methylguanine-DNA Methyltransferase Status Research Articles

Prognostic or predictive value of MGMT promoter methylation in gliomas depends on IDH1 mutation

To explore whether the isocitrate dehydrogenase 1 (IDH1) or 1p/19q status determines the prognostic vs predictive role of O(6)-methylguanine-DNA methyltransferase (MGMT) promoter methylation in the Neuro-Oncology Working Group of the German Cancer Society (NOA)-04 trial anaplastic glioma biomarker cohort. Patients (n = 183) of the NOA-04 trial with known MGMT and IDH1 status were analyzed for interdependency of the prognostic vs predictive role of MGMT promoter methylation from IDH1 or 1p/19q status and treatment, using progression-free survival (PFS) as an endpoint. An independent validation cohort of the German Glioma Network (n = 75) and the NOA-08 trial (n = 34) served as a confirmation cohort. In tumors with IDH1 mutation, MGMT promoter methylation was associated with prolonged PFS with chemotherapy ± radiotherapy (RT) or RT-only groups, and is thus prognostic. In tumors without IDH1 mutation, MGMT promoter methylation was associated with increased PFS in patients treated with chemotherapy, but not in those who received RT alone as the first-line treatment, and is thus chemotherapy-predictive. In contrast, 1p/19q codeletions showed no such association with the prognostic vs predictive value of MGMT. MGMT promoter methylation is a predictive biomarker for benefit from alkylating agent chemotherapy in patients with IDH1-wild-type, but not IDH1-mutant, malignant gliomas of World Health Organization grades III/IV. Combined IDH1/MGMT assessment may help to individualize clinical decision-making in neuro-oncology.

O6-methylguanine-DNA methyltransferase in the defense against N-nitroso compounds and colorectal cancer

Colorectal cancer (CRC) is among the leading causes of cancer death worldwide, involving multiple dietary and non-dietary risk factors. A growing body of evidence suggests that N-nitroso compounds (NOC) play a pivotal role in the etiology of CRC. NOC are present in food and are also formed endogenously in the large intestine. Upon metabolic activation and also spontaneously, they form electrophilic species that methylate the DNA, producing N-methylated purines and O(6)-methylguanine, the latter of which bears high mutagenic and carcinogenic potential. Methylated DNA bases are removed by base excision repair initiated by the alkyladenine-DNA glycosylase, the family of AlkB homologs proteins, and the suicide enzyme O(6)-methylguanine-DNA methyltransferase (MGMT), which is the main focus of this review. We present animal models with a deficiency of MGMT that display a tremendously enhanced sensitivity toward alkylation-induced colorectal carcinogenesis, highlighting its role in the protection against the cytotoxic and mutagenic effects of alkylating agents. In line with these studies, MGMT was linked to the formation of human sporadic CRC. Colorectal tumors and precursor lesions frequently display epigenetic inactivation of MGMT resulting from promoter hypermethylation, which is tightly associated with the occurrence of G:C to A:T transition mutations in the KRAS oncogene. We also discuss clinical data, which identified the MGMT status of CRC patients as promising parameter for the treatment of metastasized CRC using alkylating anticancer drugs such as temozolomide.

O6-methylguanine-DNA methyltransferase (MGMT) immunohistochemistry as a predictor of resistance to temozolomide in primary CNS lymphoma

Temozolomide, an alkylating agent, has shown promise in treating primary central nervous system lymphoma (PCNSL). The enzyme O(6)-methylguanine-DNA methyltransferase (MGMT) repairs alkylating damage, such as that induced by temozolomide. We hypothesized that MGMT immunohistochemistry would predict resistance to temozolomide in PCNSL. A retrospective study of newly-diagnosed and recurrent PCNSL patients treated at our institution was conducted to study the predictive value of MGMT immunohistochemistry for response to temozolomide. 20 patients who were treated with temozolomide as a single agent were identified during the study time period. 6/20 patients demonstrated a response, corresponding to an objective response rate of 30 % (95 % CI 8-52). Five patients with low MGMT level (<30 %) showed a response to temozolomide. Only one of 10 patients (10 %) with high MGMT level (≥30 %) exhibited a response to temozolomide. Small sample numbers precluded formal statistical comparisons. Two patients with complete response remain alive without progressive disease 6.7 and 7.2 years after temozolomide initiation. Immunohistochemistry can be performed on small biopsies to selectively assess MGMT status in tumor versus surrounding inflammation. MGMT analysis by immunohistochemistry may predict response to temozolomide in PCNSL and should be prospectively investigated.

