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Small interfering RNAs (siRNAs) are potential candidates for gene regulation with efficient activity, but off-target effects and limited systemic delivery. Herein, we report the design and synthesis of the branched siRNA nanostructures with highly improved resistance against exonucleases. Also, these branched siRNAs showed suppression of off-target gene silencing through selection of the passenger strand as the branching unit. The physical characterization of branched siRNAs showed that they form a compact assembly with a hydrodynamic diameter of 6.9 nm against 2.8 nm of the duplex. We demonstrated that a branched siRNA synthesized with a trebling solid-support selectively exhibits RNAi activity and suppresses the off-target effect.

Design, synthesis and evaluation of novel, branched trident small interfering RNA nanostructures for sequence-specific RNAi activity