Combined Effects of Fluoxetine and Temozolomide on Apoptosis, Autophagy and Migration in Human Glioblastoma Cells

Abstract

Glioblastoma multiforme (GBM) is an aggressive central nervous system tumor associated with poor prognosis and resistance to conventional therapies. This study investigated the cytotoxic and mechanistic effects of the selective serotonin reuptake inhibitor fluoxetine (FLT), alone and in combination with the alkylating agent temozolomide (TMZ), in human glioblastoma T98-G cells and non-tumorigenic human embryonic kidney HEK-293 cells. Cell viability, apoptotic cell death, cell migration, autophagic activity, and mitochondrial membrane potential were assessed using the MTT assay, Annexin V-FITC/PI staining, wound healing assay, monodansylcadaverine (MDC), and JC-1 staining, respectively. Quantitative image analyses were performed with ImageJ, and drug interaction effects were assessed using ComboSyn software. FLT significantly reduced cell viability in T98-G cells (IC₅₀: 36 µM), whereas HEK-293 cells exhibited markedly lower sensitivity (IC₅₀: 320 µM). In glioblastoma cells, FLT and TMZ, particularly in combination, predominantly induced late-stage apoptosis, while early apoptotic responses were more pronounced in HEK-293 cells. Combined treatment resulted in 36.4% late apoptosis in T98-G cells and 16.5% early apoptosis in HEK-293 cells. Both agents significantly inhibited cell migration, with the strongest suppression observed following combination treatment. At 24 hours, all treatments induced autophagic alterations and mitochondrial membrane depolarization. At 48 hours, autophagic activity persisted mainly in single-agent treatments, whereas mitochondrial dysfunction remained evident across all groups. Overall, FLT exerts cytotoxic effects in glioblastoma cells through apoptosis induction, mitochondrial dysfunction, modulation of autophagy, and inhibition of cell migration. These effects are enhanced when combined with TMZ, suggesting that fluoxetine may represent a promising adjuvant candidate for GBM therapy and warrants further in vivo investigation.