Phase 2 study of dose-intense temozolomide in recurrent glioblastoma

Among patients with glioblastoma (GBM) who progress on standard temozolomide, the optimal therapy is unknown. Resistance to temozolomide is partially mediated by O(6)-methylguanine-DNA methyltransferase (MGMT). Because MGMT may be depleted by prolonged temozolomide administration, dose-intense schedules may overcome resistance. This was a multicenter, phase 2, single-arm study of temozolomide (75-100 mg/m(2)/day) for 21 days of each 28-day cycle. Patients had GBM in first recurrence after standard therapy. The primary end point was 6-month progression-free survival (PFS6). Fifty-eight participants were accrued, 3 of whom were ineligible for analysis; one withdrew before response assessment. There were 33 men (61%), with a median age of 57 years (range, 25-79 years) and a median Karnofsky performance score of 90 (range, 60-100). Of 47 patients with MGMT methylation results, 36 (65%) had methylated tumors. There were 7 (13%) partial responses, and PFS6 was only 11%. Response and PFS did not depend on MGMT status; MSH2, MLH1, or ERCC1 expression; the number of prior temozolomide cycles; or the time off temozolomide. Treatment was well tolerated, with limited grade 3 neutropenia (n = 2) or thrombocytopenia (n = 2). Dose-intense temozolomide on this schedule is safe in recurrent GBM. However, efficacy is marginal and predictive biomarkers are needed.

Aldehyde dehydrogenase 1A1—a new mediator of resistance to temozolomide in glioblastoma

Implementation of chemotherapy with the drug temozolomide increased the overall survival of patients with glioblastoma multiforme (GBM; WHO grade IV), in particular when the O(6)-methylguanine DNA methyltransferase (MGMT) promoter is epigenetically silenced. Nevertheless, the prognosis remains poor, and relapse in GBM occurs regularly. This clinical behavior seems to be due to the existence of a therapy-resistant subpopulation of cells that induce tumor regrowth. The objective of this work was to analyze the role of aldehyde dehydrogenase (ALDH) 1A1 in mediating temozolomide resistance and its value as a predictor of clinical outcome in GBM patients. Nine GBM cell lines were treated with temozolomide alone or in combination with 4-diethylaminobenzaldehyde (DEAB), an inhibitor of ALDH1A1, or with ALDH1A1 short hairpin (sh)RNA. ALDH1A1 expression and MGMT status of 70 primary GBM patients were correlated with median survival. ALDH1A1 overexpression predicted temozolomide resistance in vitro. Sensitivity of ALDH1A1 positive/MGMT-positive cells to temozolomide could be restored by inhibition of ALDH1A1 by DEAB or by knockdown with shRNA, as indicated by increased cytotoxicity, reduced clonogenicity, and accumulation in the G2/M cell-cycle phase. The prognosis of patients with a high level of ALDH1A1 expression was poor compared with that of patients with low levels (P < .0001). ALDH1A1 is a new mediator for resistance of GBM to temozolomide and a reliable predictor of clinical outcome and may serve as a potential target to improve treatment of human GBM.

