CRISPR-Cas Systems: Mechanisms, Variants, and Biomedical Applications: A Comprehensive Review

Objectives: This review aims to provide a comprehensive synthesis of the mechanisms, classification, emerging variants, and biomedical applications of CRISPR-Cas systems, while critically evaluating current limitations and future directions in genome editing. Data Sources: Peer-reviewed literature was sourced from PubMed, Google Scholar, and institutional databases. Sources included primary research articles, systematic reviews, clinical trial reports, and authoritative commentary up till 2025. Study Selection: Studies were selected based on relevance to CRISPR-Cas mechanism, classification, therapeutic application, and emerging technologies. Priority was given to high-impact journals in molecular biology, genetics, and clinical medicine. Summary: CRISPR-Cas systems, originally characterized as adaptive immune mechanisms in prokaryotes, have been repurposed as highly precise genome engineering platforms. The two major system classes, defined by multi-protein versus single-effector complexes, encompass diverse types with distinct nuclease activities and target specificities. Key variants, including base editors, prime editors, and diagnostic platforms such as SHERLOCK and DETECTR, have substantially expanded functional capabilities. Biomedical applications span therapeutic gene correction in monogenic disorders, cancer immunotherapy, antiviral strategies, functional genomics, and disease modelling. Persistent challenges include off-target effects, delivery limitations, immune responses, and ethical concerns surrounding germline editing. Conclusion: CRISPR-Cas technology represents a paradigm shift in molecular biology. Continued refinement of editing fidelity, delivery systems, and ethical frameworks will be essential for its safe and equitable clinical translation. Keywords: CRISPR-Cas systems, genome editing, Cas9 nuclease, gene knockout, gene therapy, base editing, prime editing