Japanese Journal of Clinical Oncology Advance Access originally published online on December 21, 2007
Japanese Journal of Clinical Oncology 2007 37(12):897-906; doi:10.1093/jjco/hym132
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
© 2007 Foundation for Promotion of Cancer Research
Prognostic Significance of O6-Methylguanine-DNA Methyltransferase Protein Expression in Patients with Recurrent Glioblastoma Treated with Temozolomide
1 Department of Neurosurgery, Kyorin University School of Medicine, Tokyo
2 Department of Neuro-Oncology, Saitama Medical University International Medical Center, Hidaka, Saitama
3 Brainpier Minamiohta, Ibaragi, Japan
For reprints and all correspondence: Motoo Nagane, Department of Neurosurgery, Kyorin University School of Medicine, 6-20-2 Shinkawa, Mitaka, Tokyo 181-8611, Japan. E-mail: nagane-nsu{at}umin.ac.jp
Received August 8, 2007; accepted September 10, 2007
 |
Abstract |
|---|
 TOP Abstract INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONAcknowledgementsReferences |
|---|
Background: Temozolomide (TMZ) is active against newly diagnosed glioblastoma (GBM), and O6-methylguanine-DNA methyltransferase (MGMT) is implicated in resistance to TMZ and nitrosoureas. We evaluated the efficacy and safety of the standard 5-day TMZ regimen in patients with recurrent GBM after initial therapy including nitrosourea-based chemotherapy, in conjunction with an analysis of the prognostic value of MGMT protein expression regarding response to TMZ and survival.
Methods: From September 2003 to January 2007, 30 patients having recurrent GBM received 150–200 mg/m2/day of TMZ for five consecutive days every 28 days. Tumor tissue from 19 patients was analysed for MGMT protein expression using western blotting, and 17 of them were assessable for a response.
Results: The overall response rate was 23.5% (one complete response and three partial responses). Six patients had stable disease (35.3%). Median progression-free survival (PFS) time was 2.2 months, and median overall survival (OS) time was 9.9 months from the initiation of TMZ therapy. Patients with low MGMT protein expression had a significantly improved PFS (P = 0.016) and OS (P = 0.019) compared to those with high expression. Both low MGMT expression (P = 0.040) and re-resection at relapse (P = 0.014) persisted as significant independent favorable prognostic factors for OS. The most common grade 3 and 4 hematological toxicity was lymphopenia (22.2%).
Conclusions: The standard 5-day TMZ regimen resulted in moderate antitumor activity with an acceptable safety profile in patients with nitrosourea-pretreated recurrent GBM, and protein expression of MGMT is an important prognostic factor for patients treated with TMZ even after recurrence.
Key Words: glioblastoma • temozolomide • O6-methylguanine-DNA methyltransferase • western blot • recurrence
|
INTRODUCTION |
|---|
|
TOPAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONAcknowledgementsReferences |
|---|
Patients with malignant gliomas, the most common primary neoplasms arising in the central nervous system, have yet a poor prognosis, despite multimodal intensive treatments including maximum surgical resection, irradiation and chemotherapy. The median survival time (MST) of patients with glioblastoma multiforme (GBM), the most malignant form of glioma, remains 12–15 months from initial diagnosis (1). This disappointing outcome results, at least in part, from low responsiveness to chemotherapy. The therapeutic benefit of chemotherapy using nitrosoureas, alkylating agents such as carmustine, lomustine and nimustine, has been controversial, even in combination with procarbazine, lomustine and vincristine (PCV), until the Medical Research Council reported a large-scale meta-analysis of 12 selected randomized clinical trials demonstrating a significant improvement in risk of progression and survival (2). However, the survival benefit remained unsatisfactory, with only a 15% reduction in the hazard ratio.
Temozolomide (TMZ) is an alkylating agent that is rapidly absorbed after oral administration, and penetrates well into the cerebrospinal fluid at a concentration up to 40% of that measured in plasma (3–6). TMZ has been shown to have clinical antitumor activity against malignant gliomas and a relatively good safety profile (7,8). A recent phase III clinical trial conducted by the European Organisation for Research and Treatment of Cancer (EORTC) and the National Cancer Institute of Canada (NCIC) demonstrated that concomitant radiotherapy (RT) plus TMZ followed by six cycles of TMZ significantly prolonged survival at the hazard ratio of 65% compared with RT alone in patients with newly diagnosed GBM (9). The RT plus concomitant and adjuvant TMZ regimen has thus been considered as the new standard care for this group of patients. However, no standard of care exists for recurrent GBM yet.
The mechanism of action by TMZ is thought to be methylation at the O6 position of guanine in DNA, with additional methylation at the N7 position (10,11). These alkylating lesions are effectively repaired by a DNA repair enzyme O6-methylguanine-DNA methyltransferase (MGMT). MGMT catalyzes the stoichiometric, covalent transfer of the alkyl group to an internal cysteine residue, and is finally inactivated (12). A direct relationship between MGMT activity or expression and resistance to alkylating nitrosoureas, which also exert their antitumor effects by alkylating the O6 position of guanine, has been well documented in cell lines and xenografts derived from a variety of human tumors, including gliomas (13,14). Likewise, a significant correlation was recently found between methylation status of the MGMT gene promoter determined by the methylation-specific polymerase-chain reaction (PCR) (MSP) assay and survival of patients with newly diagnosed GBM treated with RT plus concomitant and adjuvant TMZ (15). The methylation of the promoter region of MGMT gene has been shown to be a major mechanism to turn off gene transcription, thus is speculated to reduce the intracellular level of MGMT expression (16). MGMT protein expression determined by immunohistochemistry (IHC) was also found to correlate with survival or response to TMZ treatment in newly diagnosed GBM patients (17,18). However, other clinical studies failed to show any relationship between MGMT promoter methylation status and survival or response to TMZ, even for dose-intensified regimens, in patients with recurrent GBM (19,20). These contradictory results could be due to the timing of TMZ administration, i.e. at initial diagnosis versus at recurrence, or might be dependent on assay methods for detection of MGMT status (21,22).
