RNA-Targeting CRISPR/Cas13d System Alleviates Disease-Related Phenotypes in Pre-clinical Models of Huntington’s Disease
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ABSTRACT:
ORGANISM(S): Mus musculus Homo sapiens
PROVIDER: GSE214110 | GEO | 2022/11/15
REPOSITORIES: GEO
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Project description:Huntington's disease (HD) is a fatal, dominantly inherited neurodegenerative disorder caused by a CAG trinucleotide expansion in exon 1 of the huntingtin (HTT) gene. Although the pathogenesis of HD remains complex, the CAG-expanded (CAGEX) HTT mRNA and protein ultimately causes disease through a toxic gain-of-function mechanism. As the reduction of pathogenic mutant HTT mRNA is beneficial as a treatment, we developed a mutant allele-sensitive CAGEX RNA-targeting CRISPR-Cas13d system (Cas13d/CAGEX) that eliminates toxic CAGEX RNA in HD patient-derived fibroblasts and iPSC-derived striatal neurons. We show that intrastriatal delivery of Cas13d/CAGEX via a single adeno-associated viral vector, serotype 9 (AAV9) mediates significant and selective reduction of mutant HTT mRNA and protein levels within the striatum of heterozygous zQ175 mice, an established mouse model of HD. Moreover, the reduction of mutant HTT mRNA renders a sustained partial reversal of HD phenotypes, including improved motor coordination, attenuated striatal atrophy, and reduction of mutant HTT protein aggregates. Importantly, phenotypic improvements were durable for at least 8 months without gross or behavioral adverse effects, and with minimal off-target interactions of Cas13d/CAGEX in the mouse transcriptome. Taken together, we demonstrate a proof-of-principle of an RNA-targeting CRISPR/Cas13d system as a therapeutic approach for HD, a strategy with broad implications for the treatment of other dominantly inherited neurodegenerative disorders.
2022-11-15 | GSE214108 | GEO
Project description:Huntington's disease (HD) is a fatal, dominantly inherited neurodegenerative disorder caused by a CAG trinucleotide expansion in exon 1 of the huntingtin (HTT) gene. Although the pathogenesis of HD remains complex, the CAG-expanded (CAGEX) HTT mRNA and protein ultimately causes disease through a toxic gain-of-function mechanism. As the reduction of pathogenic mutant HTT mRNA is beneficial as a treatment, we developed a mutant allele-sensitive CAGEX RNA-targeting CRISPR-Cas13d system (Cas13d/CAGEX) that eliminates toxic CAGEX RNA in HD patient-derived fibroblasts and iPSC-derived striatal neurons. We show that intrastriatal delivery of Cas13d/CAGEX via a single adeno-associated viral vector, serotype 9 (AAV9) mediates significant and selective reduction of mutant HTT mRNA and protein levels within the striatum of heterozygous zQ175 mice, an established mouse model of HD. Moreover, the reduction of mutant HTT mRNA renders a sustained partial reversal of HD phenotypes, including improved motor coordination, attenuated striatal atrophy, and reduction of mutant HTT protein aggregates. Importantly, phenotypic improvements were durable for at least 8 months without gross or behavioral adverse effects, and with minimal off-target interactions of Cas13d/CAGEX in the mouse transcriptome. Taken together, we demonstrate a proof-of-principle of an RNA-targeting CRISPR/Cas13d system as a therapeutic approach for HD, a strategy with broad implications for the treatment of other dominantly inherited neurodegenerative disorders.
2022-11-15 | GSE214109 | GEO
Project description: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.
| S-EPMC9829537 | biostudies-literature
Project description:RNA-Targeting CRISPR/Cas13d System Alleviates Disease-Related Phenotypes in Pre-clinical Models of Huntington’s Disease
| PRJNA883961 | ENA
Project description:RNA-Targeting CRISPR/Cas13d System Alleviates Disease-Related Phenotypes in Pre-clinical Models of Huntington’s Disease (Human).
| PRJNA883962 | ENA
Project description:RNA-Targeting CRISPR/Cas13d System Alleviates Disease-Related Phenotypes in Pre-clinical Models of Huntington’s Disease (Mouse).
