CRISPR/Cas system: A game changing genome editing technology, to treat human genetic diseases

Genome editing technologies promise therapeutic advances for genetic diseases. We discuss the ethical and societal issues raised by these technologies, including their use in preclinical research, their potential to address mutations in somatic cells, and their potential to make germ line alterations that may be passed to subsequent generations. We call for a proportionate response from health leaders based on a realistic assessment of benefits, risks, and timelines for clinical translation.

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External Stimuli-Responsive Nanoparticles for Spatially and Temporary Controlled Delivery of CRISPR-Cas Genome Editors

Biomaterials Science ◽

10.1039/d1bm00558h ◽

2021 ◽

Author(s):

Ruosen Xie ◽

Yuyuan Wang ◽

Shaoqin Gong

Keyword(s):

Genome Editing ◽

Genetic Diseases ◽

Delivery Systems ◽

Controlled Delivery ◽

Stimuli Responsive ◽

New Therapies ◽

External Stimuli

The CRISPR–Cas9 system is a powerful tool for genome editing, which can potentially lead to new therapies for genetic diseases. Up to date, various viral and non-viral delivery systems have...

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CRISPR base editors: genome editing without double-stranded breaks

Biochemical Journal ◽

10.1042/bcj20170793 ◽

2018 ◽

Vol 475 (11) ◽

pp. 1955-1964 ◽

Cited By ~ 79

Author(s):

Ayman Eid ◽

Sahar Alshareef ◽

Magdy M. Mahfouz

Keyword(s):

Genome Editing ◽

Dna Sequences ◽

Genetic Diseases ◽

Great Promise ◽

Cytidine Deaminases ◽

Strand Breaks ◽

Base Editing ◽

Potential Applications ◽

Short Palindromic Repeat ◽

Basic Biology

The CRISPR (clustered regularly interspaced short palindromic repeat)/Cas9 adaptive immunity system has been harnessed for genome editing applications across eukaryotic species, but major drawbacks, such as the inefficiency of precise base editing and off-target activities, remain. A catalytically inactive Cas9 variant (dead Cas9, dCas9) has been fused to diverse functional domains for targeting genetic and epigenetic modifications, including base editing, to specific DNA sequences. As base editing does not require the generation of double-strand breaks, dCas9 and Cas9 nickase have been used to target deaminase domains to edit specific loci. Adenine and cytidine deaminases convert their respective nucleotides into other DNA bases, thereby offering many possibilities for DNA editing. Such base-editing enzymes hold great promise for applications in basic biology, trait development in crops, and treatment of genetic diseases. Here, we discuss recent advances in precise gene editing using different platforms as well as their potential applications in basic biology and biotechnology.

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Duchenne muscular dystrophy: genome editing gives new hope for treatment

Postgraduate Medical Journal ◽

10.1136/postgradmedj-2017-135377 ◽

2018 ◽

Vol 94 (1111) ◽

pp. 296-304 ◽

Cited By ~ 2

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.

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Application of CRISPR/Cas9 Nuclease in Amphioxus Genome Editing

Genes ◽

10.3390/genes11111311 ◽

2020 ◽

Vol 11 (11) ◽

pp. 1311

Author(s):

Liuru Su ◽

Chenggang Shi ◽

Xin Huang ◽

Yiquan Wang ◽

Guang Li

Keyword(s):

