Structure-guided chemical modification of guide RNA enables potent non-viral in vivo genome editing
RNA Therapeutics Institute; Department of Molecular, Cell and Cancer Biology; Department of Biochemistry and Molecular Pharmacology; Program in Molecular Medicine
Biochemistry, Biophysics, and Structural Biology | Cell and Developmental Biology | Genetics and Genomics | Therapeutics
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.
DOI of Published Version
Nat Biotechnol. 2017 Dec;35(12):1179-1187. doi: 10.1038/nbt.4005. Epub 2017 Nov 13. Link to article on publisher's site
Yin, Hao; Song, Chun-Qing; Mintzer, Esther; Bolukbasi, Mehmet F.; Zhu, Lihua Julie; Mou, Haiwei; Kwan, Suet-Yan; Wolfe, Scot A.; Xue, Wen; and Anderson, Daniel G., "Structure-guided chemical modification of guide RNA enables potent non-viral in vivo genome editing" (2017). RNA Therapeutics Institute Publications. 6.