The Changes in MGMT Promoter Methylation Status in Initial and Recurrent Glioblastomas

To evaluate the mechanism of the development of therapeutic resistance after temozolomide treatment, we focused on changes in O(6)-methylguanine DNA methyltransferase (MGMT) and mismatch repair (MMR) between initial and recurrent glioblastomas. Tissue samples obtained from 24 paired histologically confirmed initial and recurrent adult glioblastoma patients who were initially treated with temozolomide were used for MGMT and MMR gene promoter methylation status and protein expression analysis using methylation-specific multiplex ligation probe amplification (MS-MLPA), methylation-specific polymerase chain reaction (MSP), and immunohistochemical staining. There was a significant decrease in the methylation ratio of the MGMT promoter determined by MS-MLPA, which was not detectable with MSP, and MGMT protein expression changes were not remarkable. However, there was no epigenetic variability in MMR genes, and a relatively homogeneous expression of MMR proteins was observed in initial and recurrent tumors. We conclude that the development of reduced methylation in the MGMT promoter is one of the mechanisms for acquiring therapeutic resistance after temozolomide treatment in glioblastomas.

New treatment options with cytotoxic agents in neuroendocrine tumours

There are numerous treatment options for patients with advanced digestive neuroendocrine tumours (NETs). Medical treatment includes systemic chemotherapies, targeted therapies, somatostatin analogs, liver-directed therapies such as (chemo)embolization or thermoablation, and peptide receptor radionuclide therapy. Cytotoxic chemotherapies can help control tumour progression in patients with non-resectable tumours and may improve symptoms by reducing tumour bulk. In addition, tumour response is usually greater than that obtained with targeted therapies. This should be taken into consideration in neoadjuvant strategies. Efficacy of temozolomide depends on the O(6) methylguanine DNA methyl transferase status, and thus, this drug will likely have to be considered in the future in patients with a favourable enzyme profile. Because numerous treatment options are available for patients with advanced digestive NETs, and thanks to their long survival, successive drugs should be used. Careful attention should be paid to the adverse events in order to maintain the quality of life in these patients who have with a long life expectancy.

Salvage treatment with temozolomide in refractory or relapsed primary central nervous system lymphoma and assessment of the MGMT status

High-dose methotrexate (HD-MTX) is effective in the initial treatment of primary central nervous system lymphoma (PCNSL). Because treatment options in patients with progressive or recurrent PCNSL are limited, prognosis is poor. Temozolomide, a well-tolerated oral alkylating agent that permeates the blood brain barrier (BBB), is effective against malignant glioma and recurrent PCNSL. The gene for the deoxyribonucleic acid (DNA) repair enzyme O(6)-methylguanine-DNA methyltransferase (MGMT), which is closely related to cellular sensitivity to alkylating agents, is inactivated by promoter hypermethylation. We evaluated the results of temozolomide treatment and the methylation status of the promoter region of the MGMT gene in 17 patients (median age 68 years) with refractory or relapsed PCNSL. They were immunocompetent and had received initial treatment with HD-MTX (3.5 g/m(2)) with or without irradiation. All were treated with temozolomide 150-200 mg/m(2), for 5 days in the course of 28 days; treatment was continued until disease progression. We observed five complete remissions, five partial responses (PRs) with stable disease (SD), and seven with disease progression. Median overall survival after the temozolomide treatment was 6.7 months. One patient manifested grade 3 neutropenia and thrombocytopenia. Eleven tumor specimens were available for MGMT analysis. MGMT promoter methylation (mMGMT) in the tumor tissue was found in 4 (36.4%), the other seven harbored a non-methylated MGMT promoter (nmMGMT). There was no statistically significant difference in median overall survival between patients with mMGMT (11.1 months) and nmMGMT (6.7 months) (P = 0.63). Although some patients were elderly and had been heavily pre-treated, temozolomide resulted in a complete response (CR) in 29% and was well tolerated without any major toxicity.

O(6)-methylguanine DNA-methyltransferase (MGMT) in glioblastoma: Analysis on first and following surgeries.