Animal Model ◽

Thermal Treatment ◽

Genome Editing ◽

Phylogenetic Position ◽

Transcription Activator ◽

Cell Stage ◽

System A ◽

Cas9 Nuclease ◽

Cas9 Protein ◽

Amphioxus Genome

The cephalochordate amphioxus is a promising animal model for studying the origin of vertebrates due to its key phylogenetic position among chordates. Although transcription activator-like effector nucleases (TALENs) have been adopted in amphioxus genome editing, its labor-intensive construction of TALEN proteins limits its usage in many laboratories. Here we reported an application of the CRISPR/Cas9 system, a more amenable genome editing method, in this group of animals. Our data showed that while co-injection of Cas9 mRNAs and sgRNAs into amphioxus unfertilized eggs caused no detectable mutations at targeted loci, injections of Cas9 mRNAs and sgRNAs at the two-cell stage, or of Cas9 protein and sgRNAs before fertilization, can execute efficient disruptions of targeted genes. Among the nine tested sgRNAs (targeting five genes) co-injected with Cas9 protein, seven introduced mutations with efficiency ranging from 18.4% to 90% and four caused specific phenotypes in the injected embryos. We also demonstrated that monomerization of sgRNAs via thermal treatment or modifying the sgRNA structure could increase mutation efficacies. Our study will not only promote application of genome editing method in amphioxus research, but also provide valuable experiences for other organisms in which the CRISPR/Cas9 system has not been successfully applied.

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CRISPR Cas9 Tip of an Iceberg: It’s Potential in Genome Editing and Many More

10.20944/preprints202107.0182.v1 ◽

2021 ◽

Author(s):

Jalal Ahmad ◽

Nayyer Siddique

Keyword(s):

Genome Editing ◽

Genetic Diseases ◽

Adaptive Immune System ◽

Biological Research ◽

Adaptive Immune ◽

Scientific World ◽

A Genome ◽

Foreign Dna ◽

Immense Potential ◽

Potential Use

Clustered regularly interspaced short palindromic repeats or CRISPR, one of the major technological tools from nature's toolbox, has revolutionized the scientific world with its potential use in humans and plants. CRISPR Cas9 was first known as an adaptive immune system of bacteria. It is a system that cleaves foreign DNA. It has been exploited to be used as a genome editing tool for correcting genetic diseases in humans, for plants to create stress-resistant plants, and for a variety of different purposes. This review provides a basic overview of its applications in different areas of biological research. It has immense potential for a variety of researches, but it's still a mystery for science. It feels like scientists just know a tip of an iceberg.

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CRISPR-Cas9 genome editing in human cells works via the Fanconi Anemia pathway

10.1101/136028 ◽

2017 ◽

Cited By ~ 17

Author(s):

Chris D Richardson ◽

Katelynn R Kazane ◽

Sharon J Feng ◽

Nicholas L Bray ◽

Axel J Schäfer ◽

...

Keyword(s):

Dna Repair ◽

Genome Editing ◽

Fanconi Anemia ◽

High Efficiency ◽

Genetic Diseases ◽

Dermal Fibroblasts ◽

Human Cells ◽

Individual Gene ◽

Break Site ◽

Repair Pathways

AbstractCRISPR-Cas9 genome editing creates targeted double strand breaks (DSBs) in eukaryotic cells that are processed by cellular DNA repair pathways. Co-administration of single stranded oligonucleotide donor DNA (ssODN) during editing can result in high-efficiency (>20%) incorporation of ssODN sequences into the break site. This process is commonly referred to as homology directed repair (HDR) and here referred to as single stranded template repair (SSTR) to distinguish it from repair using a double stranded DNA donor (dsDonor). The high efficacy of SSTR makes it a promising avenue for the treatment of genetic diseases1,2, but the genetic basis of SSTR editing is still unclear, leaving its use a mostly empiric process. To determine the pathways underlying SSTR in human cells, we developed a coupled knockdown-editing screening system capable of interrogating multiple editing outcomes in the context of thousands of individual gene knockdowns. Unexpectedly, we found that SSTR requires multiple components of the Fanconi Anemia (FA) repair pathway, but does not require Rad51-mediated homologous recombination, distinguishing SSTR from repair using dsDonors. Knockdown of FA genes impacts SSTR without altering break repair by non-homologous end joining (NHEJ) in multiple human cell lines and in neonatal dermal fibroblasts. Our results establish an unanticipated and central role for the FA pathway in templated repair from single stranded DNA by human cells. Therapeutic genome editing has been proposed to treat genetic disorders caused by deficiencies in DNA repair, including Fanconi Anemia. Our data imply that patient genotype and/or transcriptome profoundly impact the effectiveness of gene editing treatments and that adjuvant treatments to bias cells towards FA repair pathways could have considerable therapeutic value.