Herein, we report the efficacy and safety of the standard 5-day on/28-day cycle regimen of TMZ in patients with recurrent GBM after failure of multimodal intensive initial therapy including nitrosourea-based chemotherapy. In addition, MGMT protein expression determined by western blotting was analysed for a correlation with response to TMZ and survival.
|
MATERIALS AND METHODS |
|---|
|
TOPAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONAcknowledgementsReferences |
|---|
Patient Eligibility
Patients were eligible for this study if they had been diagnosed with recurrent GBM. All patients were
3 years of age with a Karnofsky performance status (KPS)60. They had to show objective evidence of tumor recurrence or progression and to have adequate hematologic, renal and hepatic function as follows: absolute neutrophil count1500/µl, platelet count100 000/µl, hemoglobin8 g/dl, serum creatinine and bilirubin levels less than 1.5 times the upper limit of laboratory normal and AST or ALT
3 times the upper limit of laboratory normal. Patients were required to have a life expectancy greater than 3 months, to have prior chemotherapy or radiation 4 weeks or more before, and to have provided written informed consent. The treatment protocol was approved by the Institutional Ethics Committees.
Treatment
TMZ was provided by Schering-Plough Corp. It was administered for a maximum of 30 cycles or until unacceptable toxicity or tumor progression occurred. Patients received TMZ (150 mg/m2/day) for 5 days every 4 weeks in the first cycle (750 mg/m2/cycle). In the absence of grade 3 or 4 hematologic toxicity, dosing for the following cycle could be increased to 200 mg/m2/day for 5 days (1000 mg/m2/cycle). Repeat cycles were administered on schedule only if, before the first day of the next cycle, grade 2 or greater neutropenia or thrombocytopenia was absent. In the case of grade 3 or greater hematologic toxicity or reversible grade 3 non-hematologic toxicity (except for nausea/vomiting), the dose of TMZ was reduced by 25%, with the lowest dose being 100 mg/m2/day. In cases of non-hematologic grade 4 toxicity, chemotherapy was interrupted. Prophylactic anti-emetics were given routinely. Neurologic stability was provided with the lowest corticosteroid dose when required.
Patient Evaluation
Gadolinium-enhanced magnetic resonance imaging (MRI) or contrast-enhanced computed tomography was performed at either Kyorin University Hospital or the original investigator's institution. Response evaluation conformed to the RECIST system (23) for tumor size, and also to Macdonald's criteria (24). Patients were closely monitored for toxicity in all cycles and graded according to National Cancer Institute Common Terminology Criteria for Adverse Events (CTCAE) version 3.0. Hematologic and biochemical evaluations were repeated at least once per cycle.
Clinical Specimens
Malignant glial tumors were surgically removed at either Kyorin University Hospital or Tokyo Metropolitan Fuchu Hospital and stored at –80°C prior to use. Patient material was obtained with informed consent on approval from the Institutional Ethics Committees.
Western Blot Analysis
Whole lysates from tumor specimens were prepared in RIPA buffer [50 mM Tris, pH 7.4, 150 mM NaCl, 2 mM EDTA, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, 1x complete protease inhibitor (Roche), 5 mg/ml pepstatin, 2 mM sodium orthovanadate, 0.5 M sodium fluoride and 0.2 M sodium pyrophosphate] and subjected to western blot analyses as previously described (25). Proteins on PVDF membranes were probed with a monoclonal antibody against MGMT (MT3.1, NeoMarker), and β-actin (monoclonal, Sigma), and were detected by chemiluminescence and quantified (LAS 1000, Fuji). The loading of lysates on membranes was evaluated by β-actin blotting.
Assessment of MGMT Protein Expression Level (MGMT Score)
MGMT protein expression was detected and quantified by western blotting as described earlier. MGMT expression was normalized relative to the β-actin level of the tumor. Tumor ‘MGMT score (%)’ was calculated by standardizing the MGMT level of each tumor relative to that of the T98G glioma cell line, which expresses a high level of MGMT protein and is resistant to TMZ and nitrosourea (26).
