review · Biologics
The CRISPR-Cas9 system has emerged as a prominent genome-editing tool with the potential to address various genetic disorders. At its core, the system relies on a single-guide RNA containing a user-defined twenty-base spacer sequence paired with the Cas9 endonuclease. This guide directs the Cas9 nuclease to cut specific genomic targets adjacent to a protospacer adjacent motif matching the spacer sequence. Despite its promise, clinical translation faces substantial hurdles due to off-target effects, where unintended DNA cleavage occurs at incorrect sites. Such non-specific activity limits the therapeutic effectiveness of CRISPR-Cas9 and risks confusing scientific investigations into gene function. Recent technological developments have concentrated on mitigating these off-target cleavage events to improve the precision and clinical reliability of the editing process.
Targeting genetic disorders directly at the DNA level requires exceptional precision. Off-target cuts by CRISPR-Cas9 can introduce unintended mutations, posing safety risks in medical therapies and clouding laboratory discoveries. Addressing these off-target errors is essential for developing safer clinical treatments and ensuring reliable biological research.
For biotechnology and clinical therapeutics developers, minimising off-target cutting is crucial for regulatory approval and patient safety in genetic therapies. While the abstract reviews advancements aimed at resolving this barrier, it does not detail specific commercial tools or clinical trial stages, reflecting an early-to-applied research focus that requires further validation before reaching routine therapeutic deployment.
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The CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR)) and the associated protein (Cas9) system, a young but well-studied genome-editing tool, holds plausible solutions to a wide range of genetic disorders. The single-guide RNA (sgRNA) with a 20-base user-defined spacer sequence and the Cas9 endonuclease form the core of the CRISPR-Cas9 system. This sgRNA can direct the Cas9 nuclease to any genomic region that includes a protospacer adjacent motif (PAM) just downstream and matches the spacer sequence. The current challenge in the clinical applications of CRISPR-Cas9 genome-editing technology is the potential off-target effects that can cause DNA cleavage at the incorrect sites. Off-target genome editing confuses and diminishes the therapeutic potential of CRISPR-Cas9 in addition to potentially casting doubt on scientific findings regarding the activities of genes. In this review, we summarize the recent technological advancements in reducing the off-target effect of CRISPR-Cas9 genome editing.
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DOI: 10.2147/btt.s429411
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