Gymnotic delivery and gene silencing activity of reduction-responsive siRNAs bearing lipophilic disulfide-containing modifications at 2′-position

Modified oligoribonucleotides used as siRNAs bearing biolabile disulfide-containing groups at some 2'-positions were synthesized following a post-synthesis transformation of solid-supported 2'-O-acetylthiomethyl RNA, previously described. Thus, the reduction-responsive and lipophilic benzyldithiomethyl (BnSSM) modification was introduced at different locations into siRNAs targeting the Ewing sarcoma EWS-Flil protein. Thermal stability, serum stability and response to glutathione treatment of modified siRNAs were thoroughly investigated. Among 17 modified siRNAs, significant gene silencing activities were demonstrated for the 8 most stable siRNAs in serum (half-life > 1 h) when using a transfection reagent. Of special interest, two naked 2'-O-BnSSM siRNAs transfection exhibited a remarkable gene silencing activity after 24 h incubation. These inhibitions are consistent with an efficient gymnotic delivery demonstrated by the presence of the corresponding fluorescent siRNAs within cells.