A rapid ex vivo co-culture of memory-like NK and activated T cells enhances anti-tumor response in gastric cancer.

[BACKGROUND] Adoptive transfer of NK or T cells alone requires a long preparation time. We developed a rapid co-culture system, including cytokine-induced memory-like NK (CIML NK) cells and activated T cells (Ac-T), as well as tumor-penetrating peptide iRGD.

[METHODS] PBMCs were pretreated with IL-12/15/18 for 16 h, then washed to induce a memory NK phenotype. K562-CD48-41BBL-mbIL-21 cells were generated by lentiviral transduction and used as feeder cells to expand and activate NK cells from PBMCs for 7 days. On day 7, an anti-CD3 monoclonal antibody (OKT3) and an anti-CD28 monoclonal antibody (CD28.2) were added and washed overnight for 24 h. The in vitro (cytotoxicity, cytokines) and in vivo (tumor inhibition, survival) functions of iRGD-modified CIML NK&Ac-T cells (CIML NK&Ac-T-iRGD) were compared to CIML NK&T-iRGD without addition of CD3/CD28.

[RESULTS] We established a one-week rapid co-culture system comprising NK and T cells. This system markedly increased cytokine secretion and demonstrated potent in vitro cytotoxicity against gastric cancer cell lines, significantly inhibited tumor growth, and prolonged survival in a gastric cancer xenograft model. Notably, iRGD-modified CIML NK&Ac-T cells showed superior efficacy compared with CIML NK&T cells.

[CONCLUSION] Our data suggest that the rapid co-culture system of iRGD-modified CIML NK&Ac-T cells used for adoptive cell transfer demonstrates potent anti-tumor effects in gastric cancer. This approach is time-efficient and cost-saving, with potential for broader clinical application compared with conventional NK cell therapies.