Gabriele Schackert

Molecular predictors of progression-free and overall survival in patients with newly diagnosed glioblastoma: A prospective translational study of the German Glioma Network

PURPOSE The prognostic value of genetic alterations characteristic of glioblastoma in patients treated according to present standards of care is unclear. PATIENTS AND METHODS Three hundred one patients with glioblastoma were prospectively recruited between October 2004 and December 2006 at the clinical centers of the German Glioma Network. Two hundred fifty-eight patients had radiotherapy, 199 patients had temozolomide, 189 had both, and seven had another chemotherapy as the initial treatment. The tumors were investigated for TP53 mutation, p53 immunoreactivity, epidermal growth factor receptor, cyclin-dependent kinase CDK 4 or murine double minute 2 amplification, CDKN2A homozygous deletion, allelic losses on chromosome arms 1p, 9p, 10q, and 19q, O(6)-methylguanine methyltransferase (MGMT) promoter methylation, and isocitrate dehydrogenase 1 (IDH1) mutations. RESULTS Median progression-free (PFS) and overall survival (OS) were 6.8 and 12.5 months. Multivariate analysis revealed younger age, higher performance score, MGMT promoter methylation, and temozolomide radiochemotherapy as independent factors associated with longer OS. MGMT promoter methylation was associated with longer PFS (relative risk [RR], 0.5; 95% CI, 0.38 to 0.68; P < .001) and OS (RR, 0.39; 95% CI, 0.28 to 0.54; P < .001) in patients receiving temozolomide. IDH1 mutations were associated with prolonged PFS (RR, 0.42; 95% CI, 0.19 to 0.91; P = .028) and a trend for prolonged OS (RR, 0.43; 95% CI, 0.15 to 1.19; P = .10). No other molecular factor was associated with outcome. CONCLUSION Molecular changes associated with gliomagenesis do not predict response to therapy in glioblastoma patients managed according to current standards of care. MGMT promoter methylation and IDH1 mutational status allow for stratification into prognostically distinct subgroups.

Primary Central Nervous System Lymphoma: Results of a Pilot and Phase II Study of Systemic and Intraventricular Chemotherapy With Deferred Radiotherapy

PURPOSE To evaluate response rate, response duration, overall survival (OS), and toxicity in primary CNS lymphoma (PCNSL) after systemic and intraventricular chemotherapy with deferred radiotherapy. PATIENTS AND METHODS From September 1995 to July 2001, 65 consecutive patients with PCNSL (median age, 62 years) were enrolled onto a pilot and phase II study evaluating chemotherapy without radiotherapy. A high-dose methotrexate (MTX; cycles 1, 2, 4, and 5) and cytarabine (ARA-C; cycles 3 and 6)-based systemic therapy (including dexamethasone, vinca-alkaloids, ifosfamide, and cyclophosphamide) was combined with intraventricular MTX, prednisolone, and ARA-C. RESULTS Sixty-one of 65 patients were assessable for response. Of these, 37 patients (61%) achieved complete response, six (10%) achieved partial response, and 12 (19%) progressed under therapy. Six (9%) of 65 patients died because of treatment-related complications. Follow-up is 0 to 87 months (median, 26 months). The Kaplan-Meier estimates for median time to treatment failure (TTF) and median OS were 21 months and 50 months, respectively. For patients older than 60 years, median survival was 34 months, and the median TTF was 15 months. In patients younger than 61 years, median survival and median TTF have not been reached yet; the 5-year survival fraction is 75%. Systemic toxicity was mainly hematologic. Ommaya reservoir infection occurred in 12 patients (19%), and permanent cognitive dysfunction possibly as a result of treatment occurred in only two patients (3%). CONCLUSION Primary chemotherapy based on high-dose MTX and ARA-C is highly efficient in PCNSL. Response rate and response duration in this series are comparable to the response rates and durations reported after combined radiotherapy and chemotherapy. Neurotoxicity was infrequent.

Resection and survival in glioblastoma multiforme: An RTOG recursive partitioning analysis of ALA study patients

The benefit of cytoreductive surgery for glioblastoma multiforme (GBM) is unclear, and selection bias in past series has been observed. The 5-aminolevulinic acid (ALA) study investigated the influence of fluorescence-guided resections on outcome and generated an extensive database of GBM patients with optimized resections. We evaluated whether the Radiation Therapy Oncology Group recursive partitioning analysis (RTOG-RPA) would predict survival of these patients and whether there was any benefit from extensive resections depending on RPA class. A total of 243 per-protocol patients with newly diagnosed GBM were operated on with or without ALA and treated by radiotherapy. Postoperative MRI was obtained in all patients. Patients were allocated into RTOG-RPA classes III-V based on age, KPS, neurological condition, and mental status (as derived from the NIH Stroke Scale). Median overall survival among RPA classes III, IV, and V was 17.8, 14.7, and 10.7 months, respectively, with 2-year survival rates of 26%, 12%, and 7% (p = 0.0007). Stratified for degree of resection, survival of patients with complete resections was clearly longer in RPA classes IV and V (17.7 months vs. 12.9 months, p = 0.0015, and 13.7 months vs. 10.4 months, p = 0.0398; 2-year rates: 21.0% vs. 4.4% and 11.1% vs. 2.6%, respectively), but was not in the small subgroup of RPA class III patients (19.3 vs. 16.3 months, p = 0.14). Survival of patients from the ALA study is correctly predicted by the RTOG-RPA classes. Differences in survival depending on resection status, especially in RPA classes IV and V, support a causal influence of resection on survival.

