Glycogen synthase kinase 3β inhibition sensitizes human glioblastoma cells to temozolomide by affecting O6-methylguanine DNA methyltransferase promoter methylation via c-Myc signaling
| Autor: | Lei Teng, Kazuyuki Kawakami, Jun-ichiro Hamada, Natsuki Furuyama, Ilya V. Pyko, Hemragul Sabit, Аliaksandr S. Fedulau, Toshinari Minamoto, Yutaka Hayashi, Mitsutoshi Nakada |
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| Rok vydání: | 2013 |
| Předmět: |
Cancer Research
Cell Survival Antineoplastic Agents DNA methyltransferase DNA Methyltransferase 3A Proto-Oncogene Proteins c-myc Glycogen Synthase Kinase 3 O(6)-Methylguanine-DNA Methyltransferase GSK-3 Glioma Cell Line Tumor medicine Temozolomide Humans DNA (Cytosine-5-)-Methyltransferases Phosphorylation Promoter Regions Genetic neoplasms GSK3B Protein Kinase Inhibitors Regulation of gene expression Glycogen Synthase Kinase 3 beta Chemistry Drug Synergism General Medicine Methylation DNA Methylation medicine.disease digestive system diseases Dacarbazine Gene Expression Regulation Neoplastic Drug Resistance Neoplasm DNA methylation Cancer research Glioblastoma medicine.drug Protein Binding Signal Transduction |
| Zdroj: | Carcinogenesis. 34(10) |
| ISSN: | 1460-2180 |
| Popis: | Glycogen synthase kinase 3β (GSK3β) is a serine/threonine protein kinase involved in human cancers including glioblastoma. We have previously demonstrated that GSK3β inhibition enhances temozolomide effect in glioma cells. In this report, we investigated the molecular mechanisms of sensitization of glioblastoma cells to temozolomide by GSK3β inhibition, focusing on O(6)-methylguanine DNA methyltransferase (MGMT) gene silencing. Glioblastoma tissues from patients treated with the GSK3β-inhibiting drugs were subjected to immunohistochemistry and methylation-specific PCR assay. Human glioblastoma cell lines T98G, U138, U251 and U87 were treated with a small-molecule GSK3β inhibitor, AR-A014418 or GSK3β-specific small interfering RNA. The combined effect of temozolomide and AR-A014418 on cell proliferation was determined by AlamarBlue assay and an isobologram method. MGMT promoter methylation was estimated by methylation-specific PCR and MethyLight assay. MGMT gene expression was evaluated by real-time quantitative reverse transcriptase-PCR. c-Myc and DNA (cytosine-5)-methyltransferase 3A binding to the MGMT promoter was estimated by chromatin immunoprecipitation assay. GSK3β inhibition decreased phosphorylation of glycogen synthase and reduced MGMT expression and increased MGMT promoter methylation in clinical tumors. In glioblastoma cell lines, GSK3β inhibition decreased cell viability, enhanced temozolomide effect and downregulated MGMT expression with relevant changes in the methylation levels of the MGMT promoter. Here, we showed for the first time that c-Myc binds to the MGMT promoter with consequent recruitment of DNA (cytosine-5)-methyltransferase 3A, regulating the levels of MGMT promoter methylation. The results of this study suggest that GSK3β inhibition enhances temozolomide effect by silencing MGMT expression via c-Myc-mediated promoter methylation. |
| Databáze: | OpenAIRE |
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