Abstract LB-071: Functional NK cells generated from human iPS cells with 3D-bioreactor for immuno-oncology

NK cells are cytotoxic cells critical for innate immune system. Early investigation using NK cells carrying CAR specific targeting tumor antigens have shown promising antitumor activities. CAR-T therapy requires either autologous or MHC matching cells to avoid graft-verse-host disease (GVHD), whereas allogeneic NK cells showed substantial anti-tumor activity without GVHD. Therefore, the “ready-to-kill” NK population is regarded as the ideal Off-The-Shelf immune cell therapy products. The development of NK-based immunotherapy, however, has been limited by unsatisfactory sources of human NK cells. Human induced pluripotent stem cells (iPSCs) offer unlimited source for manufacturing NK cells suitable for immunotherapy. Previously NK cell derivation methods from human iPS cells used feeder cells with multiple scale-up limited processes which are not suitable for industrial scale production. In this study we described a novel 3D-bioreactor platform that can continuously generate highly pure NK cells with strong cytotoxic activity for 2 weeks. First, undifferentiated human iPS cells were cultured as 3D-spheres with mesoderm inducing conditions in bioreactor, which converted 60-70% iPS cells into hemogenic endothelial progenitor (HEP, CD31 + CD144 + CD34 + ) in a week. HEP cells were then cultured under NK cell differentiation condition for up to 35 days. Starting from day 20, NK cells were started to release from these 3D-spheres which can be harvested daily by centrifugation. These iPS-NK cells can be cryopreserved for long time. Human iPS-NK cells display a distinctively homogenous morphology. Over 95% of collected cells express CD56, NKG2D, NKp44 and NKp46 markers; Approximately 30-50% of CD56 + iPS-NK cells express KIR2DL/DS1 and KIR2DS4 receptors; CD56 + PSC-NK cells do not express TCR and CD3, and CD19, which are specific markers for T and B cells, respectively. Surprisingly, we discovered that more than 80% CD56 + iPS-NK cells also expressed CD8 as compared to ≈30% of peripheral blood CD56 + NK cells, the increase of which is associated with disease regression in AIDS patients. More importantly, when iPS-NK cells and K562 leukemia cells were mixed together, even at a ratio of 1 to 1, more than 80% of leukemia cells were killed within hours as demonstrated both by flow cytometry analysis and time-lapse record. With the establishment of permanent master iPS-CAR cell lines, our novel technology platform will provide inexhaustible cell sources for the generation of truly off-the-shelf CAR-NK cells suitable for treatment of a large numbers of patients, which can revolutionize the field of immuno-oncology. Citation Format: Qiang Feng, Miao-yun Zhang, Shi-Jiang Lu. Functional NK cells generated from human iPS cells with 3D-bioreactor for immuno-oncology [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr LB-071.