Methylguanine Methyltransferase Promoter Research Articles

O6 -methylguanine methyltransferase promoter methylation status of glioblastoma cell line clonal population.

Glioblastoma (GBM) remains a treatment-resistant malignant brain tumor in large part because of its genetic heterogeneity and epigenetic plasticity. In this study, we investigated the epigenetic heterogeneity of GBM by evaluating the methylation status of the O6 -methylguanine methyltransferase (MGMT) promoter in individual clones of a single cell derived from GBM cell lines. The U251 and U373 GBM cell lines, from the Brain Tumour Research Centre of the Montreal Neurological Institute, were used for the experiments. To evaluate the methylation status of the MGMT promoter, pyrosequencing and methylation-specific PCR (MSP) were used. Moreover, mRNA and protein expression levels of MGMT in the individual GBM clones were evaluated. The HeLa cell line, which hyper-expresses MGMT, was used as control. A total of 12 U251 and 12 U373 clones were isolated. The methylation status of 83 of 97 CpG sites in the MGMT promoter were evaluated by pyrosequencing, and 11 methylated CpG sites and 13 unmethylated CpG sites were evaluated by MSP. The methylation status by pyrosequencing was relatively high at CpG sites 3-8, 20-35, and 7-83, in both the U251 and U373 clones. Neither MGMT mRNA nor protein was detected in any clone. These findings demonstrate tumor heterogeneity among individual clones derived from a single GBM cell. MGMT expression may be regulated, not only by methylation of the MGMT promoter but by other factors as well. Further studies are needed to clarify the mechanisms underlying the epigenetic heterogeneity and plasticity of GBM.

Carboplatin Plus Vincristine as an Alternative Chemotherapeutic Scheme in Patients With Glioblastoma.

Background and objectiveAlternative chemotherapy regimens, including cisplatin, carmustine, or other agents, have been shown to be effective; however, the use of carboplatin plus vincristine (C/V) has not been studied before. In this study, we aimed to determine the survival rates in patients treated with C/V, by comparing our findings with treatments based on temozolomide (TMZ), and to explore a possible relationship with the methylation status of the methylguanine methyltransferase (MGMT) promoter in patients with glioblastoma (GB).MethodsA retrospective cohort study was conducted involving 45 surgically treated patients diagnosed with GB. Fresh tissue samples were examined by the DNA bisulfite conversion method to determine methylation status. After surgery, different chemotherapy regimens were employed as adjuvants. Follow-up of participants was performed as outpatients at three-month intervals to determine overall survival (OS), by comparing the use of TMZ versus C/V.ResultsMGMT promoter methylation status could only be determined in 35 samples; 20 patients received adjuvant chemotherapy, of which 14 were treated with C/V and six with TMZ-based schemes. The median OS (mOS) was eight months (range: 1-24 months). OS was 57.25% at six months, 48.7% at 12 months, and 28.5% at 24 months. In the TMZ group, an OS of 83% was observed at 24 months. In the C/V group, OS was 71.4% at six months, 57.1% at 12 months, and 35.7% at 24 months. Patients who did not receive adjuvant chemotherapy treatment had the lowest survival rates with an OS of 39.9% at six months, 26.6% at 12 months, and 19.9% ​​at 24 months.ConclusionsBased on our findings, C/V offers an accessible and effective alternative treatment when the TMZ-based scheme is not accessible, providing higher rates of OS compared to patients without chemotherapy management. The methylation status of the MGMT promoter is a significant prognostic factor, resulting in higher survival rates among patients when it is methylated.

Radiological model based on the standard magnetic resonance sequences for detecting methylguanine methyltransferase methylation in glioma using texture analysis.

