methylguanine-DNA Methyltransferase Methylation Research Articles

MGMT promoter methylation prediction based on multiparametric MRI via vision graph neural network.

Glioblastoma (GBM) is aggressive and malignant. The methylation status of the -methylguanine-DNA methyltransferase (MGMT) promoter in GBM tissue is considered an important biomarker for developing the most effective treatment plan. Although the standard method for assessing the MGMT promoter methylation status is via bisulfite modification and deoxyribonucleic acid (DNA) sequencing of biopsy or surgical specimens, a secondary automated method based on medical imaging may improve the efficiency and accuracy of those tests. We propose a deep vision graph neural network (ViG) using multiparametric magnetic resonance imaging (MRI) to predict the MGMT promoter methylation status noninvasively. Our model was compared to the RSNA radiogenomic classification winners. The dataset includes 583 usable patient cases. Combinations of MRI sequences were compared. Our multi-sequence fusion strategy was compared with those using single MR sequences. Our best model [Fluid Attenuated Inversion Recovery (FLAIR), T1-weighted pre-contrast (T1w), T2-weighted (T2)] outperformed the winning models with a test area under the curve (AUC) of 0.628, an accuracy of 0.632, a precision of 0.646, a recall of 0.677, a specificity of 0.581, and an F1 score of 0.661. Compared to the winning models with single MR sequences, our ViG utilizing fused-MRI showed a significant improvement statistically in AUC scores, which are FLAIR (), T1w (), T1wCE (), and T2 (). Our model is superior to challenge champions. A graph representation of the medical images enabled good handling of complexity and irregularity. Our work provides an automatic secondary check pipeline to ensure the correctness of MGMT methylation status prediction.

Outcomes of Fluorescence-Guided vs White Light Resection of Glioblastoma in a Single Institution.

Background Glioblastoma (GBM) is the most common malignant primary brain tumour and confers a very poor prognosis. Maximal safe resection of tumour is the goal of neurosurgical intervention and may be more easily achieved through the use of surgical adjuncts such as fluorescence-guided surgery (FGS). 5-Aminolevulinic acid (5-ALA) accumulates in GBM tissue and fluorescered, distinguishing tumour cells from the surrounding tissue and therefore making resection easier. 5-ALA-guided resection in GBM has been shown to increase resection rates and prolongprogression-free survival without impacting post-operative morbidity. Radiotherapy and concomitant chemotherapy also improve survival in GBM. Other factors such as patient age and molecular status of the tumour also impact prognosis. Aims The aim of this study was to compare the outcomes of 5-ALA vs white light-guided resection for glioblastoma in the west of Scotland. Methods This was a retrospective analysis of baseline characteristics (age, sex, tumour molecular markers, radiotherapy, chemotherapy, anatomical location of tumour and treatment group) and outcomes (mortality, survival, degree of resection and performance status) of 239 patients who underwent primary resection of glioblastoma over a four-year period (2017-2020). A variety of statistical methods were used to analyse the relationship between each variable and surgical technique; multivariate Cox regression and the Kaplan-Meier method were used in survival analysis. Results 5-ALA-guided resection substantiallyimproved resection rates (74.0% vs 40.2%). Mortality at 15 months was 5.1% lower in the 5-ALA group (52.0% vs 57.1%, p = 0.53), and patients lived an average of 68 days longer compared to the white light group (444 days vs 376 days, p = 0.21). There were negligible differences between treatment groups in terms of post-operative performance status (PS) and post-operative complications. In our multivariate Cox regression model, six factors were statistically significant at a level of p ≤ 0.05: age, radiotherapy, chemotherapy, O(6)-methylguanine-DNA methyltransferase (MGMT) methylation, anatomical location and >90% resection. Receiving chemotherapy and radiotherapy, MGMT methylation and undergoing >90% resection conferred a survival benefit at 15 months. Older age and multi-focal disease were related to a worsened mortality rate. Undergoing radiotherapy and maximal resection were the two greatest predictors of improved survival, reducing mortality risk by 58% and 51%, respectively. Conclusion 5-ALA-guided resection improved resection rates without impacting post-operative morbidity. 5-ALA-guided resection was associated with improved survival and lower mortality rate, but this was not statistically significant. Receiving chemoradiotherapy, MGMT methylation and undergoing maximal resection conferred a survival benefit, whilst older age and multi-focal disease were associated with a poorer prognosis.

