Precise gene manipulation by gene editing approaches facilitates the potential to cure several debilitating genetic disorders. Gene modification stimulated by engineered nucleases induces a double-stranded break (DSB) in the target genomic locus, thereby activating DNA repair mechanisms. DSBs triggered by nucleases are repaired either by the nonhomologous end-joining or the homology-directed repair pathway, enabling efficient gene editing. While there are several ongoing ex vivo genome editing clinical trials, current research underscores the therapeutic potential of CRISPR/Cas-based (clustered regularly interspaced short palindrome repeats-associated Cas nuclease) in vivo gene editing. In this review, we provide an overview of the CRISPR/Cas-mediated in vivo genome therapy applications and explore their prospective clinical translatability to treat human monogenic disorders. In addition, we discuss the various challenges associated with in vivo genome editing technologies and strategies used to circumvent them. Despite the robust and precise nuclease-mediated gene editing, a promoterless, nuclease-independent gene targeting strategy has been utilized to evade the drawbacks of the nuclease-dependent system, such as off-target effects, immunogenicity, and cytotoxicity. Thus, the rapidly evolving paradigm of gene editing technologies will continue to foster the progress of gene therapy applications.
Keywords: AAV vectors; CRISPR/Cas9; HDR pathway; gene editing; gene therapy; in vivo; monogenic.