Restoring NK Cell Cytotoxicity Post‐Cryopreservation via Synthetic Cells

Natural killer (NK) cells are critical components of the first‐line immune defense, responsible for eliminating tumorigenic cells. NK cell‐based adoptive immunotherapy has gained increasing attention; however, cryopreservation, a standard technique for NK cell storage, significantly impairs NK cell cytotoxicity, particularly in physiological 3D environments. Here, we demonstrate that short‐term co‐culture with effector T cells markedly enhances NK cell motility and killing functionality. Notably, a brief 1‐day co‐culture is sufficient to restore cryopreservation‐impaired NK cell functionality in 3D environments. This enhancement requires direct contact between T cells and NK cells, which facilitates localized high concentrations of IL‐2 at the cell contact sites. To develop a controled, donor‐independent solution, we demonstrate that synthetic T cells with surface‐bound IL‐2 exhibit superior efficiency in revitalizing cryopreserved NK cells. These findings uncover a previously unrecognized role for physical contact‐mediated local IL‐2 signaling and provide an efficient, cost‐effective, and tunable strategy to rescue NK cell functionality post‐cryopreservation, paving the way for more scalable, potent, and clinically viable NK cell‐based immunotherapies.