Poor drug distribution as a possible explanation for the results of the PRECISE trial

OBJECT Convection-enhanced delivery (CED) is a novel intracerebral drug delivery technique with considerable promise for delivering therapeutic agents throughout the CNS. Despite this promise, Phase III clinical trials employing CED have failed to meet clinical end points. Although this may be due to inactive agents or a failure to rigorously validate drug targets, the authors have previously demonstrated that catheter positioning plays a major role in drug distribution using this technique. The purpose of the present work was to retrospectively analyze the expected drug distribution based on catheter positioning data available from the CED arm of the PRECISE trial. METHODS Data on catheter positioning from all patients randomized to the CED arm of the PRECISE trial were available for analyses. BrainLAB iPlan Flow software was used to estimate the expected drug distribution. RESULTS Only 49.8% of catheters met all positioning criteria. Still, catheter positioning score (hazard ratio 0.93, p = 0.043) and the number of optimally positioned catheters (hazard ratio 0.72, p = 0.038) had a significant effect on progression-free survival. Estimated coverage of relevant target volumes was low, however, with only 20.1% of the 2-cm penumbra surrounding the resection cavity covered on average. Although tumor location and resection cavity volume had no effect on coverage volume, estimations of drug delivery to relevant target volumes did correlate well with catheter score (p < 0.003), and optimally positioned catheters had larger coverage volumes (p < 0.002). Only overall survival (p = 0.006) was higher for investigators considered experienced after adjusting for patient age and Karnofsky Performance Scale score. CONCLUSIONS The potential efficacy of drugs delivered by CED may be severely constrained by ineffective delivery in many patients. Routine use of software algorithms and alternative catheter designs and infusion parameters may improve the efficacy of drugs delivered by CED.

Identification of SOX2 as a novel glioma-associated antigen and potential target for T cell-based immunotherapy.

Prognosis for patients suffering from malignant glioma has not substantially improved. Specific immunotherapy as a novel treatment concept critically depends on target antigens, which are highly overexpressed in the majority of gliomas, but the number of such antigens is still very limited. SOX2 was identified by screening an expression database for transcripts that are overexpressed in malignant glioma, but display minimal expression in normal tissues. Expression of SOX2 mRNA was further investigated in tumour and normal tissues by real-time PCR. Compared to cDNA from pooled normal brain, SOX2 was overexpressed in almost all (9 out of 10) malignant glioma samples, whereas expression in other, non-malignant tissues was almost negligible. SOX2 protein expression in glioma cell lines and tumour tissues was verified by Western blot and immunofluorescence. Immunohistochemistry demonstrated SOX2 protein expression in all malignant glioma tissues investigated ranging from 6 to 66% stained tumour cells. Human leucocyte antigen-A*0201-restricted SOX2-derived peptides were tested for the activation of glioma-reactive CD8+ cytotoxic T lymphocytes (CTLs). Specific CTLs were raised against the peptide TLMKKDKYTL and were capable of lysing glioma cells. The abundant and glioma-restricted overexpression of SOX2 and the generation of SOX2-specific and tumour-reactive CTLs may recommend this antigen as target for T-cell-based immunotherapy of glioma.

Predictive impact of MGMT promoter methylation in glioblastoma of the elderly

O6‐methylguanine‐DNA‐methyltransferase (MGMT) promoter methylation identifies a subpopulation of glioblastoma patients with more favorable prognosis and predicts a benefit from alkylating agent chemotherapy (CT). Little is known about its prevalence and clinical significance in older glioblastoma patients. We studied 233 glioblastoma patients aged 70 years or more (144 males, 89 females, median age: 74 years, range: 70.0–86.6 years), who were prospectively enrolled in the German Glioma Network, for MGMT promoter methylation by methylation‐specific PCR (MSP) in all patients and DNA pyrosequencing in 166 patients. MGMT data were correlated with patient outcome. Median progression‐free survival (PFS) was 4.8 months (95% CI: 4.3–5.3) and median overall survival (OS) was 7.7 months (95% CI: 6.3–9.0). MGMT promoter methylation was detected by MSP in 134 patients (57.5%). For the whole cohort, PFS was 5.2 versus 4.7 months (p = 0.207) and OS was 8.4 versus 6.4 months (p = 0.031) in patients with versus without MGMT promoter methylation. Patients with MGMT methylated tumors had longer PFS when treated with radiotherapy (RT) plus CT or CT alone compared to patients treated with RT alone. Patients with MGMT unmethylated tumors appeared to derive no survival benefit from CT, regardless of whether given at diagnosis together with RT or as a salvage treatment. Patients treated with RT plus CT or CT alone demonstrated longer OS when pyrosequencing revealed >25% MGMT methylated alleles. Taken together, MGMT promoter methylation may be a useful biomarker to stratify elderly glioblastoma patients for treatment with versus without alkylating agent CT.

