CRISPR-Cas9, derived from bacterial adaptive immunity, has revolutionized genome editing since its 2012 adaptation for eukaryotic cells. The system uses a guide RNA (gRNA) to direct the Cas9 endonuclease to specific DNA sequences, creating double-strand breaks (DSBs) repaired via non-homologous end joining (NHEJ) for knockouts or homology-directed repair (HDR) for insertions/corrections. This precision surpasses earlier tools like ZFNs and TALENs, enabling applications in treating monogenic disorders (e.g., sickle cell disease via BCL11A disruption), cancers (e.g., PD-1 knockout in T-cells), and infections (e.g., HIV CCR5 excision).
Therapeutic progress includes FDA-approved ex vivo therapies like Casgevy (2023) for β-thalassemia and in vivo trials such as NTLA-2001 (LNP-delivered for ATTR amyloidosis, >80% protein reduction). Variants like base editors (for point mutations without DSBs) and prime editors (for diverse edits) mitigate off-target risks, with high-fidelity Cas9 mutants (e.g., HypaCas9) enhancing specificity. Delivery challenges are addressed by viral vectors (AAV, lentivirus) for non-dividing cells and non-viral LNPs/exosomes for immune evasion.
Prospects include multiplex editing for polygenic diseases and epigenetic modulation via dCas9 fusions (e.g., CRISPRa for gene activation). Ethical considerations guide germline restrictions, focusing on somatic therapies. Ongoing trials target muscular dystrophy, LCA blindness, and obesity.
Sources: Signal Transduction and Targeted Therapy (2023) for progress, PMC (2020) for applications. Word count: 268.
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