Immune-evading universal donor iPSC-derived natural killer cells enable off-the-shelf cancer immunotherapy

Background Natural killer (NK) cells are a promising modality for cancer immunotherapy due to their intrinsic ability to eliminate malignant cells and their safety profile, while their clinical application remains limited by donor variability, immune rejection, and challenges in large-scale manufacturing. Methods We developed a multi–gene-edited universal donor induced pluripotent stem cell (iPSC)-derived NK cell (UDC-NK) platform. First, the engineered iPSC line was generated by disrupting B2M and RFXANK to eliminate HLA class I and II molecules and by introducing a single-chain HLA-E for immune evasion. In addition, a safety switch system was incorporated to ensure controllability. We also developed a novel feeder-free, scalable manufacturing process for cryopreserved UDC-NK cells for off-the-shelf cellular therapy. Results We succeeded in generating large numbers of UDC-NK cells with an activated phenotype. UDC-NK cells exhibited robust cytotoxicity and cytokine production against CD20 + lymphoma cell lines in vitro , which were further enhanced in the presence of Rituximab. UDC-NK cells effectively demonstrated immune evasion from allogeneic T cell– and NK cell–mediated responses. In addition, UDC-NK cells could be selectively eliminated via a herpes simplex virus thymidine kinase–based safety switch upon ganciclovir treatment in vitro . In disseminated B-cell lymphoma models, combination treatment with UDC-NK cells and Rituximab resulted in near-complete tumor growth inhibition and significantly prolonged survival. Conclusions These findings establish UDC-NK cells as a scalable off-the-shelf cell therapy platform with enhanced anti-tumor efficacy and built-in safety control, supporting their potential clinical application in combination with antibody-based cancer therapies.