An RNA-targeting CRISPR-Cas13d system alleviates disease-related phenotypes in Huntington's disease models
Kathryn H Morelli1, Qian Wu2,3, Maya L Gosztyla1
1Department of Cellular and Molecular Medicine, University of California San Diego, La Jolla, CA, USA.
Insights
This study introduces a CRISPR-Cas13d system targeting toxic RNA in Huntington's disease (HD). The system effectively reduced mutant huntingtin (HTT) RNA and protein, improving motor function in a mouse model.
Area of Science:
- Neuroscience
- Genetics
- Biotechnology
Background:
- Huntington's disease (HD) is a fatal, inherited neurodegenerative disorder.
- It is caused by a CAG trinucleotide expansion in the huntingtin (HTT) gene.
- Reducing mutant HTT messenger RNA is a potential therapeutic strategy.
Purpose of the Study:
- To develop and evaluate a mutant allele-specific RNA-targeting CRISPR-Cas13d system for Huntington's disease.
- To assess the therapeutic efficacy of this system in cellular and animal models of HD.
Main Methods:
- Developed a CRISPR-Cas13d system (Cas13d-CAG EX) targeting expanded CAG repeats in HTT RNA.
- Tested the system in patient-derived fibroblasts and iPSC-derived neurons.
- Administered Cas13d-CAG EX via adeno-associated virus vector in zQ175 mice (HD model).
Main Results:
- Selective reduction of mutant HTT mRNA and protein in the striatum of HD mice.
- Significant improvements in motor coordination and attenuation of striatal atrophy.
- Sustained therapeutic effects for at least eight months with minimal off-target effects.
Conclusions:
- Demonstrated proof of principle for an RNA-targeting CRISPR-Cas13d system as a therapeutic approach for HD.
- This strategy holds promise for treating other dominantly inherited disorders caused by similar genetic expansions.
Abstract:
Huntington's disease (HD) is a fatal, dominantly inherited neurodegenerative disorder caused by CAG trinucleotide expansion in exon 1 of the huntingtin (HTT) gene. Since the reduction of pathogenic mutant HTT messenger RNA is therapeutic, we developed a mutant allele-sensitive CAGEX RNA-targeting CRISPR-Cas13d system (Cas13d-CAGEX) that eliminates toxic CAGEX RNA in fibroblasts derived from patients with HD and induced pluripotent stem cell-derived neurons. We show that intrastriatal delivery of Cas13d-CAGEX via an adeno-associated viral vector selectively reduces mutant HTT mRNA and protein levels in the striatum of heterozygous zQ175 mice, a model of HD. This also led to improved motor coordination, attenuated striatal atrophy and reduction of mutant HTT protein aggregates. These phenotypic improvements lasted for at least eight months without adverse effects and with minimal off-target transcriptomic effects. Taken together, we demonstrate proof of principle of an RNA-targeting CRISPR-Cas13d system as a therapeutic approach for HD, a strategy with implications for the treatment of other dominantly inherited disorders.
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