Mide. MGMT straight demethylates O6-meG and is downregulated in about
Mide. MGMT straight demethylates O6-meG and is downregulated in about 45 of glioblastoma patients with MGMT promoter methylation within the tumor and enhanced temozolomide sensitivity [15]. A reported mechanism of temozolomide chemosensitization by disulfiram has been identified in pituitary adenoma stem-like cells [51] and in glioblastoma cell lines [44]: disulfiram covalently modifies MGMT, major for the proteasomal degradation of the DNA repair enzyme. Furthermore, disulfiram has been proposed in glioblastoma spheroid cultures to facilitate the DNA-damaging temozolomide impact by impairing DNA repair [12]. Temozolomide-mediated DNA DSBs reportedly trigger a G2 /M arrest of cell cycle [55]. In our present experiments (see Figures four and 5), a temozolomide-mediated G2 /M arrest couldn’t be SIRT2 Inhibitor list detected in unirradiated LK7 and LK17 cells. Given the doubling times of exponentially increasing LK7 and LK17 pGSCs in NSC medium of 1.7 and 1.0 days, respectively, (see Figure 1C) it could be assumed that all cells (LK17) or possibly a substantial fraction of cells (LK7) underwent two rounds of DNA replication (necessary for temozolomidetriggered MMR-mediated DNA damage) in the course of the selected incubation period (48 h) on the flow cytometry experiments (see Figures 4 and five). Furthermore, temozolomide in the chosen concentration (30 ) has been demonstrated in our previous experiments to exert a high tumoricidal effect in MGMT promotor-methylated pGSCs (unpublished personal observations). As a result, the flow cytometry data on cell cycle and cell death of the present study confirms the relative temozolomide resistance of MGMT promoter-unmethylated glioblastoma. This was also evident from the statistically insignificant effects of temozolomide on mGluR2 Agonist manufacturer clonogenic survival in each pGSC cultures (see Figures 6A and 7A). Even though confirming the tumoricidal action of disulfiram/Cu2+ in temozolomide-resistant glioblastoma stem-cell cultures, our present study did not observe a temozolomidesensitizing effect of disulfiram/Cu2+ (see Figures 6A and 7A). Rather the contrary, in both cell models, temozolomide markedly or had a tendency to attenuate the inhibitoryBiomolecules 2021, 11,16 ofeffect of disulfiram on clonogenic survival. Such a disulfiram effect-diminishing action of temozolomide was also suggested by our flow cytometry experiments on the cell cycle (see Figures 4 and five). One might speculate that temozolomide interferes with lethal pathways triggered by disulfiram. Independent of the underlying molecular mechanisms, the present observations do not assistance future therapy approaches pursuing a concomitant disulfiramtemozolomide chemotherapy. Also, this observation suggests that the tumoricidal effect of disulfiram may well be sensitive to pharmaco-interactions with co-medications. The understanding of such pharmaco-interactions, even so, is usually a prerequisite for the success of future clinical trials using disulfiram for second-line therapy in glioblastoma sufferers with tumor progression in the course of temozolomide upkeep therapy. The evaluation from the molecular mechanism of such pharmaco-interactions (right here, the temozolomide-disulfiram interaction), having said that, goes beyond the scope in the present study. four.two. Disulfiram as a Radiosensitizer Likewise, our present study did not identify any radiosensitization of each glioblastoma stem-cell cultures by disulfiram/Cu2+ . This is in seeming contrast to earlier research that show a disulfiram/Cu2+ -mediated radiosensitization in patient-derived spheroid glioblas.
