title: Gene Silencing Using 4'-thioDNA as an Artificial Template to Synthesize Short Hairpin RNA Without Inducing a Detectable Innate Immune Response.
abstract: The development of a versatile technique to induce RNA interference (RNAi) without immune stimulation in vivo is of interest as existing approaches to trigger RNAi, such as small interfering RNA (siRNA) and plasmid DNA (pDNA) expressing short hairpin RNA (shRNA), present drawbacks arising from innate immune stimulation. To overcome them, an intelligent shRNA expression device (iRed) designed to induce RNAi was developed. The minimum sequence of iRed encodes only the U6 promoter and shRNA. A series of iRed comprises a polymerase chain reaction (PCR)-amplified 4'-thioDNA in which any one type of adenine (A), guanine (G), cytosine (C), or thymine (T) nucleotide unit was substituted by each cognate 4'-thio derivatives, i.e., dSA iRed, dSG iRed, dSC iRed, and ST iRed respectively. Each modified iRed acted as a template to transcribe shRNA with RNAi activity. The highest shRNA yield was generated using dSC iRed that exerted gene silencing activity in an orthotopic mouse model of mesothelioma. Reducing the minimal structure required to transcribe shRNA and the presence of the 4'-thiomodification synergistically function to abrogate innate immune response induced by dsDNA. The iRed will introduce a new approach to induce RNAi without inducing a detectable innate immune response.
authors.affiliation: Graduate School of Pharmaceutical Sciences, Tokushima University, Tokushima, Japan.
authors.forename: Noriko
authors.initials: N
authors.lastname: Tarashima
authors.affiliation: Graduate School of Pharmaceutical Sciences, Tokushima University, Tokushima, Japan.
authors.forename: Hidenori
authors.initials: H
authors.lastname: Ando
authors.affiliation: Graduate School of Pharmaceutical Sciences, Tokushima University, Tokushima, Japan.
authors.forename: Takamitsu
authors.initials: T
authors.lastname: Kojima
authors.affiliation: Graduate School of Pharmaceutical Sciences, Tokushima University, Tokushima, Japan.
authors.forename: Nozomi
authors.initials: N
authors.lastname: Kinjo
authors.affiliation: Graduate School of Pharmaceutical Sciences, Tokushima University, Tokushima, Japan.
authors.forename: Yosuke
authors.initials: Y
authors.lastname: Hashimoto
authors.affiliation: Graduate School of Pharmaceutical Sciences, Tokushima University, Tokushima, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Furukawa
authors.affiliation: Graduate School of Pharmaceutical Sciences, Tokushima University, Tokushima, Japan.
authors.forename: Tatsuhiro
authors.initials: T
authors.lastname: Ishida
authors.affiliation: Graduate School of Pharmaceutical Sciences, Tokushima University, Tokushima, Japan.
authors.forename: Noriaki
authors.initials: N
authors.lastname: Minakawa