RNA interference-mediated simultaneous silencing of four genes using cross-shaped RNA.

The structural flexibility of RNA interference (RNAi)-triggering nucleic acids suggests that the design of unconventional RNAi trigger structures with novel features is possible. Here, we report a cross-shaped RNA duplex structure, termed quadruple interfering RNA (qiRNA), with multiple target gene silencing activity. qiRNA triggers the simultaneous down-regulation of four cellular target genes via an RNAi mechanism.

Enhanced intracellular delivery and multi-target gene silencing triggered by tripodal RNA structures.

Background: The development of gene interfering RNA (iRNA) molecules such as small interfering RNAs (siRNAs) and antagomirs provides promising therapeutic modalities for targeting specific mRNAs and microRNAs (miRNAs) involved in disease mechanisms. Therapeutic iRNA strategy against cancer or hypermutable viruses prefers targeting multiple genes simultaneously to achieve synergistic inhibition and to prevent resistance.

Methods: In the present study, we report chemically synthesized, multi-target gene interfering RNA structures based upon branched, tripodal interfering RNAs (termed T-tiRNAs).

Branched, tripartite-interfering RNAs silence multiple target genes with long guide strands.

Structural modifications could provide classical small interfering RNA (siRNA) structure with several advantages, including reduced off-target effects and increased silencing activity. Thus, RNA interference (RNAi)-triggering molecules with diverse structural modifications have been investigated by introducing variations on duplex length and overhang structure. However, most of siRNA structural variants are based on the linear duplex structure.

Long double-stranded RNA-mediated RNA interference and immunostimulation: long interfering double-stranded RNA as a potent anticancer therapeutics.

In most applications, small interfering RNAs are designed to execute specific gene silencing via RNA interference (RNAi) without triggering nonspecific responses such as immunostimulation. However, in anticancer therapeutics, immunostimulation combined with specific oncogene silencing could be beneficial, resulting in the synergistic inhibition of cancer cell growth. In this study, we report an immunostimulatory long double-stranded RNA (dsRNA) structure with the ability to trigger RNAi-mediated specific target gene silencing, termed as long interfering dsRNA (liRNA).

Structural diversity repertoire of gene silencing small interfering RNAs.

Since the discovery of double-stranded (ds) RNA-mediated RNA interference (RNAi) phenomenon in Caenorhabditis elegans, specific gene silencing based upon RNAi mechanism has become a novel biomedical tool that has extended our understanding of cell biology and opened the door to an innovative class of therapeutic agents. To silence genes in mammalian cells, short dsRNA referred to as small interfering RNA (siRNA) is used as an RNAi trigger to avoid nonspecific interferon responses induced by long dsRNAs. An early structure-activity relationship study performed in Drosophila melanogaster embryonic extract suggested the existence of strict siRNA structural design rules to achieve optimal gene silencing.

Dual-target gene silencing by using long, synthetic siRNA duplexes without triggering antiviral responses.

Chemically synthesized small interfering RNAs (siRNAs) can specifically knock-down expression of target genes via RNA interference (RNAi) pathway. To date, the length of synthetic siRNA duplex has been strictly maintained less than 30 bp, because an early study suggested that double-stranded RNAs (dsRNAs) longer than 30 bp could not trigger specific gene silencing due to the induction of nonspecific antiviral interferon responses. Contrary to the current belief, here we show that synthetic dsRNA as long as 38 bp can result in specific target gene silencing without nonspecific antiviral responses.

A structure-activity relationship study of siRNAs with structural variations.

Specific knock-down of cellular gene expression using small interfering RNAs (siRNAs) is a powerful gene silencing technique in mammalian cells. Early siRNAs were double stranded, and 19-21bp in length, but several variations in siRNA structure have been introduced to achieve better silencing efficiency. In addition, siRNA modules have been incorporated into higher-order RNA structures to generate multi-functional RNA molecules.