Statistical Analysis
PFS was evaluated from the first day of treatment to relapse, progression or death, or to the last date of follow-up, and overall survival (OS) was calculated as MST from the first day of treatment of the recurrent tumor to death for any reason or to the last date of follow-up. PFS and OS were calculated according to the Kaplan–Meier method, and differences in progression and survival according to prognostic factors were evaluated with the log-rank test. Parameters possibly correlated with disease progression and survival were age, gender, KPS at relapse, frontal lobe localization, presence or absence of re-resection at relapse, prior use of nitrosourea, response to treatment and expression of MGMT protein. A multivariate analysis with the Cox model, used to assess truly independent prognostic factors, was performed only for variables for which P < 0.1 was obtained in the univariate analysis. For continuous variables, the cut-off level chosen for KPS and MGMT score was the median value, and that for age was the mean value. All the probability values were two-sided, and all statistical analyses were done at a significance level of P = 0.05, using the statistical package SPSS 15.0J (SPSS, Inc., Chicago, IL, USA). The correlation of tumor MGMT protein expression with clinical variables was evaluated by the chi-square test and the Fisher's exact test. The length of follow-up was described as the median and range.
|
RESULTS |
|---|
|
TOPAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONAcknowledgementsReferences |
|---|
Patients' Characteristics
From September 2003 to January 2007, a total of 30 patients with recurrent GBM were treated with TMZ. Among them, 19 patients (11 male and 8 female, a mean age of 48.2 years) were analysed for tumor MGMT status and were included in this study; their characteristics are outlined in Table 1. One patient was lost to follow-up after initiation of treatment cycle 2 and was not assessable for response, survival and safety. Another patient was not assessable for response because of no residual tumor on MRI after re-resection. All patients had undergone surgical resection as well as RT at the time of initial diagnosis. Seventeen patients (89.5%) had received nitrosourea-based chemotherapy prior to initiation of TMZ therapy. Re-resection was performed in 10 patients (52.6%) when the tumor relapsed.
|
MGMT Protein Expression Analysis
The MGMT status of the tumors was analysed in 19 patients (63.3%) whose frozen tumor tissues were available using western blotting. The MGMT protein level was determined by comparing it with that of the T98G human glioma cell line having high MGMT expression and nitrosourea resistance, and was calculated as the ‘MGMT score’ as described in ‘Materials and Methods’. The level of MGMT protein in the GBMs varied substantially (Fig. 1), ranging from almost undetectable (MGMT score 3.4) to very high (104), with a median value of 12.8 (Table 1). MGMT protein expression was not related to any clinical variables (Fisher's exact test) (Table 2). Their characteristics were similar to those of the entire patient population.
|
|
Among the 19 patients in this study, there were six patients whose primary and recurrent tumor specimens were available for MGMT protein analysis. In four cases (67%), MGMT expression was at a similar level in samples obtained from primary and recurrent tumors of the same patients. In two cases (33%), however, it increased from a low to a high level, suggesting that recurrent tumor might have resulted from selection of tumor cells expressing high levels of MGMT in a minority of cases.
Response
Of 19 patients whose tumor MGMT expression was determined, 17 patients were assessable for response. One had a complete response (CR) (5.9%), three had a partial response (PR) (17.6%; overall response: 23.5%), six had stable disease (SD) (35.3%) and seven had progressive disease (PD) (35.3%). TMZ treatment resulted in an overall disease control rate (CR + PR + SD) of 58.8%. All responses were confirmed mostly after a 1-month interval or at least 2 months apart. The response rates of this subpopulation were almost identical to those of the overall patient population. In the univariate analysis, none of the categoric variables was found to be significantly correlated with response (CR + PR) or overall disease control (CR + PR + SD). MGMT score tended to be lower in the responders (patients with CR/PR) than non-responders (those with SD/PD), but the difference was not significant (Mann–Whitney U-test) (Fig. 2), perhaps due to an insufficient number of patients.
|
Progression-free Survival
After a median follow-up of 7.1 months (range: 2.4 – 16.7), one patient (5.6%) remained free of progression. PFS from the initiation of TMZ therapy for the 18 assessable patients in the MGMT analysis was 2.2 months (95% CI: 0.0–5.6 months).
In the univariate analysis (Table 3), only low MGMT expression was found to be significantly correlated with PFS (P = 0.016) (Fig. 3A). Unlike age, KPS, frontal localization and re-resection at relapse, response to treatment (CR + PR) (P = 0.070) was nearly significant. In the multivariate analysis (Table 4), low MGMT expression remained as the only independent prognostic factor close to the significant level (P = 0.060).
|
|
|
Overall Survival
Four of the 19 patients (21.1%) remained alive at the end of follow-up. MST from the initiation of TMZ therapy for the patients in the MGMT analysis was 9.9 months (95% CI: 5.5–14.3 months).
In the univariate analysis (Table 3), re-resection at relapse (P = 0.002) and low MGMT expression (P = 0.019) were found to be significantly correlated with OS (Fig. 3B). KPS (80 versus <80) was near to the level of significance (P = 0.09), unlike age, gender, frontal localization and response to treatment. Both re-resection at relapse and KPS also remained as significant factors when analysed in the entire patient population. In the multivariate analysis (Table 4), only re-resection at relapse (P = 0.014) and low MGMT expression (P = 0.040) persisted as significant independent prognostic factors. These factors were not associated with each other (chi-square: P = 0.463). Because most patients had received prior nitrosourea-based chemotherapy, there was no correlation between survival and history of nitrosourea use.
There were 10 patients whose recurrent tumors were available for MGMT protein analysis. The median value for the MGMT score of these tumors was 10.2, which was slightly lower than that of all 19 tumors analysed (12.8). Using this median value, patients with low MGMT protein expression still had nearly significantly improved OS than those with high expression (log-rank, P = 0.050), despite a small number of patients analysed.