Outcomes and Treatment Algorithm in Glioblastoma Patients 80 Years and Older

The current standard of care for patients with glioblastoma (GBM) is maximal safe resection followed by adjuvant radiation therapy with concurrent temozolomide chemotherapy. Previous studies that identified this treatment regimen focused on younger patients with GBM. The proportion of patients with GBM over the age of 80 years is increasing. We investigate whether elderly patients benefit from the current standard of care with additional maximal safe resection. Clinical, operative, radiographic, demographic, genetic, and outcomes data were retrospectively collected for patients treated for histologically confirmed World Health Organization grade 4 GBM at University of Pittsburgh Medical Center from 2009 to 2020. Only patients 80 years and older were included (n= 123). Statistically significant values were set at P < 0.05. A univariate Cox proportional hazards analysis of GBM patients aged >80 years identified the use of temozolomide, radiation, Karnofsky Performance Status (KPS) > 70, and methylguanine DNA methyltransferase methylation with increased overall survival (OS). Further multivariate Cox proportional hazards model analysis showed that the variables identified in the univariate analysis passed multicollinearity testing, and that use of temozolomide, KPS >70, and gross total resection were shown to significantly impact survival. Survival analysis showed that patients with biopsy alone had a shorter median OS compared with patients who received resection, temozolomide, and radiation (P < 0.0001, median OS 1.6 vs. 7.5 months). Additionally, patients who underwent biopsy and then received temozolomide and radiation had a shorter median OS when compared with patients who received resection, temozolomide, and radiation (P= 0.0047, median OS 3.6 vs. 7.5 months). For elderly patients with KPS >70, GTR followed by radiation and temozolomide is associated with maximum OS.

Identification of MGMT Downregulation Induced by miRNA in Glioblastoma and Possible Effect on Temozolomide Sensitivity.

Glioblastoma multiforme (GBM) remains one of the tumors with the worst prognosis. In recent years, a better overall survival (OS) has been described in cases subjected to Gross Total Resection (GTR) that were presenting hypermethylation of Methylguanine-DNA methyltransferase (MGMT) promoter. Recently, also the expression of specific miRNAs involved in MGMT silencing has been related to survival. In this study, we evaluate MGMT expression by immunohistochemistry (IHC), MGMT promoter methylation and miRNA expression in 112 GBMs and correlate the data to patients' clinical outcomes. Statistical analyses demonstrate a significant association between positive MGMT IHC and the expression of miR-181c, miR-195, miR-648 and miR-767.3p between unmethylated cases and the low expression of miR-181d and miR-648 and between methylated cases and the low expression of miR-196b. Addressing the concerns of clinical associations, a better OS has been described in presence of negative MGMT IHC, in methylated patients and in the cases with miR-21, miR-196b overexpression or miR-767.3 downregulation. In addition, a better progression-free survival (PFS) is associated with MGMT methylation and GTR but not with MGMT IHC and miRNA expression. In conclusion, our data reinforce the clinical relevance of miRNA expression as an additional marker to predict efficacy of chemoradiation in GBM.

Peritumoral ADC Values Correlate with the MGMT Methylation Status in Patients with Glioblastoma

Different results have been reported concerning the relationship of the apparent diffusion coefficient (ADC) values and the status of methylation as the promoter gene for the enzyme methylguanine-DNA methyltransferase (MGMT) in patients with glioblastomas (GBs). The aim of this study was to investigate if there were correlations between the ADC values of the enhancing tumor and peritumoral areas of GBs and the MGMT methylation status. In this retrospective study, we included 42 patients with newly diagnosed unilocular GB with one MRI study prior to any treatment and histopathological data. After co-registration of ADC maps with T1-weighted sequences after contrast administration and dynamic susceptibility contrast (DSC) perfusion, we manually selected one region-of-interest (ROI) in the enhancing and perfused tumor and one ROI in the peritumoral white matter. Both ROIs were mirrored in the healthy hemisphere for normalization. In the peritumoral white matter, absolute and normalized ADC values were significantly higher in patients with MGMT-unmethylated tumors, as compared to patients with MGMT-methylated tumors (absolute values p = 0.002, normalized p = 0.0007). There were no significant differences in the enhancing tumor parts. The ADC values in the peritumoral region correlated with MGMT methylation status, confirmed by normalized ADC values. In contrast to other studies, we could not find a correlation between the ADC values or the normalized ADC values and the MGMT methylation status in the enhancing tumor parts.

Immunohistochemical Expression of PD-L1 and IDH1 with Detection of MGMT Promoter Methylation in Astrocytoma

Programmed death ligand 1 (PD-L1) expression was suggested as a poor prognostic predictor for glioblastoma. While isocitrate dehydrogenase (IDH) has been linked to enhanced overall survival in glioma cells. In glioblastoma patients receiving treatment with alkylating drugs, the methylguanine-DNA methyltransferase (MGMT) promoter's methylation status has been discovered as a potent and distinct predictor of good survival. In this study, we aimed to investigate the expression rate of PD-L1, IDH1, and MGMT methylation in patients with different grades of astrocytoma. The present retrospective study retrieved the data and archived paraffin blocks of 60 cases of astrocytoma. Immunohistochemical evaluation was done to assess the expressions of PD-L1 and IDH1, Methylation-specific-PCR was used to investigate the MGMT promoter. This study included astrocytoma grade II 18% (11/60), grade III 22% (13/60), grade IV 60% (36 cases). PD-L1 expression was detected in 82% of all studied cases (49/60) while IDH1 mutant astrocytoma were 73% (44/60) & methylation was reported in 58.3% (35 cases). High grade astrocytoma showed highrer expression of PD-L1 & IDH1 but with insignificant correlation (p=0.989). There is a relatively high expression of PD-L1 and IDH1 in patients with astrocytoma. More than half of the patients presented with MGMT promoter methylation. Further studies with larger sample size are required to investigate the association between these biomarkers and characteristics of patients with astrocytoma.

