NANOTECHNOLOGY-ENABLED NON-VIRAL DELIVERY SYSTEMS FOR CRISPR/CAS9 GENE THERAPY: CURRENT PROGRESS AND TRANSLATIONAL CHALLENGES

Abstract

CRISPR/Cas9 has become a versatile platform for programmable genome editing and has generated substantial interest as a therapeutic modality for inherited and acquired diseases. However, clinical translation depends not only on editing specificity but also on safe and efficient delivery of the Cas9 nuclease and guide RNA to the intended cells. Viral vectors can provide high transduction efficiency, but concerns related to immunogenicity, payload constraints, manufacturing complexity and, for some vectors, genomic integration have encouraged development of non-viral alternatives. This review examines nanotechnology-enabled non-viral delivery strategies for CRISPR/Cas9, with emphasis on plasmid DNA, messenger RNA and ribonucleoprotein cargo formats. Major delivery barriers – including extracellular degradation, protein-corona formation, limited tissue penetration, cellular uptake, endosomal entrapment, intracellular trafficking and nuclear access – are discussed together with strategies for overcoming them. Lipid nanoparticles, polymeric nanoparticles, lipid–polymer hybrids, dendrimers, inorganic nanoparticles and emerging vesicular or virus-like systems are compared in terms of cargo compatibility, targeting, endosomal escape, safety and translational potential. Recent advances in liver-directed in vivo editing, non-liver targeting, stimuli-responsive delivery and clinical translation are highlighted. Particular attention is given to the need for tissue-selective delivery, transient editor exposure, reproducible manufacturing and long-term safety monitoring. Overall, rationally engineered, biodegradable and multifunctional nanocarriers represent an important route toward safer and more effective CRISPR/Cas9 therapeutics.