In Vivo CRISPR/Cas9 Gene Editing Corrects Retinal Dystrophy in the S334ter-3 Rat Model of Autosomal Dominant Retinitis Pigmentosa

Bakondi, Benjamin; Lv, Wenjian; Lü, Bin; Jones, Melissa K.; Tsai, Yu‐Lin; Kim, Kevin J.; Levy, Rachelle; Akhtar, Aslam Abbasi; Breunig, Joshua J.; Svendsen, Clive N.; Wang, Shaomei

doi:10.1038/mt.2015.220

Cited by 256 publications

(199 citation statements)

References 25 publications

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“…Recently, SpCas9 has been proven to be a powerful tool for making precise genomic perturbations in vivo. It has been used in vivo to study the function of genes in the liver and brain [42,43] and to cure metabolic and genetic diseases in animal models [44]. Here, we demonstrated that SpCas9 can be used to disrupt the genes in adult mouse islets.…”

Section: Discussionmentioning

confidence: 78%

RNA-binding protein CUGBP1 regulates insulin secretion via activation of phosphodiesterase 3B in mice

Zhai

Yang

et al. 2016

Diabetologia

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Aims/hypothesis CUG-binding protein 1 (CUGBP1) is a multifunctional RNA-binding protein that regulates RNA processing at several stages including translation, deadenylation and alternative splicing, as well as RNA stability. Recent studies indicate that CUGBP1 may play a role in metabolic disorders. Our objective was to examine its role in endocrine pancreas function through gain-and loss-of-function experiments and to further decipher the underlying molecular mechanisms. Methods A mouse model in which type 2 diabetes was induced by a high-fat diet (HFD; 60% energy from fat) and mice on a standard chow diet (10% energy from fat) were compared. Pancreas-specific CUGBP1 overexpression and knockdown mice were generated. Different lengths of the phosphodiesterase subtype 3B (PDE3B) 3′ untranslated region (UTR) were cloned for luciferase reporter analysis. Purified CUGBP1 protein was used for gel shift experiments.Results CUGBP1 is present in rodent islets and in beta cell lines; it is overexpressed in the islets of diabetic mice. Compared with control mice, the plasma insulin level after a glucose load was significantly lower and glucose clearance was greatly delayed in mice with pancreas-specific CUGBP1 overexpression; the opposite results were obtained upon pancreas-specific CUGBP1 knockdown. Glucose-and glucagon-like peptide1 (GLP-1)-stimulated insulin secretion was significantly attenuated in mouse islets upon CUGBP1 overexpression. This was associated with a strong decrease in intracellular cAMP levels, pointing to a potential role for cAMP PDEs. CUGBP1 overexpression had no effect on the mRNA levels of PDE1A, 1C, 2A, 3A, 4A, 4B, 4D, 7A and 8B subtypes, but resulted in increased PDE3B expression. CUGBP1 was found to directly bind to a specific ATTTGTT sequence residing in the 3′ UTR of PDE3B and stabilised PDE3B mRNA. In the presence of the PDE3 inhibitor cilostamide, glucose-and GLP-1-stimulated insulin secretion was no longer reduced by CUGBP1 overexpression. Similar to CUGBP1, PDE3B was overexpressed in the islets of diabetic mice. Conclusions/interpretation We conclude that CUGBP1 is a critical regulator of insulin secretion via activating PDE3B. Repressing this protein might provide a potential strategy for treating type 2 diabetes.

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Section: Discussionmentioning

confidence: 78%

RNA-binding protein CUGBP1 regulates insulin secretion via activation of phosphodiesterase 3B in mice

Zhai

Yang

et al. 2016

Diabetologia

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“…Partial phenotypic recovery has been observed in these studies Nelson et al 2016;Tabebordbar et al 2016). The use of CRISPR in vivo to ablate the rhodopsin gene carrying the dominant S334ter mutation in rats with severe autosomal dominant retinitis pigmentosa also highlights the use of genetic correction in disease (Bakondi et al 2016). These proof-of-principle experiments may be the first steps towards overcoming many currently incurable genetic diseases.…”

