The present study describes the synthesis and anticancer activity of novel octahedral Pt(IV) complexes with cyclohexyl functionalized ethylenediamine-N,N'-diacetate-type ligands. Molecular mechanics calculations and density functional theory analysis revealed that s-cis is the preferred geometry of these Pt(IV) complexes with tetradentate-coordinated (S,S)-ethylenediamine-N,N'-di-2-(3-cyclohexyl)propanoate. The viability of cancer cell lines (U251 human glioma, C6 rat glioma, L929 mouse fibrosarcoma, and B16 human melanoma) was assessed by measuring mitochondrial dehydrogenase activity and lactate dehydrogenase release. Cell-cycle distribution, oxidative stress, caspase activation, and induction of autophagy were analyzed by flow cytometry using appropriate fluorescent reporter dyes. The cytotoxic activity of novel Pt(IV) complexes against various cancer cell lines (IC(50) range: 1.9-8.7 microM) was higher than that of cisplatin (IC(50) range: 10.9-67.0 microM) and proceeded through completely different mechanisms. Cisplatin induced caspase-dependent apoptosis associated with the cytoprotective autophagic response. In contrast, the new Pt(IV) complexes caused rapid, caspase-independent, oxidative stress-mediated non-apoptotic cell death characterized by massive cytoplasmic vacuolization, cell membrane damage, and the absence of protective autophagy.