2097 Background: Glioblastoma (GBM) is the most common malignant primary brain tumor in adults. Outcome has recently improved with the introduction of post-operative concomitant chemoradiotherapy (CRT) followed by 6 cycles of temozolomide (TMZ). MGMT expression has already been encoded as an independent prognostic factor and predictive for TMZ sensitivity, but few data are available on the modifications of MGMT status after CRT. Methods: From January 2005 to November 2010 at Modena University Hospital and Nuovo Ospedale Civile S. Agostino-Estense, 15 consecutive patients (pts) with GBM and treated with at least 2 subsequent surgeries were included in this retrospective study. Aim of the present analysis is to evaluate if methylation status may change between first and following determinations. Results: Fifteen pts (M/F=10/5; median age: 48, range: 40-67 years) underwent a first surgery for GBM. MGMT status was: methylated in 5 (33%); unmethylated in 10 pts (77%). Two unmethylated pts presented an early relapse and underwent a second surgery after 41 and 46 days, respectively. MGMT analysis confirmed the absence of methylation. One pt with multifocal GBM was treated after the first surgery on right frontal lobe with TMZ, operated on left frontal lobe, submitted to CRT, then re-operated for a relapse in the first site of surgery (right frontal lobe). In this case MGMT status evaluated in the site of relapse (right frontal lobe) changed from unmethylated to methylated. MGMT on the lesion of left frontal lobe was unmethylated. The remaining 12 pts underwent post-operative concomitant CRT followed by monthly TMZ (median number of cycles: 8.5; range: 2-24). MGMT status on second surgery was: 5 methylated (42%) and 7 unmethylated (58%), with a 100% concordance. Two pts received a third surgery: in one case MGMT status changed from unmethylated to methylated. Overall median survival was 15.8 months. The two patients with “MGMT switch” have a median survival of 35.8 months. Conclusions: in 2 on 13 cases (15%) after CRT, MGMT status changed from unmethylated to methylated and these pts have a median overall survival similar to methylated pts. This observation may be confirmed on larger series.

Prognostic and predictive value of epigenetic silencing of MGMT in patients with high grade gliomas: a systematic review and meta-analysis

Epigenetic silencing of the O(6)-methylguanine-DNA methyltransferase (MGMT) gene is associated with improved survival in patients with high-grade gliomas (HGG), with varying estimates of magnitude. The objective of this meta-analysis is to determine the prognostic value of MGMT silencing, and assess its predictive value by treatment type. MEDLINE and EMBASE databases were searched for studies relating to gliomas and MGMT. Studies reporting overall survival (OS) by MGMT status in patients with HGG were considered potentially eligible. We excluded studies that did not control for potential confounding variables. A meta-analysis of studies was performed via random-effects modelling. Subgroup meta-analyses by treatment were performed according to a priori hypotheses. Twenty studies were ultimately eligible, including 2,018 patients. In the pooled analysis, MGMT silencing was associated with improved OS (HR = 0.436; 95% CI: 0.333-0.571; P < 0.001). The prognostic utility of MGMT status varies significantly by treatment type (P = 0.001): the HR for OS for MGMT silenced tumors is 0.190 (0.047-0.770), 0.403 (0.282-0.576), 0.743 (0.579-0.954), and 1.070 (0.722-1.585) for studies using surgery plus the addition of either: chemotherapy (CT), chemoradiotherapy (CRT), radiotherapy (RT), and nothing (surgery alone), respectively. Epigenetic silencing of MGMT is associated with markedly improved survival in patients with HGG who receive adjuvant therapy. MGMT silencing serves as a predictive marker, with the largest benefit seen in patients receiving CT as a component of adjuvant treatment, an intermediate benefit in patients receiving adjuvant RT, and no evidence to support benefit in those receiving surgery alone.

