Abstract 4053: Disulfiram-copper combination induces cell death time- and schedule-dependently in glioblastoma (GBM) and medulloblastoma cell lines, and enhances temozolomide sensitivity of methylator+ve GBM cells in vitro and in vivo
Authors: Gabr Mostafa , Marcus Stephen , Madala Hanumantha R. , Srivenugopal Kalkunte S. , Ali-Osman Francis 
Published: July 2017

Abstract
Background. Disulfiram (DSF), a carbamate derivative and a potent aldehyde dehydrogenate inhibitor, is used clinically to treat alcoholism. Although, relatively nontoxic, DSF exhibits significant anticancer effects, when administered with heavy metals (Cu and Zn). Because of this and the fact that DSF crosses the blood-brain barrier, DSF is being investigated as a potential agent for brain tumor therapy. In a previous study, we showed that, in glioblastoma (GBM) cells, a copper-DSF complex inhibited activity of the DNA repair protein, O6methyguanine-DNA methyltransferase, MGMT, involved in tumor resistance to temozolomide. This study investigates, in greater detail, the anti-tumor efficacy of DSF and copper (as the gluconate), alone and in combination, in a series of GBM cell lines (UW18, UW28, UW35, UW14 and MGR3) and one medulloblastoma cell line (UW228). We examined the dose-, time- and schedule-dependence of the anti-tumor activity (growth rates in a real-time cell, doubling times and apoptotic induction) and correlated the results with cellular levels of caspase activation, production of reactive oxygen species (ROS) and activated MAP kinases. The effects of DSF, copper and DSF-copper on temozolomide resistance was examined in vitro and in vivo in two methylator+ve GBM cell lines, UW18 and UW28.
Results. DSF and copper, as single agents, up to 100 uM for 4-7 days, had little to no toxicity on any of the GBM cell lines. In contrast, all the cell lines were highly sensitive to DSF, when combined with copper. The efficacy of the combination was highly dependent on the time (minimum 6 hrs) and schedule (DSF prior to copper) of cellular exposure. The UW228 medulloblastoma cell line was approx. 4-fold more sensitive to DSF and to the DSF-copper combination than GBM cells. We observed a direct correlation between the levels of cellular ROS production and MAP activation, and the level of cytotoxicity of the DSF-copper combination. The temozolomide sensitivity of the cell lines was significantly enhanced by the DSF-copper combination and in vivo xenograft growth of two temozolomide-resistant, methylator+ve GBM cell lines was significantly inhibited and their temozolomide sensitivity enhanced by the DSF-copper combination and was associated with significant inhibition of MGMT in the tumor cells.
Conclusion: The results demonstrate significant sensitivity of GBM and, for the first time, medulloblastoma, cells to DSF-copper, which is dependent on both the duration of cellular exposure to and the scheduling of DSF and copper. The DSF-copper combination increases the sensitivity of temozolomide-resistant, methylator+ve GBM cells to temozolomide. The findings support DSF-copper as a treatment strategy for GBM and medulloblastoma.
Citation Format: Mostafa Gabr, Stephen Marcus, Hanumantha R. Madala, Kalkunte S. Srivenugopal, Francis Ali-Osman. Disulfiram-copper combination induces cell death time- and schedule-dependently in glioblastoma (GBM) and medulloblastoma cell lines, and enhances temozolomide sensitivity of methylator+ve GBM cells in vitro and in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4053. doi:10.1158/1538-7445.AM2017-4053