| PRJNA883965 | ENA
Project description:Recently discovered Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas13 proteins are programmable RNA-guided ribonucleases that target single-stranded RNA (ssRNA). CRISPR/Cas13-mediated RNA targeting has emerged as a powerful tool for detecting and eliminating RNA viruses. Here, we demonstrate the effectiveness of CRISPR/Cas13d to inhibit HIV-1 replication. We designed guide RNAs (gRNAs) targeting highly conserved regions of HIV-1. RfxCas13d (CasRx) in combination with HIV-specific gRNAs efficiently inhibited HIV-1 replication in cell line models. Furthermore, simultaneous targeting of four distinct, non-overlapping sites in the HIV-1 transcript resulted in robust inhibition of HIV-1 replication. We also show the effective HIV-1 inhibition in primary CD4+ T-cells and suppression of HIV-1 reactivated from latently infected cells using the CRISPR/Cas13d system. Our study demonstrates the utility of the CRISPR/Cas13d nuclease system to target acute and latent HIV infection and provides an alternative treatment modality against HIV.
| S-EPMC8473377 | biostudies-literature
Project description:In the second edition of the Huntington’s Disease Clinical Trials Corner we list all currently registered and ongoing clinical trials, summarise the top-line results of the recently-announced IONIS-HTTRX trial (NCT02519036), expand on Wave Life Sciences’ PRECISION-HD1 (NCT03225833) and PRECISION-HD2 (NCT03225846), and cover one recently finished trial: the FIRST-HD deutetrabenazine trial (NCT01795859).
| S-EPMC5836398 | biostudies-literature
Project description:RNA-guided and RNA-targeting type IV-D CRISPR/Cas systems (CRISPR/Cas13d) have recently been identified and employed for efficient and specific RNA knockdown in mammalian and plant cells. Cas13d possesses dual RNase activities and is capable of processing CRISPR arrays and cleaving target RNAs in a protospacer flanking sequence (PFS)-independent manner. These properties make this system a promising tool for multiplex gene expression regulation in microbes. Herein, we aimed to establish a CRISPR/Cas13d-mediated RNA knockdown platform for bacterial chassis. CasRx, Cas13d from Ruminococcus flavefaciens XPD3002, was selected due to its high activity. However, CasRx was found to be highly toxic to both Escherichia coli and Corynebacterium glutamicum, especially when it cooperated with its guide and target RNAs. After employing a low copy number vector, a tightly controlled promoter, and a weakened ribosome binding site, we successfully constructed an inducible expression system for CasRx and applied it for repressing the expression of a green fluorescent protein (GFP) in E. coli. Despite our efforts to optimize inducer usage, guide RNA (gRNA) architecture and combination, and target gene expression level, the highest gene repression efficiency was 30-50% at the protein level and ∼70% at the mRNA level. The moderate RNA knockdown is possibly caused by the collateral cleavage activity toward bystander RNAs, which acts as a mechanism of type IV-D immunity and perturbs microbial metabolism. Further studies on cellular response to CRISPR/Cas13d and improvement in RNA knockdown efficiency are required prior to practical application of this system in microbes.
| S-EPMC7406568 | biostudies-literature
Project description:To dissect the impact of nuclear and extranuclear mutant htt on the initiation and progression of disease, we generated a series of transgenic mouse lines in which nuclear localization (NLS) or nuclear export sequences (NES) have been placed N-terminal to the htt exon 1 protein carrying 144 glutamines. Our data indicate that the exon 1 mutant protein is present in the nucleus as part of an oligomeric or aggregation complex. Increasing the concentration of the mutant transprotein in the nucleus is sufficient for, and dramatically accelerates the onset and progression of behavioral phenotypes. Furthermore, nuclear exon 1 mutant protein is sufficient to induce cytoplasmic neurodegeneration and transcriptional dysregulation. However, our data suggests that cytoplasmic mutant exon 1 htt, if present, contributes to disease progression. Keywords: Trangenic mouse brain gene expression
2005-09-07 | GSE3248 | GEO