Enhancing NK Cells Antitumor Activity by CRISPR-Cas9 Mediated Knockout of Inhibitory Receptors

Cancer is still one of the leading causes of death globally, and current treatment methods such as chemotherapy and CAR-T cell therapy are commonly used but are restricted by systemic toxicity, high costs, and reduced efficacy in many solid tumors. CRISPR-Cas9 technology offers a versatile platform to engineer natural killer cells to overcome these limitations by precise genomic edits that enhance cytotoxicity and enable off the shelf manufacturing. Recent approaches include knockout of inhibitory receptors including NKG2A, TIGIT and TIM 3, high throughput target discovery, and combinatorial strategies such as overexpression of activating receptors or integration of chimeric antigen receptors to broaden antigen specificity. Preclinical studies report improved target cell killing, resistance to tumor microenvironment induced dysfunction, and efficacy in hematologic and solid tumor models, while outstanding challenges remain in delivery efficiency, off target editing, functional persistence and safety. Addressing these challenges will be critical for clinical translation. This review aims to integrate current preclinical and clinical evidence on CRISPR-edited natural killer (NK) cells, assess the efficacy and safety of inhibitory receptor disruption and combined engineering strategies, and outline priorities for translational research to accelerate the process of developing safer and more effective NK cell therapies.