[Spermine induces ferroptosis-related phenotype in acute myeloid leukemia cells through reactive oxygen species]

Polyamines, including putrescine, spermidine, and spermine, are widely involved in various physiological processes in vivo. Their metabolic alterations are closely associated with tumor development, and interfering with polyamine metabolism has become a therapeutic strategy under investigation for cancer. However, studies have also demonstrated that spermine can inhibit the growth of castration-resistant prostate cancer, indicating its antitumor effects. This study aimed to investigate the effects of spermine on acute myeloid leukemia (AML) cells. CCK8 assay, cell counting, and lactate dehydrogenase release assay demonstrated that spermine reduced the viability of MOLM-13 and HL-60 cells, inhibited their proliferation, and induced cytotoxicity. Flow cytometry analysis of Annexin V and 7-AAD binding in MOLM-13 cells revealed an increase in the proportion of Annexin V⁺ cells and Annexin V⁺7-AAD⁺ late apoptotic cells as spermine concentration increased. Proteomic and transcriptomic analyses revealed altered expression of ferroptosis pathway-related proteins and genes in MOLM-13 cells following spermine treatment. Using Liperfluo, BDP 581/591 C11, and FerroOrange probes to detect cellular lipid peroxidation and intracellular free Fe²⁺ levels, combined with transmission electron microscopy to observe cell ultrastructure, it was confirmed that spermine induced ferroptosis-related phenotype in MOLM-13 cells; CM-H₂DCFDA and MitoSO^TM Red probes further detected elevated levels of intracellular and mitochondrial reactive oxygen species (ROS), and Western blot showed that spermine caused changes in the expression of ferroptosis-related proteins. Ferroptosis inhibitor liproxstatin-1 partially reversed the spermine-induced ferroptosis-related phenotype in MOLM-13 cells, but did not reverse the decline in cell viability. In contrast, the ROS scavenger N-acetylcysteine completely reversed both the ferroptosis-related phenotype and the reduction in cell viability. These results suggest that spermine induces ferroptosis-related phenotype in AML cells, and its mechanism is associated with ROS generation.