Cell-specific effects of methoxy-substituted iron(III) salophene complexes

[N,N'-Bis(salicylidene)-1,2-phenylenediamine]iron(III) complexes bearing methoxy substituents at the 3-, 4-, 5-, or 6-positions of the salicylidene moieties (C1 - C4) were previously shown to reach maximum cytotoxic activity within 48 h. This raised the question of cellular processes initiated during shorter incubation times. Therefore, the effects of C1 - C4 were evaluated after 24 h in MDA-MB 231 breast cancer, HL-60 acute myeloid leukemia, and nontumorigenic MCF-10A mammary epithelial cells using cell-based and mitochondria-related assays. All complexes accumulated in MDA-MB 231 and HL-60 cells within 4 h. Despite efficient uptake, the 4-methoxy-substituted complex C2 showed minimal biological activity after 24 h, whereas C1, C3, and C4 induced pronounced, cell type-dependent effects. In MDA-MB 231 cells, these complexes caused mitochondrial membrane depolarization and increased reactive oxygen species (ROS) levels in both mitochondria and cytosol. The effect on oxygen consumption of MDA-MB 231 cells was relatively low. In contrast, C1, C3, and C4 significantly impaired mitochondrial respiration of HL-60 cells. While all three complexes increased cytosolic ROS levels within 4 h, only C4 elevated mitochondrial ROS in HL-60 cells. After 24 h, cell death - induced by both apoptosis and necrosis - was detected in both malignant cell lines. On the contrary, nontumorigenic MCF-10A cells were largely unaffected, with only C1 reducing viability at the highest concentration tested. This study shows that C1, C3, and C4 rapidly induce distinct, cell-specific mitochondrial responses that contribute to early cytotoxic effects, whereas C2 remains largely inactive despite efficient cellular uptake.