Toxicity
A total of 107 treatment cycles of TMZ were administered (median: six cycles per patient; range: 1–13). Treatment was generally well tolerated (Table 5). Among 18 patients who received more than one treatment cycle, the dose was escalated to 200 mg/m2/day in all patients. Two patients received a reduction in dose to 150 mg/m2/day and one patient, to 100 mg/m2/day, due to nausea or infection. Regarding hematologic toxicity, grade 4 neutropenia or thrombocytopenia did not occur. Grade 4 lymphopenia was found in only one patient (5.6%). A total of five incidents of grade 3 hematologic toxicity were found in four patients (neutropenia: n = 1; thrombocytopenia: n = 1; lymphopenia: n = 3). No patients received a blood transfusion. The most common, probably treatment-related, hematologic toxicity was lymphopenia (grade 1–4, 77.8%). The incidence of non-hematologic grade 3 or 4 toxicity was low, including elevated transamidases levels (22.2%), nausea/vomiting (11.1%) and fatigue (5.6%). Two patients (11.1%) discontinued treatment because of adverse events, which consisted of grade 3 liver dysfunction (n = 1) and grade 2 pneumonia (n = 1). This pulmonary event resolved without sequelae with standard treatment. Pneumocystis carinii pneumonia was not confirmed in any patient despite that the prophylactic trimethoprim was not administered. There was no significant correlation between tumor MGMT protein expression and toxicity, except for grade 3 and 4 lymphopenia that occurred only in patients whose GBM expressed little MGMT protein.
|
|
DISCUSSION |
|---|
|
TOPAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONAcknowledgementsReferences |
|---|
Despite recent progress in the medical management of many malignant tumors, high-grade glioma, especially its most malignant form GBM, remains incurable and patients with GBM usually do not survive more than 1 year, necessitating the development of new active agents. TMZ, an orally administrable alkylating agent, has become the first significantly active therapeutic drug against newly diagnosed GBM, extending survival time by a median value of 2.7 months when concomitantly combined with conventional fractionated RT of 60 Gy, compared with RT alone (9). The survival benefits derived from the concomitant TMZ therapy were shown to correlate with methylation status of the gene of the DNA repair enzyme MGMT (15), which appeared reasonable because MGMT can sequester TMZ-induced genotoxic methyl adducts from guanines in DNA (27). These results prompted us to analyse whether TMZ could also have a similar effect on recurrent GBM, in conjunction with an analysis of the influence of tumor MGMT expression, in patients who had previously received multimodal treatment regimens including ACNU whose antitumor activity is also known to be hampered by MGMT (14,28).
In the present study, we demonstrated that the standard 5-day on/28-day cycle TMZ regimen was safe and feasible even in patients with chemotherapy-pretreated recurrent GBM. Most patients received more than two cycles of TMZ, and the major reason for the discontinuation of treatment was tumor progression, not toxicity. The TMZ therapy resulted in a MST from the initiation of therapy of 9.9 months, an OS rate at 6 months of 78.0%, a PFS rate at 6 months of 22.2%, an overall response rate of 23.5% and a disease control rate of 58.8%. These results are slightly better than those reported in patients with GBM at either first or second relapse in several phase II clinical trials, where OS at 6 months ranged from 46% to 60%, MST was 5.4–6.5 months, PFS at 6 months was 18–24%, the response rate was 8–19% and the disease control rate was 40–51% (29–32). These studies contained more patients who were chemonaïve at the initiation of TMZ therapy, suggesting that activity with the standard TMZ regimen might not be substantially affected by preceding chemotherapy with nitrosoureas.
Although molecular determinants of the sensitivity of glioma cells to TMZ may include the mismatch repair function and protein expression (18), and G2/M checkpoint proteins (33–35), MGMT has been indicated as the major factor conferring resistance to TMZ because of its direct repair of methylated guanine residues (27). As expected, patients with recurrent GBM, which expressed a low level of MGMT protein, had a significantly better OS and PFS after undergoing TMZ therapy than those with a high level of tumor MGMT expression in the univariate analyses, even though the number of patients analysed was small (Fig. 3). The low MGMT protein expression was further identified as an independent favorable prognostic factor in terms of OS, and also a nearly significant factor for PFS (P = 0.069) by multivariate analyses (Tables 3 and 4). These results were consistent with the findings in the large phase III EORTC/NCIC trial conducted for patients with newly diagnosed GBM (15), and further extend the notion that MGMT plays a key role in determining the efficacy of TMZ against GBM not only in chemonaïve but also in nitrosourea-pretreated patients. Indeed, among patients whose recurrent tumors were analysed for MGMT expression, those with low MGMT expression still had improved OS than those with high expression. However, a recent phase II study by Brandes et al. (20) showed no correlation between MGMT promoter methylation status and response rate, PFS or OS in patients treated with the intensified 3-week on/1-week off TMZ regimen for recurrent GBM after the first line chemotherapy with TMZ. In addition to differences in the preceding alkylating agent, i.e. TMZ or nitrosourea, TMZ dose settings and time since the last cycle of the previous chemotherapy, this finding raises the question of whether methods to be used for detection of MGMT status influence these correlation analyses, and might suggest a potential pitfall to the use of the MSP assay for analysing MGMT status.
