The sustained expression of Cas9 targeting toxic RNAs reverses disease phenotypes in mouse models of myotonic dystrophy type 1

Ranjan Batra; David A Nelles; Daniela M Roth; Florian Krach; Curtis A Nutter; Takahiro Tadokoro; James D Thomas; Łukasz J Sznajder; Steven M Blue; Haydee L Gutierrez; Patrick Liu; Stefan Aigner; Oleksandr Platoshyn; Atsushi Miyanohara; Martin Marsala; Maurice S Swanson; Gene W Yeo

doi:10.1038/s41551-020-00607-7

The sustained expression of Cas9 targeting toxic RNAs reverses disease phenotypes in mouse models of myotonic dystrophy type 1

Nat Biomed Eng. 2021 Feb;5(2):157-168. doi: 10.1038/s41551-020-00607-7. Epub 2020 Sep 14.

Authors

Ranjan Batra^#^{1

2

3

4}, David A Nelles^#^{1

2

3

4}, Daniela M Roth⁴, Florian Krach^{1

2

3}, Curtis A Nutter⁵, Takahiro Tadokoro⁶, James D Thomas⁵, Łukasz J Sznajder⁵, Steven M Blue^{1

2

3}, Haydee L Gutierrez⁴, Patrick Liu⁴, Stefan Aigner^{1

2

3}, Oleksandr Platoshyn⁷, Atsushi Miyanohara⁷, Martin Marsala⁷, Maurice S Swanson⁵, Gene W Yeo^{8

9

10}

Affiliations

¹ Department of Cellular and Molecular Medicine, University of California San Diego, La Jolla, CA, USA.
² Stem Cell Program, University of California San Diego, La Jolla, CA, USA.
³ Institute for Genomic Medicine, University of California San Diego, La Jolla, CA, USA.
⁴ Locanabio, La Jolla, CA, USA.
⁵ Department of Molecular Genetics and Microbiology, Center for NeuroGenetics and the Genetics Institute, University of Florida, College of Medicine, Gainesville, FL, USA.
⁶ Department of Anesthesiology, School of Medicine, University of California San Diego, La Jolla, CA, USA.
⁷ Neuroregeneration Laboratory, Department of Anesthesiology, University of California San Diego, La Jolla, CA, USA.
⁸ Department of Cellular and Molecular Medicine, University of California San Diego, La Jolla, CA, USA. geneyeo@ucsd.edu.
⁹ Stem Cell Program, University of California San Diego, La Jolla, CA, USA. geneyeo@ucsd.edu.
¹⁰ Institute for Genomic Medicine, University of California San Diego, La Jolla, CA, USA. geneyeo@ucsd.edu.

^# Contributed equally.

Abstract

Myotonic dystrophy type I (DM1) is a multisystemic autosomal-dominant inherited human disorder that is caused by CTG microsatellite repeat expansions (MREs) in the 3' untranslated region of DMPK. Toxic RNAs expressed from such repetitive sequences can be eliminated using CRISPR-mediated RNA targeting, yet evidence of its in vivo efficacy and durability is lacking. Here, using adult and neonatal mouse models of DM1, we show that intramuscular or systemic injections of adeno-associated virus (AAV) vectors encoding nuclease-dead Cas9 and a single-guide RNA targeting CUG repeats results in the expression of the RNA-targeting Cas9 for up to three months, redistribution of the RNA-splicing protein muscleblind-like splicing regulator 1, elimination of foci of toxic RNA, reversal of splicing biomarkers and amelioration of myotonia. The sustained reversal of DM1 phenotypes provides further support that RNA-targeting Cas9 is a viable strategy for treating DM1 and other MRE-associated diseases.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Adenoviridae / physiology
Animals
CRISPR-Associated Protein 9 / metabolism*
CRISPR-Cas Systems*
Disease Models, Animal
Female
Gene Editing / methods*
Genetic Vectors / physiology
Male
Mice, Transgenic
Muscle, Skeletal / metabolism
Myotonic Dystrophy / genetics
Myotonic Dystrophy / metabolism*
Phenotype
RNA / metabolism*

Substances

RNA
CRISPR-Associated Protein 9

Abstract

Publication types

MeSH terms

Substances

Grants and funding