Investigation of thioridazine-mediated cell death types in glioma spheres

It has been suggested that monolayer (2B) cell culture method used in classical in vitro studies is inadequate to obtain preliminary data in in vivo studies. Spheroid (3B) cell culture method, which has been introduced in recent years, is a method aimed to form micro-environment and cell-to-cell interaction of cells in in vivo conditions. In this way, the data obtained from in vitro conditions is shown with more similarities to in vivo conditions. Glioblastoma Multiforme (GBM) is the IV Stage Astrocytoma which one of the malignant brain tumors that observed in adults. Despite the treatments through surgical operation, chemotherapy and radiotherapy, the survival rate is 12-15 months. Thioridazine (THZ), an anti-psychotic agent, has been shown to be an anti-cancer agent in the course of its studies. Starting from this, cytotoxic effects of THZ on GBM cell lines T98G and U-87 MG was aimed to determine comparatively in monolayer and spheroid cell cultures. Hanging drop method was used to form spheroid cell cultures. The cytotoxic effect of THZ in monolayer and spheroid cultures was measured by MTT method. In order to determine the mechanism of cell death tests, which are compatible with the Muse cell analyzer and specific inhibitors were used. The IC50 values were determined as 12,67 μM (monolayer T98G 24 hours), 29.30 μM (spheroid T98G 24 hours), 12.80 μM (monolayer U-87 MG 24 hours) 28.68 μM (spheroid U-87 MG 24 hours) as a result of cytotoxicity experiments performed separately on monolayer and spheroid cell cultures. It has been determined that THZ stimulates the cytotoxic effect on monolayer and spheroid cell cultures through the autophagy mechanism by Muse cell analysis method and death pathways specific inhibitors. In our study, it has shown for the first time that cellular effects of THZ differentiate in monolayer and spheroid GBM cultures. It is suggested in the light of this study's findings that in vitro cytotoxicity studies should perform with not only monolayer cell cultures but also with spheroid cell cultures for obtaining data in vivo-like. After all THZ has been shown to be cytotoxic effective in the GBM cell lines via the autophagic pathway, suggesting that this drug is an agent that can be used in the treatment of GBM, which is resistant to apoptosis-inducing chemotherapeutics. Keywords: Spheroid (3B) cell culture, Monolayer (2B) cell culture, Glioblastoma, Thioridazine, Cytotoxicity, Apoptosis, Authophagy.