Hot spots in dynamic18FET-PET delineate malignant tumor parts within suspected WHO grade II gliomas

Molecular imaging studies have recently found inter- and intratumoral heterogeneity in World Health Organization (WHO) grade II gliomas. A correlative analysis with tumor histology, however, is still lacking. For elucidation we conducted the current prospective study. Fifty-five adult patients with an MRI-based suspicion of a WHO grade II glioma were included. [F-18]Fluoroethyltyrosine ((18)FET) uptake kinetic studies were combined with frame-based stereotactic localization techniques and used as a guide for stepwise (1-mm steps) histopathological evaluation throughout the tumor space. In tumors with heterogeneous PET findings, the O(6)-methylguanine-DNA methyltransferase (MGMT) promoter methylation status and expression of mutated protein isocitrate dehydrogenase variant R132H (IDH1) were determined inside and outside of hot spot volumes. Metabolic imaging revealed 3 subgroups: the homogeneous WHO grade II glioma group (30 patients), the homogeneous malignant glioma group (10 patients), and the heterogeneous group exhibiting both low- and high-grade characteristics at different sites (15 patients). Stepwise evaluation of 373 biopsy samples indicated a strong correlation with analyses of uptake kinetics (p < 0.0001). A homogeneous pattern of uptake kinetics was linked to homogeneous histopathological findings, whereas a heterogeneous pattern was associated with histopathological heterogeneity; hot spots exhibiting malignant glioma characteristics covered 4-44% of the entire tumor volumes. Both MGMT and IDH1 status were identical at different tumor sites and not influenced by heterogeneity. Maps of (18)FET uptake kinetics strongly correlated with histopathology in suspected grade II gliomas. Anaplastic foci can be accurately identified, and this finding has implications for prognostic evaluation and treatment planning.

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

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.

Mismatch repair deficiency is an uncommon mechanism of alkylator resistance in pediatric malignant gliomas: A report from the children's oncology group

Alkylating agents are commonly used in the treatment of childhood malignant gliomas. Overexpression of O(6)-methylguanine-DNA methyltransferase (MGMT) constitutes an important mechanism for resistance to such agents, and MGMT status has been associated with outcome in several recent trials. Deficiency in mismatch repair (MMR) function has been implicated in preclinical studies as an additional potential mechanism of resistance to methylating agents, such as temozolomide, independent of tumor MGMT status. However, the frequency of this abnormality as a clinical resistance mechanism in childhood malignant gliomas has not been well characterized. To address this issue, we examined the frequency of microsatellite instability (MSI), a marker of defective MMR, in a series of 68 tumors, derived from newly diagnosed patients treated on the Children's Cancer Group 945 study, and the Children's Oncology Group ACNS0126 and 0423 studies. MSI was assessed using a panel of six microsatellite markers, including BAT-25, BAT-26, CAT-25, D2S123, D5S346, and D17S250. MGMT immunoreactivity was assessed in parallel to allow comparison of the relative incidence of MGMT overexpression and MSI. Only three tumors had high-level MSI involving three or more markers; the remainder had no MSI at any of the loci examined. These children did not have unusual features in terms of their outcome. In contrast to the infrequency of MSI, 25 tumors (37%) exhibited MGMT overexpression as assessed by immunohistochemistry. None of the tumors with MSI exhibited overexpression of MGMT. MMR deficiency is an infrequent contributor to initial alkylator resistance in children with malignant gliomas.

Absence of the MGMT protein as well as methylation of the MGMT promoter predict the sensitivity for temozolomide

Background:The DNA repair protein O6-methylguanine-DNA methyltransferase (MGMT) can cause resistance to the alkylating drug temozolomide (TMZ). The purpose of this study was to determine the relationship between the MGMT status, determined by means of several techniques and methods, and the cytotoxic response to TMZ in 11 glioblastoma multiforme (GBM) cell lines and 5 human tumour cell lines of other origins.Methods:Cell survival was analysed by clonogenic assay. The MGMT protein levels were assessed by western blot analysis. The MGMT promoter methylation levels were determined using methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) and quantitative real-time methylation-specific PCR (qMSP). On the basis of the results of these techniques, six GBM cell lines were selected and subjected to bisulphite sequencing.Results:The MGMT protein was detected in all TMZ-resistant cell lines, whereas no MGMT protein could be detected in cell lines that were TMZ sensitive. The MS-MLPA results were able to predict TMZ sensitivity in 9 out of 16 cell lines (56%). The qMSP results matched well with TMZ sensitivity in 11 out of 12 (92%) glioma cell lines. In addition, methylation as detected by bisulphite sequencing seemed to be predictive of TMZ sensitivity in all six cell lines analysed (100%).Conclusion:The MGMT protein expression more than MGMT promoter methylation status predicts the response to TMZ in human tumour cell lines.