Several methods have been used to determine MGMT status in glioma tissues; a MSP assay for methylation status of the MGMT gene promoter region (15,16), reverse-transcription (RT)–PCR for MGMT mRNA expression (36), IHC (37), DNA repair enzymatic activity assays for MGMT (37,38) and a quantitative protein expression assay using western blotting as shown herein. These assays, however, have not been standardized or validated for clinical use, because contradicting results have been reported regarding the correlation between MGMT promoter methylation and MGMT protein expression (21,39,40). The MSP assay requires small quantities of DNA, and the presence of a methylated MGMT allele can be attributed solely to the neoplastic cells, thus contamination of non-neoplastic tissue would have little influence (22). However, the MSP assay has potential technical difficulties, since its success rate with biopsied material was low in the study by Hegi et al. (15). Furthermore, promoter methylation status does not always reflect protein expression, thus limiting its application to use as an epigenetic marker, rather than as a substitute for MGMT enzymatic activity. Indeed, a recent study by Maxwell et al. (37) showed not only no correlation between MSP results and MGMT enzymatic activity, but also a substantial disagreement between the grouping of patients by IHC and MSP based on the assigned prognostic criteria. IHC is a feasible method to detect MGMT protein even in archived paraffin-embedded samples. Although it has the advantage of allowing tumor cells to be identified from non-neoplastic brain components, its major disadvantages include low sensitivity leading to an underestimation of MGMT positivity (41), and wide variation in the positive rate and threshold level among studies (37) rendering meaningful comparison difficult. While requiring a large amount of starting material, the MGMT enzymatic activity assay would be the best in terms of a theoretical connection with the resistance mechanism. A good correlation was reported between MGMT activity and MGMT protein expression quantified by both IHC (37) and western blotting (42). Considering the laborious nature of the activity assay, these findings suggest that the quantification of MGMT protein expression by western blotting may well substitute for the MGMT activity assay for predictions of the response to TMZ treatment. A western blot analysis can be performed with a relatively small amount of starting material (
20 µg tumor lysate), and the expression level is easily standardized using the lysate of a glioma cell line with known MGMT expression and sensitivity to nitrosoureas and TMZ such as T98G (26). The observation that the MGMT status of gliomas determined by either the MGMT activity assay or IHC showed a good correlation indicates that potential contamination by a small amount of normal cells in tumor tissues may not significantly affect the result of the assays (37). Accordingly, we show a correlation, for the first time, between the tumor MGMT protein expression level and the OS or PFS of patients with recurrent GBM after TMZ treatment, whereas MSP was not associated with survival or response to TMZ (20). Further investigation of the relationship among MGMT assays in our cohort of patients is ongoing.
Since gliomas with a high MGMT expression level may respond to TMZ poorly, attempts to increase the antitumor activity of TMZ have been encouraged. One such approach is to administer TMZ via dose-dense regimens, which have been shown to effectively deplete cellular MGMT activity (43). The 7-day on/7-day off schedule at 150 mg/m2/day or the 21-day on/7-day off schedule at 75 mg/m2/day provides theoretically a 2.1- or 1.5-fold greater dose of TMZ, respectively, than the cumulative dose obtained with the standard 5-day regimen (20,44). These treatments achieved a 30–48% PFS rate at 6 months with acceptable safety profiles in patients with recurrent GBM (20,44), which were higher than the values obtained in our series. However, patients with low MGMT expression had a significantly higher response rate to the 7-day on/7-day off TMZ regimen, than those with high MGMT expression (17), suggesting that the depletion of MGMT may be still insufficient. Another approach is the use of MGMT-depleting agents other than TMZ itself. One of the most potent agents, O6-benzylguanine, has been investigated in combination with TMZ in a couple of phase I studies with evidence of activity against refractory malignant gliomas (45,46). Similarly, clinical trials to evaluate the activity of procarbazine or cisplatin combined with TMZ have been conducted in patients with recurrent GBM (47,48). Whether these approaches can overcome resistance to TMZ through down-regulation of MGMT activity needs to be verified in comparison with the standard TMZ regimen in patients with GBM having high MGMT expression.
Another factor found to be significantly favorable for survival in the multivariate analysis was re-resection at relapse of the tumor. Although a recent study has demonstrated that the extent of resection at initial surgery correlates with OS in patients with newly diagnosed GBM (49), this finding might have resulted from potential selection bias, because re-resection is usually considered only when the recurrent tumor bulk is located outside of the eloquent areas, not infiltrating into deep brain structures and contralateral parenchyma, reflecting a better prognosis from the beginning.
In conclusion, the standard 5-day TMZ regimen resulted in moderate antitumor activity with an acceptable safety profile in patients with recurrent GBM even after pretreatment with nitrosourea. This study also provides additional evidence that MGMT protein expression is an important prognostic factor for patients treated with TMZ even after recurrence. Further prospective studies are needed to determine subgroups of patients for whom the standard 5-day TMZ regimen may be beneficial, or those who require more intensive TMZ regimens to overcome TMZ resistance.
|
Acknowledgements |
|---|
|
TOPAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONAcknowledgementsReferences |
|---|
The authors are grateful to the following investigators who referred patients to our institution for temozolomide therapy: T. Mizutani, Tokyo Metropolitan Fuchu Hospital, Tokyo, Japan; K. Tsutsumi, Showa General Hospital, Tokyo, Japan; N. Kaneko, Saitama Cardiovascular and Respiratory Center, Saitama, Japan; S. Okabe, Seirei Memorial Hospital, Ibaragi, Japan; J. A. Takahashi, Kyoto University Hospital, Kyoto, Japan; H. Takahashi, Tokyo Metropolitan Neurological Hospital, Tokyo, Japan; T. Kohno, Brainpier Minamiohta, Ibaragi, Japan; Y. Fujii, Kugayama Hospital, Tokyo, Japan; T. Kimura, Kitahara Neurosurgical Institute, Tokyo, Japan; T. Maeda, Maeda Hospital, Tokyo, Japan; T. Tokime, Tenri Hospital, Nara, Japan; H. Tomita, Musashino Red Cross Hospital, Tokyo, Japan; N. Ito, Akiru Municipal Medical Center, Tokyo, Japan; E. Sato, Inagi Municipal Hospital, Tokyo, Japan; K. Okamura, Fuji Brain Institute and Hospital, Shizuoka, Japan; T. Fukushima, Duke University, NC, USA; S. Oikawa, Kofu Municipal Hospital, Yamanashi, Japan; M. Morimoto, Kochi Health Sciences Center, Kochi, Japan. The authors thank members of Department of Neurosurgery, Kyorin University School of Medicine for patient care.