RTID-05. TRIAL IN PROGRESS: DOSE-FINDING STUDY AND EVALUATION OF [177LU]LU-DOTA-TATE IN COMBINATION WITH STANDARD OF CARE IN NEWLY DIAGNOSED GLIOBLASTOMA AND AS A SINGLE AGENT IN RECURRENT GLIOBLASTOMA

Abstract Patients with glioblastoma have a poor prognosis despite extensive efforts to develop new treatments. Standard of care for newly diagnosed patients is surgery followed by radiotherapy plus temozolomide. However, temozolomide only provides a clinical benefit to patients who have glioblastomas with a methylated O(6)-methylguanine-DNA methyltransferase (MGMT) gene promotor. Disease recurrence is inevitable and subsequent treatments have poor outcomes due to limited efficacy and a lack of established therapeutic regimens. [68Ga]Ga-DOTA-TATE is a radioligand imaging agent that selectively binds to somatostatin receptors (SSTRs) and is used with positron emission tomography (PET) for the molecular imaging of SSTR-positive neuroendocrine tumors. Glioblastoma tumoral PET scans have shown moderate uptake of [68Ga]Ga-DOTA-TATE. The radioligand therapy [177Lu]Lu-DOTA-TATE has the same SSTR-targeted ligand and demonstrated promising clinical activity in a pilot study in patients with Grade III/IV gliomas. This Phase 1b open-label dose-finding study (NCT05109728) will determine the recommended [177Lu]Lu-DOTA-TATE dose in three different groups of glioblastoma (newly diagnosed with methylated MGMT promoter [group 1], newly diagnosed with unmethylated MGMT promoter [group 2], and recurrent disease [group 3]). All participants will be scanned with [68Ga]Ga-DOTA-TATE imaging during screening and, for each group, approximately 15 participants will be enrolled in consecutive cohorts of 3–6 participants. All participants will receive up to 6 administrations of [177Lu]Lu-DOTA-TATE (initial dose 150 mCi with 3 provisional dose levels up to 250 mCi) using the Bayesian Optimal Interval design. Group 1 will receive [177Lu]Lu-DOTA-TATE every 4 weeks with concomitant radiotherapy and temozolomide, followed by temozolomide maintenance. Group 2 will receive [177Lu]Lu-DOTA-TATE every 4 weeks for 3 doses with radiotherapy then every 3 weeks as a single agent. Group 3 will receive [177Lu]Lu-DOTA-TATE only, every 3 weeks. The primary endpoint is incidence of dose limiting toxicity. Secondary endpoints include overall objective status (modified RANO criteria), progression free survival, overall survival and safety.

P11.60.A Quantitative analysis of the MGMT methylation status of glioblastomas in the light of the 2021 WHO classification

Abstract Background Glioblastomas with methylation of the promotor region of the O(6)-methylguanine-DNA methyltransferase (MGMT) gene exhibit increased sensitivity to alkylating chemotherapy. Quantitative assessment of the MGMT promotor methylation status might provide additional prognostic information. The aim of our study was to determine a quantitative threshold of MGMT methylation for better survival among patients with isocitrate dehydrogenase (IDH) wildtype glioblastomas. Material and Methods We included consecutive patients treated at our department between 11/2010 and 08/2018. Inclusion criteria were i) diagnosis of glioblastoma, IDH wildtype according to the 2021 WHO classification of tumors of the central nervous system, ii) availability of a quantitative MGMT promotor methylation analysis, iii) absence of an IDH mutation as determined by immunohistochemistry and/or sequencing, and iii) age ≥ 18 years. The primary endpoint was overall survival. To evaluate a threshold for a benefit from temozolomide (TMZ), a secondary analysis included only tumors exposed to TMZ, which were compared to a historical cohort treated by radiotherapy only with a median survival of 13.0 months. Results Between 11/2010 and 08/2018, 321 patients diagnosed with a glioblastoma, IDH wildtype meeting our inclusion criteria were included. Mean age at diagnosis was 64.1 years (± 11.4). Median overall survival was 12.6 months. Kaplan-Meier analysis showed better survival for all groups with ≥ 16% methylation. Similarly, Cox regression analysis confirmed significantly better survival in the groups with 16-30%, 31-60%, and 61-100% methylation. In contrast, survival in the group with 1-15% methylation was similar to that with unmethylated promotor regions. When the percentage of methylation of each patient treated with TMZ was plotted on an ROC curve using survival &amp;gt; 13.0 months as outcome parameter, the Youden index was maximal at ≥ 16% methylation. Conclusion Better survival is seen in patients with glioblastomas with ≥ 16% methylation of the MGMT promotor region than with &amp;lt; 16% methylation. Survival with tumors with 1-15% methylation is similar to with unmethylated tumors. Above 16% methylation, we found no additional benefit with increasing methylation.

