EACR25-3169

DHODH inhibition overcomes macrophage-mediated resistance to cytarabine in acute myeloid leukemia (AML)

C. Wang1, Y. Wang1, C. Benetti1, P. Hyroššová2, X. Lin1, E. Ayoub3, J. Rohlena2, M. Andreeff3, K. Rohlenova2, S. Bhatt1,4
1National University of Singapore, Department of Pharmacy and Pharmaceutical Sciences, Singapore, Singapore
2Czech Academy of Sciences, Institute of Biotechnology, Prague-West, Czech Republic
3The University of Texas MD Anderson Cancer Center, Department of Leukemia, Houston, United States
4Emory University, Department of Hematology and Medical Oncology, Atlanta, United States
Introduction:

The standard induction therapy for acute myeloid leukemia (AML) consists of 7 days of cytarabine (AraC) and 3 days of an anthracycline (the “7+3” regimen). Although this regimen induces remission in many patients, most eventually relapse due to chemoresistance. Macrophages, which are abundant in both solid and hematologic malignancies, have been implicated in therapeutic resistance, but their specific role in AML remains unclear.

Material and method:

To investigate the role of macrophages in AML chemoresistance, we analyzed paired diagnosis and relapse samples from six AML patients using single-cell RNA sequencing. To assess the functional contribution of macrophages to AraC resistance, we depleted macrophages in both AML patient-derived xenograft (PDX) and syngeneic mouse models. In vitro experiments were used to assess the effects of macrophage-derived metabolites on AraC resistance. Global metabolomic profiling identified resistance-associated metabolites. Genetic and pharmacological inhibition of key metabolic enzymes was used to explore strategies to overcome AraC resistance. Finally, we tested the efficacy of combining DHODH inhibitors with AraC in vivo using mouse AML models.

Result and discussion:

Relapsed AML samples showed increased monocyte/macrophage abundance. Depletion of macrophages sensitized AML cells to AraC in both PDX and mouse syngeneic models. In vitro, macrophage-conditioned media induced AraC resistance via secretion of small (<3 kDa) soluble factors. Metabolomics identified deoxycytidine (dC) as the key resistance-inducing metabolite. Mechanistically, dC inhibits deoxycytidine kinase (DCK), impairing AraC activation in AML cells. Further, we found that macrophages express high levels of SAM domain and HD domain-containing protein 1 (SAMHD1), a critical enzyme in dC biosynthesis. Inhibiting SAMHD1 or dihydroorotate dehydrogenase (DHODH) reduced dC levels and restored AraC sensitivity in vitro. In vivo, DHODH inhibitors Brequinar and Leflunomide decreased dC concentrations in bone marrow and enhanced AraC efficacy.

Conclusion:

Our study identifies macrophage-derived dC as a novel mediator of AraC resistance in AML. Targeting pyrimidine metabolism in macrophages may provide a strategy to overcome chemoresistance and improve therapeutic outcomes in AML patients.