CRISPR-Cas Systems in Functional Genomics and Therapeutics

Abstract

CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeats and CRISPR-associated proteins) systems
have revolutionised functional genomics and therapeutic development by providing programmable, precise, and efficient
tools for genome editing, transcriptional regulation, epigenome remodelling, and base editing across virtually all organism
types. This review comprehensively surveys the mechanistic diversity of CRISPR-Cas systems--from the nuclease-active
SpCas9 and SaCas9 to the RNA-targeting CasRx, the single-base editors BE3 and ABE, and the versatile prime editors
PE2 and PE3--and evaluates their application across two transformative domains: functional genomics, including
genome-wide CRISPR screens, CRISPRi/CRISPRa transcriptional perturbation libraries, and epigenome editing for gene
regulatory network dissection; and therapeutics, covering ex-vivo haematopoietic stem cell editing for sickle cell disease
and beta-thalassaemia, in-vivo liver-targeted delivery for transthyretin amyloidosis (NTLA-2001), CAR-T cell engineering,
and emerging in-vivo CNS and muscle delivery approaches. Clinical trial progress as of 2025 is reviewed, including the
landmark FDA and EMA approval of Casgevy (exa-cel) for sickle cell disease and beta-thalassaemia in December
2023--the first approved CRISPR therapeutic--and the expanding pipeline of 47 active clinical trials. Delivery systems,
off-target safety profiles, immunogenicity, and regulatory considerations are critically evaluated. The review concludes by
identifying the five most impactful near-term advances: prime editing clinical translation, in-vivo muscle CRISPR delivery,
epigenome-only editing, pan-genome CRISPR libraries, and AI-guided guide RNA design.