Natural killer (NK) cell-based therapies exhibit potent antitumor efficacy and a favorable safety profile. However, their clinical potential is constrained by limited in vivo persistence. In this study, we investigated the clinical relevance of the NK cell population and the expression levels of BIM and NOXA—important apoptosis regulators in NK cells—in relation to the survival of pancreatic ductal adenocarcinoma (PDAC) patients. Our analyses revealed that a low NK score and high expression of these genes correlate with poorer patient survival. Thus, we aimed to enhance NK cell survival and cytotoxic activity by targeting apoptosis through CRISPR-Cas9 gene editing of BIM and NOXA in human peripheral blood NK cells. Although BIM ablation did not yield additional benefits, functional assays demonstrated that NOXA knockout (KO) markedly enhanced NK cell cytotoxicity in serial cancer-killing experiments using PDAC-derived cell lines and in killing assays employing patient-derived organoid models with post-cryopreservation functionality. Furthermore, NOXA-KO NK cells exhibited augmented proliferative capacity, elevated expression of activation markers, and enhanced metabolic activities even in a TME-mimicking environment. These findings underscore NOXA as a critical regulator of apoptosis and cellular metabolism within human NK cells and highlight its potential for augmenting NK-based immunotherapy. Collectively, targeting NOXA can serve as a promising therapeutic modality for achieving sustained cytotoxicity and improved clinical outcomes in cancer immunotherapy.