A microarray-based DNA methylation study of glioblastoma multiforme.

Glioblastoma multiforme (GBM) is the most frequent and devastating primary brain tumor in adults. The presence of epigenetic lesions, like hypermethylation of known tumor suppressor genes such as MGMT, has been widely described in GBM, but to our knowledge, a genome-wide profile of DNA methylation changes in these lethal tumors is not yet available. In the present analysis, we have quantified the DNA methylation level of 1,505 CpG dinucleotides (807 genes) in 87 consecutive GBMs using universal BeadArrays. Supervised cluster analyses identified 25 and seven genes that were respectively hypermethylated and hypomethylated in more than 20% of the cases studied. The most frequently hypermethylated genes were HOXA11, CD81, PRKCDBP, TES, MEST, TNFRSF10A and FZD9, being involved in more than half of the cases. Studying the biological features of hypermethylated genes, we found that the group of genes hypermethylated in GBM was highly enriched (41%, P

Promoter methylation and expression of MGMT and the DNA mismatch repair genes MLH1, MSH2, MSH6 and PMS2 in paired primary and recurrent glioblastomas

Epigenetic silencing of the O6‐methylguanine‐DNA methyltransferase (MGMT) gene promoter is associated with prolonged survival in glioblastoma patients treated with temozolomide (TMZ). We investigated whether glioblastoma recurrence is associated with changes in the promoter methylation status and the expression of MGMT and the DNA mismatch repair (MMR) genes MLH1, MSH2, MSH6 and PMS2 in pairs of primary and recurrent glioblastomas of 80 patients, including 64 patients treated with radiotherapy and TMZ after the first operation. Among the primary tumors, the MGMT promoter was methylated in 31 patients and unmethylated in 49 patients. In 71 patients (89%), the MGMT promoter methylation status of the primary tumor was retained at recurrence. MGMT promoter methylation, but not MGMT protein expression, was associated with longer progression‐free survival, overall survival and postrecurrence survival (PRS). Moreover, PRS was increased under salvage chemotherapy. Investigation of primary and recurrent glioblastomas of 43 patients did not identify promoter methylation in any of the four MMR genes. However, recurrent glioblastomas demonstrated significantly lower MSH2, MSH6 and PMS2 protein expression as detected by immunohistochemistry. In conclusion, reduced expression of MMR proteins, but not changes in MGMT promoter methylation, is characteristic of glioblastomas recurring after the current standards of care.

Genesis of cerebral aneurysms--an update.

Summary To elucidate the molecular pathogenesis of diseases has become a crucial step in the development of new treatment strategies. Although the pathogenesis of cerebral aneurysms has been studied intensively, it is poorly understood. Endogenous factors like elevated arterial blood pressure, the special anatomy of the Circle of Willis or the effect of haemodynamic factors, particularly originating at vessel bifurcation, are all known to be involved in the growth and rupture of an aneurysm. There is an ongoing discussion as to whether these factors also contribute to the very early steps of pathogenesis. Arteriosclerosis and secondary inflammatory reactions are thought to be elementary preconditions. Exogenous factors like cigarette smoking, heavy alcohol consumption or certain medications known to help generate arteriosclerosis and elevated blood pressure have also been found to be related to the occurrence of cerebral aneurysms. Furthermore, there has been a long-lasting debate on whether aneurysms might develop as a result of an inborn genetic defect. First-degree relatives of patients with cerebral aneurysms have a higher risk of having an aneurysm. In addition, the elevated prevalence of cerebral aneurysms in patients suffering from various inherited diseases points to a genetic background in the development of an aneurysm. Recent advances in molecular biology provide evidence that genetic variants of different candidate genes are associated with the occurrence of cerebral aneurysms. The aim of this review is to expose the current status of these various hypotheses and their contribution to the pathogenesis of cerebral aneurysms in order to provide a basis for future investigations in this field.

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

Objective: To explore whether the isocitrate dehydrogenase 1 (IDH1) or 1p/19q status determines the prognostic vs predictive role of O6-methylguanine-DNA methyltransferase (MGMT) promoter methylation in the Neuro-Oncology Working Group of the German Cancer Society (NOA)-04 trial anaplastic glioma biomarker cohort. Methods: 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. Results: 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. Conclusions: 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.