Quantitative Analysis of the MGMT Methylation Status of Glioblastomas in Light of the 2021 WHO Classification.

Simple SummaryThe status of a DNA repair protein called MGMT is a prognostic marker in patients with glioblastomas, the most frequent malignant brain tumor. Epigenetic silencing of this gene predicts increased sensitivity to chemotherapy. Silencing is assessed by analyzing modifications, so-called methylation, of a certain gene region. Whereas most studies report such information only in a qualitative manner, quantitative testing might provide additional information. The aim of our study was to determine a quantitative threshold for better survival among patients with glioblastomas. Among 321 patients suffering from glioblastoma, we found better survival in patients with glioblastomas that have ≥16% methylation of a particular region of the MGMT gene. Above 16% methylation, we found no additional benefit with increasing degree of methylation. We suggest using this threshold for selection in clinical trials and for patient counseling.Background: Glioblastomas with methylation of the promoter region of the O(6)-methylguanine-DNA methyltransferase (MGMT) gene exhibit increased sensitivity to alkylating chemotherapy. Quantitative assessment of the MGMT promoter methylation status might provide additional prognostic information. The aim of our study was to determine a quantitative methylation threshold for better survival among patients with glioblastomas. Methods: We included consecutive patients ≥18 years treated at our department between 11/2010 and 08/2018 for a glioblastoma, IDH wildtype, undergoing quantitative MGMT promoter methylation analysis. The primary endpoint was overall survival. Results: A total of 321 patients were included. Median overall survival was 12.6 months. Kaplan–Meier and adjusted Cox regression analysis showed better survival for the groups with 16–30%, 31–60%, and 61–100% methylation. In contrast, survival in the group with 1–15% methylation was similar to those with unmethylated promoter regions. A secondary analysis confirmed this threshold. Conclusions: Better survival is observed in patients with glioblastomas with ≥16% methylation of the MGMT promoter region than with <16% methylation. Survival with tumors with 1–15% methylation is similar to with unmethylated tumors. Above 16% methylation, we found no additional benefit with increasing methylation.

CTNI-21. PHASE 2 STUDY OF VAL-083 AND RADIOTHERAPY IN NEWLY DIAGNOSED MGMT-UNMETHYLATED GBM

Abstract Approximately 60% of glioblastoma multiforme (GBM) patients possess an unmethylated methylguanine DNA-methyltransferase (MGMT) gene promoter, which confers a limited clinical response to standard-of-care treatment with temozolomide (TMZ), resulting in shorter median survival when compared to patients with a methylated MGMT promoter. VAL-083 is a novel bi-functional DNA targeting agent that induces interstrand DNA cross-links at N7-guanine, leading to DNA double-strand breaks and ultimately cell death. VAL-083 circumvents MGMT-mediated TMZ resistance in vitro and in vivo. A Phase 2 study has been conducted to evaluate efficacy and safety of VAL-083 when administered concurrently with radiation therapy (RT) in newly diagnosed MGMT unmethylated GBM. The study was conducted in 2 stages: Stage 1 was a dose-escalation phase to confirm the dose of VAL-083 in this setting. Patients received VAL-083 at 20, 30, or 40 mg/m2/day x 3 days every 21 days along with standard radiation treatment (RT) (2 Gy/day, 5 days/week for 6 weeks). At the end of this stage, 30 mg/m2/day of VAL-083 in combination with RT was generally safe and well-tolerated. Stage 2 was an expansion phase to enroll up to 20 additional patients at the 30 mg/m2/day of VAL-083 in combination with RT. All patients have been enrolled, with a total of 29 patients in the study, and 25 patients receiving 30 mg/m2/day VAL-083. All 29 patients have completed treatment and patients are in the follow-up phase of the study. Consistent with our prior experience, myelosuppression was the most common adverse event. As of March 2021, 22/29 (75.9%) subjects had disease progression. The median progression free survival for all patients enrolled was 9.3 (95%CI: 6.4-12.0) months. Sixteen (16/29; 55.2%) patients had died, and median overall survival for all patients enrolled was 19.6 (95%CI: 14.0-22.4) months. Further safety and efficacy updates will be presented at the meeting. Clinicaltrials.gov identifier: NCT03050736.

Predictors of Survival in Turkish Patients with Primary Glioblastoma.

