Abstract 3151: A novel method to produce clinical scale induced pluripotent stem cell-derived natural killer (iPSC-NK) cells with improved anti-tumor activity for next-generation allogenic cell therapies
Authors: Bernareggi Davide , Gonsalves Caryn , Schabla Max , Garate-Carrilo Alejandra , O’Gorman Eleanor , Chang Nathan , Solchaga Luis , El-Kalay Mohammad , Sandborn William , Kaufman Dan S. , Zhu Huang 
Published: June 2022

Abstract
Natural killer (NK) cells are innate immune cells that play a key role in tumor immune surveillance. NK cells have self-tolerance to healthy cells and can kill allogeneic tumor cells- both hematological and solid malignancies. Recent advances to derive immune cells from human induced pluripotent stem cells (iPSCs) allow the production of NK cells that can be used for immunotherapy. Recent clinical trials suggest that high doses of NK cells (approx. 0.5-1 x 109 cells per dose) and multiple doses are both safe and likely necessary for clinical efficacy. Manufacturing large number of NK cells from a clonal master iPSC line provides a promising strategy to enable off-the-shelf, next generation allogenic cell therapies. To do this, we have developed a novel method to produce clinical scale iPSC-NK cells and improved the process to expand NK cells efficiently and consistently. Briefly, hematopoietic progenitor cells were induced using an improved spin embryoid body (EB) method and the hematopoietic progenitor cells were subsequently differentiated into mature NK cells without cell sorting step as well as stroma cells support. Using this novel protocol, differentiated NK cells expanded more than 3,000-fold to enable us to potentially produce 1 x 1012 iPSC-NK cells starting from 1 x 106 undifferentiated iPSC. This cell production scale can supply &gt;200 doses of 109 cells per dose. Importantly, iPSC-NK cells produced using this method display the typical phenotype of NK cell activating receptor including NKG2D, NKp44, NKp46, and DNAM-1. These iPSC-NK cells demonstrate better anti-tumor activity (against AML) and higher levels of IFNγ and TNFα compared to primary peripheral blood-derived NK cells isolated from healthy donors. Additionally, our iPSC-NK cells demonstrated prolonged “serial killing” of AML cells up to six days. Moreover, this protocol has been tested in large scale using not only unmodified iPSC-NK cells, but also genetically engineered iPSC-NK cells including those with deletion of CISH in iPSC-derived NK cells us that demonstrate improved in vivo anti-tumor activity, in vivo persistence, metabolic fitness, polyfunctionality and resistance to cell exhaustion to create next-generation CAR-NK cell-based immunotherapies. Overall, this novel cell production strategy paves the way for clinical trials using higher doses of more potent iPSC-derived NK cells.
Citation Format: Davide Bernareggi, Caryn Gonsalves, Max Schabla, Alejandra Garate-Carrilo, Eleanor O’Gorman, Nathan Chang, Luis Solchaga, Mohammad El-Kalay, William Sandborn, Dan S. Kaufman, Huang Zhu. A novel method to produce clinical scale induced pluripotent stem cell-derived natural killer (iPSC-NK) cells with improved anti-tumor activity for next-generation allogenic cell therapies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3151.