negative variant in mouse models of von Willebrand disease type 2A

2020 ◽  
Vol 19 (1) ◽  
pp. 139-146
Author(s):  
Matteo Campioni ◽  
Paulette Legendre ◽  
Cécile Loubiere ◽  
Barbara Lunghi ◽  
Mirko Pinotti ◽  
...  
Keyword(s):  
Mouse Models ◽  
Disease Type ◽  
Von Willebrand Disease ◽  
Dominant Negative ◽  
Dominant Negative Variant ◽  
Von Willebrand

Type 1 von Willebrand disease mutation Cys1149Arg causes intracellular retention and degradation of heterodimers: a possible general mechanism for dominant mutations of oligomeric proteins

Blood ◽  
2001 ◽  
Vol 98 (10) ◽  
pp. 2973-2979 ◽  
Author(s):  
Imre Bodó ◽  
Akira Katsumi ◽  
Elodee A. Tuley ◽  
Jeroen C. J. Eikenboom ◽  
Zhengyu Dong ◽  
...  
Keyword(s):  
Full Range ◽  
Disease Type ◽  
Von Willebrand Disease ◽  
Dominant Negative ◽  
General Mechanism ◽  
Oligomeric Proteins ◽  
Dominant Phenotype ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Some families affected by von Willebrand disease type 1 show high penetrance with exceptionally low von Willebrand factor (VWF) levels. Previously, a mutation associated with this dominant phenotype, Cys1149Arg, was found to decrease the secretion of coexpressed normal VWF, and the mutation was proposed to cause intracellular retention of pro-VWF heterodimers. To demonstrate heterodimer formation, a model was developed in which subunits could be distinguished immunologically and by size. Recombinant VWF lacking domain A1 (dA1), A3 (dA3), or both (dA13) was secreted efficiently as a full range of multimers. Cotransfection of Cys1149Arg and dA13 resulted in the secretion of multimeric VWF containing about 250 kd (Cys1149Arg) and about 210 kd (dA13). Cell lysates contained pro-VWF forms of Cys1149Arg and dA13. Immunoprecipitation with an antidomain A1 antibody recovered both subunits in heterodimers, and subunit ratios were consistent with random dimerization. Similar results were obtained for cotransfection of Cys1149Arg and dA1. Normal VWF has a Cys1149-Cys1169 intrachain bond. When cotransfected with normal VWF, Cys1149Arg or the double mutant Cys1149Arg+Cys1169Ser caused a similar decrease in VWF secretion, suggesting that an unpaired Cys1169 does not explain the intracellular retention of Cys1149Arg. VWF Cys1149Arg was not secreted from BHK cells but was degraded intracellularly within about 4 hours, and the proteasome inhibitor lactacystin delayed its clearance more than 16 hours. Thus, dominant von Willebrand disease type 1 may be caused by heterodimerization of mutant and normal subunits in the endoplasmic reticulum followed by proteasomal degradation in the cytoplasm. A similar dominant negative mechanism could cause quantitative deficiencies of other multisubunit proteins.

scholarly journals Ex vivo Improvement of a von Willebrand Disease Type 2A Phenotype Using an Allele-Specific Small-Interfering RNA

2020 ◽  
Vol 120 (11) ◽  
pp. 1569-1579 ◽  
Author(s):  
Annika de Jong ◽  
Richard J. Dirven ◽  
Johan Boender ◽  
Ferdows Atiq ◽  
Seyed Yahya Anvar ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Coagulation Factor ◽  
Endothelial Activation ◽  
Von Willebrand Disease ◽  
Dominant Negative ◽  
Nucleotide Polymorphisms ◽  
Small Interfering Rnas ◽  
Collagen Binding ◽  
Allele Specific ◽  
Von Willebrand

AbstractVon Willebrand disease (VWD) is the most common inherited bleeding disorder and is mainly caused by dominant-negative mutations in the multimeric protein von Willebrand factor (VWF). These mutations may either result in quantitative or qualitative defects in VWF. VWF is an endothelial protein that is secreted to the circulation upon endothelial activation. Once secreted, VWF multimers bind platelets and chaperone coagulation factor VIII in the circulation. Treatment of VWD focuses on increasing VWF plasma levels, but production and secretion of mutant VWF remain uninterrupted. Presence of circulating mutant VWF might, however, still affect normal hemostasis or functionalities of VWF beyond hemostasis. We hypothesized that inhibition of the production of mutant VWF improves the function of VWF overall and ameliorates VWD phenotypes. We previously proposed the use of allele-specific small-interfering RNAs (siRNAs) that target frequent VWF single nucleotide polymorphisms to inhibit mutant VWF. The aim of this study is to prove the functionality of these allele-specific siRNAs in endothelial colony-forming cells (ECFCs). We isolated ECFCs from a VWD type 2A patient with an intracellular multimerization defect, reduced VWF collagen binding, and a defective processing of proVWF to VWF. After transfection of an allele-specific siRNA that specifically inhibited expression of mutant VWF, we showed amelioration of the laboratory phenotype, with normalization of the VWF collagen binding, improvement in VWF multimers, and enhanced VWF processing. Altogether, we prove that allele-specific inhibition of the production of mutant VWF by siRNAs is a promising therapeutic strategy to improve VWD phenotypes.