An increasing number of biomarkers of primary glioblastoma (GBM) have recently been described. We aimed to investigate the biological and clinical factors that affect survival in Turkish patients with primary GBM. The clinical and demographic data of all patients with primary GBM diagnosed between 2007 and 2016 were evaluated. In all the patients? pathological specimens, O6 methylguanine-DNA methyltransferase (MGMT) methylation and isocitrate dehydrogenase (IDH) 1 mutation were detected retrospectively by immunohistochemistry. Kaplan-Meier survival analysis, log-rank test, and multivariate analyses of the Cox hazard proportional model for all the variables were performed using the SPSS statistical package. The treatment details and other patient-related factors were identified, and their correlations were analyzed. We enrolled 137 primary GBM patients to the study. Median progression free survival (PFS) was 8.57 months (95% CI:6.8-9.5) and median overall survival (OS) was 12 months (95% CI:10.8-13.3). IDH-1 mutations were detected in 21 primary GBMs (15.3%). PFS was 15.43 ± 1.95 months. Survival rates were higher, but no statistically significant difference (p=0.074). MGMT methylation was detected in 40 primary GBMs (29.2%). OS and PFS of MGMT (+) cases were higher than MGMT(-) cases (p=0.001; p=0.001 respectively). Ki67 (%) measurement (10%-90%) average is 32.64 ± 16.56. No statistically significant between higher and lower ki67 levels (p=0.510, p=0.505 respectively). KPS (%) more than 70 at the time of diagnosis statistically significant longer median OS and PFS (p=0.001). PFS and OS were higher in all treatment modalities. The most important factors that affected survival were performance score, MGMT methylation status, systemic oncologic therapy, and IDH mutation in the Turkish population with primary GBM. We demonstrated that MGMT methylation and higher KPS levels were associated with significiantly longer OS and PFS.

NIMG-37. DELAYED PSEUDOPROGRESSION IN TWO PATIENTS UNDERGOING TTFIELDS TREATMENT FOR NEWLY DIAGNOSED GLIOBLASTOMA

Abstract PURPOSE Tumor treatment fields (TTFields) are approved by the FDA for newly diagnosed as well as recurrent glioblastoma (GBM). TTFields have been shown to extend survival by 4.9 months in newly diagnosed GBM, and a landmark overall survival rate of 13% at 5 years. However, the specific effects remain widely unknown, which has prevented widespread clinical use of this treatment. METHODS This case study examines two glioblastoma patients, IDH-1 wildtype, MGMT unmethylated, that received TTFields (Optune) in addition to maintenance temozolomide (TMZ) following radiation (RT). Both cases were followed using standard MR imaging. Second resections were performed due to radiographic progression of contrast enhancement. RESULTS Although imaging was concerning for tumor progression, pathology showed only treatment effect, ultimately leading to the diagnosis of pseudoprogression. Both patients fell outside the normal expected timeline for chemo-radiation induced pseudoprogression. Based on the pathology, both patients resumed treatment with TMZ/TTFields. One patient expired at 25 months and one is still alive. CONCLUSIONS Pathologic confirmation was essential for guiding further treatment and allowed patients to continue treatment that was effective despite contrary indications on imaging. These findings suggest that pathological confirmation should be strongly considered in patients receiving TMZ/TTFields who develop radiographic progression, potentially with a less invasive biopsy procedure. Future studies should look to characterize the underlying mechanism of interaction between TTFields/TMZ and quantify the prevalence, associated risk factors and whether there is a genotype more susceptible. Both patients reported here had O(6)-methylguanine-DNA methyltransferase (MGMT) unmethylated GBM, and while about 60% of glioblastomas are diagnosed likewise, it is possible that MGMT methylation status plays a role in TTFields response. Better characterization of this phenomenon will improve treatment guidance, potentially reducing unnecessary surgeries and the discontinuance of successful therapies.

CTNI-76. PHASE 2 CLINICAL TRIAL OF VAL-083 IN NEWLY DIAGNOSED MGMT-UNMETHYLATED GBM

Abstract Approximately 60% of glioblastoma multiforme (GBM) patients possess an unmethylated methylguanine DNA-methyltransferase (MGMT) promoter region, which confers a limited response to standard-of-care treatment with temozolomide (TMZ), resulting in shorter median survival when compared to patients with methylated MGMT promoter. VAL-083 is a novel bi-functional DNA targeting agent that induces inter-strand cross-links at N7-guanine, leading to DNA double-strand breaks and ultimately cell death. VAL-083 circumvents MGMT-mediated TMZ resistance in vitro and in vivo. A Phase 2 study has been initiated for VAL-083 in newly diagnosed MGMT unmethylated GBM. The study has 2 stages: Stage 1 is a dose-escalation safety and tolerability phase to confirm the phase 2 dose of VAL-083 when administered concurrently with radiation therapy (RT). Patients received VAL-083 at 20, 30, or 40 mg/m2/day x 3 days every 21 days along with standard radiation treatment (RT) (2 Gy/day, 5 days/week). The dose escalation stage is complete, and 30 mg/m2/day of VAL-083 in combination with RT was generally safe and well-tolerated. Stage 2 comprises an expansion phase to enroll up to 20 additional patients at the 30 mg/m2/day of VAL-083 in combination with RT. As of June 2, 2020, all patients have been enrolled, with a total of 29 patients in the study, and 25 patients receiving 30 mg/m2/day VAL-083. Of the 29 patients enrolled, 27 have completed their prospectively planned MRI scans and had their initial assessment for tumor response. Two additional patients died prior to their post-cycle 3 MRI. Consistent with our prior experience, myelosuppression was the most common adverse event. Three patients have experienced dose-limiting toxicities - one (1/3; 33%) at the 40 mg/m2/day and two (2/25; 8%) at the 30 mg/m2/day dose. Further safety and efficacy updates will be presented at the meeting. Clinicaltrials.gov identifier: NCT03050736.

