Abstract A022: Synergizing hypomethylating agents with off-the-shelf CD70-targeted CAR-engineered natural killer T cells for the treatment of acute myeloid leukemia

Abstract Acute myeloid leukemia (AML) is an aggressive hematologic malignancy characterized by the uncontrolled proliferation of immature myeloid precursors in the bone marrow, leading to bone marrow failure and poor clinical outcomes. Despite advances in chemotherapy and hematopoietic stem cell transplantation, relapse and therapeutic resistance remain major challenges. Hypomethylating agents (HMAs) such as azacitidine and decitabine are widely used in AML therapy and are known to reverse aberrant DNA methylation, promote leukemic cell differentiation, and enhance immune recognition. However, their efficacy as single agents remains limited, underscoring the need for combinatorial approaches that can potentiate their immunomodulatory effects. In this study, we discovered that prolonged, low-dose HMA treatment markedly upregulated CD70, CD1d, and multiple natural killer (NK) receptor ligands on AML cells, rendering them more susceptible to immune-mediated cytotoxicity. Leveraging these findings, we developed two CD70-targeting chimeric antigen receptor–engineered invariant natural killer T (CAR-NKT) cell products: cord blood hematopoietic stem and progenitor cell (HSPC)-derived allogeneic CAR70-NKT (AlloCAR70-NKT) cells and peripheral blood mononuclear cell (PBMC)-derived CAR70-NKT (PBMCCAR70-NKT) cells. Both CAR70-NKT products exhibited potent cytotoxicity against AML cells in vitro and demonstrated synergistic antitumor activity when combined with HMAs. Notably, AlloCAR70-NKT cells showed enhanced tumor targeting, multi-antigen recognition, and sustained in vivo persistence compared to PBMCCAR70-NKT cells. In multiple xenograft AML models, AlloCAR70-NKT cells achieved durable tumor clearance without inducing graft-versus-host disease, cytokine release syndrome, or long-term organ toxicity. Together, these findings establish AlloCAR70-NKT cells as a safe, potent, and truly off-the-shelf cellular immunotherapy. When combined with hypomethylating agents, this strategy offers a promising and mechanistically synergistic approach to improve treatment outcomes for patients with AML and potentially other myeloid malignancies. Citation Format: Yan-Ruide Li, Xinyuan Shen, Yuning Chen, Lili Yang. Synergizing hypomethylating agents with off-the-shelf CD70-targeted CAR-engineered natural killer T cells for the treatment of acute myeloid leukemia [abstract]. In: Proceedings of the AACR Immuno-Oncology Conference (AACR IO): Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2026 Feb 18-21; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2026;14(2 Suppl):Abstract nr A022.