This study aims to build a radiological model based on standard MR sequences for detecting methylguanine methyltransferase (MGMT) methylation in gliomas using texture analysis. A retrospective cross‐sectional study was undertaken in a cohort of 53 glioma patients who underwent standard preoperative magnetic resonance (MR) imaging. Conventional visual radiographic features and clinical factors were compared between MGMT promoter methylated and unmethylated groups. Texture analysis extracted the top five most powerful texture features of MR images in each sequence quantitatively for detecting the MGMT promoter methylation status. The radiomic signature (Radscore) was generated by a linear combination of the five features and estimates in each sequence. The combined model based on each Radscore was established using multivariate logistic regression analysis. A receiver operating characteristic (ROC) curve, nomogram, calibration, and decision curve analysis (DCA) were used to evaluate the performance of the model. No significant differences were observed in any of the visual radiographic features or clinical factors between different MGMT methylated statuses. The top five most powerful features were selected from a total of 396 texture features of T1, contrast‐enhanced T1, T2, and T2 FLAIR. Each sequence’s Radscore can distinguish MGMT methylated status. A combined model based on Radscores showed differentiation between methylated MGMT and unmethylated MGMT both in the glioblastoma (GBM) dataset as well as the dataset for all other gliomas. The area under the ROC curve values for the combined model was 0.818, with 90.5% sensitivity and 72.7% specificity, in the GBM dataset, and 0.833, with 70.2% sensitivity and 90.6% specificity, in the overall gliomas dataset. Nomogram, calibration, and DCA also validated the performance of the combined model. The combined model based on texture features could be considered as a noninvasive imaging marker for detecting MGMT methylation status in glioma.

Prognostic and predictive impact of MGMT promoter methylation in grade 3 gliomas

BackgroundGrade 3 gliomas are aggressive primary brain tumors. Promoter methylation of methyl guanine methyl transferase (MGMT) has been associated with a favorable prognosis in patients with glioblastoma, but the impact of MGMT promoter methylation in patients with grade 3 gliomas is less clear. The purpose of the present study was to evaluate the utilization of MGMT testing in patients with Grade 3 glioma, as well its prognostic and predictive value. MethodsThe National Cancer Database (NCDB) was queried (2004–2016) for patients with newly diagnosed grade 3 glioma without 1p19q codeletion. Statistics included Kaplan-Meier overall survival (OS) analysis, along with Cox proportional hazards modeling. ResultsOf 20,488 total patients, 1,209 (5.0%) had MGMT testing. Of these patients, 561 (46.4%) were MGMT methylated (mMGMT), and 648 (53.6%) were MGMT unmethylated (uMGMT). mMGMT patients experienced greater median overall survival (OS) than both uMGMT patients as well as patients with no MGMT status reported (p < 0.05 for both). mMGMT was associated with improved OS for patients receiving adjuvant chemoradiation or adjuvant radiation, but not for patients receiving adjuvant chemotherapy or no adjuvant treatment. ConclusionsThis is the largest study to date describing the utilization of and outcomes for mMGMT patients with grade 3 glioma. The present results demonstrate that mMGMT is a prognostic factor and possibly a predictive biomarker, and is currently under-utilized within the US. MGMT methylation status could be used to risk-stratify and select patients for treatment intensification. Importance of studyThe present study is the largest of its kind to examine the prognostic and predictive impact of MGMT methylation (mMGMT) amongst patients with Grade 3 Glioma. The results suggest that mMGMT is prognostic, as amongst all patients, mMGMT was associated with improved overall survival. These results also suggest that mMGMT is predictive, as patients treated with adjuvant chemoradiation or adjuvant radiation therapy did have improved overall survival with mMGMT, though there was no difference in overall survival observes amongst patients receiving adjuvant chemotherapy or those patients receiving no adjuvant treatment. The study also found that only 5% of patients nationwide with Grade 3 Glioma are tested for MGMT.

Overcoming the Odds: Toward a Molecular Profile of Long-Term Survival in Glioblastoma.

For over a century, gliomas were characterized solely by histologic features. With the publication of the WHO Classification of Tumours of the Central Nervous System, Revised 4th Edition in 2016, integrated histologic and molecular diagnosis became the norm, providing improved tumor grading and prognosis with IDH1/2 (isocitrate dehydrogenase 1 and 2) mutation being the most significant prognostic feature in all grades of adult diffuse glioma. Since then, much work has been done to identify additional molecular prognostic features, but the bulk of the progress has been made in defining aggressive features in lower grade astrocytoma. Although there have been several large case series of glioblastomas with long-term survival (LTS; overall survival ≥36 months), less is known about the clinical and molecular features of these cases. Herein, we review 19 studies examining LTS glioblastoma patients from 2009 to 2020 that include variable molecular analysis, including 465 cases with survival of 36 months or more (total n = 2328). These studies suggest that while there is no definitive molecular signature of long survival, younger age, IDH mutation, and MGMT (methyl guanine methyl transferase) promoter hypermethylation are associated with longer overall survival, and in IDH-wildtype tumors, chromosome 19/20 co-gain and lack of EGFR amplification, chromosome 7 gain/10 loss, and TERT promoter mutation are associated with LTS.