Conflict of interest statement
None declared.
|
Footnotes |
|---|
Abbreviations: MST, medial survival time; GBM, glioblastoma multiforme; TMZ, temozolomide; EORTC, European Organisation for Research and Treatment of Cancer; NCIC, National Cancer Institute of Canada; RT, radiotherapy; MGMT, O6-methylguanine-DNA methyltransferase; PCR, polymerase-chain reaction; MSP, methylation-specific PCR; IHC, immunohistochemistry; KPS, Karnofsky performace status; MRI, magnetic resonance imaging; PFS, progression-free survival; OS, overall survival; CR, complete response; PR, partial response; SD, stable disease; PD, progressive disease.
|
References |
|---|
|
TOPAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONAcknowledgementsReferences |
|---|
1 Laperriere N, Zuraw L, Cairncross G. Radiotherapy for newly diagnosed malignant glioma in adults: a systematic review. Radiother Oncol (2002) 64:259–73.[CrossRef][Web of Science][Medline]
2 Stewart LA. Chemotherapy in adult high-grade glioma: a systematic review and meta-analysis of individual patient data from 12 randomised trials. Lancet (2002) 359:1011–8.[CrossRef][Web of Science][Medline]
3 Hickman JA, Stevens MF, Gibson NW, Langdon SP, Fizames C, Lavelle F, et al. Experimental antitumor activity against murine tumor model systems of 8-carbamoyl-3-(2-chloroethyl)imidazo[5,1-d]-1,2,3,5-tetrazin-4(3 H)-one (mitozolomide), a novel broad-spectrum agent. Cancer Res (1985) 45:3008–13.
4 Stevens MF, Hickman JA, Langdon SP, Chubb D, Vickers L, Stone R, et al. Antitumor activity and pharmacokinetics in mice of 8-carbamoyl-3-methyl-imidazo[5,1-d]-1,2,3,5-tetrazin-4(3H)-one (CCRG 81045; M & B 39831), a novel drug with potential as an alternative to dacarbazine. Cancer Res (1987) 47:5846–52.
5 Tsang LL, Farmer PB, Gescher A, Slack JA. Characterisation of urinary metabolites of temozolomide in humans and mice and evaluation of their cytotoxicity. Cancer Chemother Pharmacol (1990) 26:429–36.[Web of Science][Medline]
6 Yung WK. Temozolomide in malignant gliomas. Semin Oncol (2000) 27:27–34.[Medline]
7 van den Bent MJ, Taphoorn MJ, Brandes AA, Menten J, Stupp R, Frenay M, et al. Phase II study of first-line chemotherapy with temozolomide in recurrent oligodendroglial tumors: the European Organization for Research and Treatment of Cancer Brain Tumor Group Study 26971. J Clin Oncol (2003) 21:2525–8.
8 Yung WK, Prados MD, Yaya-Tur R, Rosenfeld SS, Brada M, Friedman HS, et al. Multicenter phase II trial of temozolomide in patients with anaplastic astrocytoma or anaplastic oligoastrocytoma at first relapse. Temodal Brain Tumor Group. J Clin Oncol (1999) 17:2762–71.
9 Stupp R, Mason WP, van den Bent MJ, Weller M, Fisher B, Taphoorn MJ, et al. Radiotherapy plus concomitant and adjuvant temozolomide for glioblastoma. N Engl J Med (2005) 352:987–96.
10 Clark AS, Deans B, Stevens MF, Tisdale MJ, Wheelhouse RT, Denny BJ, et al. Antitumor imidazotetrazines. 32. Synthesis of novel imidazotetrazinones and related bicyclic heterocycles to probe the mode of action of the antitumor drug temozolomide. J Med Chem (1995) 38:1493–504.[CrossRef][Web of Science][Medline]
11 Tisdale MJ. Antitumor imidazotetrazines–XV. Role of guanine O6 alkylation in the mechanism of cytotoxicity of imidazotetrazinones. Biochem Pharmacol (1987) 36:457–62.[CrossRef][Web of Science][Medline]
12 Gerson SL. Clinical relevance of MGMT in the treatment of cancer. J Clin Oncol (2002) 20:2388–99.
13 Pegg AE, Dolan ME, Moschel RC. Structure, function, and inhibition of O6-alkylguanine-DNA alkyltransferase. Prog Nucleic Acid Res Mol Biol (1995) 51:167–223.[Web of Science][Medline]
14 Nagane M, Asai A, Shibui S, Nomura K, Matsutani M, Kuchino Y. Expression of O6-methylguanine-DNA methyltransferase and chloroethylnitrosourea resistance of human brain tumors. Jpn J Clin Oncol (1992) 22:143–9.
