CC BY-NC-ND 4.0 · Thromb Haemost 2020; 120(11): 1569-1579
DOI: 10.1055/s-0040-1715442
New Technologies, Diagnostic Tools and Drugs

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

1  Department of Internal Medicine, Division of Thrombosis and Hemostasis, Einthoven laboratory for Vascular and Regenerative Medicine, Leiden University Medical Center, Leiden, the Netherlands
,
1  Department of Internal Medicine, Division of Thrombosis and Hemostasis, Einthoven laboratory for Vascular and Regenerative Medicine, Leiden University Medical Center, Leiden, the Netherlands
,
Johan Boender
2  Department of Hematology, Erasmus University Medical Center, Rotterdam, The Netherlands
,
2  Department of Hematology, Erasmus University Medical Center, Rotterdam, The Netherlands
,
Seyed Yahya Anvar
3  Department of Human Genetics, Leiden University Medical Center, Leiden, The Netherlands
,
Frank W. G. Leebeek
2  Department of Hematology, Erasmus University Medical Center, Rotterdam, The Netherlands
,
Bart J. M. van Vlijmen
1  Department of Internal Medicine, Division of Thrombosis and Hemostasis, Einthoven laboratory for Vascular and Regenerative Medicine, Leiden University Medical Center, Leiden, the Netherlands
,
1  Department of Internal Medicine, Division of Thrombosis and Hemostasis, Einthoven laboratory for Vascular and Regenerative Medicine, Leiden University Medical Center, Leiden, the Netherlands
› Author Affiliations
Funding This study was financially supported by a research grant from the Landsteiner Foundation for Blood Transfusion Research (grant 1504).
  

Abstract

Von 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.

Authors’ Contributions

A.d.J. designed the study, performed the experiments and data analyses, and wrote the manuscript. R.J.D. performed the experiments. J.B., F.A., and F.W.G.L. were involved in the control and patient inclusions. S.Y.A. contributed to the analysis of PacBio sequencing data. B.J.M.v.V. contributed to discussions and reviewing of the manuscript. J.E. designed the study, interpreted the data, and contributed to writing of the manuscript. All authors revised and approved the final version of the manuscript.


Supplementary Material



Publication History

Received: 14 April 2020

Accepted: 25 June 2020

Publication Date:
15 August 2020 (online)

© 2020. The Author(s). This is an open access article published by Thieme under the terms of the Creative Commons Attribution-NonDerivative-NonCommercial-License, permitting copying and reproduction so long as the original work is given appropriate credit. Contents may not be used for commercial purposes, or adapted, remixed, transformed or built upon. (https://creativecommons.org/licenses/by-nc-nd/4.0/).

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