The Effects of O6-methyl Guanine DNA-methyl Transferase Promotor Methylation and CpG1, CpG2, CpG3 and CpG4 Methylation on Treatment Response and their Prognostic Significance in Patients with Glioblastoma.

This retrospective study examined the prognostic significance and treatment effect of promoter methylation of O6- methyl guanine methyl transferase (MGMT) and meth-ylation of CpG 1, CpG2, CpG3 and CpG4 in glioblastoma (GB) patients received postoperative radiotherapy (PORT), with or without adjuvant temozolomide (TMZ). One hundred patients with GB who received PORT with concomitant TMZ plus adjuvant TMZ or PORT alone, were included. The MGMT promoter methylation of CpG1, CpG2, CpG3 and CpG4 islands were examined. Overall, MGMT-methylation emerged as a significant prognostic factor for better overall survival (OS) and progression-free survival (PFS) [odds ratio (OR): 0.609, 95% confidence interval (95% CI): 0.395-0.939, p = 0.02; OR: 0.662,95% CI: 0.430-1019, p = 0.5, respectively]. The methylation of each CpG1, CpG2, CpG3 and CpG4 islands was found to have no significant effects on OS and the methylation of each CpGl, CpG2 and CpG4 islands had no significant effect on PFS (p <0.05 for all). On the other hand, the methylation of CpG3 had a positive prognostic effect on PFS (OR: 2.1, 95% CI: 0.99-4.67, p = 0.04). In the group that only received radiotherapy (RT), CpG1 and CpC3 methylations were found to have a positive prognostic significance in terms of PFS (OR: 266, 95% CI: 1.05-6.75, p -0.03 for CpG1; OR: 2.4, 95% CI: 1.01-5.92, p = 0.04 for CpG3). The MGMT promoter methylation represents an important biomarker for predicting response to therapy. Individual islands, particularly CpG3, deserves further investigation as a prognostic marker. Further studies need to be done with larger sample sizes to clarify the results.

Craniotomy for recurrent glioblastoma: Is it justified? A comparative cohort study with outcomes over 10 years

ObjectiveThe role of repeat resection for recurrent glioblastoma multiforme (rGBM) is unclear. This large comparative cohort study assessed overall survival (OS), survival since recurrence (SSR), quality of life, and complications in reoperated versus non-reoperated patients for rGBM. Patients and methodsAll patients with rGBM between 2005 and 2015, who were discussed by our institution’s multi-disciplinary team, and who either did or did not undergo reoperation, were prospectively followed up with data collected and compared. Survival and prognostic factors were analysed using Kaplan–Meier and Cox regression methods. Results312 patients (reoperated, n = 145; non-reoperated, n = 167) were analysed. Median SSR was 10.8 months and 6.9 months in the reoperated and non-reoperated groups respectively (Log-rank test: p = 0.02). Median OS was 24.1 months and 20.4 months in the reoperated and non-reoperated groups, respectively (Log-rank test: p = 0.04). Quality of life as measured by Short Form 36 scores were 59 versus 54 at baseline and 62 versus 51 at four-month follow-up for re-operated and non-reoperated groups, respectively (p < 0.05). Age < 60 years, Karnofsky Performance Status (KPS) ≥ 80, recurrence ≥ 9 months from initial diagnosis, methylguanine methyltransferase (MGMT) promoter methylation, and extent of resection (EOR) > 80 %, each were significant predictors of SSR and OS. Complication rates were 5.5 % and 6.2 % following repeat resection and primary resection, respectively (p > 0.05). ConclusionThis is the first large prospective comparative cohort study of rGBM and demonstrates that repeat resection confers a small but significant benefit in survival and quality of life over non-operative treatment. Best prognosis is associated with: younger age, KPS ≥ 80, late recurrence, MGMT promoter methylation and EOR > 80 %.

