Phosphorothioate backbone modifications of nucleotide-based drugs are potent platelet activators

• DOI: 10.1084/jem.20140391
• Publication Date: 2015-02-03T06:01:14Z
• Authors: Ulrike Flierl; Tracy L. Nero; Bock Lim; Jane F. Arthur; Yu Yao; Stephanie M. Jung; Eelo Gitz; Alice Y. Pollitt; Maria T.K. Zaldivia; Martine Jandrot-P...; Andreas Schäfer; Bernhard Nieswandt; Robert K. Andrews; Michael W. Parker; Elizabeth E. Gard...; Karlheinz Peter
• Keywords: antisense oligonucleotide; collagen; oligonucleotide; Blood Platelets; Male; Models, Molecular; 570; Keywords: 4 amino 7 tert butyl 5 (4 chlorophenyl)pyrazolo[3,4 d]pyrimidine; Platelet Aggregation; integrin; Molecular Conformation; 610; Phosphorothioate Oligonucleotides; Platelet Membrane Glycoproteins; glycoprotein VI; Mice; 03 medical and health sciences; Platelet Adhesiveness; Mesenteric Vascular Occlusion; Animals; Humans; 0303 health sciences; C reactive protein; Brief Definitive Report; 500; nucleotide; Oligonucleotides, Antisense; Platelet Activation; Mesenteric Arteries; 3. Good health; Disease Models, Animal; oligodeoxynucleotide phosphorothioate; reactive oxygen meta; RNA Interference; Pulmonary Embolism; Reactive Oxygen Species; Fc receptor IIa; Protein Binding

Nucleotide-based drug candidates such as antisense oligonucleotides, aptamers, immunoreceptor-activating nucleotides, or (anti)microRNAs hold great therapeutic promise for many human diseases. Phosphorothioate (PS) backbone modification of nucleotide-based drugs is common practice to protect these promising drug candidates from rapid degradation by plasma and intracellular nucleases. Effects of the changes in physicochemical properties associated with PS modification on platelets have not been elucidated so far. Here we report the unexpected binding of PS-modified oligonucleotides to platelets eliciting strong platelet activation, signaling, reactive oxygen species generation, adhesion, spreading, aggregation, and thrombus formation in vitro and in vivo. Mechanistically, the platelet-specific receptor glycoprotein VI (GPVI) mediates these platelet-activating effects. Notably, platelets from GPVI function–deficient patients do not exhibit binding of PS-modified oligonucleotides, and platelet activation is fully abolished. Our data demonstrate a novel, unexpected, PS backbone–dependent, platelet-activating effect of nucleotide-based drug candidates mediated by GPVI. This unforeseen effect should be considered in the ongoing development programs for the broad range of upcoming and promising DNA/RNA therapeutics.