Abstract 891: Therapeutic role of NK cells in treatment of myeloid malignancies using hypomethylating agents

We previously analyzed serial bone marrow (BM) aspirates from a patient with high-risk myelodysplastic syndrome (HR-MDS) who responded to a hypomethylating agent (HMA), azacitidine using single-cell RNA sequencing. Our finding showed an increased proportion of NK cells in the BM, with elevated CXCR4 expression following azacitidine treatment (Hong J, et al. AACR 2023). Building on these observations, we tested our hypothesis that HMA can potentiate BM homing and cytotoxic effect of NK cells in myeloid leukemia. To test our hypothesis, we will carry out in vitro assays, animal experiments, and patients’ serial BM samples. First, we assessed NK cells changes in BM aspirates from AML or HR-MDS patients before and after HMA treatment. Flow cytometric analysis revealed that patients who achieved complete remission after HMA treatment exhibited a significant increase in NK cells proportion in the BM compared to non-responders (add statistics). Additionally, CXCR4 expression on NK cells was higher in patients with a favorable response, suggesting a link between NK cell dynamics and treatment outcomes (add statistics). To carry out mechanistics studies, C1498 mouse AML cells were transplanted into C57/BL6 mice, which were treated with decitabine or vehicle for 5 days. Decitabine not only inhibited in vivo AML proliferation but also increased the proportion of NK cells cells in the BM. Of interest, CXCR4, a chemokine receptor that has been known for BM homing, was also upregulated on the surface of NK cells. In addition, CXCR4 expression was first increased in NK cells, and then NK cell homing was followed. To investigate whether HMA treatment increases CXCR4 expression and induces NK cell migration, an in vitro Transwell migration assay was performed using NK92 cells (a human NK cell line) with AML cell lines (MV4-11 and OCI-AML3) and mesenchymal stromal cells (MSC) from AML patients. CMFDA-labeled NK92 cells showed higher migration and increased CXCR4 expression in decitabine-treated wells compared to vehicle controls, with the effect being more pronounced in the presence of MSCs. Taken together, these findings suggest that HMA-induced NK cell recruitment to the BM via CXCR4 upregulation may play a key role in the anti-leukemic effects of HMAs in AML and HR-MDS. Increased NK cell migration and CXCR4 expression are associated with better treatment responses, highlighting the potential of enhancing NK cell immunity as a therapeutic strategy. Citation Format: Junshik Hong, Suji Min, Jihyun Park, Carly Fielder, Qianni Hu, Seula Kim, Sung-Soo Yoon, Tae Kon Kim. Therapeutic role of NK cells in treatment of myeloid malignancies using hypomethylating agents [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 891.