Details

Autor(en) / Beteiligte

• Bramsen, Jesper B
• Pakula, Malgorzata M
• Hansen, Thomas B
• Bus, Claus
• Langkjær, Niels
• Odadzic, Dalibor
• Smicius, Romualdas
• Wengel, Suzy L
• Chattopadhyaya, Jyoti
• Engels, Joachim W
• Herdewijn, Piet
• Wengel, Jesper
• Kjems, Jørgen

Titel

A screen of chemical modifications identifies position-specific modification by UNA to most potently reduce siRNA off-target effects

Ist Teil von

• Nucleic acids research, 2010-09, Vol.38 (17), p.5761

Ort / Verlag

England

Erscheinungsjahr

2010

Quelle

MEDLINE

Beschreibungen/Notizen

• Small interfering RNAs (siRNAs) are now established as the preferred tool to inhibit gene function in mammalian cells yet trigger unintended gene silencing due to their inherent miRNA-like behavior. Such off-target effects are primarily mediated by the sequence-specific interaction between the siRNA seed regions (position 2-8 of either siRNA strand counting from the 5'-end) and complementary sequences in the 3'UTR of (off-) targets. It was previously shown that chemical modification of siRNAs can reduce off-targeting but only very few modifications have been tested leaving more to be identified. Here we developed a luciferase reporter-based assay suitable to monitor siRNA off-targeting in a high throughput manner using stable cell lines. We investigated the impact of chemically modifying single nucleotide positions within the siRNA seed on siRNA function and off-targeting using 10 different types of chemical modifications, three different target sequences and three siRNA concentrations. We found several differently modified siRNAs to exercise reduced off-targeting yet incorporation of the strongly destabilizing unlocked nucleic acid (UNA) modification into position 7 of the siRNA most potently reduced off-targeting for all tested sequences. Notably, such position-specific destabilization of siRNA-target interactions did not significantly reduce siRNA potency and is therefore well suited for future siRNA designs especially for applications in vivo where siRNA concentrations, expectedly, will be low.