Prognostic Markers in Gliomas

In recent years, extensive molecular studies have identified diagnostic and prognostic markers in gliomas that reinforce the WHO histological classification. 1p19q codeletion, O(6)-methylguanine DNA methyltransferase (MGMT) status, and mutations of isocitrate dehydrogenases 1 and 2 (IDH1/IDH2) are currently the three most pertinent markers in diffuse gliomas. Ongoing clinical trials already stratify or select patients according to their 1p19q codeletion or MGMT status. Mutations of IDH1 have been recently identified as a major prognostic factor in diffuse gliomas. Future studies should determine whether IDH1/IDH2 mutations are predictive of response to treatments. Routine use of these markers considerably contributes to a more objective classification of gliomas based on both histological and molecular features.

O(6)-methylguanine DNA-methyltransferase methylation status can change between first surgery for newly diagnosed glioblastoma and second surgery for recurrence: clinical implications.

O(6)-methylguanine DNA-methyltransferase (MGMT) promoter methylation status is a prognostic factor in newly diagnosed glioblastoma patients. However, it is not yet clear whether, and if so how, MGMT methylation status may change. Moreover, it is unknown whether the prognostic role of this epigenetic feature is retained during the disease course. A retrospective analysis was made using a database of 614 glioblastoma patients treated prospectively from January 2000 to August 2008. We evaluated only patients who met the following inclusion criteria: age > or = 18 years; performance status 0-2; histological diagnosis of glioblastoma at both first and second surgery for recurrence; postoperative treatment consisting of: (i) radiotherapy (RT) followed by adjuvant temozolomide (TMZ) until 2005 and (ii) TMZ concurrent with and adjuvant to RT after 2005; a time interval > or = 3 months between first and second surgery. MGMT status was evaluated at first and second surgery in all 44 patients (M:F 32:12, median age: 49 years, range: 27-67 years). In 38 patients (86.4%), MGMT promoter status was assessable at both first and second surgery. MGMT methylation status, changed in 14 patients (37%) of second surgery samples and more frequently in methylated than in unmethylated patients (61.5% vs 24%, P = .03). The median survival was significantly influenced only by MGMT methylation status determined at first surgery (P = .04). Significant changes in MGMT methylation status during the course of GBM occur more frequently in MGMT methylated than unmethylated cases. MGMT methylation status determined at first surgery appears to be of prognostic value; however, it is not predictive of outcome following second surgery.

MGMT promoter methylation in malignant gliomas: ready for personalized medicine?

The DNA repair enzyme O(6)-methylguanine-DNA methyltransferase (MGMT) antagonizes the genotoxic effects of alkylating agents. MGMT promoter methylation is the key mechanism of MGMT gene silencing and predicts a favorable outcome in patients with glioblastoma who are exposed to alkylating agent chemotherapy. This biomarker is on the verge of entering clinical decision-making and is currently used to stratify or even select glioblastoma patients for clinical trials. In other subtypes of glioma, such as anaplastic gliomas, the relevance of MGMT promoter methylation might extend beyond the prediction of chemosensitivity, and could reflect a distinct molecular profile. Here, we review the most commonly used assays for evaluation of MGMT status, outline the prerequisites for standardized tests, and evaluate reasons for difficulties in reproducibility. We critically discuss the prognostic and predictive value of MGMT silencing, reviewing trials in which patients with different types of glioma were treated with various chemotherapy schedules, either up-front or at recurrence. Standardization of MGMT testing requires comparison of different technologies across laboratories and prospectively validated cut-off values for prognostic or predictive effects. Moreover, future clinical trials will need to determine, for each subtype of glioma, the degree to which MGMT promoter methylation is predictive or prognostic, and whether testing should become routine clinical practice.

