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Integrating CRISPR technology with exosomes: Revolutionizing gene delivery systems.
CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) serves as an adaptive immune system in bacteria and archaea, offering a defense mechanism against invading genetic elements such as viruses (bacteriophages) and plasmids. Today, CRISPR has evolved into a powerful gene-editing technology that enables highly specific and rapid modifications of DNA within a genome. It has a broad range of applications across various fields, including medicine, agriculture, and fundamental research. One of the significant challenges facing this technology is the efficient transfer of CRISPR constructs into target cells for gene editing. There are several methods to deliver this system into target cells, which can be classified as viral and non-viral methods. Each of these approaches has its own advantages and disadvantages. Recently, the use of extracellular vesicles for delivery has garnered particular attention. Exosomes are nano-sized extracellular vesicles that have emerged as promising carriers for drug delivery due to their unique properties. These naturally occurring vesicles, typically ranging from 30 to 150 nm in diameter, facilitate intercellular communication by transferring bioactive molecules such as proteins, lipids, and nucleic acids between cells. Exosome therapy has surfaced as a promising strategy in regenerative medicine, utilizing small extracellular vesicles to deliver therapeutic molecules to target cells. One of the emerging options for transferring the CRISPR system is exosomes. The integration of these two advanced technologies holds significant potential for developing efficient and targeted gene editing and advancing precision medicine. In contemporary medicine, there is an increasing focus on personalized and targeted treatments that cater to the distinct genetic and molecular profiles of individual patients. The synergy of CRISPR technology and exosome therapy presents a remarkable opportunity to develop highly targeted and effective therapeutic strategies customized to individual patient requirements. This review article examines the potential of incorporating CRISPR technology within exosomes for precision therapeutic applications.
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CRISPR/Cas9-Based Multiplex Genome Editing in Monocot and Dicot Plants.
The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9-mediated genome targeting system has been applied to a variety of organisms, including plants. Compared to other genome-targeting technologies such as zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs), the CRISPR/Cas9 system is easier to use and has much higher editing efficiency. In addition, multiple "single guide RNAs" (sgRNAs) with different target sequences can be designed to direct the Cas9 protein to multiple genomic sites for simultaneous multiplex editing. Here, we present a procedure for highly efficient multiplex genome targeting in monocot and dicot plants using a versatile and robust CRISPR/Cas9 vector system, emphasizing the construction of binary constructs with multiple sgRNA expression cassettes in one round of cloning using Golden Gate ligation. We also describe the genotyping of targeted mutations in transgenic plants by direct Sanger sequencing followed by decoding of superimposed sequencing chromatograms containing biallelic or heterozygous mutations using the Web-based tool DSDecode. © 2016 by John Wiley & Sons, Inc.
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Is CAR-T Really Putting Us On Road to Gene Therapy?
Some say gene editing platforms like CRISPR are a truer version of gene therapy because they are designed to home in on a particular genomic location.
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[CRISPR-Cas system as molecular scissors for gene therapy].
Since the discovery of the CRISPR-Cas system as the adaptive immune system of prokaryotes, the underlying mechanism has proven to be a precise molecular tool for the targeted editing of genetic information in various cell types. By using the CRISPR-Cas9 system DNA sequences can be cut out at any site in the genome and changed in a sequence-specific manner. In the long term this provides the opportunity to cure diseases caused by gene mutations.
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Antiviral Goes Viral: Harnessing CRISPR/Cas9 to Combat Viruses in Humans.
The clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) systems are RNA-guided sequence-specific prokaryotic antiviral immune systems. In prokaryotes, small RNA molecules guide Cas effector endonucleases to invading foreign genetic elements in a sequence-dependent manner, resulting in DNA cleavage by the endonuclease upon target binding. A rewired CRISPR/Cas9 system can be used for targeted and precise genome editing in eukaryotic cells. CRISPR/Cas has also been harnessed to target human pathogenic viruses as a potential new antiviral strategy. Here, we review recent CRISPR/Cas9-based approaches to combat specific human viruses in humans and discuss challenges that need to be overcome before CRISPR/Cas9 may be used in the clinic as an antiviral strategy.
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CRISPR for Neuromuscular Disorders: Gene Editing and Beyond.
This is a review describing advances in CRISPR/Cas-mediated therapies for neuromuscular disorders (NMDs). We explore both CRISPR-mediated editing and dead Cas approaches as potential therapeutic strategies for multiple NMDs. Last, therapeutic considerations, including delivery and off-target effects, are also discussed.
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Keeping crispr in check: diverse mechanisms of phage-encoded anti-crisprs.
CRISPR-Cas represents the only adaptive immune system of prokaryotes known to date. These immune systems are widespread among bacteria and archaea, and provide protection against invasion of mobile genetic elements, such as bacteriophages and plasmids. As a result of the arms-race between phages and their prokaryotic hosts, phages have evolved inhibitors known as anti-CRISPR (Acr) proteins to evade CRISPR immunity. In the recent years, several Acr proteins have been described in both temperate and virulent phages targeting diverse CRISPR-Cas systems. Here, we describe the strategies of Acr discovery and the multiple molecular mechanisms by which these proteins operate to inhibit CRISPR immunity. We discuss the biological relevance of Acr proteins and speculate on the implications of their activity for the development of improved CRISPR-based research and biotechnological tools.
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Gene editing: from technologies to applications in research and beyond.
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Hunting for Treasure Chests in Microbial Defense Islands.
Microbes exist in a constant struggle with viruses and have evolved multiple defense mechanisms. Some of these, such as restriction endonucleases and CRISPR-Cas, have revolutionized genome editing methodology. A systematic strategy combining computation and experiment led to discovery of ten new defense systems that remain to be characterized mechanistically.
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[Progress and application of genome-edited pigs in biomedical research].
Genome editing technologies (GETs) can precisely alter the genomic sequences and modify the genetic information at the target site of an organism. Since the beginning of the 21st century, the GETs, including zinc finger nucleases (ZFN), transcription-activating-like receptor factor (TALEN), and clustered regularly interspaced short palindromic repeats/Cas endonucleases (CRISPR/Cas), have been successively developed. The GETs can easily engineer the targeted genomic site of animals to exhibit a desired phenotype(s), thereby providing valuable tools in biomedical research. The pigs are closely related to human, in terms of similarities in physiological properties and pathogenic characters. Thus, pigs have been used as important animal models in studies of human disease, xenotransplantation, and humanized organs regeneration. In this review, we summarize the development of the three GETs, research progress of genome-edited pigs as disease models and organ donors for xenotransplantation, and the prospects of their applications in future biomedical research.
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