Validation of a PCR-Based MGMT Promoter Methylation Assay and Comparison With Reference Lab Results

Abstract Introduction Glioblastoma multiforme (GBM) is the most aggressive and most common primary brain tumor. Treatment of GBM and other high-grade gliomas consists of surgical resection followed by radiation and temozolomide (TMZ) chemotherapy. TMZ alkylates purine DNA bases, causing DNA damage and eventually cell death. Alkylation at O6-gauanine can be reversed by the DNA repair enzyme methylguanine methyltransferase (MGMT), inhibiting the desired cytotoxic effect. MGMT is epigenetically silenced by methylation of a CpG island in its promoter. Many high-grade gliomas are hypomethylated at the MGMT promoter. MGMT promoter methylation has emerged as a biomarker for response to alkylating chemotherapy. Multiple clinical trials demonstrate that the patients most likely to benefit from TMZ therapy have a hypermethylated MGMT promoter. MGMT promoter status is now incorporated into adjuvant therapy guidelines. Our institution receives approximately 70 to 80 high-grade glioma resections per year. The goal of this study is to validate a PCR-based MGMT promoter methylation assay and to compare results to those previously obtained at a well-known reference lab. Methods We extracted DNA from high-grade gliomas specimens (FFPE) or used commercially available methylated and unmethylated DNA controls. DNA was treated with bisulfite to convert unmethylated cytosine into uracil. Promoter methylation was detected with real-time quantitative PCR using a primer/probe set specific for methylated CpG sites within the MGMT promoter (meMGMT, published in Ida et al 2017) or to a housekeeping gene (COL2A1). Results Using control methylated DNA, we demonstrated that our assay consistently detects meMGMT and COL2A1 at DNA inputs greater than 5 ng. Both within- and between-run imprecision was minimal with little variation in crossing thresholds (Cts) for both COL2A1 (%CV at 50 ng input: 7.9% [within-run], 0.3% [between-run]) and MGMT (%CV at 50 ng input: 0.6% [within-run], 0.4% [between-run]). Delta Ct values were consistent across different levels of DNA input. Analysis of 15 cases from our archives (8 positive, 7 negative) demonstrates that our results (delta Ct values) correlate (R2 = 0.9) with the reference lab methylation score (&lt;2 considered unmethylated). Separation between positive and negative cases was good with minimal overlap; positive cases had delta Cts of 0.2 to 7.6, while negative cases had delta Cts of 7.1 to 13.6 or had no meMGMT amplification. ROC curve analysis showed an AUC of 0.96, and using the cutoff of delta Ct &lt;7.6, our assay had 93% (14/15) agreement with the reference lab, a sensitivity of 100%, and a specificity of 71%. The single negative case that was misclassified had a reference lab methylation score of 1.86, near their cutoff of &lt;2. Additional patient samples are pending. Conclusions Our PCR-based MGMT methylation assay has excellent analytical sensitivity and low imprecision. Preliminary analysis of patient samples shows that our results are concordant with historic reference lab results.

Diagnostic and Prognostic Potential of Serum Cell-Free microRNA-214 in Glioma

We aimed to expound feasibility of serum cell-free microRNA-214 (miR-214) as a noninvasive biomarker for glioma in this study. We detected expression of miR-214 in medium from 2 glioma cell lines to confirm whether it is secretory in screening phase. Then, we verified cell-free miR-214 expression in serum samples from an independent set of 100 preoperative patients with glioma, 30 matching postoperative patients, and 100 healthy controls. MiR-214 was secreted from glioma cell lines. Extracellular miR-214 levels were significantly overexpressed in preoperative serum from glioma patients with glioma, whereas its expression significantly decreased in matched postoperative serum. Upregulated cell-free miR-214 in serum was significantly associated with higher tumor grade, absence of isocitrate dehydrogenase, and unmethylated methylguanine methyltransferase promoter. Extracellular miR-214 in serum could effectively distinguish patients with glioma from healthy control (area under the curve= 0.885; 95% confidence interval, 0.833-0.926). Moreover, serum cell-free miR-214 was an independent prognostic indicator of overall survival for patients with glioma. Serum cell-free miR-214 could serve as a promising noninvasive biomarker of glioma in tumor stratification, early diagnosis, and prognostic evaluation.