The Impact of Surgery in IDH 1 Wild Type Glioblastoma in Relation With the MGMT Deregulation.

Object: The treatment of choice in glioblastoma (GBM) is the maximal surgical extent of resection (EOR) followed by adjuvant chemo-radiotherapy. Furthermore, methylguanine-DNA methyltransferase (MGMT) promoter methylation is associated with prolonged overall survival (OS) and progression free survival (PFS). The objective of the present study is correlate the biomolecular aspects in relation with EOR.Materials and methods: We analyzed a series of 116 patients with IDH-1 wild type GBM and different EOR (Gross Total Resection—GTR-, Partial Resection—PR- and Biopsy), treated with adjuvant chemo-radiotherapy. The MGMT status was analyzed in terms of promoter methylation and protein expression.Results: When GTR was possible, OS and PFS were significantly better compared to the other two groups (p = 0.001 and p = 0.035, respectively). MGMT methylation was significantly associated with better OS in the biopsy group (p = 0.022) and better OS and PFS in PR (p = 0.02 and p = 0.012, respectively), but not in the GTR group (p = 0.252 for OS, p = 0.256 for PFS) nor the PFS in the biopsy group (p = 0.259). MGMT protein expression levels do not show any association with OS and PFS, regardless of the type of surgery.Conclusions: Our study confirms the positive association of a safe maximal EOR with better OS and PFS, and indicates a positive prognostic value of MGMT methylation status only in case of the presence of residual tumor tissue. MGMT protein expression seems not to play a clinical role in relation with the type of surgery.

Noninvasive O6 Methylguanine-DNA Methyltransferase Status Prediction in Glioblastoma Multiforme Cancer Using Magnetic Resonance Imaging Radiomics Features: Univariate and Multivariate Radiogenomics Analysis

This study aimed to predict methylation status of the O6 methylguanine-DNA methyltransferase (MGMT) gene promoter status by using magnetic resonance imaging radiomics features, as well as univariate and multivariate analysis. Eighty-two patients who had an MGMT methylation status were included in this study. Tumors were manually segmented in the 4 regions of magnetic resonance images, 1) whole tumor, 2) active/enhanced region, 3) necrotic regions, and 4) edema regions. About 7000 radiomics features were extracted for each patient. Feature selection and classifier were used to predict MGMT status through different machine learning algorithms. The area under the curve (AUC) of the receiver operating characteristic curve was used for model evaluations. Regarding univariate analysis, the Inverse Variance feature From Gray Level Co-occurrence Matrix in whole tumor segment with 4.5 mm Sigma of Laplacian of Gaussian filter with AUC of 0.71 (P value= 0.002) was found to be the best predictor. For multivariate analysis, the Decision Tree classifier with Select from Model feature selector and LOG (Laplacian of Gaussian) filter in edema region had the highest performance (AUC, 0.78), followed by Ada-Boost classifier with Select from Model feature selector and LOG filter in edema region (AUC, 0.74). This study showed that radiomics using machine learning algorithms is a feasible noninvasive approach to predict MGMT methylation status in patients with glioblastoma multiforme cancer.

The prevalence and persistence of aberrant promoter DNA methylation in benzene-exposed Chinese workers

Aberrant DNA methylation patterns are common in cancers and environmental pollutant exposed subjects. Up to date, few studies have examined the aberrant DNA methylation patterns in benzene exposed workers. We recruited 141 benzene-exposed workers, including 83 benzene-exposed workers from a shoe factory in Wenzhou and 58 workers from a painting workshop in Wuhu, 35 workers in Wuhu were followed from 2009 to 2013, and 48 indoor workers as controls from Wenzhou. We used high-resolution melting (HRM) to quantitate human samples of DNA methylation in long interspersed nuclear element-1 (LINE-1), (6)-methylguanine-DNA methyltransferase (MGMT), and DNA mismatch repair gene human mutator L homologue 1 (hMLH1). AML-5 cells were treated with benzoquinone (BQ) and hydroquinone (HQ), and the promoter methylation of MGMT and hMLH1 was detected using the bisulfite sequencing PCR method. The degree of LINE-1 methylation in benzene-exposed workers was significantly lower than that of the controls (p<0.001), and the degree of MGMT (p<0.001) and hMLH1 (p = 0.01) methylation was significantly higher than that of the controls. The in vitro study validated the aberrant hypermethylation of hMLH1 after treatment with BQ. Among the cohort workers who were followed from 2009 to 2013, the LINE1 methylation elevated in 2013 than 2009 (p = 0.004), and premotor methylation in hMLH1 reduced in 2013 than 2009 (p = 0.045) with the reduction of the benzene exposure. This study provides evidence that benzene exposure can induce LINE-1 hypomethylation and DNA repair gene hypermethylation.

