The synthesis and in vitro evaluation of short interfering RNAs that contain internal modified alkyl spacers

Abstract

The ability to silence genes effectively at the pre-translational level through the use of short interfering ribonucleic acid (siRNA) has become a widely studied area since the discovery of the RNA interference pathway in 1998. The ability to silence genes at this point in the central dogma of cell biology offers the chance for excellent gene knockdown specificity as the gene silencing tool is tailored for an exact messenger RNA sequence. There are some inherent problems associated with this type of technology which arise from the chemical structure of RNA including poor cell permeability due to the polyanionic backbone or due to the fact that RNA is a natural substrate for nucleases. In an attempt to mitigate these problems there has been considerable focus on chemical modification of siRNAs such as alteration of the ribose ring or backbone. Backbone alterations such as abasic alkyl linkers have been shown to retain, if not improve, gene-silencing capability while providing a means of centrally destabilizing the siRNA duplex, even when occupying the site of the catalytic protein, Argonaute2, whose action was thought to be an essential part of the RNA interference pathway. This study herein reports the synthesis of novel abasic alkyl linkers and an evaluation of their dose-dependent ability to silence genes in vitro while occupying central regions within the sense stand.