structure-guided chemical modification of guide RNA enables potent non-viral in vivo genome editing

Hao Yin, Chun Qing Song, Sneha Suresh, Qiongqiong Wu, Stephen Walsh, Luke Hyunsik Rhym, Esther Mintzer, Mehmet Fatih Bolukbasi, Lihua Julie Zhu, Kevin Kauffman, Haiwei Mou, Alicia Oberholzer, Junmei Ding, Suet Yan Kwan, Roman L. Bogorad, Timofei Zatsepin, Victor Koteliansky, Scot A. Wolfe, Wen Xue, Robert LangerDaniel G. Anderson

    Research output: Contribution to journalArticlepeer-review

    241 Citations (Scopus)

    Abstract

    Efficient genome editing with Cas9-sgRNA in vivo has required the use of viral delivery systems, which have limitations for clinical applications. Translational efforts to develop other RNA therapeutics have shown that judicious chemical modification of RNAs can improve therapeutic efficacy by reducing susceptibility to nuclease degradation. Guided by the structure of the Cas9-sgRNA complex, we identify regions of sgRNA that can be modified while maintaining or enhancing genome-editing activity, and we develop an optimal set of chemical modifications for in vivo applications. Using lipid nanoparticle formulations of these enhanced sgRNAs (e-sgRNA) and mRNA encoding Cas9, we show that a single intravenous injection into mice induces >80% editing of Pcsk9 in the liver. Serum Pcsk9 is reduced to undetectable levels, and cholesterol levels are significantly lowered about 35% to 40% in animals. This strategy may enable non-viral, Cas9-based genome editing in the liver in clinical settings.

    Original languageEnglish
    Pages (from-to)1179-1187
    Number of pages9
    JournalNature Biotechnology
    Volume35
    Issue number12
    DOIs
    Publication statusPublished - 2017

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