Novel tetrameric bispecific KK-LC-1×CD16A-armed memory-like NK cells enhance antitumor efficacy in gastric cancer.

Background: Cytokine-induced memory-like natural killer (CIML NK) cells demonstrate potent antitumor efficacy against hematological malignancies. Nevertheless, the therapeutic potential of adoptive NK cell transfer remains constrained in solid tumors due to insufficient tumor infiltration efficiency. Developing a novel tetravalent bispecific killer engager (BiKE) to modify CIML NK cells would be a promising strategy to enhance therapeutic efficacy.

Methods: We employed SpyTag-SpyCatcher technology to engineer tetravalent BiKEs, specifically KK-LC-1×CD16A, and to equip CIML NK cells with this protein. Flow cytometry and a luciferase reporter gene system were used to evaluate the activation and cytotoxicity of CIML NK cells. Histological analysis and in vivo real-time fluorescence imaging were employed in the xenograft tumor model to confirm the tumor-infiltrating effectiveness of KK-LC-1×CD16A-armed CIML NK cells. The in vivo antitumor efficacy of armed CIML NK cells was evaluated in subcutaneous gastric cancer xenograft models.

Results: We successfully developed a tetravalent NK cell engager, KK-LC-1×CD16A, which exhibits excellent binding affinity to its targets. CIML NK cells armed with this protein exhibit enhanced activation and augmented cytotoxicity against tumor cells. In xenograft tumor models, the accumulation of KK-LC-1×CD16A-armed CIML NK cells at the tumor site was significantly increased, resulting in marked suppression of tumor growth and prolonged survival in mice.

Conclusion: KK-LC-1×CD16A-armed CIML NK cells demonstrate significant clinical potential as a multifunctional therapeutic platform, effectively overcoming both tumor infiltration limitations and functional suppression in conventional NK cell therapies. This approach provides valuable insights for optimizing the therapeutic efficacy of CIML NK cell therapy.