Nat Biotechnol. 2016 Mar;34(3):328-33. doi: 10.1038/nbt.3471. Epub 2016 Feb 1.

Therapeutic genome editing by combined viral and non-viral delivery of CRISPR system components in vivo.

Yin H¹, Song CQ^2,³, Dorkin JR^1,⁴, Zhu LJ^3,^5,⁶, Li Y⁷, Wu Q¹, Park A², Yang J¹, Suresh S¹, Bizhanova A², Gupta A⁵, Bolukbasi MF^5,⁸, Walsh S¹, Bogorad RL¹, Gao G⁹, Weng Z⁶, Dong Y¹⁰, Koteliansky V^11,¹², Wolfe SA^5,⁸, Langer R^1,^13,^14,¹⁵, Xue W^2,^3,⁵, Anderson DG^1,^13,^14,¹⁵.

Author information

1: David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
2: RNA Therapeutics Institute, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
3: Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
4: Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
5: Department of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
6: Program in Bioinformatics and Integrative Biology, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
7: Department of Bioinformatics, School of Life Science and Technology, Tongji University, Shanghai, P.R. China.
8: Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
9: Gene Therapy Center, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
10: College of Pharmacy, the Ohio State University, Columbus, Ohio, USA.
11: Skolkovo Institute of Science and Technology, Skolkovo, Russia.
12: Department of Chemistry, M.V. Lomonosov Moscow State University, Leninskie Gory, Russia.
13: Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
14: Harvard-MIT Division of Health Sciences &Technology, Cambridge, Massachusetts, USA.
15: Institute of Medical Engineering and Science, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.

Abstract

The combination of Cas9, guide RNA and repair template DNA can induce precise gene editing and the correction of genetic diseases in adult mammals. However, clinical implementation of this technology requires safe and effective delivery of all of these components into the nuclei of the target tissue. Here, we combine lipid nanoparticle-mediated delivery of Cas9 mRNA with adeno-associated viruses encoding a sgRNA and a repair template to induce repair of a disease gene in adult animals. We applied our delivery strategy to a mouse model of human hereditary tyrosinemia and show that the treatment generated fumarylacetoacetate hydrolase (Fah)-positive hepatocytes by correcting the causative Fah-splicing mutation. Treatment rescued disease symptoms such as weight loss and liver damage. The efficiency of correction was >6% of hepatocytes after a single application, suggesting potential utility of Cas9-based therapeutic genome editing for a range of diseases.