RNAi has rapidly become a powerful tool for
drug target discovery and validation in cell culture, and now has largely displaced efforts with antisense and
ribozymes. Consequently, interest is rapidly growing for extension of its application to in vivo systems, such as
animal disease models and human
therapeutics. Studies on RNAi have resulted in two basic methods for its use for gene selective inhibition: 1) cytoplasmic delivery of short dsRNA
oligonucleotides (
siRNA), which mimics an active intermediate of an endogenous RNAi mechanism and 2) nuclear delivery of gene expression cassettes that express a
short hairpin RNA (
shRNA), which mimics the micro interfering
RNA (
miRNA) active intermediate of a different endogenous RNAi mechanism. Non-viral gene delivery systems are a diverse collection of technologies that are applicable to both of these forms of RNAi. Importantly, unlike antisense and
ribozyme systems, a remarkable trait of
siRNA is a lack of dependence on chemical modifications blocking enzymatic degradation, although chemical protection methods developed for the earlier systems are being incorporated into
siRNA and are generally compatible with non-viral delivery systems. The use of
siRNA is emerging more rapidly than for
shRNA, in part due to the increased effort required to construct
shRNA expression systems before selection of active sequences and verification of
biological activity are obtained. In contrast, screens of many
siRNA sequences can be accomplished rapidly using synthetic oligos. It is not surprising that the use of
siRNA in vivo is also emerging first. Initial in vivo studies have been reported for both viral and non-viral delivery but viral delivery is limited to
shRNA. This review describes the emerging in vivo application of non-viral delivery systems for RNAi for functional genomics, which will provide a foundation for further development of
RNAi therapeutics. Of interest is the rapid adaptation of
ligand-targeted plasmid-based nanoparticles for RNAi agents. These systems are growing in capabilities and beginning to pose a serious rival to viral vector based gene delivery. The activity of
siRNA in the cytoplasm may lower the hurdle and thereby accelerate the successful development of
therapeutics based on targeted non-viral delivery systems.