Abstract 553: Combined use of human iPSC-derived natural killer cells with macrophages and anti-CD47 blockade to improve killing of acute myeloid leukemia
Authors: Goldenson Benjamin , Fierro Manuel , Pouyanfard Somayeh , Kaufman Dan S. 
Published: June 2022

Abstract
Despite many recent advances, treatment of acute myeloid leukemia (AML) remains challenging. To generate an improved cell-based therapy against AML, our group has produced NK cells from induced pluripotent stem cells (iPSCs). iPSC-derived NK cells effectively kill AML cells, but may benefit from combination with other therapies. Based on studies showing that targeting the CD47 pathway on macrophages (Mϕ) improves anti-tumor activity in AML, we investigated the combination of iPSC-NK cells with iPSC-Mϕ with and without CD47 blockade for the treatment of AML. To determine if the addition of iPSC-Mϕ can improve the cytotoxicity of NK cells against AML blasts, we co-cultured iPSC-NK cells and MOLM13 or MV-4-11 AML cells with iPSC-Mϕ in standard cytotoxicity assays. While Mϕ alone did not kill AML blasts, the addition of iPSC-Mϕ to iPSC-NK cells significantly improved killing of AML blasts by 50% (p&lt;0.01). Addition of an anti-CD47 mAb (B6H12) further increased killing of AML blasts by an additional 23% (p&lt;0.01). Addition of the CD47 mAb to iPSC-Mϕ without NK cells did not result in increased killing. We also demonstrated that blockade of SIRPα (receptor for CD47) significantly increased NK cell killing of AML blasts by 16% (p&lt;0.05). Furthermore, the combination of iPSC-NK cells + iPSC-Mϕ + SIRPα mAb led to a 37% increase in cytotoxicity compared to iPSC-NK cells + iPSC-Mϕ alone (p&lt;0.01). To confirm that addition of anti-CD47 or anti-SIRPα antibodies increased killing via loss of the inhibitory CD47-SIRPα interaction on NK cells and not via another mechanism, we tested the effect of adding anti-CD47 mAb and SIRPα antibodies simultaneously. Compared to addition of either mAb alone, the combination induced no additional increase in anti-AML activity by the NK cells. We also found the addition of the SIRPα mAb to iPSC-Mϕ alone did not increase cytotoxicity. We next evaluated if antibody dependent cellular cytotoxicity (ADCC) mediated by anti-CD47 mAb binding AML cells could account for the increase in cytotoxicity and found that blockade of Fc-receptors on NK cells does not diminish the increase in cytotoxicity, indicating no role for ADCC. Finally, we showed that direct contact between NK cells and Mϕ is required to increase killing of AML in a transwell assay. Separation of iPSC-Mϕ from iPSC-NK cells by a transwell insert abrogated the increase in anti-AML cytotoxicity seen with addition of iPSC-Mϕ to NK cells without the insert. In vivo studies testing the combination of iPSC-NK cells, iPSC-Mϕ and CD47 mAb against human AML in mouse xenograft-models are ongoing. Together our results indicate that CD47 blockade combined with iPSC-Mϕ leads to increased NK cell-mediated anti-tumor activity. iPSC-NK cells and iPSC-Mϕ provide an important, standardized, allogeneic cell therapy approach.
Citation Format: Benjamin Goldenson, Manuel Fierro, Somayeh Pouyanfard, Dan S. Kaufman. Combined use of human iPSC-derived natural killer cells with macrophages and anti-CD47 blockade to improve killing of acute myeloid leukemia [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 553.