title: Local RNA target structure influences siRNA efficacy: a systematic global analysis.
abstract: The efficiency with which small interfering RNAs (siRNAs) down-regulate specific gene expression in living cells is variable and a number of sequence-governed, biochemical parameters of the siRNA duplex have been proposed for the design of an efficient siRNA. Some of these parameters have been clearly identified to influence the assembly of the RNA-induced silencing complex (RISC), or to favour the sequence preferences of the RISC endonuclease. For other parameters, it is difficult to ascertain whether the influence is a determinant of the siRNA per se, or a determinant of the target RNA, especially its local structural characteristics. In order to gain an insight into the effects of local target structure on the biological activity of siRNA, we have used large sets of siRNAs directed against local targets of the mRNAs of ICAM-1 and survivin. Target structures were classified as accessible or inaccessible using an original, iterative computational approach and by experimental RNase H mapping. The effectiveness of siRNA was characterized by measuring the IC50 values in cell culture and the maximal extent of target suppression. Mean IC50 values were tenfold lower for accessible local target sites, with respect to inaccessible ones. Mean maximal target suppression was improved. These data illustrate that local target structure does, indeed, influence the activity of siRNA. We suggest that local target screening can significantly improve the hit rate in the design of biologically active siRNAs.
authors.affiliation: Universität zu Lübeck, Institut für Molekulare Medizin, Ratzeburger Allee 160, D-23538 Lübeck, Germany.
authors.forename: Marita
authors.initials: M
authors.lastname: Overhoff
authors.forename: Martina
authors.initials: M
authors.lastname: Alken
authors.forename: Rosel Kretschmer-Kazemi
authors.initials: RK
authors.lastname: Far
authors.forename: Marc
authors.initials: M
authors.lastname: Lemaitre
authors.forename: Bernard
authors.initials: B
authors.lastname: Lebleu
authors.forename: Georg
authors.initials: G
authors.lastname: Sczakiel
authors.forename: Ian
authors.initials: I
authors.lastname: Robbins
mesh_terms: D002460:Cell Line; D019295:Computational Biology; D006801:Humans; D020128:Inhibitory Concentration 50; D018799:Intercellular Adhesion Molecule-1 / Q000235:genetics; D008869:Microtubule-Associated Proteins / Q000235:genetics; D009690:Nucleic Acid Conformation; D034622:RNA Interference; D012333:RNA, Messenger / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism*; D016914:Ribonuclease H / Q000378:metabolism; D013379:Substrate Specificity; D014162:Transfection
chemical_list: D008869:Microtubule-Associated Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D018799:Intercellular Adhesion Molecule-1; D016914:Ribonuclease H