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A toolmaker's perspective on CRISPR-directed gene editing as a therapeutic strategy for leukemia and beyond.
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CRISPR-Pass: Gene Rescue of Nonsense Mutations Using Adenine Base Editors.
A nonsense mutation is a substitutive mutation in a DNA sequence that causes a premature termination during translation and produces stalled proteins, resulting in dysfunction of a gene. Although it usually induces severe genetic disorders, there are no definite methods for inducing read through of premature termination codons (PTCs). Here, we present a targeted tool for bypassing PTCs, named CRISPR-pass, that uses CRISPR-mediated adenine base editors. CRISPR-pass, which should be applicable to 95.5% of clinically significant nonsense mutations in the ClinVar database, rescues protein synthesis in patient-derived fibroblasts, suggesting potential clinical utility.
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The CRISPR Toolbox: The End of the Beginning.
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Challenges of in vitro genome editing with CRISPR/Cas9 and possible solutions: A review.
Microbial production of bio-based ingredients often requires metabolically engineered bacterial strains with the edited genome. Genome editing tools are also essential for gene identification and investigating genotype-phenotype connections. Currently, one of the most common tools of genome editing is based on a natural bacterial adaptive immune system known as CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)/Cas9 (CRISPR-associated protein 9) due to its simple, rapid, and efficient activities. Although successful in some in vitro systems, its application as an approach of metabolic engineering and genome editing is still not so extensive. Here, we discuss existing barriers and challenges of the CRISPR/Cas9 editing tool for in vitro systems. Firstly, we aim to briefly introduce the CRISPR/Cas9 method as an in vitro gene editing tool. Next, we discuss existing obstacles to CRISPR-based editing in bacterial and in vitro model systems and offer guidelines to help achieve editing in an expanded range of in vitro systems.
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CRISPR/Cas Systems: Biological Basis and Genome Editing Applications.
Clustered regularly interspaced short palindromic repeats (CRISPR) and associated (Cas) systems have revolutionized the field of genome engineering by providing versatile, efficient, and programmable tools for precise genetic manipulation. Originally identified as an adaptive immune mechanism in prokaryotes, CRISPR/Cas systems have been extensively repurposed for a wide range of applications across molecular biology, biotechnology, and medicine. This chapter provides a comprehensive overview of the molecular mechanisms underlying CRISPR/Cas immunity. Furthermore, the classification of CRISPR/Cas systems into distinct types and subtypes is discussed, highlighting their structural and functional diversity. Advances in genome editing technologies, including CRISPR-mediated knockout, base editing, and prime editing, are explored with an emphasis on their mechanisms and applications. The chapter also examines emerging CRISPR-based platforms for transcriptional regulation, epigenome editing, and RNA targeting, which enable precise and reversible modulation of gene expression without altering genomic DNA. In addition, the transformative impact of CRISPR technologies on functional genomics is addressed, particularly through high-throughput screening approaches that facilitate the identification of gene function and genetic vulnerabilities. CRISPR-based diagnostic tools and therapeutic strategies are also reviewed, underscoring their potential in disease detection and treatment. Despite significant progress, challenges such as off-target effects, delivery limitations, and safety concerns remain critical considerations. Overall, this chapter highlights the expanding capabilities of CRISPR/Cas systems and their growing importance in both fundamental research and clinical applications.
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CRISPR challenges in clinical developments.
CRISPR-Cas (clustered regularly interspaced short palindromic repeats and associated proteins) is a novel genome editing technology with potential applications in treating diseases. Currently, its use in humans is restricted to clinical trials, although its growth rate is significant, and some have received initial FDA approval. It is crucial to examine and address the challenges for this technology to be implemented in clinical settings. This review aims to identify and explore new research ideas to increase of CRISPR's efficiency in treating genetic diseases and cancer, as well as its future prospects. Given that a substantial amount of previous research has focused on CRISPR-Cas delivery strategies and materials, this overview introduces specific conditions and strategies. It also discusses some of the challenges and opportunities in this field, offering a unique perspective.
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Recent Advances in CRISPR Base Editing: From A to RNA.
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Gene editing in hemophilia: a "CRISPR" choice?
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Charting the development and engineering of CRISPR base editors: lessons and inspirations.
CRISPR base editors (BEs) have introduced a new chapter in precise genome editing. The brief but fruitful history of BE development documents many case studies that not only lay the foundation of base-editing technology but are also instrumental to future protein engineering efforts. In this review, we summarize the development and engineering of various BEs with a focus on recent progress. These include traditional cytosine and adenine base editors (CBEs and ABEs), novel TadA-derived CBEs, transversion BEs, dual BEs, and CRISPR-free BEs. We discuss each aspect of the workflow and highlight the successes and challenges encountered in the engineering process.
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Tearing CRISPR apart.
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