scholarly journals Multifaceted Pathomolecular Mechanism of a VWF Large Deletion Involved in the Pathogenesis of Severe VWD

2021 ◽  
Author(s):  
Hamideh Yadegari ◽  
Muhammad Ahmer Jamil ◽  
Jens Müller ◽  
Natascha Marquardt ◽  
Orla Rawley ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Signaling Pathways ◽  
Large Deletion ◽  
Von Willebrand Disease ◽  
Dominant Negative ◽  
Cell Junctions ◽  
Nuclear Accumulation ◽  
Severe Bleeding ◽  
Rna Seq ◽  
Von Willebrand

An in-frame heterozygous large deletion of exons 4-34 of the von Willebrand factor (VWF) gene was identified in a type 3 von Willebrand disease (VWD) index patient (IP), as the only VWF variant. The IP exhibited severe bleeding episodes despite prophylaxis treatment, with a short VWF half-life after infusion of VWF/FVIII concentrates. Transcript analysis confirmed transcription of normal VWF mRNA besides an aberrant deleted transcript. The IP endothelial colony-forming cells (ECFCs) exhibited a defect in the VWF multimers and Weibel-Palade bodies (WPBs) biogenesis, although demonstrating normal VWF secretion, as compared with healthy cells. Immunostaining of IP-ECFCs revealed subcellular mislocalization of WPBs pro-inflammatory cargos angiopoietin-2 (Ang2, nuclear accumulation) and P-selectin. Besides, the RNA-sequencing (RNA-seq) analysis showed upregulation of pro-inflammatory and proangiogenic genes, P-selectin, IL8, IL6, and GROα, co-packaged with VWF into WPBs. Further, whole-transcriptome RNA-seq and subsequent Gene Ontology (GO) enrichment analysis indicated the most enriched GO-Biological Process terms among the differentially expressed genes in IP-ECFCs were regulation of cell differentiation, cell adhesion, leukocyte adhesion to vascular endothelial, blood vessel morphogenesis and angiogenesis, which resemble downstream signaling pathways associated with inflammatory stimuli and Ang2 priming. Accordingly, our functional experiments exhibited an increased endothelial cell adhesiveness and interruption in endothelial cell-cell junctions of the IP-ECFCs. In conclusion, the deleted VWF has a dominant-negative impact on multimer assembly and the biogenesis of WPBs, leading to altered trafficking of their pro-inflammatory cargos uniquely, which, in turn, causes changes in cellular signaling pathways, phenotype, and function of the endothelial cells.

The dominant-negative von Willebrand factor gene deletion p.P1127_C1948delinsR: molecular mechanism and modulation

Blood ◽  
2010 ◽  
Vol 116 (24) ◽  
pp. 5371-5376 ◽  
Author(s):  
Caterina Casari ◽  
Mirko Pinotti ◽  
Stefano Lancellotti ◽  
Elena Adinolfi ◽  
Alessandra Casonato ◽  
...  
Keyword(s):  
Von Willebrand Factor ◽  
Molecular Mechanisms ◽  
Von Willebrand Disease ◽  
Dominant Negative ◽  
Dominant Negative Effect ◽  
Wild Type ◽  
Collagen Binding ◽  
Cellular Models ◽  
Von Willebrand ◽  
Willebrand Factor

