Effect of North Bicyclo[3.1.0]hexane 2′-Deoxy-pseudosugars on RNA Interference: A Novel Class of siRNA Modification

North bicyclo methanocarba thymidine (T-N) nucleosides were substituted into siRNAs to investigate the effect of bicyclo-[3.1.0]hexane 2'-deoxy-pseudosugars on RNA interference activity. Here we provide evidence that these modified siRNAs are compatible with the intracellular RNAi machinery. We studied the effect of the T-N modification in a screen involving residue- specific changes in an siRNA targeting Renilla luciferase and we applied the most effective pattern of modification to the knockdown of murine tumor necrosis factor (TNF-alpha). We also showed that incorporation of T-N units into siRNA duplexes increased their thermal stabilities, substantially enhanced serum stabilities, and decreased innate immunostimulation. Comparative RNAi studies involving the T-N substitution and locked nucleic acids (LNAs) showed that the gene-silencing activities of T-N-modified siRNAs were comparable to those obtained with the LNA modification. An advantage of the North 2'-deoxy-methanocarba modification is that it may be explored further in the future by changing the 2'-position. The results from these studies suggest that this modification might be valuable for the development of siRNAs for therapeutic applications.