Quantitation of O6-Methylguanine-DNA Methyltransferase Gene Messenger RNA in Gliomas by Means of Real-Time RT-PCR and Clinical Response to Nitrosoureas

Tanaka, Satoshi; Oka, Hiroyuki; Fujii, Kiyotaka; Watanabe, Kaoru; Nagao, Kumi; Kakimoto, Atsushi

doi:10.1007/s10571-005-8475-0

Cited by 17 publications

(18 citation statements)

References 14 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…The survival benefits derived from concomitant TMZ therapy were correlated with genetic methylation status of the DNA repair enzyme MGMT 9. Several authors have assessed the impact of MGMT status on outcome and response to other alkylating agents like carmustine in high‐grade gliomas with conflicting results 15, 19‐27. However, to our knowledge, no study has assessed the potential impact of MGMT methylation status in patients with recurrent GBM who received treatment with a biodegradable alkylating agent, namely, carmustine wafers.…”

Section: Discussionmentioning

confidence: 99%

Prognostic impact of O⁶‐methylguanine‐DNA methyltransferase silencing in patients with recurrent glioblastoma multiforme who undergo surgery and carmustine wafer implantation

Métellus¹,

Coulibaly²,

Nanni³

et al. 2009

Cancer

View full text Add to dashboard Cite

BACKGROUND: O6‐methylguanine‐DNA methyltransferase (MGMT) is a key enzyme in the DNA repair process after alkylating agent action. Epigenetic silencing of the MGMT gene by promoter methylation has been associated with longer survival in patients with newly diagnosed glioblastoma multiforme (GBM) who receive alkylating agents. In this study, the authors evaluated the prognostic value of different biomarkers in recurrent GBM and analyzed the changes in MGMT status between primary tumors and recurrent tumors. METHODS: Twenty‐two patients who had recurrent GBM and who underwent surgery with carmustine wafer implantation were enrolled prospectively between 2005 and 2007. The authors investigated the correlation between MGMT silencing in the tumor at recurrence and survival taking into account other clinically recognized prognostic factors. MGMT status was determined by using methylation‐specific polymerase chain reaction analysis, a high‐throughput quantitative methylation assay, and immunohistochemistry. In addition, expression analyses of human mutL homolog 1, human mutS homolog 2, and tumor necrosis factor α‐induced protein 3 at recurrence were conducted with regard to their prognostic impact. RESULTS: The median progression‐free survival (PFS) and overall survival (OS) rates after recurrence were 3.6 months and 9.9 months, respectively, and the 6‐month PFS rate after recurrence was 27.2%. On multivariate analysis, only age (P = .04) and MGMT promoter hypermethylation at recurrence, as determined by MethyLight technology (P = .0012) and methylation‐specific polymerase chain reaction (MSP) analysis (P = .004), were correlated with better PFS. On multivariate analysis, only MGMT promoter hypermethylation at recurrence, as determined by using MethyLight technology (P = .019) and MSP analysis (P = .046), was associated with better OS. CONCLUSIONS: MGMT methylation status was an important prognostic factor in patients with recurrent GBM who underwent surgery plus carmustine wafer implantation; therefore, it was useful in predicting the outcome of GBM therapy at recurrence. Cancer 2009. © 2009 American Cancer Society.

show abstract

Section: Discussionmentioning

confidence: 99%

Prognostic impact of O⁶‐methylguanine‐DNA methyltransferase silencing in patients with recurrent glioblastoma multiforme who undergo surgery and carmustine wafer implantation

Métellus¹,

Coulibaly²,

Nanni³

et al. 2009

Cancer

View full text Add to dashboard Cite

show abstract

“…Enzymatically methylated human male genomic DNA (CpGenome Universal Methylated DNA; Chemicon, Millipore Corp.) and DNA from normal lymphocytes were included in each set of the PCR as methylated and unmethylated controls, respectively. (20,21). Briefly, the real-time PCR reaction mixture was prepared using a TaqMan Universal Master Mix (Applied Biosystems, CA, USA), 120 nM of each MGMT primer, 200 nM probe (5'-CGA GCA GTG GGA GGA GCA ATG AGA-3'), and 2.5 μl of each cDNA sample.…”

Section: Methodsmentioning

confidence: 99%

Gene expression profiling predicts response to temozolomide in malignant gliomas

Yoshino

Ogino²,

Yachi³

et al. 2010

Int J Oncol

View full text Add to dashboard Cite

Abstract. Malignant gliomas are highly lethal neoplasms that cannot be cured with currently available therapies. Temozolomide (TMZ) is a recently introduced alkylating agent that has yielded significant benefits and become a key agent in the treatment of high-grade gliomas. However, its survival benefit remains unsatisfactory. Understanding the molecular basis of TMZ sensitivity/resistance is necessary for improving the treatment outcome by devising strategies that are able to circumvent primary drug resistance. We therefore combined the in vitro TMZ response with microarray gene expression data to identify genes that could potentially be used to predict the response of malignant gliomas to TMZ therapy. We first obtained the individual IC 50 values for TMZ in seven malignant glioma cell lines (A-172, AM-38, T98G, U-87MG, U-138MG, U-251MG and YH-13) and then identified the genes whose expression correlated most highly with TMZ sensitivity employing a cDNA microarray. We present here a list of the most highly up-regulated and down-regulated genes which may be involved in conferring TMZ sensitivity/resistance in malignant gliomas, although most of the genes have not been implicated as a causal factor in the TMZ response except MGMT. We also demonstrated and confirmed the MGMT methylation status, quantitative MGMT mRNA levels, and MGMT protein expression levels in TMZ resistant glioma cells in vitro. Our results are thus consistent with previous studies and suggest that a dominant mechanism conferring sensitivity/resistance to TMZ exists in malignant glioma cells. Although the present study dose have several limitations, our reported candidate genes could represent not only potential molecular markers for TMZ sensitivity/resistance but also chemotherapy targets. Furthermore, the present study could provide a foundation for alternative therapeutic strategies including novel combination treatments that incorporate additional reagents directed at overcoming resistance to TMZ.

