MGMT activity, promoter methylation and immunohistochemistry of pretreatment and recurrent malignant gliomas: a comparative study on astrocytoma and glioblastoma

Abstract

The DNA repair protein O 6 -methylguanine-DNA methyltransferase (MGMT) is a key player in tumor cell resistance. Promoter methylation, MGMT activity and immunohistochemistry are used for determining the MGMT status. However, it is unclear whether MGMT promoter methylation correlates with MGMT activity and whether MGMT promoter methylation of the pretreatment tumor predicts the MGMT status of recurrences. To address these questions, we determined MGMT activity promoter methylation and immunoreactivity in pretreatment and recurrent glioblastomas (GB, WHO Grade IV), and in astrocytomas (WHO Grade III). We show that GB that were promoter methylated display a range of 0-62 fmol/mg MGMT and tumors that were nonmethylated 0-423 fmol/mg protein. For astrocytomas, promoter-methylated samples displayed 0-28 fmol/mg and, nonmethylated samples, 23-107 fmol/mg. No correlation was found between the intensity of promoter methylation and MGMT activity. Given a threshold level of 30 fmol/mg of protein, we found a correlation between promoter methylation and no/low MGMT activity in 82.4% of the tumors. This high correlation level was only observed when tumors were excluded showing a hemimethylated promoter (20%). Therefore, classification of hemimethylated tumors remains questionable. Further, we show that 39.1% of pretreatment GB and 5.3% of recurrences were promoter methylated, which is in line with the observed increase of MGMT activity in recurrences. Although individual exceptions were found, the data show an overall correlation between promoter methylation and lack/low MGMT activity in GB and astrocytomas. We also show that promoter methylation assay is superior over immunohistochemistry in determining the MGMT status defined by a given MGMT activity level.Glioblastoma (GB, WHO Grade IV) is the most aggressive brain tumor. For decades, the standard GB treatment was surgery followed by radiotherapy (RT). In 2005, the Phase III trial of Stupp et al.1 showed a significantly longer 2-year survival by applying concomitant and adjuvant treatment with temozolomide (TMZ) plus RT compared with RT alone. Also, the final results of this study with a median follow-up of more than 5 years showed benefits of adjuvant therapy using TMZ and RT.2 Based on these studies, chemoradiotherapy is considered the current standard in GB management. However, despite TMZ, almost 80% of the patients die within the first 2 years after diagnosis, underlining the need for further improved chemotherapy protocols. Hence, intensified TMZ 3 or the combination with other anticancer drugs such as CCNU are currently under investigation in clinical Phase II trials, showing acceptable toxicity and promising survival data. 4 The DNA repair protein O 6 -methylguanine-DNA methyltransferase (MGMT) is presumably the best-established molecular marker that correlates with the response of tumor cells to alkylating agents. MGMT removes methyl and chloroethyl groups from the O 6 position of guanine in a damage reversal reaction (for review see Refs. 5 and 6). If not repaire...

Temozolomide Treatment for Aggressive Pituitary Tumors: Correlation of Clinical Outcome with O6-Methylguanine Methyltransferase (MGMT) Promoter Methylation and Expression

“…In contrast to other published findings (31,32), our data suggest that MGMT promoter methylation and MGMT expression by IHC are not clinically useful in predicting tumor response to temozolomide therapy. The intraoperator heterogeneity in MGMT IHC scores in our series is in line with recent large studies that have evaluated MGMT expression in gliomas; the intraoperator variability seen across the various scoring methods used in these studies reflects the significant intratumor variability of immunoreactivity patterns (multiple regions within a single tumor can vastly differ in regard to the frequency and the relative intensity of the immunoreactivity) (33)(34)(35). Furthermore, our MSP data suggest that MGMT promoter methylation is rare among invasive pituitary tumors and does not explain the wide variability of IHC expression seen in our cohort of tumors.…”

Targeting O 6-methylguanine-DNA methyltransferase with specific inhibitors as a strategy in cancer therapy

“…The same is true for MGMT activity with high MGMT-expressing gliomas ([30 fmol/mg protein) responding poorly to O 6 -alkylating drug-based therapy [29]. Since low MGMT activity in gliomas (asctocytomas WHO III and glioblastomas) is considered to correlate with promoter methylation [37], and several trials showed MGMT promoter methylation correlates with better outcome of therapy (for review, see [38]), the methylation status of the MGMT promoter is currently being used to predict those patients who are likely to have successful temozolomide or combined temozolomide/CCNU/ACNU chemotherapy. It should be noted that, in current glioma therapy, temozolomide is administered concomitantly with radiotherapy (RT), the outcome of which also correlates with MGMT promoter methylation [35].…”

Distinct molecular mechanisms of acquired resistance to temozolomide in glioblastoma cells

“…Few, if any, patients exhibit a change of MGMT promoter methylation status at progression Christmann et al 2010;Felsberg et al 2011).…”