Is a reduction of radiation dose feasible in patients affected by glioblastoma undergoing radio-chemotherapy according to MGMT promoter methylation status without jeopardizing survival?

ObjectiveTo explore therapeutic results of different radiotherapy (RT) dose schedules combined to Temozolomide (TMZ)-RT treatment in newly diagnosed glioblastoma (GB), according to the O (6)-methylguanine-DNA methyltransferase (MGMT) methylation status. Patients and methodsPatients with newly diagnosed GB received either standard (60-59.4 Gy) or reduced (54-52 Gy) dose radiation therapy (RT) with concurrent and adjuvant TMZ between June 2010 and October 2016. We retrospectively evaluated the therapeutic effectiveness of the RT ranges schedules in terms of overall survival (OS) with univariate and multivariate analysis, after analyzing the MGMT methylation status. ResultsOne hundred and seventeen patients were selected for the present analysis out of 146 total treated patients accrued. Seventy-two out of the selected cases received the standard RT-TMZ course (SDRT-TMZ) whereas the remaining 45 underwent the reduced dose schedule (RDRT-TMZ). The analysis according to the MGMT promoter methylation status showed that, in methylated-MGMT GB patients, SDRT-TMZ and RDRT-TMZ groups did not show different median OS (p = ns) according to the two RT schedules, independently by the extent of surgical resection. Instead, a difference in survival outcomes was confirmed in unmethylated-MGMT GB patients with better survival for patients undergoing to SDRT, particularly in sub-total resection. ConclusionIn our experience, a reduction of radiation dose schedule does not seem to jeopardize survival in methylated-MGMT patients independently by the extent of resection.A therapeutic approach to a standard reduction of RT dose for the methylated subset of patients may be feasible and could deserve prospective trials for validation.

Prognostic value of test(s) for O6 -methylguanine-DNA methyltransferase (MGMT) promoter methylation for predicting overall survival in people with glioblastoma treated with temozolomide

Background Glioblastoma is an aggressive form of brain cancer. Approximately five in 100 people with glioblastoma survive for five years past diagnosis. Glioblastomas that have a particular modification to their DNA (called methylation) in a particular region (the O6-methylguanine-DNA methyltransferase (MGMT) promoter) respond better to treatment with chemotherapy using a drug called temozolomide. Objectives To determine which method for assessing MGMT methylation status best predicts overall survival in people diagnosed with glioblastoma who are treated with temozolomide. Search methods We searched MEDLINE, Embase, BIOSIS, Web of Science Conference Proceedings Citation Index to December 2018, and examined reference lists. For economic evaluation studies, we additionally searched NHS Economic Evaluation Database (EED) up to December 2014. Selection criteria Eligible studies were longitudinal (cohort) studies of adults with diagnosed glioblastoma treated with temozolomide with/without radiotherapy/surgery. Studies had to have related MGMT status in tumour tissue (assessed by one or more method) with overall survival and presented results as hazard ratios or with sufficient information (e.g. Kaplan-Meier curves) for us to estimate hazard ratios. We focused mainly on studies comparing two or more methods, and listed brief details of articles that examined a single method of measuring MGMT promoter methylation. We also sought economic evaluations conducted alongside trials, modelling studies and cost analysis. Data collection and analysis Two review authors independently undertook all steps of the identification and data extraction process for multiple-method studies. We assessed risk of bias and applicability using our own modified and extended version of the QUality In Prognosis Studies (QUIPS) tool. We compared different techniques, exact promoter regions (5'-cytosine-phosphate-guanine-3' (CpG) sites) and thresholds for interpretation within studies by examining hazard ratios. We performed meta-analyses for comparisons of the three most commonly examined methods (immunohistochemistry (IHC), methylation-specific polymerase chain reaction (MSP) and pyrosequencing (PSQ)), with ratios of hazard ratios (RHR), using an imputed value of the correlation between results based on the same individuals. Main results We included 32 independent cohorts involving 3474 people that compared two or more methods. We found evidence that MSP (CpG sites 76 to 80 and 84 to 87) is more prognostic than IHC for MGMT protein at varying thresholds (RHR 1.31, 95% confidence interval (CI) 1.01 to 1.71). We also found evidence that PSQ is more prognostic than IHC for MGMT protein at various thresholds (RHR 1.36, 95% CI 1.01 to 1.84). The data suggest that PSQ (mainly at CpG sites 74 to 78, using various thresholds) is slightly more prognostic than MSP at sites 76 to 80 and 84 to 87 (RHR 1.14, 95% CI 0.87 to 1.48). Many variants of PSQ have been compared, although we did not see any strong and consistent messages from the results. Targeting multiple CpG sites is likely to be more prognostic than targeting just one. In addition, we identified and summarised 190 articles describing a single method for measuring MGMT promoter methylation status. Authors' conclusions PSQ and MSP appear more prognostic for overall survival than IHC. Strong evidence is not available to draw conclusions with confidence about the best CpG sites or thresholds for quantitative methods. MSP has been studied mainly for CpG sites 76 to 80 and 84 to 87 and PSQ at CpG sites ranging from 72 to 95. A threshold of 9% for CpG sites 74 to 78 performed better than higher thresholds of 28% or 29% in two of three good-quality studies making such comparisons.