15 Hegi ME, Diserens AC, Gorlia T, Hamou MF, de Tribolet N, Weller M, et al. MGMT gene silencing and benefit from temozolomide in glioblastoma. N Engl J Med (2005) 352:997–1003.
16 Esteller M, Garcia-Foncillas J, Andion E, Goodman SN, Hidalgo OF, Vanaclocha V, et al. Inactivation of the DNA-repair gene MGMT and the clinical response of gliomas to alkylating agents. N Engl J Med (2000) 343:1350–4.
17 Chinot OL, Barrie M, Fuentes S, Eudes N, Lancelot S, Metellus P, et al. Correlation between O6-methylguanine-DNA methyltransferase and survival in inoperable newly diagnosed glioblastoma patients treated with neoadjuvant temozolomide. J Clin Oncol (2007) 25:1470–5.
18 Friedman HS, McLendon RE, Kerby T, Dugan M, Bigner SH, Henry AJ, et al. DNA mismatch repair and O6-alkylguanine-DNA alkyltransferase analysis and response to Temodal in newly diagnosed malignant glioma. J Clin Oncol (1998) 16:3851–7.
19 Paz MF, Yaya-Tur R, Rojas-Marcos I, Reynes G, Pollan M, Aguirre-Cruz L, et al. CpG island hypermethylation of the DNA repair enzyme methyltransferase predicts response to temozolomide in primary gliomas. Clin Cancer Res (2004) 10:4933–8.
20 Brandes AA, Tosoni A, Cavallo G, Bertorelle R, Gioia V, Franceschi E, et al. Temozolomide 3 weeks on and 1 week off as first-line therapy for recurrent glioblastoma: phase II study from gruppo italiano cooperativo di neuro-oncologia (GICNO). Br J Cancer (2006) 95:1155–60.[CrossRef][Web of Science][Medline]
21 Brell M, Tortosa A, Verger E, Gil JM, Vinolas N, Villa S, et al. Prognostic significance of O6-methylguanine-DNA methyltransferase determined by promoter hypermethylation and immunohistochemical expression in anaplastic gliomas. Clin Cancer Res (2005) 11:5167–74.
22 Stupp R, Hegi ME. Methylguanine methyltransferase testing in glioblastoma: when and how? J Clin Oncol (2007) 25:1459–60.
23 Therasse P, Arbuck SG, Eisenhauer EA, Wanders J, Kaplan RS, Rubinstein L, et al. New guidelines to evaluate the response to treatment in solid tumors. European Organization for Research and Treatment of Cancer, National Cancer Institute of the United States, National Cancer Institute of Canada. J Natl Cancer Inst (2000) 92:205–16.
24 Macdonald DR, Cascino TL, Schold SC Jr, Cairncross JG. Response criteria for phase II studies of supratentorial malignant glioma. J Clin Oncol (1990) 8:1277–80.[Abstract]
25 Nagane M, Coufal F, Lin H, Bogler O, Cavenee WK, Huang HJS. A common mutant epidermal growth factor receptor confers enhanced tumorigenicity on human glioblastoma cells by increasing proliferation and reducing apoptosis. Cancer Res (1996) 56:5079–86.
26 Kanzawa T, Bedwell J, Kondo Y, Kondo S, Germano IM. Inhibition of DNA repair for sensitizing resistant glioma cells to temozolomide. J Neurosurg (2003) 99:1047–52.[Web of Science][Medline]
27 Gerson SL. MGMT: its role in cancer aetiology and cancer therapeutics. Nat Rev Cancer (2004) 4:296–307.[CrossRef][Web of Science][Medline]
28 Nagane M, Asai A, Shibui S, Nomura K, Kuchino Y. Application of antisense ribonucleic acid complementary to O6-methylguanine-deoxyribonucleic acid methyltransferase messenger ribonucleic acid for therapy of malignant gliomas. Neurosurgery (1997) 41:434–40. Discussion in 1997;41:40–1.[CrossRef][Web of Science][Medline]
29 Teixeira MM, Garcia I, Portela I, Cernuda M, Oliveira C, Albano J, et al. Temozolomide in second-line treatment after prior nitrosurea-based chemotherapy in glioblastoma multiforme: experience from a Portuguese institution. Int J Clin Pharmacol Res (2002) 22:19–22.[Web of Science][Medline]
30 Brandes AA, Ermani M, Basso U, Paris MK, Lumachi F, Berti F, et al. Temozolomide in patients with glioblastoma at second relapse after first line nitrosourea-procarbazine failure: a phase II study. Oncology (2002) 63:38–41.[CrossRef][Web of Science][Medline]
31 Brada M, Hoang-Xuan K, Rampling R, Dietrich PY, Dirix LY, Macdonald D, et al. Multicenter phase II trial of temozolomide in patients with glioblastoma multiforme at first relapse. Ann Oncol (2001) 12:259–66.
32 Yung WK, Albright RE, Olson J, Fredericks R, Fink K, Prados MD, et al. A phase II study of temozolomide vs. procarbazine in patients with glioblastoma multiforme at first relapse. Br J Cancer (2000) 83:588–93.[CrossRef][Web of Science][Medline]
33 Hirose Y, Katayama M, Mirzoeva OK, Berger MS, Pieper RO. Akt activation suppresses Chk2-mediated, methylating agent-induced G2 arrest and protects from temozolomide-induced mitotic catastrophe and cellular senescence. Cancer Res (2005) 65:4861–9.
