Recent Advancements in Reducing the Off-Target Effect of CRISPR-Cas9 Genome Editing

Recent Advancements in Reducing the Off-Target Effect of CRISPR-Cas9 Genome Editing

18 January 2024 | Misganaw Asmamaw Mengstie, Muluken Teshome Azezew, Tadesse Asmamaw Dejenie, Assefa Agegnehu Teshome, Fitalew Tadele Admasu, Awgichew Behaile Teklemariam, Anemut Tilahun Mulu, Melaku Mekonnen Agidew, Dagnew Getnet Adugna, Habtamu Geremew, Endeshaw Chekol Abebe
The CRISPR-Cas9 genome-editing technology, a powerful tool for addressing genetic disorders, faces significant challenges due to its potential off-target effects, which can lead to unwanted genetic modifications and diminish its therapeutic potential. This review summarizes recent advancements aimed at reducing these off-target effects. Key methods include optimizing sgRNA specificity through GC content, length, and chemical modifications, developing improved Cas9 variants like eSpCas9 and SpCas9-HF1, employing CRISPR nickase to reduce double-strand breaks, and utilizing prime editing, which does not require donor DNA or double-strand breaks. Additionally, anti-CRISPR proteins and the discovery of SuperFi-Cas9, a highly specific Cas9 variant, offer further strategies to enhance specificity. These advancements collectively aim to improve the precision and safety of CRISPR-Cas9 genome editing, making it more viable for clinical applications.The CRISPR-Cas9 genome-editing technology, a powerful tool for addressing genetic disorders, faces significant challenges due to its potential off-target effects, which can lead to unwanted genetic modifications and diminish its therapeutic potential. This review summarizes recent advancements aimed at reducing these off-target effects. Key methods include optimizing sgRNA specificity through GC content, length, and chemical modifications, developing improved Cas9 variants like eSpCas9 and SpCas9-HF1, employing CRISPR nickase to reduce double-strand breaks, and utilizing prime editing, which does not require donor DNA or double-strand breaks. Additionally, anti-CRISPR proteins and the discovery of SuperFi-Cas9, a highly specific Cas9 variant, offer further strategies to enhance specificity. These advancements collectively aim to improve the precision and safety of CRISPR-Cas9 genome editing, making it more viable for clinical applications.
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