Pancreatic cancer, characterized by aggressive progression and poor prognosis, is a highly lethal malignancy, ranking among the leading causes of cancer-related deaths. Conventional treatments provide limited efficacy, presenting ongoing therapeutic challenges. Advances in immunotherapy have recognized natural killer (NK) cells as a promising avenue for cancer treatment owing to their ability to selectively target tumor cells. However, despite progress in ex vivo expansion techniques for NK-cell-based therapies, their cytotoxic efficacy can vary significantly depending on the functional variability among individual donors, which presents a notable limitation. In this study, we optimized a large-scale ex vivo expansion protocol for NK cells derived from the peripheral blood of patients. On implementation, the expanded NK cells demonstrated high purity and viability after 20 days of culture and had potent cytotoxic activity against pancreatic cancer cell lines and patient-derived cells (PDCs). Through transcriptomic analysis, the gene expression patterns and functional characteristics associated with differences in inhibition efficacy were investigated. Furthermore, the addition of cetuximab, a monoclonal antibody targeting the epidermal growth factor receptor (EGFR), significantly enhanced antibody-dependent cellular cytotoxicity (ADCC), particularly in low-activity NK cells (NK-LA)—resulting in performance comparable to high-activity NK cells. In xenograft models using cell lines and PDCs, the combination therapy with cetuximab had enhanced tumor-suppressive effects of NK-LA compared to monotherapy. These findings highlight the therapeutic potential of integrating expanded patient-derived NK cells with cetuximab in the treatment of pancreatic cancer and underscore the requirement for clinical translation of this strategy.