Determining a cut-off residual tumor volume threshold for patients with newly diagnosed glioblastoma treated with temozolomide chemoradiotherapy: A multicenter cohort study

Standard-of-care treatment of glioblastomas involves maximal safe resection and adjuvant temozolomide chemo-radiotherapy. Although extent of resection (EOR) is a well-known surgical predictor for overall survival most lesions cannot be completely resected. We hypothesize that in the event of incomplete resection, residual tumor volume (RTV) may be a more significant predictor than EOR. This was a multicenter retrospective review of 147 adult glioblastoma patients (mean age 53 years) that underwent standard treatment. Semiautomatic magnetic resonance imaging segmentation was performed for pre- and postoperative scans for volumetric analysis. Cox proportional hazards regression and Kaplan-Meier survival analyses were performed for prognostic factors including: age, Karnofsky performance score (KPS), O(6)-methylguanine methyltransferase (MGMT) promoter methylation status, EOR and RTV. EOR and RTV cut-off values for improved OS were determined and internally validated by receiver operator characteristic (ROC) analysis for 12-month overall survival. Half of the tumors had MGMT promoter methylation (77, 52%). The median tumor volume, EOR and RTV were 43.20 cc, 93.5%, and 3.80 cc respectively. Gross total resection was achieved in 52 patients (35%). Cox proportional hazards regression, ROC and maximum Youden index analyses for RTV and EOR showed that a cut-off value of <3.50 cc (HR 0.69; 95% CI 0.48–0.98) and ≥84% (HR 0.64; 95% CI 0.43–0.96) respectively conferred an overall survival advantage. Independent overall survival predictors were MGMT promoter methylation (adjusted HR 0.35; 95% CI 0.23–0.55) and a RTV of <3.50 cc (adjusted HR 0.53; 95% CI 0.29–0.95), but not EOR for incompletely resected glioblastomas.

A Comparative Analysis of the Usefulness of Survival Prediction Models for Patients with Glioblastoma in the Temozolomide Era: The Importance of Methylguanine Methyltransferase Promoter Methylation, Extent of Resection, and Subventricular Zone Location

Several survival prediction models for patients with glioblastoma have been proposed, but none is widely used. This study aims to identify the predictors of overall survival (OS) and to conduct an independent comparative analysis of 5 prediction models. Multi-institutional data from 159 patients with newly diagnosed glioblastoma who received adjuvant temozolomide concomitant chemoradiotherapy (CCRT) were collected. OS was assessed by Cox proportional hazards regression and adjusted for known prognostic factors. An independent CCRT patient cohort was used to externally validate the 1) RTOG (Radiation Therapy Oncology Group) recursive partitioning analysis (RPA) model, 2) Yang RPA model, and 3) Wee RPA model, Chaichana model, and the RTOG nomogram model. The predictive accuracy for each model at 12-month survival was determined by concordance indices. Calibration plots were performed to ascertain model prediction precision. The median OS for patients who received CCRT was 19.0 months compared with 12.7 months for those who did not (P < 0.001). Independent predictors were:1)subventricular zone II tumors (hazard ratio [HR], 1.6; 95% confidence interval [CI], 1.0-2.5); 2) methylguanine methyltransferase promoter methylation (HR, 0.36; 95% CI, 0.2-0.6); and 3) extent of resection of >85% (HR, 0.59; 95% CI, 0.4-0.9). For 12-month OS prediction, the RTOG nomogram model was superior to the RPA models with a c-index of 0.70. Calibration plots for 12-month survival showed that none of the models was precise, but the RTOG nomogram performed relatively better. The RTOG nomogram best predicted 12-month OS. Methylguanine methyltransferase promoter methylation status, subventricular zone tumor location, and volumetric extent of resection should be considered when constructing prediction models.

Imaging Correlates of Adult Glioma Genotypes.

Primary brain tumors, most commonly gliomas, are histopathologically typed and graded as World Health Organization (WHO) grades I-IV according to increasing degrees of malignancy. These grades provide prognostic information and guidance on treatment such as radiation therapy and chemotherapy after surgery. Despite the confirmed value of the WHO grading system, results of a multitude of studies and prospective interventional trials now indicate that tumors with identical morphologic criteria can have highly different outcomes. Molecular markers can allow subtypes of tumors of the same morphologic type and WHO grade to be distinguished and are, therefore, of great interest in personalization of brain tumor treatment. Recent genomic-wide studies have resulted in a far more comprehensive understanding of the genomic alterations in gliomas and provide suggestions for a new molecularly based classification. Magnetic resonance (MR) imaging phenotypes can serve as noninvasive surrogates for tumor genotypes and can provide important information for diagnosis, prognosis, and, eventually, personalized treatment. The newly emerged field of radiogenomics allows specific MR imaging phenotypes to be linked with gene expression profiles. In this article, the authors review the conventional and advanced imaging features of three tumoral genotypes with prognostic and therapeutic consequences: (a) isocitrate dehydrogenase mutation; (b) the combined loss of the short arm of chromosome 1 and the long arm of chromosome 19, or 1p19q codeletion; and (c) methylguanine methyltransferase promoter methylation. © RSNA, 2017.