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Tissue Specific DNA Repair Outcomes Shape the Landscape of Genome Editing

Frontiers in Genetics ◽

10.3389/fgene.2021.728520 ◽

2021 ◽

Vol 12 ◽

Author(s):

Mathilde Meyenberg ◽

Joana Ferreira da Silva ◽

Joanna I. Loizou

Keyword(s):

Dna Repair ◽

Genome Editing ◽

Genetic Diseases ◽

Dna Double Strand Breaks ◽

Dna Breaks ◽

Dna Damage And Repair ◽

Tissue Specific ◽

Precise Control ◽

Repair Processes ◽

Recent Developments

The use of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-Cas9 has moved from bench to bedside in less than 10years, realising the vision of correcting disease through genome editing. The accuracy and safety of this approach relies on the precise control of DNA damage and repair processes to achieve the desired editing outcomes. Strategies for modulating pathway choice for repairing CRISPR-mediated DNA double-strand breaks (DSBs) have advanced the genome editing field. However, the promise of correcting genetic diseases with CRISPR-Cas9 based therapies is restrained by a lack of insight into controlling desired editing outcomes in cells of different tissue origin. Here, we review recent developments and urge for a greater understanding of tissue specific DNA repair processes of CRISPR-induced DNA breaks. We propose that integrated mapping of tissue specific DNA repair processes will fundamentally empower the implementation of precise and safe genome editing therapies for a larger variety of diseases.

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CRISPR-Cas Genome editing system: a versatile tool for developing disease resistant crops

Plant Stress ◽

10.1016/j.stress.2022.100056 ◽

2022 ◽

pp. 100056

Author(s):

Ashwini Talakayala ◽

Srinivas Ankanagari ◽

Mallikarjuna Garladinne

Keyword(s):

Genome Editing ◽

System A ◽

Disease Resistant ◽

Versatile Tool

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Allele specific repair of splicing mutations in cystic fibrosis through AsCas12a genome editing

Nature Communications ◽

10.1038/s41467-019-11454-9 ◽

2019 ◽

Vol 10 (1) ◽

Cited By ~ 19

Author(s):

Giulia Maule ◽

Antonio Casini ◽

Claudia Montagna ◽

Anabela S. Ramalho ◽

Kris De Boeck ◽

...

Keyword(s):

Cystic Fibrosis ◽

Genome Editing ◽

Genetic Diseases ◽

Airway Epithelial Cells ◽

Airway Epithelial ◽

Gene Correction ◽

Splicing Mutations ◽

Mutant Sequence ◽

A Genome ◽

Allele Specific

Abstract Cystic fibrosis (CF) is an autosomal recessive disease caused by mutations in the CFTR gene. The 3272–26A>G and 3849+10kbC>T CFTR mutations alter the correct splicing of the CFTR gene, generating new acceptor and donor splice sites respectively. Here we develop a genome editing approach to permanently correct these genetic defects, using a single crRNA and the Acidaminococcus sp. BV3L6, AsCas12a. This genetic repair strategy is highly precise, showing very strong discrimination between the wild-type and mutant sequence and a complete absence of detectable off-targets. The efficacy of this gene correction strategy is verified in intestinal organoids and airway epithelial cells derived from CF patients carrying the 3272–26A>G or 3849+10kbC>T mutations, showing efficient repair and complete functional recovery of the CFTR channel. These results demonstrate that allele-specific genome editing with AsCas12a can correct aberrant CFTR splicing mutations, paving the way for a permanent splicing correction in genetic diseases.

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