AbstractUnderstanding molecular mechanisms in the dominant inheritance of von Willebrand disease would improve our knowledge of pathophysiologic processes underlying its prevalence. Cellular models of severe type 2 von Willebrand disease, caused by a heterozygous deletion in the von Willebrand factor (VWF) gene, were produced to investigate the altered biosynthesis. Coexpression of the wild-type and in-frame deleted (p.P1127_C1948delinsR) VWF forms impaired protein secretion, high molecular weight multimer formation and function (VWF collagen-binding 1.9% ± 0.5% of wild-type), which mimicked the patient's phenotype. mRNA, protein, and cellular studies delineated the highly efficient dominant-negative mechanism, based on the key role of heterodimers as multimer terminators. The altered VWF, synthesized in large amounts with the correctly encoded “cysteine knot” domain, formed heterodimers and heterotetramers with wild-type VWF, in addition to deleted homodimers. Impaired multimerization was associated with reduced amounts of VWF in late endosomes. Correction of the dominant-negative effect was explored by siRNAs targeting the mRNA breakpoint, which selectively inhibited the in-frame deleted VWF expression. Although the small amount of the deleted protein synthesized after inhibition still exerted dominant, even though weakened, negative effects, the siRNA treatment restored secretion of large multimers with improved function (VWF collagen-binding 28.0% ± 3.3% of wild-type).

Morbidity and mortality of patients with haemophilia in Germany 2007/2008

2009 ◽  
Vol 29 (S 01) ◽  
pp. S7-S12
Author(s):  
M. Spannagl ◽  
W. Schramm ◽  
H. Krebs ◽  
Keyword(s):  
Causes Of Death ◽  
Severity Of Illness ◽  
Von Willebrand Disease ◽  
Morbidity And Mortality ◽  
Haemophilia A ◽  
Hiv Status ◽  
Von Willebrand ◽  
Existing Data

SummarySince 1978 an annual multicentric survey regarding the epidemiology of patients suffering of haemophilia is performed with support of haemophilia treating centres of any size. Again the actual compilation is resting upon a broad database returning to over 30 years of inquiry well representing both the actual and retrospective status of mortality. Prompted was exclusively information about patients with haemophilia A, B and von Willebrand disease. In particular anonymous data concerning the last 12 months about number of treated patients, type and severity of illness, HIV-status and detailed information about causes of death was inquired. This data was merged with existing data and analyzed statistically. In the 2007/2008 survey, a total

Platelet Activation and Aggregation Induced by Recombinant von Willebrand Factors Reproducing Four Type 2B von Willebrand Disease Missense Mutations

1998 ◽  
Vol 79 (01) ◽  
pp. 211-216 ◽  
Author(s):  
Lysiane Hilbert ◽  
Claudine Mazurier ◽  
Christophe de Romeuf
Keyword(s):  
Platelet Aggregation ◽  
Platelet Activation ◽  
Von Willebrand Disease ◽  
Platelet Glycoprotein ◽  
Missense Mutations ◽  
Inhibit Platelet Aggregation ◽  
Wild Type ◽  
A1 Domain ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryType 2B of von Willebrand disease (vWD) refers to qualitative variants with increased affinity of von Willebrand factor (vWF) for platelet glycoprotein Ib (GPIb). All the mutations responsible for type 2B vWD have been located in the A1 domain of vWF. In this study, various recombinant von Willebrand factors (rvWF) reproducing four type 2B vWD missense mutations were compared to wild-type rvWF (WT-rvWF) for their spontaneous binding to platelets and their capacity to induce platelet activation and aggregation. Our data show that the multimeric pattern of each mutated rvWF is similar to that of WT-rvWF but the extent of spontaneous binding and the capacity to induce platelet activation and aggregation are more important for the R543Q and V553M mutations than for the L697V and A698V mutations. Both the binding of mutated rvWFs to platelets and platelet aggregation induced by type 2B rvWFs are inhibited by monoclonal anti-GPIb and anti-vWF antibodies, inhibitors of vWF binding to platelets in the presence of ristocetin, as well as by aurin tricarboxylic acid. On the other hand, EDTA and a monoclonal antibody directed against GPIIb/IIIa only inhibit platelet aggregation. Furthermore, the incubation of type 2B rvWFs with platelets, under stirring conditions, results in the decrease in high molecular weight vWF multimers in solution, the extent of which appears correlated with that of plasma vWF from type 2B vWD patients harboring the corresponding missense mutation. This study supports that the binding of different mutated type 2B vWFs onto platelet GPIb induces various degrees of platelet activation and aggregation and thus suggests that the phenotypic heterogeneity of type 2B vWD may be related to the nature and/or location of the causative point mutation.

A New (K1518E) Candidate Mutation Detected by Universal Heteroduplex Generator Analysis in a Patient with Type 2A (Phenotype IIA) von Willebrand Disease

1998 ◽  
Vol 79 (01) ◽  
pp. 240-240 ◽  
Author(s):  
Bimal Theophilus ◽  
Frank Hill ◽  
Peter Rose ◽  
Doris Culpan ◽  
Jeffery Bidwell ◽  
...  
Keyword(s):  
Von Willebrand Disease ◽  
Candidate Mutation ◽  
Von Willebrand
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