Effective and Accurate Gene Silencing by a Recombinant AAV-Compatible MicroRNA Scaffold

Short hairpin RNAs that are delivered by recombinant adeno-associated virus (rAAV) have the potential to elicit long-term RNAi therapy for human disease. However, the discovery that short hairpin sequences can cause truncation of the rAAV genome calls into question the efficiency and gene-silencing specificity of this strategy in humans. Here, we report that embedding the guide strand of a small silencing RNA into an artificial microRNA (miRNA) scaffold derived from mouse miRNA-33 ensures rAAV genomic integrity and reduces off-targeting by 10-fold, while maintaining effective in vivo target gene repression in mice. [Display omitted] Efficient and specific gene silencing is a challenge for rAAV-mediated RNAi. Short hairpin-induced vector genomic truncations raised concerns for clinical transition. Xie and colleagues report that an artificial miRNA scaffold derived from mouse miR-33 ensures rAAV genomic integrity and reduces off-targeting 10-fold, while maintaining effective target gene repression in vivo in mice.