SNAI2 orchestrates a hypo-oxidative mitochondrial phenotype with elevated FAO dependence in acute myeloid leukemia

Abstract Introduction Acute myeloid leukemia (AML) is a heterogeneous malignancy with poor long-term survival despite intensive therapeutic regimens. Metabolic rewiring has emerged as a hallmark of AML, with leukemia stem cells and chemotherapy-resistant cells exhibiting a preference for mitochondrial oxidative phosphorylation (OXPHOS) and fatty acid oxidation (FAO)(Blood. 2023;141(10):1119-1135). We previously showed that AML metabolism is heterogeneous and influenced by genetic alterations and transcriptional regulation(Leukemia. 2022;36(9):2196-2207). Snail family transcriptional repressor 2 (SNAI2), a zinc finger transcription factor, is a known driver of epithelial–mesenchymal transition. Elevated expression of SNAI2 in AML is associated with poor prognosis and chemoresistance(Leukemia. 2020;34(2):380-390). Analysis of AML datasets revealed that SNAI2 expression positively correlates with metabolic gene signatures. To elucidate the metabolic role of SNAI2 in AML, we conducted integrative multi-omics analyses, metabolic flux measurements, and functional assays. Methods Human AML cell lines expressing either SNAI2 (SNAI2-OE) or an empty vector (EV) control were generated. Targeted metabolomics, transcriptomics, and proteomics were conducted to assess metabolic changes. Differentially expressed metabolites, transcripts, and proteins were subsequently identified; selected targets were validated via qRT-PCR and Western blotting. Seahorse Extracellular Flux assay was used to measure the glycolysis, mitochondrial respiration, and metabolic dependencies on glucose, glutamine, and fatty acids. Intracellular reactive oxygen species (ROS), mitochondrial membrane potential (via TMRE), and mitochondrial mass (via MitoTracker) were measured by flow cytometry. Drug sensitivity assays were conducted to determine the IC₅₀ values for rotenone (an OXPHOS inhibitor) and etomoxir (a FAO inhibitor). Statistical analyses included two-tailed t-tests and mixed-effects models with Benjamini–Hochberg correction. Results SNAI2 overexpression markedly suppressed mitochondrial respiration without altering glycolytic rates in AML cells, indicating that the decline in OXPHOS was not compensated by glycolytic upregulation. Targeted metabolomics revealed the suppression of the tricarboxylic acid (TCA) cycle, with the levels of key intermediates (α-ketoglutarate, succinate, fumarate, and malate) significantly reduced in SNAI2-OE cells. SNAI2-OE cells exhibited decreased ROS levels and mitochondrial membrane potential, despite unchanged mitochondrial mass. RNA-seq and proteomic integration showed consistent downregulation of mitochondrial enzymes—SDHB (complex II), IDH3B, and MDH2—whereas lipid metabolism enzymes (ACSM3 and MID1IP1) were upregulated. These findings were validated using qRT-PCR and immunoblotting. Seahorse substrate oxidation stress test showed a marked reduction in mitochondrial respiration upon FAO inhibition by etomoxir in SNAI2-OE cells, indicating that SNAI2 drives a heightened reliance on fatty acids as respiratory substrates. Consistently, SNAI2-OE cells exhibited heightened sensitivity to both the OXPHOS inhibitor rotenone and the FAO inhibitor etomoxir. These data suggest that SNAI2-driven AML cells exhibit increased metabolic vulnerability due to their reliance on FAO and compromised OXPHOS capacity.Conclusions Our findings uncover a novel metabolic reprogramming mechanism in AML orchestrated by SNAI2. By suppressing TCA cycle activity and OXPHOS while enhancing FAO, SNAI2 enforces a hypo-oxidative, FAO-dependent mitochondrial phenotype. This shift renders SNAI2-high AML cells metabolically fragile and highly susceptible to inhibition of OXPHOS and FAO. Clinically, SNAI2 may serve as a biomarker for identifying AML patients who are likely to benefit from metabolism-targeted therapies. In summary, SNAI2 promotes leukemic progression through metabolic rewiring, thereby exposing novel therapeutic vulnerabilities exploitable for precision medicine in AML.