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

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 effec
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.
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Author:Jesper B. Bramsen, Malgorzata M. Pakula, Thomas B. Hansen, Claus Bus, Niels Langkjær, Dalibor Odadžić, Romualdas Smicius, Suzy L. Wengel, Jyoti Chattopadhyaya, Joachim W. Engels, Piet Herdewijn, Jesper Wengel, Jørgen Kjems
URN:urn:nbn:de:hebis:30-79873
DOI:http://dx.doi.org/10.1093/nar/gkq341
ISSN:1362-4962
ISSN:0305-1048
Pubmed Id:http://www.ncbi.nlm.nih.gov/pubmed?term=20453030
Parent Title (English):Nucleic acids research
Publisher:Oxford Univ. Press
Place of publication:Oxford
Document Type:Article
Language:English
Date of Publication (online):2010/05/07
Date of first Publication:2010/05/07
Publishing Institution:Univ.-Bibliothek Frankfurt am Main
Release Date:2010/09/17
Volume:38
Issue:17
Pagenumber:13
First Page:5761
Last Page:5773
Note:
© The Author(s) 2010. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.5), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
HeBIS PPN:22845039X
Institutes:Biochemie und Chemie
Dewey Decimal Classification:540 Chemie und zugeordnete Wissenschaften
Sammlungen:Universitätspublikationen
Licence (German):License LogoCreative Commons - Namensnennung-Nicht kommerziell 3.0

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