Cilengitide modulates attachment and viability of human glioma cells, but not sensitivity to irradiation or temozolomide in vitro

Cilengitide is a cyclic peptide antagonist of integrins alphavbeta3 and alphavbeta5 that is currently being evaluated as a novel therapeutic agent for recurrent and newly diagnosed glioblastoma. Its mode of action is thought to be mainly antiangiogenic but may include direct effects on tumor cells, notably on attachment, migration, invasion, and viability. In this study we found that, at clinically relevant concentrations, cilengitide (1-100 microM) induces detachment in some but not all glioma cell lines, while the effect on cell viability is modest. Detachment induced by cilengitide could not be predicted by the level of expression of the cilengitide target molecules, alphavbeta3 and alphavbeta5, at the cell surface. Glioma cell death induced by cilengitide was associated with the generation of caspase activity, but caspase activity was not required for cell death since ectopic expression of cytokine response modifier (crm)-A or coexposure to the broad-spectrum caspase inhibitor zVAD-fmk was not protective. Moreover, forced expression of the antiapoptotic protein marker Bcl-X(L) or altering the p53 status did not modulate cilengitide-induced cell death. No consistent effects of cilengitide on glioma cell migration or invasiveness were observed in vitro. Preliminary clinical results indicate a preferential benefit from cilengitide added to temozolomide-based radiochemotherapy in patients with O(6)-methylguanine DNA methyltransferase (MGMT) gene promoter methylation. Accordingly, we also examined whether the MGMT status determines glioma cell responses to cilengitide alone or in combination with temozolomide. Neither ectopic expression of MGMT in MGMT-negative cells nor silencing the MGMT gene in MGMT-positive cells altered glioma cell responses to cilengitide alone or to cilengitide in combination with temozolomide. These data suggest that the beneficial clinical effects derived from cilengitide in vivo may arise from altered perfusion, which promotes temozolomide delivery to glioma cells.

Primary CNS lymphoma in the elderly: temozolomide therapy and MGMT status

This retrospective series explores temozolomide monotherapy in elderly patients with primary CNS lymphoma (PCNSL) and severe comorbidities. In 17 patients (62-90 years old), the complete response rate was 47%, median progression-free survival was 5 months, and median overall survival was 21 months. Five of 17 patients (29.4%) had prolonged responses for at least 12 months and survived for more than 24 months. Three of these patients had a methylated O(6)-methylguanine-DNA methyltransferase (MGMT) promoter, while the MGMT status was not assessable in the remaining two patients. Temozolomide monotherapy appears to be effective in a subgroup of elderly PCNSL patients and deserves further evaluation.

Prognostic value of O6-methylguanine-DNA methyltransferase status in glioblastoma patients, assessed by five different methods

This multicenter study assesses the value of O(6)-methylguanine-DNA methyltransferase (MGMT) status for predicting overall survival in glioblastoma patients. Five methods are used, to identify the approach with the best prognostic value. Eighty-one tumors were obtained from patients with glioblastomas treated by surgery and radiotherapy with concomitant temozolomide (TMZ) followed by adjuvant TMZ. MGMT promoter methylation was assessed by qualitative methyl-specific polymerase chain reaction (MSP), semiquantitative methyl-specific polymerase chain reaction (SQ-MSP), and pyrosequencing, while MGMT expression was measured at the RNA level by quantitative real-time PCR (Q-RT-PCR) and at the protein level by immunohistochemistry (IHC). MGMT promoter methylation as evaluated by MSP, SQ-MSP, and pyrosequencing was significantly correlated with overall survival. The best predictive value was obtained by pyrosequencing of one specific CpG position. Overall survival was 14 and 25 months for patients with percentages of methylation below and above the median, respectively. In contrast, MGMT status determined by Q-RT-PCR and IHC showed little or no correlation with overall survival, respectively. These results confirm the prognostic value of MGMT promoter methylation in glioblastoma patients initially treated with TMZ. SQ-MSP allowed better discrimination than classical MSP, and pyrosequencing represented a good option.