scholarly journals Cardiac Myoediting Attenuates Cardiac Abnormalities in Human and Mouse Models of Duchenne Muscular Dystrophy

2021 ◽  
Author(s):  
Ayhan Atmanli ◽  
Andreas C Chai ◽  
Miao Cui ◽  
Zhaoning Wang ◽  
Takahiko Nishiyama ◽  
...  
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Dilated Cardiomyopathy ◽  
Genome Editing ◽  
Rna Sequencing ◽  
Reading Frame ◽  
Transcriptional Dysregulation ◽  
Cardiac Abnormalities ◽  
Dmd Gene ◽  
Novel Therapeutic

Rationale: Absence of dystrophin in Duchenne muscular dystrophy (DMD) results in the degeneration of skeletal and cardiac muscles. Owing to advances in respiratory management of DMD patients, cardiomyopathy has become a significant aspect of the disease. While CRISPR/Cas9 genome editing technology holds great potential as a novel therapeutic avenue for DMD, little is known about the potential of DMD correction using CRISPR/Cas9 technology to mitigate cardiac abnormalities in DMD. Objective: To define the effects of CRISPR/Cas9 genome editing on structural, functional and transcriptional abnormalities in DMD-associated cardiac disease. Methods and Results: We generated induced pluripotent stem cells (iPSCs) from a patient with a deletion of exon 44 of the DMD gene (ΔEx44) and his healthy brother. We targeted exon 45 of the DMD gene by CRISPR/Cas9 genome editing to generate corrected DMD (cDMD) iPSC lines, wherein the DMD open reading frame was restored via reframing (RF) or exon skipping (ES). While DMD cardiomyocytes (CMs) demonstrated morphologic, structural and functional deficits compared to control CMs, CMs from both cDMD lines were similar to control CMs. Bulk RNA-sequencing of DMD CMs showed transcriptional dysregulation consistent with dilated cardiomyopathy, which was mitigated in cDMD CMs. We then corrected dysfunctional DMD CMs by adenoviral delivery of Cas9/gRNA and showed that correction of DMD CMs post-differentiation reduces their arrhythmogenic potential. Single-nucleus RNA-sequencing of hearts of DMD mice showed transcriptional dysregulation in CMs and fibroblasts, which in corrected mice was reduced to similar levels as wildtype mice. Conclusions: We show that CRISPR/Cas9-mediated correction of DMD ΔEx44 mitigates structural, functional and transcriptional abnormalities consistent with dilated cardiomyopathy irrespective of how the protein reading frame is restored. We show that these effects extend to postnatal editing in iPSC-CMs and mice. These findings provide key insights into the utility of genome editing as a novel therapeutic for DMD-associated cardiomyopathy.

scholarly journals CRISPR-Generated Animal Models of Duchenne Muscular Dystrophy

Genes ◽  
2020 ◽  
Vol 11 (3) ◽  
pp. 342 ◽  
Author(s):  
Kenji Rowel Q. Lim ◽  
Quynh Nguyen ◽  
Kasia Dzierlega ◽  
Yiqing Huang ◽  
Toshifumi Yokota
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Animal Models ◽  
Mechanical Stress ◽  
Muscle Cell ◽  
Neuromuscular Disorder ◽  
Reading Frame ◽  
Advantages And Disadvantages ◽  
Short Palindromic Repeat ◽  
Dmd Gene

Duchenne muscular dystrophy (DMD) is a fatal X-linked recessive neuromuscular disorder most commonly caused by mutations disrupting the reading frame of the dystrophin (DMD) gene. DMD codes for dystrophin, which is critical for maintaining the integrity of muscle cell membranes. Without dystrophin, muscle cells receive heightened mechanical stress, becoming more susceptible to damage. An active body of research continues to explore therapeutic treatments for DMD as well as to further our understanding of the disease. These efforts rely on having reliable animal models that accurately recapitulate disease presentation in humans. While current animal models of DMD have served this purpose well to some extent, each has its own limitations. To help overcome this, clustered regularly interspaced short palindromic repeat (CRISPR)-based technology has been extremely useful in creating novel animal models for DMD. This review focuses on animal models developed for DMD that have been created using CRISPR, their advantages and disadvantages as well as their applications in the DMD field.

