Role of epigenetic and metabolic reprogramming in normal and neoplastic hematopoiesis

Rationale: Acute myeloid leukemia (AML) is an aggressive malignancy of hematopoietic stem and progenitor cells (HSPCs), marked by uncontrolled proliferation and blocked differentiation. A rare subtype of acute promyelocytic leukemia (APL), driven by the PLZF::RARA (ZBTB16-RARA) fusion, is resistant to standard therapies such as all-trans retinoic acid (ATRA) and arsenic trioxide (ATO). Recent evidence points to a key interplay between chromatin remodelling and cellular metabolism in regulating normal and leukemic myeloid differentiation. General Objectives: To elucidate the connection between chromatin organization and metabolic regulation in normal and aberrant myelopoiesis, with the aim of identifying novel therapeutic targets in AML. Experimental Design and Methods: We analysed leukemic blasts from patients with PLZF::RARa-APL, PML::RARa-APL, AML, and normal bone marrow (NBM), as well as a ZnSO₄-inducible PLZF::RARa cell line (U937-B412) and its control (U937-MT). Mitochondrial respiration was assessed using the Seahorse XFe96 Analyzer; ROS levels via MitoSOX. Gene/protein expression and functional studies were performed using Q-RT-PCR, Western blot, immunofluorescence, co-immunoprecipitation, and proximity ligation assays (PLA). miR-223 activity on chromatin was examined via chromatin immunoprecipitation after mimic transfection. Apoptosis and proliferation was evaluated via mitochondrial membrane potential (BD™ MitoScreen), cleaved caspase-3/PARP, and MTT assays. Results: AML blasts display a flexible metabolic profile and enhanced mitochondrial capacity compared to normal progenitors, correlating with poor patient survival. In PLZF::RARa-APL, PLZF::RARa enhances mitochondrial respiration and disrupts redox balance, increasing ROS levels. Mechanistically, PLZF::RARa forms a repressive nuclear complex with NRF2, downregulate G6PDH, and reduces expression of SIRT1, a key metabolic and chromatin regulator. miR-223 was found to associate with chromatin regions regulating metabolic genes, including SIRT1. Loss of SIRT1 in B412 cells led to FOXO3a hyperacetylation, impairing SOD1 and Catalase transcription, and increasing oxidative stress sensitivity. High-dose ascorbate (a pro-oxidant) induced apoptosis in PLZF::RARa+ cells. Combined treatment with Azacitidine + Ascorbate or Azacitidine + Venetoclax further enhanced apoptosis and mitochondrial dysfunction. Conclusions: Our study identifies: 1. A pivotal role of metabolic reprogramming in AML biology; 2. A redox vulnerability in PLZF::RARa+ APL, due to antioxidant system impairment.