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Glioblastoma (GBM) is the most frequent malignant glioma. Treatment of GBM patients is multimodal with maximum surgical resection, followed by concurrent radiation and chemotherapy with the alkylating drug temozolomide (TMZ). The present study aims to identify genes implicated in the acquired resistance of two human GBM cells of astrocytic origin, T98G and U373, to TMZ. Resistance to TMZ was induced by culturing these cells in vitro for months with incremental TMZ concentrations up to 1 mM. Only partial resistance to TMZ has been achieved and was demonstrated in vivo in immunocompromised mice bearing orthotopic U373 and T98G xenografts. Our data show that long-term treatment of human astroglioma cells with TMZ induces increased expression of facilitative glucose transporter/solute carrier GLUT/SLC2A family members, mainly GLUT-3, and of the AKR1C family of proteins. The latter proteins are phase 1 drug-metabolizing enzymes involved in the maintenance of steroid homeostasis, prostaglandin metabolism, and metabolic activation of polycyclic aromatic hydrocarbons. GLUT-3 has been previously suggested to exert roles in GBM neovascularization processes, and TMZ was found to exert antiangiogenic effects in experimental gliomas. AKR1C1 was previously shown to be associated with oncogenic potential, with proproliferative effects similar to AKR1C3 in the latter case. Both AKR1C1 and AKR1C2 proteins are involved in cancer pro-proliferative cell chemoresistance. Selective targeting of GLUT-3 in GBM and/or AKR1C proteins (by means of jasmonates, for example) could thus delay the acquisition of resistance to TMZ of astroglioma cells in the context of prolonged treatment with this drug.

Gliomas account for more than 50% of all primary brain tumors. The worst prognosis is associated with gliomas of astrocytic origin, whereas gliomas with an oligodendroglial origin offer higher sensitivity to chemotherapy, especially when oligodendroglioma cells display 1p19q deletions. Temozolomide (TMZ) provides therapeutic benefits and is commonly used with radiotherapy in highly malignant astrocytic tumors, including glioblastomas. The actual benefits of TMZ during long-term treatment in oligodendroglioma patients have not yet been clearly defined. In this study, we have investigated the effects of such a long-term TMZ treatment in the unique Hs683 oligodendroglioma model. We have observed increased TMZ sensitivity of Hs683 orthotopic tumors that were previously treated in vitro with months of progressive exposure to increasing TMZ concentrations before being xenografted into the brains of immunocompromised mice. Whole-genome and proteomic analyses have revealed that this increased TMZ sensitivity of Hs683 oligodendroglioma cells previously treated for long periods with TMZ can be explained, at least partly, by a TMZ-induced p38-dependant dormancy state, which in turn resulted in changes in amino acid metabolism balance, in growth delay, and in a decrease in Hs683 oligodendroglioma cell-invasive properties. Thus, long-term TMZ treatment seems beneficial in this Hs683 oligodendroglioma model, which revealed itself unable to develop resistance against TMZ.

We have previously reported that galectin 1 (Gal-1) plays important biological roles in astroglial as well as in oligodendroglial cancer cells. As an oligodendroglioma model, we make use of the Hs683 cell line that has been previously extensively characterized at cell biology, molecular biology, and genetic levels. Galectin 1 has been shown to be involved in Hs683 oligodendroglioma chemoresistance, neoangiogenesis, and migration. Down-regulating Gal-1 expression in Hs683 cells through targeted small interfering RNA provokes a marked decrease in the expression of the brain-expressed X-linked gene: BEX2. Accordingly, the potential role of BEX2 in Hs683 oligodendroglioma cell biology has been investigated. The data presented here reveal that decreasing BEX2 expression in Hs683 cells increases the survival of Hs683 orthotopic xenograft-bearing mice. Furthermore, this decrease in BEX2 expression impairs vasculogenic mimicry channel formation in vitro and angiogenesis in vivo, and modulates glioma cell adhesion and invasive features through the modification of several genes previously reported to play a role in cancer cell migration, including MAP2, plexin C1, SWAP70, and integrin β6. We thus conclude that BEX2 is implicated in oligodendroglioma biology.

Purpose: The aims of the present work were to investigate the in vitro and in vivo antiangiogenic effects of chronic temozolomide treatment on various glioma models and to demonstrate whether bevacizumab (Avastin) increased the therapeutic benefits contributed by temozolomide in glioma. Experimental Design: The expression levels of various antiangiogenic factors in four glioma cell lines were evaluated after chronic in vitro treatment with temozolomide by Western blot. Proliferation and migration assays were performed on human endothelial cells incubated with supernatants of glioma cells treated with and without temozolomide. Orthotopic glioma models were used to evaluate the antiangiogenic effects of temozolomide in vivo and the therapeutic benefits of different temozolomide treatment schedules used alone or in combination with bevacizumab. Results: Temozolomide, a proautophagic and proapoptotic drug, decreased the expression levels of HIF-1α, ID-1, ID-2, and cMyc in the glioma models investigated, all of which playing major roles in angiogenesis and the switch to hypoxic metabolism. These changes could be, at least partly, responsible for the impairment of angiogenesis observed in vitro and in vivo. Moreover, combining bevacizumab with temozolomide increased the survival of glioma-bearing mice in comparison to each compound administered alone. Conclusions: In addition to the numerous mechanisms of action already identified for temozolomide, we report here that it also exerts antitumor effects by impairing angiogenic processes. We further emphasize that bevacizumab, which is an antiangiogenic drug with a different mechanism of action, could be useful in combination with temozolomide to increase the latter's therapeutic benefit in glioma patients.