Effect of temozolomide on the U-118 glioma cell line

Carmo,A.; Carvalheiro,H.; Crespo,I.; Nunes,I.; Lopes,M. C.

doi:10.3892/ol.2011.406

Effect of temozolomide on the U-118 glioma cell line

Authors:
- A. Carmo
- H. Carvalheiro
- I. Crespo
- I. Nunes
- M. C. Lopes
View Affiliations

Affiliations: Centre for Neuroscience and Cell Biology, University of Coimbra, 3000 Coimbra, Portugal
Published online on: September 2, 2011 https://doi.org/10.3892/ol.2011.406
Pages: 1165-1170

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Glioblastomas (GBM) are the most lethal subtype of astrocytomas, with a mean patient survival rate of 12 months after diagnosis. The gold standard treatment of GBM, which includes surgery followed by the combination of radiotherapy and chemotherapy with temozolomide (TMZ), increases the survival rate to 14.6 months. The success of TMZ appears to be limited by the occurrence of chemoresistance that allows glioma cells to escape from death signaling pathways. However, the mechanism of TMZ action is yet to be clarified although some controversial results have been reported. Therefore, our aim was to evaluate the occurrence of apoptosis and autophagy in glioma cells treated with TMZ and to correlate TMZ action with the survival pathways Pi3K/Akt and ERK1/2 MAP kinase. Cell proliferation was evaluated by incorporation of bromodeoxyuridine. Apoptosis was studied by flow cytometry as well as by fluorescence confocal microscopy in order to evaluate the sub G0/G1 percentage of cells and chromatin condensation. The expression of the autophagy-associated protein, LC3, as well as Akt and ERK1/2 was performed by Western blotting. In TMZ-treated GBM cells the expression of LC3, the autophagy-associated protein, was increased and only a reduced percentage of cells underwent apoptosis. In addition, we showed that the phosphorylation status of Pi3K/Akt and ERK1/2 MAP kinase was maintained during the treatment with TMZ, suggesting that glioma cells escape from TMZ-induced cell death due to these signaling pathways. The chemoresistance of U-118 cells to TMZ was partially eradicated when cells were simultaneously treated with specific inhibitors of Pi3K/Akt and ERK1/2 MAP kinase signaling pathways and TMZ. Therefore, we hypothesized that in order to induce glioma cell death it is essential to evaluate the activation of the survival pathways and establish a combined therapy using TMZ and inhibitors of those signaling pathways.

