A factor IX variant that functions independently of factor VIII mitigates the hemophilia A phenotype in patient plasma

Strijbis, Viola J.F.; Romano, Lorenzo G R; Cheung, Ka Lei; Eikenboom, Jeroen; Liu, Ying Poi; McCreary, Andrew C.; Leebeek, Frank W.G.; Bos, Mettine H.A.

doi:10.1016/j.jtha.2023.02.019

Cited by 1 publication

(1 citation statement)

References 26 publications

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“…Plasma coagulation factor VIII (FVIII) was measured using a 2-stage modified activated partial thromboplastin time (APTT) clotting assay. 35 Samples were diluted 1:5 in Owren-Koller buffer (Stago, Leiden, The Netherlands) and mixed with equal volume (25 μL) human factor VIII–deficient plasma (STA-ImmunoDef VIII; Stago) and 50 μL APTT reagents (Triniclot Automated APTT; Stago). After a 3-minute incubation at 37°C (STA Start clotting machine; Stago) and recalcification with 50 μL prewarmed 25 mM CaCl 2 (Stago), clotting time was recorded.…”

Section: Methodsmentioning

confidence: 99%

Small interfering RNA–mediated allele-selective silencing of von Willebrand factor in vitro and in vivo

Jongejan,

Schrader Echeverri,

Dirven

et al. 2023

Blood Advances

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An imbalance in von Willebrand factor (VWF) may either lead to bleeding (von Willebrand disease, VWD) or thrombosis. Both disorders have shortcomings in the currently available treatments. VWF itself could be a potential therapeutic target because of its role in both bleeding and thrombosis. Inhibiting VWF gene expression through allele-selective silencing of VWF with small interfering RNAs (siRNAs) could be a personalized approach to specifically inhibit mutant VWF in VWD or to normalize increased VWF levels in thrombotic disorders without complete VWF knockdown. Therefore, we investigated a method to allele-selectively silence the VWF gene in mice as a therapeutic strategy. Fourteen candidate siRNAs targeting murine Vwf of either the C57BL/6J (B6) or the 129S1/SvImJ (129S) strain were tested in vitro in cells expressing B6- and 129S-Vwf on inhibitory effect and allele-selective potential. Together with a non-selective siVwf, two lead candidate siRNAs, siVwf.B6 and siVwf.129S, were further tested in vivo in B6 and 129S mice. Efficient endothelial siRNA delivery was achieved by siRNA encapsulation into 7C1 oligomeric lipid nanoparticles. Treatment with the non-selective siVwf resulted in a dose-dependent inhibition up to 80% of both lung mRNA and plasma VWF protein in both mouse strains. In contrast, the allele-selective siVwf.B6 and siVwf.129S were shown to be effective in and selective solely for their corresponding mouse strain. To conclude, we showed efficient endothelial delivery of siRNAs that are highly effective in allele-selective inhibition of Vwf in mice, which constitute an in vivo proof of principle of allele-selective VWF-silencing as a therapeutic approach.

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Section: Methodsmentioning

confidence: 99%