34 Hirose Y, Katayama M, Berger MS, Pieper RO. Cooperative function of Chk1 and p38 pathways in activating G2 arrest following exposure to temozolomide. J Neurosurg (2004) 100:1060–5.[CrossRef][Web of Science][Medline]
35 Hirose Y, Berger MS, 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 (2001) 61:5843–9.
36 Tanaka S, Kobayashi I, Utsuki S, Oka H, Fujii K, Watanabe T, et al. O6-methylguanine-DNA methyltranspherase gene expression in gliomas by means of real-time quantitative RT-PCR and clinical response to nitrosoureas. Int J Cancer (2003) 103:67–72.[CrossRef][Web of Science][Medline]
37 Maxwell JA, Johnson SP, Quinn JA, McLendon RE, Ali-Osman F, Friedman AH, et al. Quantitative analysis of O6-alkylguanine-DNA alkyltransferase in malignant glioma. Mol Cancer Ther (2006) 5:2531–9.
38 Kreklau EL, Limp-Foster M, Liu N, Xu Y, Kelley MR, Erickson LC. A novel fluorometric oligonucleotide assay to measure O(6)-methylguanine DNA methyltransferase, methylpurine DNA glycosylase, 8-oxoguanine DNA glycosylase and abasic endonuclease activities: DNA repair status in human breast carcinoma cells overexpressing methylpurine DNA glycosylase. Nucleic Acids Res (2001) 29:2558–66.
39 Esteller M, Sanchez-Cespedes M, Rosell R, Sidransky D, Baylin SB, Herman JG. Detection of aberrant promoter hypermethylation of tumor suppressor genes in serum DNA from non-small cell lung cancer patients. Cancer Res (1999) 59:67–70.
40 Mollemann M, Wolter M, Felsberg J, Collins VP, Reifenberger G. Frequent promoter hypermethylation and low expression of the MGMT gene in oligodendroglial tumors. Int J Cancer (2005) 113:379–85.[CrossRef][Web of Science][Medline]
41 Ohe N, Saio M, Kijima M, Tamakawa N, Suwa T, Kojima Y, et al. In situ detection of O6-methylguanine-DNA methyltransferase messenger RNA in paraffin-embedded human astrocytic tumor tissues by nested in situ RT-PCR is useful in predicting chemotherapy-resistance of tumors. Int J Oncol (2003) 22:543–9.[Web of Science][Medline]
42 Hongeng S, Brent TP, Sanford RA, Li H, Kun LE, Heideman RL. O6-Methylguanine-DNA methyltransferase protein levels in pediatric brain tumors. Clin Cancer Res (1997) 3:2459–63.
43 Tolcher AW, Gerson SL, Denis L, Geyer C, Hammond LA, Patnaik A, et al. Marked inactivation of O6-alkylguanine-DNA alkyltransferase activity with protracted temozolomide schedules. Br J Cancer (2003) 88:1004–11.[CrossRef][Web of Science][Medline]
44 Wick W, Steinbach JP, Kuker WM, Dichgans J, Bamberg M, Weller M. One week on/one week off: a novel active regimen of temozolomide for recurrent glioblastoma. Neurology (2004) 62:2113–5.
45 Warren KE, Aikin AA, Libucha M, Widemann BC, Fox E, Packer RJ, et al. Phase I study of O6-benzylguanine and temozolomide administered daily for 5 days to pediatric patients with solid tumors. J Clin Oncol (2005) 23:7646–53.
46 Quinn JA, Desjardins A, Weingart J, Brem H, Dolan ME, Delaney SM, et al. Phase I trial of temozolomide plus O6-benzylguanine for patients with recurrent or progressive malignant glioma. J Clin Oncol (2005) 23:7178–87.
47 Brandes AA, Basso U, Reni M, Vastola F, Tosoni A, Cavallo G, et al. First-line chemotherapy with cisplatin plus fractionated temozolomide in recurrent glioblastoma multiforme: a phase II study of the Gruppo Italiano Cooperativo di Neuro-Oncologia. J Clin Oncol (2004) 22:1598–604.
48 Newlands ES, Foster T, Zaknoen S. Phase I study of temozolamide (TMZ) combined with procarbazine (PCB) in patients with gliomas. Br J Cancer (2003) 89:248–51.[CrossRef][Web of Science][Medline]
49 Lacroix M, Abi-Said D, Fourney DR, Gokaslan ZL, Shi W, DeMonte F, et al. A multivariate analysis of 416 patients with glioblastoma multiforme: prognosis, extent of resection, and survival. J Neurosurg (2001) 95:190–8.[Web of Science][Medline]
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  S. Spiegl-Kreinecker, C. Pirker, M. Filipits, D. Lotsch, J. Buchroithner, J. Pichler, R. Silye, S. Weis, M. Micksche, J. Fischer, et al. O6-Methylguanine DNA methyltransferase protein expression in tumor cells predicts outcome of temozolomide therapy in glioblastoma patients Neuro Oncology, October 15, 2009; (2009) nop003v1. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. SHARMA, F. SALEHI, B. W. SCHEITHAUER, F. ROTONDO, L. V. SYRO, and K. KOVACS Role of MGMT in Tumor Development, Progression, Diagnosis, Treatment and Prognosis Anticancer Res, October 1, 2009; 29(10): 3759 - 3768. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||