View Figures

View References

1	Schwartzbaum JA, Fisher JL, Aldape KD and Wrensch M: Epidemiology and molecular pathology of glioma. Nat Clin Pract Neurol. 2:494–503. 2006. View Article : Google Scholar : PubMed/NCBI
2	Ohgaki H and Kleihues P: Epidemiology and etiology of gliomas. Acta Neuropathol. 109:93–108. 2005. View Article : Google Scholar : PubMed/NCBI
3	CBTRUS. Statistical Report: Primary Brain Tumors in the United States, 2004—2006. Central Brain Tumor Registry of the United States; pp. 1–65. 2010
4	Hadjipanayis CG and van Meir EG: Brain cancer propagating cells: biology, genetics and targeted therapies. Trends Mol Med. 15:519–530. 2009. View Article : Google Scholar : PubMed/NCBI
5	Stupp R, Mason WP, van den Bent MJ, et al: Radiotherapy plus concomitant and adjuvant temozolomide for glioblastoma. N Engl J Med. 352:987–996. 2005. View Article : Google Scholar : PubMed/NCBI
6	Hirose Y, Berger MS and Pieper RO: Abrogation of the Chk1-mediated G(2) checkpoint pathway potentiates temozolomide-induced toxicity in a p53-independent manner in human glioblastoma cells. Cancer Res. 61:5843–5849. 2001.
7	Kanzawa T, Germano IM, Komata T, Ito H, Kondo Y and Kondo S: Role of autophagy in temozolomide-induced cytotoxicity for malignant glioma cells. Cell Death Differ. 11:448–457. 2004. View Article : Google Scholar : PubMed/NCBI
8	Hegi M, Diserens A, Gorlia T, et al: MGMT gene silencing and benefit from temozolomide in glioblastoma. N Engl J Med. 352:997–1003. 2005. View Article : Google Scholar : PubMed/NCBI
9	Keating AK, Kim GK, Jones AE, Donson AM, Ware K, Mulcahy JM, Salzberg DB, Foreman NK, Liang X, Thorburn A and Graham DK: Inhibition of Mer and Axl receptor tyrosine kinases in astrocytoma cells leads to increased apoptosis and improved chemosensitivity. Mol Cancer Ther. 9:1298–1307. 2010. View Article : Google Scholar : PubMed/NCBI
10	Akhavan D, Cloughesy TF and Mischel PS: mTOR signaling in glioblastoma: lessons learned from bench to bedside. Neuro Oncol. 12:882–889. 2010. View Article : Google Scholar : PubMed/NCBI
11	Mrugala MM and Chamberlain MC: Mechanisms of disease: temozolomide and glioblastoma – look to the future. Nat Clin Pract Oncol. 5:476–486. 2008.
12	Hirose Y, Katayama M, Mirzoeva OK, Berger MS and Pieper RO: Akt activation suppresses Chk2-mediated, methylating agent-induced G2 arrest and protects from temozolomide-induced mitotic catastrophe and cellular senescence. Cancer Res. 65:4861–4869. 2005. View Article : Google Scholar : PubMed/NCBI
13	Carmo A, Patricio I, Cruz MT, Carvalheiro H and Lopes MC: CXCL12/CXCR4 promotes motility and proliferation of glioma cells. Cancer Biol Ther. 9:56–65. 2010. View Article : Google Scholar
14	Neves BM, Cruz MT, Francisco V, Goncalo M, Figueiredo A, Duarte CB and Lopes MC: Differential modulation of CXCR4 and CD40 protein levels by skin sensitizers and irritants in the FSDC cell line. Toxicol Lett. 177:74–82. 2008. View Article : Google Scholar : PubMed/NCBI
15	Van Meir EG, Hadjipanayis CG, Norden AD, Shu H, Wen PY and Olson JJ: Exciting new advances in neuro-oncology: the avenue to a cure for malignant glioma. CA Cancer J Clin. 60:166–193. 2010.PubMed/NCBI
16	Sathornsumetee S, Reardon DA, Desjardins A, Quinn JA, Vredenburgh JJ and Rich JN: Molecularly targeted therapy for malignant glioma. Cancer. 110:13–24. 2007. View Article : Google Scholar : PubMed/NCBI
17	Furnari F, Fenton T, Bachoo R, et al: Malignant astrocytic glioma:genetics, biology, and paths to treatment. Genes and Dev. 21:2683–2710. 2007. View Article : Google Scholar : PubMed/NCBI
18	Gladson CL, Prayson RA and Liu WM: The pathobiology of glioma tumors. Annu Rev Pathol Mech Dis. 5:33–50. 2010. View Article : Google Scholar : PubMed/NCBI
19	Hirose Y, Berger MS and Pieper RO: p53 effects both the duration of G2/M arrest and the fate of temozolomide-treated human glioblastoma cells. Cancer Res. 61:1957–1963. 2001.
20	Roos WP, Batista LF, Naumann SC, Wick W, Weller M, Menck CF and Kaina B: Apoptosis in malignant glioma cells triggered by the temozolomide-induced DNA lesion O6-methylguanine. Oncogene. 26:186–197. 2007. View Article : Google Scholar : PubMed/NCBI
21	Ogier-Denisy E and Codogno P: Autophagy: a barrier or an adaptive response to cancer. Biochim Biophys Acta. 1603:113–128. 2003.PubMed/NCBI
22	Mariño G and López-Otín C: Autophagy: molecular mechanisms, physiological functions and relevance in human pathology. Cell Mol Life Sci. 61:1439–1454. 2004.PubMed/NCBI
23	Fu J, Shao C, Chen F, Ng H and Chen Z: Autophagy induced by valproic acid is associated with oxidative stress in glioma cell lines. Neuro Oncol. 12:328–340. 2009. View Article : Google Scholar : PubMed/NCBI
24	Thorburn A: Apoptosis and autophagy: regulatory connections between two supposedly different processes. Apoptosis. 13:1–9. 2008. View Article : Google Scholar : PubMed/NCBI
25	Katayama M, Kawaguchi T, Berger MS and Pieper RO: DNA damaging agent-induced autophagy produces a cytoprotective adenosine triphosphate surge in malignant glioma cells. Cell Death Differ. 14:548–558. 2007. View Article : Google Scholar : PubMed/NCBI
26	Lorente A, Mueller W, Urdangarín E, Lázcoz P, Deimling A and Castresana JS: Detection of methylation in promoter sequences by melting curve analysis-based semiquantitative real time PCR. BMC Cancer. 8:61–75. 2008. View Article : Google Scholar : PubMed/NCBI
27	Ronellenfitsch MW, Brucker DP, Burger MC, Wolking S, Tritschler F, Rieger J, Wick W, Weller M and Steinbach JP: Antagonism of the mammalian target of rapamycin selectively mediates metabolic effects of epidermal growth factor receptor inhibition and protects human malignant glioma cells fromhypoxia-induced cell death. Brain. 13:1509–1522. 2009. View Article : Google Scholar
28	Law BK: Rapamycin: an anti-cancer immunosuppressant? Crit Rev Oncol Hematol. 56:47–60. 2005. View Article : Google Scholar : PubMed/NCBI
29	Meloche S and Pouyssegur J: The ERK1/2 mitogen-activated protein kinase pathway as a master regulator of the G1- to S-phase transition. Oncogene. 26:3227–3239. 2007. View Article : Google Scholar : PubMed/NCBI
30	Tran SE, Holmstrom TH, Ahonen M, Kahari VM and Eriksson JE: MAPK/ERK overrides the apoptotic signaling from Fas, TNF, and TRAIL receptors. J Biol Chem. 276:16484–16490. 2001. View Article : Google Scholar : PubMed/NCBI