The search for a melanoma-tailored chemotherapy in the new era of personalized therapy: a phase II study of chemo-modulating temozolomide followed by fotemustine and a cooperative study of GOIM (Gruppo Oncologico Italia Meridionale)

BackgroundIt is frequently asked whether chemotherapy can still play a role in metastatic melanoma considering the effectiveness of the available drugs today, including antiCTLA4/antiPD1 immunotherapy and antiBRAF/antiMEK inhibitors. However, only approximately half of patients respond to these drugs, and the majority progress after 6–11 months. Therefore, a need for other therapeutic options is still very much apparent.We report the first large trial of a sequential full dose of fotemustine (FM) preceded by a low dose of temozolomide (TMZ) as a chemo-modulator in order to inactivate the DNA repair action of O(6)-methylguanine DNA-methyltransferase (MGMT). Primary endpoints were overall response and safety. We also evaluated specific biological parameters aiming to tailor these chemotherapies to selected patients.MethodsA total of 69 consecutive patients were enrolled. The main features included a median age of 60 years (21–81) and M1c stage, observed in 74% of the patients, with brain metastases in 15% and high LDH levels in 42% of the patients. The following schedule was used: oral TMZ 100 mg/m2 on days 1 and 2 and FM iv 100 mg/m2 on day 2, 4 h after TMZ; A translational study aiming to analyse MGMT methylation status and base-excision repair (BER) gene expression was performed in a subset of 14 patients.ResultsWe reported an overall response rate of 30.3% with 3 complete responses and a disease control rate of 50.5%. The related toxicity rate was low and mainly of haematological types. Although our population had a very poor prognosis, we observed a PFS of 6 months and an OS of 10 months. A non-significant correlation with response was found with the mean expression level of the three genes involved in the BER pathway (APE1, XRCC1 and PARP1), whereas no association was found with MGMT methylation status.ConclusionThis schedule could represent a good alternative for patients who are not eligible for immune or targeted therapy or whose previous therapies have failed.Trial registrationEUDRACT 2009–016487-36l; date of registration 23 June 2010.

Genetics and Epigenetics of Glioblastoma: Therapeutic Challenges

Glioblastoma is a brain tumor that develops due to both genetic and epigenetic risk factors. Crosstalk between the genetic and the epigenetic offers new possibilities for therapy. Abnormal methylation of methylguanine-DNA methyltransferase (MGMT) promoter region and isocitrate dehydrogenase 1 (IDH1) mutations are prognostic and therapeutic response markers in glioblastoma. Mutations in genes such as epidermal growth factor receptor, TP53, and P16 have been reported in glioblastoma; therefore, they might associate with survival and worth to be used in estimating survival risks. MKI67 expression associates with posttreatment such as adjuvant radiotherapy results evaluation. On the other hand, monosomies, such as deletions of chromosome 10, especially q23 and q25–26, are good markers for estimating the progression and aggressiveness of glioblastoma. The profile of MGMT methylation is modified in glioblastoma and hence can be a good target for epigenetic drugs. Other useful strategies in the treatment of gliomas include several micro-RNAs (MiRs) which are alerted in glioblastoma and which affect the regulation of mRNAs are associated with gene expression profiles of the disease. Epigenetic drugs, such as azacitidine and decitabine, which belong to the DNA methyltransferases (DNMT) inhibitor 5-aza-2'deoxycytidine (5-aza-dC), can suppress DNMT1 and stimulate tumor suppressor genes expression. MGMT methylation status and IDH mutational status are two valuable prognosis and therapeutic response markers in glioblastoma. Regulation of glioblastoma through epigenetic drugs, such as not only inhibitors of EZH2, histone deacetylase, and DNMT, but also MiRs, are promising approaches in glioblastoma treatment. Improves in understanding cancer genetic and epigenetic disruptions is the key point in solving the puzzle of glioblastoma treatment.