Genome Therapy of Myotonic Dystrophy Type 1 iPS Cells for Development of Autologous Stem Cell Therapy

Gao, Yuanzheng; Guo, Xiufang; Santostefano, Katherine E.; Wang, Yanlin; Reid, Tammy; Zeng, Desmond; Terada, Naohiro; Ashizawa, Tetsuo; Xia, Guangbin

doi:10.1038/mt.2016.97

Cited by 53 publications

(65 citation statements)

References 47 publications

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“…iPSC‐derived cell models have previously been used to study other diseases in a variety of different cell types, including vascular smooth muscle cells, cardiomyocytes, and neurons . iPSC‐derived macrophages have previously been shown to be a useful model for HIV‐1 infection and we have confirmed those findings in the current study.…”

Section: Discussionsupporting

confidence: 85%

CRISPR/Cas9 knockout of USP18 enhances type I IFN responsiveness and restricts HIV-1 infection in macrophages

Taylor

Cash

Santostefano

et al. 2018

Journal of Leukocyte Biology

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The IFN-stimulated gene ubiquitin-specific proteinase 18 (USP18) encodes a protein that negatively regulates T1 IFN signaling via stearic inhibition of JAK1 recruitment to the IFN-α receptor 2 subunit (IFNAR2). Here, we demonstrate that USP18 expression is induced by HIV-1 in a T1 IFN-dependent manner. Experimental depletion of USP18 by clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) gene editing results in a significant restriction of HIV-1 replication in an induced pluripotent stem cell (iPSC)-derived macrophage model. In the absence of USP18, macrophages have increased responsiveness to stimulation with T1 IFNs with prolonged phosphorylation of STAT1 and STAT2 and increased expression of IFN-stimulated genes that are key for antiviral responses. Interestingly, HIV-1 requires some signaling through the T1 IFN receptor to replicate efficiently because a neutralizing antibody that inhibits T1 IFN activity reduces HIV-1 replication rate in monocyte-derived macrophages. USP18 induction by HIV-1 tunes the IFN response to optimal levels allowing for efficient transcription from the HIV-1 LTR promoter while minimizing the T1 IFN-induced antiviral response that would otherwise restrict viral replication and spread. Finally, iPSC and CRISPR/Cas9 gene targeting offer a powerful tool to study host factors that regulate innate immune responses.

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Section: Discussionsupporting

confidence: 85%

CRISPR/Cas9 knockout of USP18 enhances type I IFN responsiveness and restricts HIV-1 infection in macrophages

Taylor

Cash

Santostefano

et al. 2018

Journal of Leukocyte Biology

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“…We next assessed the ability of RCas9 to eliminate endogenous MRE RNA in myoblasts and fibroblasts prepared from DM1 and DM2 patient biopsies (Gao et al, 2016; Xia et al, 2013), respectively. For purposes of ensuring homogeneity of expression, we constructed lentiviral vectors based upon the LentiCRISPRV2 construct (Sanjana et al, 2014), which express U6 promoter-driven sgRNAs targeting CUG exp or CCUG exp RNAs along with elongation factor-1 short (EFS)-driven PIN-dCas9 and a puromycin resistance gene that enabled cell selection.…”

Section: Resultsmentioning

confidence: 99%

Elimination of Toxic Microsatellite Repeat Expansion RNA by RNA-Targeting Cas9

Batra

Nelles

Pirie

et al. 2017

Cell

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SUMMARY Microsatellite repeat expansions in DNA produce pathogenic RNA species that cause dominantly inherited diseases such as myotonic dystrophy type 1 and 2 (DM1/2), Huntington’s disease, and C9orf72-linked amyotrophic lateral sclerosis (C9-ALS). Means to target these repetitive RNAs are required for diagnostic and therapeutic purposes. Here, we describe the development of a programmable CRISPR system capable of specifically visualizing and eliminating these toxic RNAs. We observe specific targeting and efficient elimination of micro-satellite repeat expansion RNAs both when exogenously expressed and in patient cells. Importantly, RNA-targeting Cas9 (RCas9) reverses hallmark features of disease including elimination of RNA foci among all conditions studied (DM1, DM2, C9-ALS, polyglutamine diseases), reduction of polyglutamine protein products, relocalization of repeat-bound proteins to resemble healthy controls, and efficient reversal of DM1-associated splicing abnormalities in patient myotubes. Finally, we report a truncated RCas9 system compatible with adeno-associated viral packaging. This effort highlights the potential of RCas9 for human therapeutics.

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“…Yet another study was able to use the CRISPR/Cas9 system to eliminate the repeat expansions in human cells using a smaller version that was optimized for possible adeno-associated virus packaging [145]. Although many studies concentrate on genome modification of terminally differentiated somatic cells, some have examined the use of genome editing of induced pluripotent stem cells that could be used for developing autologous stem cell therapy [146,147]. Placement of polyA signals upstream of the CTG expansion using transcription activator-like effector nucleases, a different genome editing tool than CRISPR/Cas9, was able to stop the production of the mutant transcripts.…”

Section: Genome Editingmentioning

confidence: 99%

Myotonic Dystrophies: Targeting Therapies for Multisystem Disease

2018

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Myotonic dystrophy is an autosomal dominant muscular dystrophy not only associated with muscle weakness, atrophy, and myotonia but also prominent multisystem involvement. There are 2 similar, but distinct, forms of myotonic dystrophy; type 1 is caused by a CTG repeat expansion in the DMPK gene, and type 2 is caused by a CCTG repeat expansion in the CNBP gene. Type 1 is associated with distal limb, neck flexor, and bulbar weakness and results in different phenotypic subtypes with variable onset from congenital to very late-onset as well as variable signs and symptoms. The classically described adult-onset form is the most common. In contrast, myotonic dystrophy type 2 is adult-onset or late-onset, has proximal predominant muscle weakness, and generally has less severe multisystem involvement. In both forms of myotonic dystrophy, the best characterized disease mechanism is a RNA toxic gain-of-function during which RNA repeats form nuclear foci resulting in sequestration of RNA-binding proteins and, therefore, dysregulated splicing of premessenger RNA. There are currently no disease-modifying therapies, but clinical surveillance, preventative measures, and supportive treatments are used to reduce the impact of muscular impairment and other systemic involvement including cataracts, cardiac conduction abnormalities, fatigue, central nervous system dysfunction, respiratory weakness, dysphagia, and endocrine dysfunction. Exciting preclinical progress has been made in identifying a number of potential strategies including genome editing, small molecule therapeutics, and antisense oligonucleotide-based therapies to target the pathogenesis of type 1 and type 2 myotonic dystrophies at the DNA, RNA, or downstream target level.

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Genome Therapy of Myotonic Dystrophy Type 1 iPS Cells for Development of Autologous Stem Cell Therapy

Cited by 53 publications

References 47 publications

CRISPR/Cas9 knockout of USP18 enhances type I IFN responsiveness and restricts HIV-1 infection in macrophages

CRISPR/Cas9 knockout of USP18 enhances type I IFN responsiveness and restricts HIV-1 infection in macrophages

Elimination of Toxic Microsatellite Repeat Expansion RNA by RNA-Targeting Cas9

Myotonic Dystrophies: Targeting Therapies for Multisystem Disease

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