Long-term survival in glioblastoma: methyl guanine methyl transferase (MGMT) promoter methylation as independent favourable prognostic factor.

BackgroundIn spite of significant improvement after multi-modality treatment, prognosis of most patients with glioblastoma remains poor. Standard clinical prognostic factors (age, gender, extent of surgery and performance status) do not clearly predict long-term survival. The aim of this case-control study was to evaluate immuno-histochemical and genetic characteristics of the tumour as additional prognostic factors in glioblastoma.Patients and methodsLong-term survivor group were 40 patients with glioblastoma with survival longer than 30 months. Control group were 40 patients with shorter survival and matched to the long-term survivor group according to the clinical prognostic factors. All patients underwent multimodality treatment with surgery, postoperative conformal radiotherapy and temozolomide during and after radiotherapy. Biopsy samples were tested for the methylation of MGMT promoter (with methylation specific polymerase chain reaction), IDH1 (with immunohistochemistry), IDH2, CDKN2A and CDKN2B (with multiplex ligation-dependent probe amplification), and 1p and 19q mutations (with fluorescent in situ hybridization).ResultsMethylation of MGMT promoter was found in 95% and in 36% in the long-term survivor and control groups, respectively (p < 0.001). IDH1 R132H mutated patients had a non-significant lower risk of dying from glioblastoma (p = 0.437), in comparison to patients without this mutation. Other mutations were rare, with no significant difference between the two groups.ConclusionsMolecular and genetic testing offers additional prognostic and predictive information for patients with glioblastoma. The most important finding of our analysis is that in the absence of MGMT promoter methylation, longterm survival is very rare. For patients without this mutation, alternative treatments should be explored.

IDH mutation and MGMT promoter methylation are associated with the pseudoprogression and improved prognosis of glioblastoma multiforme patients who have undergone concurrent and adjuvant temozolomide-based chemoradiotherapy

PurposeThis study aimed to investigate the potential association between IDH mutation and O6–methyl-guanine methyl transferase (MGMT) gene promoter methylation and pseudoprogression disease (psPD) in glioblastoma multiforme (GBM) patients after concurrent temozolomide (TMZ)-based chemoradiotherapy. MethodsA total of 157 GBM patients who received concurrent TMZ-based chemoradiotherapy were included in this retrospective study. The association between psPD and a number of demographic and genetic factors, including IDH mutation and MGMT promoter methylation, were analyzed based on logistic regression, Cox regression, and multivariate analysis. ResultsOf the 157 GBM patients, 145 (92.36%) patients, including 38 patients with psPD, 38 patients with early progression (ePD), and 69 patients with non-progression (non-PD), were followed up for six to 56 months. We identified a higher rate of MGMT promoter methylation and IDH1 mutation in psPD patients compared with ePD patients (P=0.002). In addition, MGMT promoter methylation and IDH1 mutation predicted a high probability of psPD development in GBM patients (P=0.001 and P<0.001, respectively). MGMT promoter methylation, IDH1 mutation, Karnofsky performance score (KPS) ≥70, and psPD were associated with a significantly longer overall survival of GBM patients (P=0.001, 0.001, 0.002, and P<0.001, respectively). Both of MGMT promoter methylation and IDH mutation had a cumulative effect on the OS of GBM patients. GBM patients with psPD (39.2±2.1months, P<0.001) had a longer median survival (MS) than GBM patients with ePD (11.9±1.1months) or with non-PD (24.4±2.4months). ConclusionMGMT promoter methylation and IDH1 mutation were associated with PsPD and predicted a longer median survival in GBM patients after TMZ-based chemoradiotherapy. Genetic analyses of the MGMT promoter and IDH1 may allow us to effectively treat GBM patients.

