In Situ Activated NK Cell as Bio‐Orthogonal Targeted Live‐Cell Nanocarrier Augmented Solid Tumor Immunotherapy

Abstract Natural killer (NK) cell‐based immunotherapy holds prominent potential for cancer treatment. However, its application in solid tumors is limited by a rapid decline in viability and function, as well as inadequate homing and infiltration. Herein, a generalized strategy is offered to construct a bio‐orthogonal targeted live‐cell nanocarrier (N₃‐NK‐NPs) by coupling with responsive released interleukin‐21 (IL‐21) nanoparticles (ILNPs) on glyco‐engineered NK cell surfaces. Complementary bio‐orthogonal groups (azide (N₃)/bicyclo [6.1.0] nonyne (BCN), serving as an artificial ligand receptor are separately implanted into NK cells (N₃‐NK) and tumor cells (BCN‐Raji) via nondestructive metabolic glycoengineering. The bio‐orthogonal strategy effectively promotes the specific recognition and migration of NK cells, showing nearly fourfold deeper infiltration in tumor than control groups. Compared with traditional systemic administration, ILNPs hitchhiking on cell vectors selectively in situ releases the IL‐21 adjuvant in the tumor to produce effective and continuous “pseudo‐autocrine” stimulation surrounding NK cells under a low dose (5 µg kg^–1), which greatly promotes proliferation and activation of NK cells, resulting in enhanced therapeutic potential while limiting systemic toxicity. Importantly, live‐cell nanocarrier effectively activates innate immune system through IL‐21 triggered recruitment of multiple immunocytes, significantly improving tumor immune microenvironment. This in situ activated NK cell nanocarrier with bio‐orthogonal targeting provides a universal and powerful strategy for immune cells activation and solid tumor immunotherapy.