1. Cas9 gene therapy for Angelman syndrome traps Ube3a-ATS long non-coding RNA
- Author
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Justin M. Wolter, Baris Oztemiz, Hannah O. Bazick, James L. Krantz, Hanqian Mao, Giulia Fragola, Jeremy M. Simon, Jason L. Stein, and Mark J. Zylka
- Subjects
0301 basic medicine ,Male ,congenital, hereditary, and neonatal diseases and abnormalities ,Ubiquitin-Protein Ligases ,Genetic Vectors ,Biology ,Nervous System ,Article ,03 medical and health sciences ,Mice ,0302 clinical medicine ,Neurodevelopmental disorder ,Angelman syndrome ,Ube3a-ATS ,CRISPR-Associated Protein 9 ,medicine ,UBE3A ,Animals ,Humans ,Gene Silencing ,Allele ,Gene ,Genetics ,Gene Editing ,Multidisciplinary ,RNA ,Genetic Therapy ,Dependovirus ,medicine.disease ,Long non-coding RNA ,Mice, Inbred C57BL ,Disease Models, Animal ,030104 developmental biology ,Phenotype ,Paternal Inheritance ,Female ,RNA, Long Noncoding ,Angelman Syndrome ,CRISPR-Cas Systems ,030217 neurology & neurosurgery ,RNA, Guide, Kinetoplastida - Abstract
Angelman syndrome (AS) is a severe neurodevelopmental disorder caused by a mutation or deletion of the maternally inherited UBE3A allele. In neurons, the paternally inherited UBE3A allele is silenced in cis by a long non-coding RNA called UBE3A-ATS. Here, as part of a systematic screen, we found that Cas9 can be used to activate ('unsilence') paternal Ube3a in cultured mouse and human neurons when targeted to Snord115 genes, which are small nucleolar RNAs that are clustered in the 3′ region of Ube3a-ATS. A short Cas9 variant and guide RNA that target about 75 Snord115 genes were packaged into an adeno-associated virus and administered to a mouse model of AS during the embryonic and early postnatal stages, when the therapeutic benefit of restoring Ube3a is predicted to be greatest1,2. This early treatment unsilenced paternal Ube3a throughout the brain for at least 17 months and rescued anatomical and behavioural phenotypes in AS mice. Genomic integration of the adeno-associated virus vector into Cas9 target sites caused premature termination of Ube3a-ATS at the vector-derived polyA cassette, or when integrated in the reverse orientation, by transcriptional collision with the vector-derived Cas9 transcript. Our study shows that targeted genomic integration of a gene therapy vector can restore the function of paternally inherited UBE3A throughout life, providing a path towards a disease-modifying treatment for a syndromic neurodevelopmental disorder. Genomic integration of an adeno-associated virus vector in a mouse model of Angelman syndrome unsilences paternal Ube3a and rescues anatomical and behavioural phenotypes, suggesting a pathway towards the treatment of this neurodevelopmental disorder.
- Published
- 2019