Duchenne muscular dystrophy: genome editing gives new hope for treatment

2018 ◽  
Vol 94 (1111) ◽  
pp. 296-304 ◽  
Author(s):  
Vassili Crispi ◽  
Antonios Matsakas
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Genome Editing ◽  
Genetic Diseases ◽  
Wasting Disease ◽  
Cardiac Muscles ◽  
Dmd Gene ◽  
Current Lines ◽  
Dystrophin Protein ◽  
Potential Therapeutics

Duchenne muscular dystrophy (DMD) is a progressive wasting disease of skeletal and cardiac muscles, representing one of the most common recessive fatal inherited genetic diseases with 1:3500–1:5000 in yearly incidence. It is caused by mutations in the DMD gene that encodes the membrane-associated dystrophin protein. Over the years, many have been the approaches to management of DMD, but despite all efforts, no effective treatment has yet been discovered. Hope for the development of potential therapeutics has followed the recent advances in genome editing and gene therapy. This review gives an overview to DMD and summarises current lines of evidence with regard to treatment and disease management alongside the appropriate considerations.

scholarly journals Reading Frame Correction by Targeted Genome Editing Restores Dystrophin Expression in Cells From Duchenne Muscular Dystrophy Patients

2013 ◽  
Vol 21 (9) ◽  
pp. 1718-1726 ◽  
Author(s):  
David G Ousterout ◽  
Pablo Perez-Pinera ◽  
Pratiksha I Thakore ◽  
Ami M Kabadi ◽  
Matthew T Brown ◽  
...  
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Genome Editing ◽  
Reading Frame ◽  
Dystrophin Expression ◽  
In Cells

Long-term evaluation of AAV-CRISPR genome editing for Duchenne muscular dystrophy shows its not safe due to AAV - and more worryingly Cas9 - integration

2019 ◽  
Author(s):  
Sandeep Chakraborty
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Genome Editing ◽  
Mdx Mouse ◽  
Sequencing Data ◽  
Important Distinction ◽  
Adult Mice ◽  
Gene Therapy Application ◽  
Dmd Gene

Duchenne muscular dystrophy (DMD), a monogenic disorder characterized by progressive muscle degeneration, is one of the first diseases being targeted for therapeutic genome editing using nuclease- based methods (CRISPR/ZFN/TALEN). However, safety and persistence remains a concern. Long-term (1 year) persistence and safety of a single intravenous administration of an adeno-associated virus (AAV) and CRISPR was reported in mdx mouse model recently [1]. They reported that ‘AAV-CRISPR is immunogenic when administered to adult mice’, which can be ‘avoided by treating neonatal mice’, and also warned about ‘unintended genome and transcript alterations’. Here, the integration of the Cas9 protein in the exact two locations in the DMD gene which has been edited has been shown based on the same sequencing data (Accid:PRJNA485509). Transcriptomic data also shows Cas9 being expressed. There is an important distinction between AAV and Cas9 integration - while AAV integration can be tolerated, Cas9 integration is a huge, and unacceptable, danger. While there are use cases where the nuclease can be sent as as protein, any gene-therapy application for DMD would require delivery using AAV and the nuclease in a plasmid. So, there is no possible alleviation for this in the future, unless we are willing to accept transgenic humans as a trade-off for curing DMD.

scholarly journals New Approach for Antisense Oligonucleotide-Mediated Exon Skipping in Duchenne Muscular Dystrophy

2012 ◽  
Vol 16 (4) ◽  
pp. 521-526
Author(s):  
Yoshitsugu Aoki ◽  
◽  
Tetsuya Nagata ◽  
Shin’ichi Takeda
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Antisense Oligonucleotide ◽  
Antisense Oligonucleotides ◽  
Exon Skipping ◽  
New Approach ◽  
Reading Frame ◽  
Promising Therapeutic Approach ◽  
Dmd Gene ◽  
Dystrophin Protein

Duchenne Muscular Dystrophy (DMD) is a lethalmuscle disorder characterized by mutations in the DMD gene. These mutations primarily disrupt the reading frame, resulting in the absence of functional dystrophin protein. Exon skipping, which involves the use of antisense oligonucleotides is a promising therapeutic approach for DMD, and clinical trials on exon skipping are currently underway in DMD patients. Recently, stable and less-toxic antisense oligonucleotides with higher efficacy have been developed in mouse and dog models of DMD. This review highlights a new approach for antisense oligonucleotide-based therapeutics for DMD, particularly for exon skipping-based methods.