Section: Gene Editing In Vivo To Treat Genetic Diseasesmentioning

confidence: 80%

Using Genome Engineering to Understand Huntington’s Disease

Bailus

Zhang

Ellerby

2017

Research and Perspectives in Neurosciences

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Huntington's disease (HD) is a fatal, dominantly inherited neurodegenerative disorder caused by a CAG trinucleotide expansion in the Huntingtin (HTT) gene, leading to an expanded polyglutamine (polyQ) region in the encoded protein HTT. We have used homologous recombination (HR) to genetically correct HD patient-derived induced pluripotent stem cells (iPSCs) and found that this reversed HD disease phenotypes. We have utilized exploited genome editing tools including TALENs (Transcription like activator effectors) and CRISPR (Clustered Regulatory Interspaced Short Palindromic Repeats)/Cas9 technology to carry out genetic correction or expansion, and we were able to detect HR without selection in human cells. The overall goal is to use this technology to model HD-relevant cell types and better understand disease progression by leveraging system biology approaches. To understand the disease progression, isogenic iPSC lines were created. We found that the disease phenotypes only manifested in the differentiated neural stem cell (NSC) stage, not in iPSCs. Transcriptomic analysis of HD iPSCs and HD NSCs compared to isogenic controls was utilized to understand the molecular basis for the CAG repeat expansion-dependent disease phenotypes in NSCs. Differential gene expression and pathway analysis identified transforming growth factor β (TGF-β) signaling, netrin-1 signaling and medium spiny neuron (MSNs) maturation and maintenance as the top dysregulated pathways in HD NSCs. The ability to create additional isogenic cell lines through CRISPR-mediated HR will further enhance our understanding of HD progression. These lines can be manipulated with CRISPR to understand the effects of common SNPs (single nucleotide polymorphism) that modulate disease onset in HD, allowing the identification of new pathways and helping to elucidate potential therapeutic targets for HD. Beyond drug discovery, the CRISPR system could eventually be optimized to use in vivo, correcting a patient's disease-causing mutation, in the asymptomatic stages of HD.

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“…This has not been tried in human patients yet; however, in trials with rats, disruption of the dominant-negative allele of rhodopsin (S334Ter) by inducing such “knockout” mutations specifically in the mutant allele have restored normal function to photoreceptors. 29 The NHEJ repair function after cutting results in the disruption of the mutant allele, such that the misfolded rhodopsin protein does not accumulate in photoreceptors and lead to death of the cells.…”

Section: A New Gene Editing Toolmentioning

confidence: 99%

Clustered Regularly Interspaced Short Palindromic Repeats

Chrenek

Nickerson

Boatright³

2016

Asia-Pacific Journal of Ophthalmology

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Ophthalmic researchers and clinicians arguably have led the way for safe, effective gene therapy, most notably with adeno-associated viral gene supplementation in the treatment of Leber congenital amaurosis type 2 (LCA 2) patients with mutations in the RPE65 gene. These successes notwithstanding, most other genetic retinal disease will be refractory to supplementation. The ideal gene therapy approach would correct gene mutations to restore normal function in the affected cells. Gene editing in which a mutant allele is inactivated or converted to sequence that restores normal function is hypothetically one such approach. Such editing involves site-specific digestion of mutant genomic DNA followed by repair. Previous experimental approaches were hampered by inaccurate and high rates of off-site lesioning and by overall low digestion rates. A new tool, clustered regularly interspaced short palindromic repeats coupled with the nuclease Cas9 (CRISPR/Cas9), may address both shortcomings. Some of the many challenges that must be addressed in moving CRISPR/Cas9 therapies to the ophthalmic clinic are discussed here.

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In Vivo CRISPR/Cas9 Gene Editing Corrects Retinal Dystrophy in the S334ter-3 Rat Model of Autosomal Dominant Retinitis Pigmentosa

Cited by 256 publications

References 25 publications

RNA-binding protein CUGBP1 regulates insulin secretion via activation of phosphodiesterase 3B in mice

RNA-binding protein CUGBP1 regulates insulin secretion via activation of phosphodiesterase 3B in mice

Using Genome Engineering to Understand Huntington’s Disease

Clustered Regularly Interspaced Short Palindromic Repeats

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