Gamma Knife Radiosurgery in Recurrent Glioblastoma

Background: We evaluated Gamma Knife radiosurgery (GKRS) as a treatment option for patients with recurrent glioblastoma. Patients and Methods: 42 patients with histopathologically diagnosed recurrent grade IV tumor were treated with GKRS. All patients had undergone standard multimodal first-line treatment. The average time from diagnosis to GKRS was 17.0 months. The median target volume was 5.1 cm³. The median margin dose was 10 Gy and the median central dose 20 Gy. In a subset of patients, O⁶-methylguanine methyltransferase (MGMT) promoter methylation analysis by pyrosequencing was performed. Results: Most patients did not develop complications after GKRS. Time to radiological progression after initial GKRS was 4.4 months (95% CI: 3.1-5.7 months). Radiological progression mainly occurred beyond the GKRS-irradiated area. The median survival time after initial GKRS was 9.6 months (95% CI: 7.7-11.5 months). The median overall survival time from diagnosis was 25.6 months (95% CI: 21.8-29.3 months). Patients with MGMT promoter methylation survived significantly longer (33.4 months; 95% CI: 21.2-45.5 months) compared to patients without MGMT promoter methylation (16.0 months; 95% CI: 8.0-23.9 months). Conclusion: GKRS seems to be a relatively safe salvage treatment option for recurrent glioblastoma for highly selected patients but must be seen as part of a multimodal treatment algorithm.

Immune checkpoint blockade as a potential therapeutic target: surveying CNS malignancies.

Expression of programmed cell death protein 1 (PD-1)/programmed death ligand 1 (PD-L1) across glioma grades is undocumented, and their interactions with commonly expressed genetic and epigenetic alterations are undefined but nonetheless highly relevant to combinatorial treatments. Patients with CNS malignancies were profiled by Caris Life Sciences from 2009 to 2016. Immunohistochemistry findings for PD-1 on tumor-infiltrating lymphocytes (TIL) and PD-L1 on tumor cells were available for 347 cases. Next-generation sequencing, pyrosequencing, immunohistochemistry, fragment analysis, and fluorescence in situ hybridization were used to determine isocitrate dehydrogenase 1 (IDH1), phosphatase and tensin homolog (PTEN), and tumor protein 53 mutational status, O(6)-DNA methylguanine-methyltransferase promoter methylation (MGMT-Me) status, PTEN expression, plus epidermal growth factor receptor variant III and 1p/19q codeletion status. PD-1+ TIL expression and grade IV gliomas were significantly positively correlated (odds ratio [OR]: 6.363; 95% CI: 1.263, 96.236)-especially in gliosarcomas compared with glioblastoma multiforme (P = .014). PD-L1 expression was significantly correlated with tumor grade with all PD-L1+ cases (n = 21) being associated with grade IV gliomas. PD-1+ TIL expression and PD-L1 expression were significantly correlated (OR: 5.209; 95% CI: 1.555, 20.144). Mutations of PTEN, tumor protein 53, BRAF, IDH1, and epidermal growth factor receptor or MGMT-Me did not associate with increased intratumoral expression of either PD-1+ TIL or PD-L1 in glioblastoma multiforme even before false discovery rate correction for multiple comparison. Targeting immune checkpoints in combination with other therapeutics based on positive biomarker selection will require screening of large patient cohorts.

The role of neuropathology in the management of patients with diffuse low grade glioma: A systematic review and evidence-based clinical practice guideline.

Adult patients (age ≥18 years) who have suspected low-grade diffuse glioma. What are the optimal neuropathological techniques to diagnose low-grade diffuse glioma in the adult? LEVEL I: Histopathological analysis of a representative surgical sample of the lesion should be used to provide the diagnosis of low-grade diffuse glioma. Both frozen section and cytopathologic/smear evaluation should be used to aid the intra-operative assessment of low-grade diffuse glioma diagnosis. A resection specimen is preferred over a biopsy specimen, to minimize the potential for sampling error issues. Patients with histologically-proven WHO grade II diffuse glioma. In adult patients (age ≥18 years) with histologically-proven WHO grade II diffuse glioma, is testing for IDH1 mutation (R132H and/or others) warranted? If so, is there a preferred method? IDH gene mutation assessment, via IDH1 R132H antibody and/or IDH1/2 mutation hotspot sequencing, is highly-specific for low-grade diffuse glioma, and is recommended as an additional test for classification and prognosis. Patients with histologically-proven WHO grade II diffuse glioma. In adult patients (age ≥18 years) with histologically-proven WHO grade II diffuse glioma, is testing for 1p/19q loss warranted? If so, is there a preferred method? 1p/19q loss-of-heterozygosity testing, by FISH, array-CGH or PCR, is recommended as an additional test in oligodendroglial cases for prognosis and potential treatment planning. Patients with histologically-proven WHO grade II diffuse glioma. In adult patients (age ≥18 years) with histologically-proven WHO grade II diffuse glioma, is MGMT promoter methylation testing warranted? If so, is there a preferred method? There is insufficient evidence to recommend methyl-guanine methyl-transferase (MGMT) promoter methylation testing as a routine for low-grade diffuse gliomas. It is recommended that patients be enrolled in properly designed clinical trials to assess the value of this and related markers for this target population. Patients with histologically-proven WHO grade II diffuse glioma. In adult patients (age ≥18 years) with histologically-proven WHO grade II diffuse glioma, is Ki-67/MIB1 immunohistochemistry warranted? If so, is there a preferred method to quantitate results? Ki67/MIB1 immunohistochemistry is recommended as an option for prognostic assessment.

