© 2008 Oxford University Press
This article appears in the following Nucleic Acid Symposium Series issue: Joint Symposium of the 18th International Roundtable on Nucleosides, Nucleotides and Nucleic Acids and the 35th International Symposium on Nucleic Acids Chemistry [View the issue table of contents]
LNA® incorporated siRNAs exhibit lower off-target effects compared to 2'-OMethoxy in Cell Phenotypic Assays and Microarray Analysis
1Ambion Inc., An Applied Biosystems Business, 2130 Woodward Street, Austin, Texas, 78744, USA, 2Cenix BioScience GmbH, Tatzberg 47, 01307 Dresden, Germany
*Corresponding Author. E-mail: nitin.puri{at}appliedbiosystems.com; susan.magdaleno{at}appliedbiosystems.com
Abstract
Despite the promise of short interfering RNAs (siRNA), contending with off-target is a challenge for RNAi users. To alleviate these problems, we have developed locked nucleic acid (LNA®) modified siRNAs and optimized performance using cellular phenotypic assays as well as microarray analysis. During development, we compared LNA® and 2'OMethoxy (2'OMe) chemistries placed strategically throughout the siRNA molecule and found a novel pattern of LNA® placement that greatly improved the specificity of the siRNA and reduced it's toxicity in culture while preserving the potency of the siRNA. The improvements in specificity made by LNA®-modified siRNAs were developed and validated by measuring the phenotypic signatures in a high content cell-based screening assay as well as comparison of the level of differentially expressed genes observed in microarray analysis between modified and unmodified siRNAs. HT screening of a collection of genes demonstrated that the LNA®-modified siRNAs exhibits the best overall rate to elicit the expected phenotype, reduced toxicity and achieved an improved coherence of phenotype compared to 2'OMe-modified or unmodified siRNAs.