Inhibited Glioblastoma Cell Growth Research Articles

Temozolomide induced c-Myc-mediated apoptosis via Akt signalling in MGMT expressing glioblastoma cells

Purpose: We investigated the molecular mechanisms underlying the cytotoxic effect of Temozolomide (TMZ) in both O6-methylguanine-DNA methyl transferase (MGMT) depleted as well as undepleted glioblastoma cell lines. Since TMZ is used in clinics in combination with radiotherapy, we also studied the effects of TMZ in combination with ionising radiation (IR).Methods: Cell colony-forming ability was measured using a clonogenic assay. Cell cycle analysis and apoptosis were evaluated by Flow Cytometry (FCM). Proteins involved in cell cycle control were detected by Western blot and co-immunoprecipitation assays.Results: Our data showed that TMZ, independent of MGMT expression, inhibited glioblastoma cell growth via an irreversible G2 block in MGMT depleted cells or the induction of apoptosis in MGMT normal expressing cells. When TMZ was administered in combination with IR, apoptosis was greater than observed with either agent separately. This TMZ-induced apoptosis in the MGMT expressing cells occurred through Akt/Glycogen-Synthase-Kinase-3ß (GSK3ß) signalling and was mediated by Myelocytomatosis (c-Myc) oncoprotein. Indeed, TMZ phosphorylated/activated Akt led to phosphorylation/inactivation of GSK3ß which resulted in the stabilisation of c-Myc protein and subsequent modulation of the c-Myc target genes involved in the apoptotic processes.Conclusion: c-Myc expression could be considered a good indicator of TMZ effectiveness.

Abstract 3575: Angiotensin-(1-7) and temozolomide provide combinatorial inhibition of glioblastoma cell growth

Abstract Glioblastoma multiforme (GBM), an anaplastic, highly cellular, poorly differentiated tumor, is the predominant malignant brain tumor diagnosed in adults. Standard treatment for glioblastoma consists of surgical resection and radiation with the possible addition of chemotherapy. For example, temozolomide, an orally active imidazotetrazinone pro-drug that exerts its anti-neoplastic effect by interfering with DNA replication, proved to be efficacious in patients with glioblastoma. Even with aggressive therapy, the average survival rate for most patients is less than one year. This poor prognosis suggests a need for novel approaches for the treatment of glioblastoma. Angiotensin-(1-7) [Ang-(1-7)], an endogenous, seven amino acid peptide targeting the mas receptor, significantly reduced the serum-stimulated growth of human G48a glioblastoma cells isolated from a primary, high-grade astrocytoma. Incubation of the glioblastoma cells for 4 days with 100 nM Ang-(1-7) or 10 μM temozolomide caused a greater than 40% reduction in cell growth. More importantly, an additive inhibition (72.0% decrease) of glioblastoma cell growth was obtained by co-incubation of Ang-(1-7) and temozolomide. The combinatorial effect of the two drugs suggests differing mechanisms of action. Treatment of human G48a cells with 100 nM Ang-(1-7) reduced phosphorylation of the mitogen-activated protein kinases, extracellular signal-regulated kinase (ERK)1 and ERK2, by 66.8% and 45.5% (n = 3), respectively, indicating that the anti-proliferative effects may occur, at least in part, through inhibition of the ERK signal transduction pathway. The decrease in ERK activation was associated with a 4.8 ± 1.3-fold increase in the dual-specificity phosphatase DUSP1/MKP-1 (n = 4; p<0.05). This suggests that the heptapeptide regulates DUSP1 to reduce ERK1/ERK2 auto-phosphorylation and activation and inhibit glioblastoma cell growth. Ang-(1-7) treatment also caused a time-dependent decrease in both vascular endothelial growth factor (VEGF) and placental growth factor (PlGF) mRNAs with maximal attenuation at 8 and 16 h, respectively (VEGF - 1.0 ± 0.1 vs. 0.58 ± 0.14; p<0.05; n=5; PlGF - 1.0 ± 0.1 vs. 0.62 ± 0.12; p<0.05; n=5). This reduction correlates with results from our recently completed Phase I clinical trial demonstrating a decrease in plasma PlGF in four patients who exhibited stable disease for greater than 3 months following treatment with Ang-(1-7). Taken together, these data suggest that Ang-(1-7) may serve as a first-in-class, targeted, anti-angiogenic chemotherapeutic agent for glioblastoma. Since Ang-(1-7) exerts its anti-proliferative effects through activation of the G protein-coupled receptor mas, representing a unique mechanism of action distinct from other cell growth modulators, the heptapeptide could be administered singly or with other chemotherapeutic agents to provide synergistic effects. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3575.