TransKingdom RNA interference: a bacterial approach to challenges in RNAi therapy and delivery

Abstract

Since its discovery in 1998 RNA interference (RNAi), a potent and highly selective gene silencing mechanism, has revolutionized the field of biological science. The ability of RNAi to specifically down-regulate the expression of any cellular protein has had a profound impact on the study of gene function in vitro. This property of RNAi also holds great promise for in vivo functional genomics and interventions against a wide spectrum of diseases, especially those with "undruggable" therapeutic targets. Despite the enormous potential of RNAi for medicine, development of in vivo applications has met with significant problems, particularly in terms of delivery. For effective gene silencing to occur, silencing RNA must reach the cytoplasm of the target cell. Consequently, various strategies using chemically modified siRNA, liposomes, nanoparticles and viral vectors are being developed to deliver silencing RNA. These approaches, however, can be expensive and in many cases they lack target cell specificity or clinical compatibility. Recently, we have shown that RNAi can be activated in vitro and in vivo by non-pathogenic bacteria engineered to manufacture Biotechnology and Genetic Engineering Reviews -Vol. 25, 113-128 (2008) * To whom correspondence may be addressed (cli@bidmc.harvard.edu) Abbreviations: APC, adenatomous polyposis coli; dsRNA, double-stranded RNA; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; GFP, green fluorescent protein; mRNA, messenger RNA; PKR, protein kinase R; RISC, RNA-induced silencing complex; RNA, ribonucleic acid; RNAi, RNA interference; miRNA, micro RNA; MOI, multiplicity of infection; RIG-I, retinoic acid inducible gene I; shRNA, short hairpin RNA; siRNA, short interfering RNA; SNALP, stabilized nucleic acid lipid particle; tkRNAi, TransKingdom RNA interference; TLR, toll-like receptor; TRIP, TransKingdom RNA interference plasmid.

ASalmonellaTyphimurium mutant strain capable of RNAi delivery

“…In contrast to viral vectors, VNP(PhoP/Q − ) does not integrate the genetic material into the host genome and can be controlled by antibiotics. 39,40 Moreover, facultative anaerobic S. Typhimurium strains can preferentially accumulate and replicate in the hypoxic regions of solid tumors. 12,41,42 This property provides the pre-conditions for VNP(PhoP/Q − ) to deliver the fragment of RNAi to mediate antitumor effects.…”

Bacterium-Mediated RNA Interference: Potential Application in Plant Protection

“…Bacterium-mediated RNA interference (bmRNAi) refers to a delivery method wherein live bacteria expressing dsRNA are applied to and colonize an organism to produce and facilitate the uptake of dsRNA in situ, resulting in RNAi knockdown of the targeted gene [78,79]. This method has been shown to be effective in both mammalian cell culture and animal subjects and shows the potential to overcome the weaknesses of more-traditional endogenous and exogenous application strategies by increasing the duration and strength of the silencing effect, while reducing both the time and effort required to produce knockdowns.…”

“…The bacterium is then applied to the host, where it secretes, or deposits dsRNA to be taken up via endocytosis, membrane transport or secretion into the cytosol subsequently delivering the RNA payload. This method is enhanced by the deletion of rnc, the gene responsible for bacterial RNase III, failure to do so results in dramatic reductions in the dsRNA yield, due to degradation into non-active forms [78,79,92,93].…”

Gene therapy using bacterial vectors

“…The advantages of bacteria-mediated gene therapy and RNA interference are combined and can be specifically applied in some conditions such as colitis or colorectal cancer. In addition, the production of small interfering RNA can be done by bacterial and host cells and does not have to be done artificially before administration (57). This makes the treatment cheaper and the pharmacokinetics easier, while targeting of treatment depends on the bacterial strain chosen for the therapy and on the treated condition.…”