Titrating haemophilia B phenotypes using siRNA strategy: evidence that antithrombotic activity is separated from bleeding liability

Joseph M. Metzger; Marija Tadin-Strapps; Anil Thankappan; Walter R. Strapps; Marti DiPietro; Karen Leander; Zuo Zhang; Myung K. Shin; John Levorse; Kunal Desai; Yiming Xu; KehDih Lai; Weizhen Wu; Zhu Chen; Tian-Quan Cai; Nina Jochnowitz; Ross Bentley; Lizbeth Hoos; Yuchen Zhou; Laura Sepp-Lorenzino; Dietmar Seiffert; Patrick Andre

doi:10.1160/TH14-06-0505

Thrombosis and Haemostasis, Inhaltsverzeichnis

Thromb Haemost 2015; 113(06): 1300-1311
DOI: 10.1160/TH14-06-0505

Coagulation and Fibrinolysis

Schattauer GmbH

Titrating haemophilia B phenotypes using siRNA strategy: evidence that antithrombotic activity is separated from bleeding liability

Joseph M. Metzger^*

¹In vivo Pharmacology, Merck Co., Inc., Kenilworth, New Jersey, USA

,

Marija Tadin-Strapps^*

²RNA Therapeutics, Merck Co., Inc., West Point, Pennsylvania, USA

,

Anil Thankappan

²RNA Therapeutics, Merck Co., Inc., West Point, Pennsylvania, USA

,

Walter R. Strapps

²RNA Therapeutics, Merck Co., Inc., West Point, Pennsylvania, USA

,

Marti DiPietro

²RNA Therapeutics, Merck Co., Inc., West Point, Pennsylvania, USA

,

Karen Leander

²RNA Therapeutics, Merck Co., Inc., West Point, Pennsylvania, USA

,

Zuo Zhang

³Genetically Engineered Models, Kenilworth, New Jersey, USA

,

Myung K. Shin

³Genetically Engineered Models, Kenilworth, New Jersey, USA

,

John Levorse

⁴Cardiometabolic Disease Merck Co., Inc., Kenilworth, New Jersey, USA

,

Kunal Desai

⁴Cardiometabolic Disease Merck Co., Inc., Kenilworth, New Jersey, USA

,

Yiming Xu

⁴Cardiometabolic Disease Merck Co., Inc., Kenilworth, New Jersey, USA

,

KehDih Lai

⁴Cardiometabolic Disease Merck Co., Inc., Kenilworth, New Jersey, USA

,

Weizhen Wu

⁴Cardiometabolic Disease Merck Co., Inc., Kenilworth, New Jersey, USA

,

Zhu Chen

⁴Cardiometabolic Disease Merck Co., Inc., Kenilworth, New Jersey, USA

,

Tian-Quan Cai

¹In vivo Pharmacology, Merck Co., Inc., Kenilworth, New Jersey, USA

,

Nina Jochnowitz

¹In vivo Pharmacology, Merck Co., Inc., Kenilworth, New Jersey, USA

,

Ross Bentley

¹In vivo Pharmacology, Merck Co., Inc., Kenilworth, New Jersey, USA

,

Lizbeth Hoos

¹In vivo Pharmacology, Merck Co., Inc., Kenilworth, New Jersey, USA

,

Yuchen Zhou

¹In vivo Pharmacology, Merck Co., Inc., Kenilworth, New Jersey, USA

,

Laura Sepp-Lorenzino

²RNA Therapeutics, Merck Co., Inc., West Point, Pennsylvania, USA

,

Dietmar Seiffert

²RNA Therapeutics, Merck Co., Inc., West Point, Pennsylvania, USA

,

Patrick Andre

⁴Cardiometabolic Disease Merck Co., Inc., Kenilworth, New Jersey, USA

› Institutsangaben

Abstract

Summary

Haemophilia A and B are characterised by a life-long bleeding predisposition, and several lines of evidence suggest that risks of atherothrombotic events may also be reduced. Establishing a direct correlation between coagulation factor levels, thrombotic risks and bleeding propensity has long been hampered by an inability to selectively and specifically inhibit coagulation factor levels. Here, the exquisite selectivity of gene silencing combined with a gene knockout (KO) approach was used to define the relative contribution of factor IX (fIX) to thrombosis and primary haemostasis in the rat. Using a lipid nanoparticle (LNP) formulation, we successfully delivered fIX siRNAs to the liver by intravenous administration. The knockdown (KD) of target gene mRNA was achieved rapidly (within 24 hour post-siRNA dosing), sustained (maintained for at least 7 days post dosing) and not associated with changes in mRNA expression levels of other coagulation factors. We found that intermediate levels of liver fIX mRNA silencing (60–95 %) translating into a 50–99 % reduction of plasma fIX activity provided protection from thrombosis without prolonging the cuticle bleeding time. Over 99 % inhibition of fIX activity was required to observe increase in bleeding, a phenotype confirmed in fIX KO rats. These data provide substantial evidence of a participation of fIX in the mechanisms regulating thrombosis prior to those regulating primary haemostasis, therefore highlighting the potential of fIX as a therapeutic target. In addition, hepatic mRNA silencing using LNP-encapsulated siRNAs may represent a promising novel approach for the chronic treatment and prevention of coagulation-dependent thrombotic disorders in humans.

Keywords

siRNA - coagulation - factor fIX - thrombosis - haemostasis

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