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BACKGROUND Little is known about the mechanisms that render cancer cells dependent on certain nutrients from the microenvironment. Cysteine is a non-essential amino acid, since it can be synthetized from methionine through the transsulfuration pathway; moreover, cysteine is also uptake from the diet as cystine. We have investigated the metabolism of cysteine in glioma cell lines, and how cysteine/cystine-deprivation alters their antioxidant response in addition to the effect of this nutrient restriction to viability and proliferation in vitro and in vivo. METHODS Cysteine metabolism was investigated through LCMS-based 13C-tracing experiments and the expression levels of key enzymes in the transsulfuration pathway were also explored. Finally, a mouse model of IDH1 mutant glioma was subjected to a cysteine/cystine-free diet and tumor metabolism was analyzed by LCMS. RESULTS Herein, we report the dependence of glioma cells on exogenous cysteine/cystine, despite this amino acid being nonessential. Using several 13C-tracers and analysis of cystathionine synthase and cystathioninase levels, we revealed that glioma cells were not able to upregulate the transulfuration pathway cysteine, which allows methionine to be converted to cysteine in cysteine/cystine deprived conditions. We demonstrated that exogenous cysteine/cystine are crucial for glutathione synthesis, and impact growth and viability. Therefore, we explored the nutritional deprivation in a mouse model of glioma. Animals subjected to a cysteine/cystine-free diet survived longer, with concomitant reductions in glutathione and cysteine plasma levels. At the endpoint higher levels of oxidative stress were detected despite the systemic recovery of cysteine-related metabolites in the plasma. CONCLUSION The results presented herein reveal an alternative therapeutic approach combining cysteine/cysteine-deprivation diets and treatments involving ROS production by limiting the ability of glioma cells to quench oxidative stress through dietary interventions. Our study highlights a time window where cysteine deprivation can be exploited for additional therapeutic strategies.

Background Standard therapy for chordoma consists of surgical resection followed by high-dose irradiation. Protein phosphatase 2A (PP2A) is a ubiquitously expressed serine/threonine phosphatase involved in signal transduction, cell cycle progression, cell differentiation, and DNA repair. LB100 is a small-molecule inhibitor of PP2A designed to sensitize cancer cells to DNA damage from irradiation and chemotherapy. A recently completed phase I trial of LB100 in solid tumors demonstrated its safety. Here, we show the therapeutic potential of LB100 in chordoma. Methods Three patient-derived chordoma cell lines were used: U-CH1, JHC7, and UM-Chor1. Cell proliferation was determined with LB100 alone and in combination with irradiation. Cell cycle progression was assessed by flow cytometry. Quantitative γ-H2AX immunofluorescence and immunoblot evaluated the effect of LB100 on radiation-induced DNA damage. Ultrastructural evidence for nuclear damage was investigated using Raman imaging and transmission electron microscopy. A xenograft model was established to determine potential clinical utility of adding LB100 to irradiation. Results PP2A inhibition in concert with irradiation demonstrated in vitro growth inhibition. The combination of LB100 and radiation also induced accumulation at the G2/M phase of the cell cycle, the stage most sensitive to radiation-induced damage. LB100 enhanced radiation-induced DNA double-strand breaks. Animals implanted with chordoma cells and treated with the combination of LB100 and radiation demonstrated tumor growth delay. Conclusions Combining LB100 and radiation enhanced DNA damage-induced cell death and delayed tumor growth in an animal model of chordoma. PP2A inhibition by LB100 treatment may improve the effectiveness of radiation therapy for chordoma.

The mammalian target of rapamycin (mTOR) has been suggested as a target for radiosensitization. Given that radiotherapy is a primary treatment modality for glioblastoma (GBM) and that mTOR is often dysregulated in GBM, the goal of this study was to determine the effects of AZD2014, a dual mTORC1/2 inhibitor, on the radiosensitivity of GBM stem-like cells (GSCs).mTORC1 and mTORC2 activities were defined by immunoblot analysis. The effects of this mTOR inhibitor on the in vitro radiosensitivity of GSCs were determined using a clonogenic assay. DNA double strand breaks were evaluated according to γH2AX foci. Orthotopic xenografts initiated from GSCs were used to define the in vivo response to AZD2014 and radiation.Exposure of GSCs to AZD2014 resulted in the inhibition of mTORC1 and 2 activities. Based on clonogenic survival analysis, addition of AZD2014 to culture media 1 hour before irradiation enhanced the radiosensitivity of CD133+ and CD15+ GSC cell lines. Whereas AZD2014 treatment had no effect on the initial level of γH2AX foci, the dispersal of radiation-induced γH2AX foci was significantly delayed. Finally, the combination of AZD2014 and radiation delivered to mice bearing GSC-initiated orthotopic xenografts significantly prolonged survival as compared with the individual treatments.These data indicate that AZD2014 enhances the radiosensitivity of GSCs both in vitro and under orthotopic in vivo conditions and suggest that this effect involves an inhibition of DNA repair. Moreover, these results suggest that this dual mTORC1/2 inhibitor may be a radiosensitizer applicable to GBM therapy.