show abstract

“…Enzymatically methylated human male genomic DNA (CpGenome Universal Methylated DNA; Chemicon, Millipore Corp.) and DNA from normal lymphocytes were included in each set of the PCR as methylated and unmethylated controls, respectively. (18,50). Briefly, complementary DNA was synthesized from 1 μg total RNA with a random primer (Invitrogen, CA, USA), 40 U M-MLV reverse transcriptase (Invitrogen), 0.5 mM dNTP (Takara Bio, Shiga, Japan), 24 U RNase inhibitor (Takara Bio), 10 μM DTT (Invitrogen) and 5X RT buffer at 37˚C for 60 min.…”

Section: Methodsmentioning

confidence: 99%

Effect of IFN-β on human glioma cell lines with temozolomide resistance

Yoshino

Ogino²,

Yachi³

et al. 2009

Int J Oncol

View full text Add to dashboard Cite

Abstract. Human interferon-ß (IFN-ß) is known to exhibit pleiotropic biological activities including antitumor effects. On the other hand, temozolomide (TMZ), an oral bioavailable alkylating agent with excellent tolerability, has demonstrated efficacy and has become a key therapeutic agent in patients with malignant gliomas; however, its survival benefit remains unsatisfactory. More recent studies have indicated that there might be favorable therapeutic interactions between IFN-ß and TMZ, although the therapeutic advantages of such a combination have not yet been fully explored. The main aim of the present study was to determine whether an antitumor effect could be potentiated by a combination of IFN-ß and TMZ. The antitumor effect of and cell sensitivity to IFN-ß and TMZ and the synergistic potential of IFN-ß and TMZ in combination were evaluated in six malignant glioma cell lines. Correlations among the MGMT methylation status, quantitative level of MGMT mRNA, MGMT protein expression and the antitumor effect of these agents were also evaluated, since one of the most prominent resistance mechanisms to TMZ involves the DNA repair protein MGMT. The cell growth inhibitory effects of IFN-ß and TMZ on all tumor cell lines were observed in a dose-dependent manner, and the human malignant glioma-derived cell lines differed in their sensitivity to TMZ. The MGMT status, including promoter hypermethylation, quantitative mRNA expression and protein expression, was strongly correlated with TMZ sensitivity. A synergistic cell growth inhibitory effect and down-regulated MGMT mRNA levels were significantly observed when a clinically achievable CNS dose of IFN-ß was combined with TMZ, as compared to treatment with IFN-ß or TMZ alone in TMZ-resistant T98G cells. Furthermore, significant amounts of endogenous IFN-ß protein were detected in TMZ-treated T98G cells by ELISA. These results suggest that the clinical therapeutic efficacy of TMZ might be improved by a combination with IFN-ß in malignant gliomas unmethylated at the MGMT gene. The data provide an experimental basis for future strategies in TMZ chemotherapy, although further studies are needed to determine the detailed role of combined IFN-ß and TMZ chemotherapy in increasing tumor sensitivity.

show abstract

Quantitation of O6-Methylguanine-DNA Methyltransferase Gene Messenger RNA in Gliomas by Means of Real-Time RT-PCR and Clinical Response to Nitrosoureas

Cited by 17 publications

References 14 publications

Prognostic impact of O⁶‐methylguanine‐DNA methyltransferase silencing in patients with recurrent glioblastoma multiforme who undergo surgery and carmustine wafer implantation

Prognostic impact of O⁶‐methylguanine‐DNA methyltransferase silencing in patients with recurrent glioblastoma multiforme who undergo surgery and carmustine wafer implantation

Gene expression profiling predicts response to temozolomide in malignant gliomas

Effect of IFN-β on human glioma cell lines with temozolomide resistance

Contact Info

Product

Resources

About

Quantitation of O6-Methylguanine-DNA Methyltransferase Gene Messenger RNA in Gliomas by Means of Real-Time RT-PCR and Clinical Response to Nitrosoureas

Cited by 17 publications

References 14 publications

Prognostic impact of O6‐methylguanine‐DNA methyltransferase silencing in patients with recurrent glioblastoma multiforme who undergo surgery and carmustine wafer implantation

Prognostic impact of O6‐methylguanine‐DNA methyltransferase silencing in patients with recurrent glioblastoma multiforme who undergo surgery and carmustine wafer implantation

Gene expression profiling predicts response to temozolomide in malignant gliomas

Effect of IFN-β on human glioma cell lines with temozolomide resistance

Contact Info

Product

Resources

About

Prognostic impact of O⁶‐methylguanine‐DNA methyltransferase silencing in patients with recurrent glioblastoma multiforme who undergo surgery and carmustine wafer implantation

Prognostic impact of O⁶‐methylguanine‐DNA methyltransferase silencing in patients with recurrent glioblastoma multiforme who undergo surgery and carmustine wafer implantation