scholarly journals CRISPR-Generated Animal Models of Duchenne Muscular Dystrophy

2020 ◽  
Author(s):  
Kenji Rowel Q. Lim ◽  
Quynh Nguyen ◽  
Kasia Dzierlega ◽  
Yiqing Huang ◽  
Toshifumi Yokota
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Animal Models ◽  
Mechanical Stress ◽  
Muscle Cell ◽  
Muscle Cells ◽  
Neuromuscular Disorder ◽  
Reading Frame ◽  
Advantages And Disadvantages ◽  
Dmd Gene

Duchenne muscular dystrophy (DMD) is a fatal X-linked recessive neuromuscular disorder most commonly caused by mutations disrupting the reading frame of the dystrophin (DMD) gene. DMD codes for dystrophin, which is critical for maintaining the integrity of muscle cell membranes. Without dystrophin, muscle cells receive heightened mechanical stress, becoming more susceptible to damage. An active body of research continues to explore therapeutic treatments for DMD as well as to further our understanding of the disease. These efforts rely on having reliable animal models that accurately recapitulate disease presentation in humans. While current animal models of DMD have served this purpose quite well, each comes with their own limitations. To help overcome this, clustered regularly interspaced short palindromic repeats (CRISPR)-based technology has been extremely useful in creating novel animal models for DMD. This review focuses on animal models developed for DMD that have been created using CRISPR, their advantages and disadvantages as well as their applications in the DMD field.

scholarly journals Genome editing for Duchenne muscular dystrophy: a glimpse of the future?

Gene Therapy ◽  
2021 ◽  
Author(s):  
Christian Kupatt ◽  
Alina Windisch ◽  
Alessandra Moretti ◽  
Eckhard Wolf ◽  
Wolfgang Wurst ◽  
...  
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Genome Editing ◽  
Rare Event ◽  
Severe Form ◽  
Muscle Disease ◽  
End Joining ◽  
Reading Frame ◽  
Non Homologous End Joining ◽  
Large Animals

AbstractMutations in Dystrophin, one of the largest proteins in the mammalian body, are causative for a severe form of muscle disease, Duchenne Muscular Dystrophy (DMD), affecting not only skeletal muscle, but also the heart. In particular, exons 45–52 constitute a hotspot for DMD mutations. A variety of molecular therapies have been developed, comprising vectors encoding micro- and minidystrophins as well as utrophin, a protein with partially overlapping functions. With the advent of the CRISPR-Cas9-nuclease, genome editing offers a novel option of correction of the disease-cuasing mutations. Full restoration of the healthy gene by homology directed repair is a rare event. However, non-homologous end-joining (NHEJ) may restore the reading frame by causing exon excision. This approach has first been demonstrated in mice and then translated to large animals (dogs, pigs). This review discusses the potential opportunities and limitations of genome editing in DMD, including the generation of appropriate animal models as well as new developments in genome editing tools.

scholarly journals Evaluating the potential of novel genetic approaches for the treatment of Duchenne muscular dystrophy

2021 ◽  
Author(s):  
Vratko Himič ◽  
Kay E. Davies
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Structural Integrity ◽  
Viral Vector ◽  
Exon Skipping ◽  
Dystrophin Gene ◽  
Pharmacological Treatments ◽  
Genetic Sequencing ◽  
Reading Frame ◽  
Target Effects

AbstractDuchenne muscular dystrophy (DMD) is an X-linked progressive muscle-wasting disorder that is caused by a lack of functional dystrophin, a cytoplasmic protein necessary for the structural integrity of muscle. As variants in the dystrophin gene lead to a disruption of the reading frame, pharmacological treatments have only limited efficacy; there is currently no effective therapy and consequently, a significant unmet clinical need for DMD. Recently, novel genetic approaches have shown real promise in treating DMD, with advancements in the efficacy and tropism of exon skipping and surrogate gene therapy. CRISPR-Cas9 has the potential to be a ‘one-hit’ curative treatment in the coming decade. The current limitations of gene editing, such as off-target effects and immunogenicity, are in fact partly constraints of the delivery method itself, and thus research focus has shifted to improving the viral vector. In order to halt the loss of ambulation, early diagnosis and treatment will be pivotal. In an era where genetic sequencing is increasingly utilised in the clinic, genetic therapies will play a progressively central role in DMD therapy. This review delineates the relative merits of cutting-edge genetic approaches, as well as the challenges that still need to be overcome before they become clinically viable.

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