Using the apparent diffusion coefficient to identifying MGMT promoter methylation status early in glioblastoma: importance of analytical method.

IntroductionAccurate knowledge of O6-methylguanine methyltransferase (MGMT) gene promoter subtype in patients with glioblastoma (GBM) is important for treatment. However, this test is not always available. Pre-operative diffusion MRI (dMRI) can be used to probe tumour biology using the apparent diffusion coefficient (ADC); however, its ability to act as a surrogate to predict MGMT status has shown mixed results. We investigated whether this was due to variations in the method used to analyse ADC.MethodsWe undertook a retrospective study of 32 patients with GBM who had MGMT status measured. Matching pre-operative MRI data were used to calculate the ADC within contrast enhancing regions of tumour. The relationship between ADC and MGMT was examined using two published ADC methods.ResultsA strong trend between a measure of ‘minimum ADC’ and methylation status was seen. An elevated minimum ADC was more likely in the methylated compared to the unmethylated MGMT group (U = 56, P = 0.0561). In contrast, utilising a two-mixture model histogram approach, a significant reduction in mean measure of the ‘low ADC’ component within the histogram was associated with an MGMT promoter methylation subtype (P < 0.0246).ConclusionThis study shows that within the same patient cohort, the method selected to analyse ADC measures has a significant bearing on the use of that metric as a surrogate marker of MGMT status. Thus for dMRI data to be clinically useful, consistent methods of data analysis need to be established prior to establishing any relationship with genetic or epigenetic profiling.

¹⁸F-fluorodeoxyglucose and ¹¹C-methionine positron emission tomography in relation to methyl-guanine methyltransferase promoter methylation in high-grade gliomas.

Methylation status of the methyl-guanine methyltransferase (MGMT) promoter is associated with a favorable response to a DNA alkylating agent in high-grade gliomas. We analyzed PET scans of patients with high-grade gliomas to determine whether the MGMT methylation status affects the tumor metabolic characteristics. Twenty-three patients with high-grade glioma, who were initially examined with 11C-methionine (MET) and 18F-fluorodeoxyglucose (FDG) PET, were retrospectively enrolled. MET and FDG PET images were coregistered to each other and quantitative uptake of MET or FDG was assessed using tumor-to-normal uptake ratio of the cortex (TNR). TNRs for MET and FDG PET were compared between the two groups classified by MGMT promoter methylation status. Maximum TNR(FDG) of the MGMT methylated group was significantly higher than that of the MGMT unmethylated group (1.80±0.90 vs. 1.29±0.19; P=0.02). The MGMT methylated group also showed a trend for increased mean TNRFDG compared with the unmethylated group (0.85±0.21 vs. 0.72±0.11; P=0.10). There was no significant difference in TNR(MET) between the groups. In subgroup analyses with WHO grade 3 and 4, a trend for higher maximum TNR(FDG) was found in the MGMT methylated group compared with the unmethylated group. The MGMT methylated group showed higher glucose metabolism compared with the unmethylated group, whereas MET uptake did not show a significant difference. This suggests that MGMT methylation in high-grade gliomas could affect the tumor glucose metabolism. Thus, MGMT methylation status can cause a discrepancy in the prognostic prediction of high-grade gliomas by FDG PET, especially in patients scheduled for DNA alkylating chemotherapeutics.

High-grade glioma: ESMO Clinical Practice Guidelines for diagnosis, treatment and follow-up

High-grade glioma: ESMO Clinical Practice Guidelines for diagnosis, treatment and follow-up