Efficient large-scale expansion of cord blood-derived NK cells: leveraging lipopolysaccharide for enhanced NK cell production

Abstract

Growing research in the immunotherapy field has shed light on the adoptive cell transfer efficacy of ex vivo expanded natural killer (NK) cells in cancer treatment. In accordance with the advantages of using cord blood as a source of hematopoietic cells, this study aimed to establish a sustainable supply of NK cells by developing a simple method for expanding cord blood-derived NK (CBNK) cells without using feeders or cell-sorting processes. To achieve this aim, culture strategies that stimulate the proliferation of NK cells from hematopoietic stem and progenitor cells (HSPCs) would result in a high purity of CBNK cells. Here, we first compared the potential of lipopolysaccharide (LPS) and the Notch signaling agonist Yhhu-3792 to promote CBNK cell proliferation in cytokine-based conditions before applying these findings to a large-scale expansion platform. Overall, we reveal that the presence of LPS at 1 µg/mL during the first week of a 21-day expansion protocol resulted in an average total nucleated cell (TNC) count of 1.68 ± 2.92 × 10¹⁰, with 92.09 ± 3.47% of the expanded cells being NK cells (n = 5). All subsequent analyses demonstrated that the expanded CBNK cells produced by this procedure are phenotypically and functionally competent NK cells. Collectively, this study developed a simple cytokine-based and cell-sorting-free method for the large-scale expansion of CBNK cells.