Symptomatic epilepsy is a common complication of glioblastoma and requires pharmacotherapy. Several uncontrolled retrospective case series and a post hoc analysis of the registration trial for ...temozolomide indicated an association between valproic acid (VPA) use and improved survival outcomes in patients with newly diagnosed glioblastoma.
To confirm the hypothesis suggested above, a combined analysis of survival association of antiepileptic drug use at the start of chemoradiotherapy with temozolomide was performed in the pooled patient cohort (n = 1,869) of four contemporary randomized clinical trials in newly diagnosed glioblastoma: AVAGlio (Avastin in Glioblastoma; NCT00943826), CENTRIC (Cilengitide, Temozolomide, and Radiation Therapy in Treating Patients With Newly Diagnosed Glioblastoma and Methylated Gene Promoter Status; NCT00689221), CORE (Cilengitide, Temozolomide, and Radiation Therapy in Treating Patients With Newly Diagnosed Glioblastoma and Unmethylated Gene Promoter Status; NCT00813943), and Radiation Therapy Oncology Group 0825 (NCT00884741). Progression-free survival (PFS) and overall survival (OS) were compared between: (1) any VPA use and no VPA use at baseline or (2) VPA use both at start of and still after chemoradiotherapy. Results of Cox regression models stratified by trial and adjusted for baseline prognostic factors were analyzed. The same analyses were performed with levetiracetam (LEV).
VPA use at start of chemoradiotherapy was not associated with improved PFS or OS compared with all other patients pooled (PFS: hazard ratio HR, 0.91; 95% CI, 0.77 to 1.07; P = .241; OS: HR, 0.96; 95% CI, 0.80 to 1.15; P = .633). Furthermore, PFS and OS of patients taking VPA both at start of and still after chemoradiotherapy were not different from those without antiepileptic drug use at both time points (PFS: HR, 0.92; 95% CI, 0.74 to 1.15; P = .467; OS: HR, 1.10; 95% CI, 0.86 to 1.40; P = .440). Similarly, no association with improved outcomes was observed for LEV use.
The results of this analysis do not justify the use of VPA or LEV for reasons other than seizure control in patients with newly diagnosed glioblastoma outside clinical trials.
Summary Primary brain tumours are heterogeneous in histology, genetics, and outcome. Although WHO's classification of tumours of the CNS has greatly helped to standardise diagnostic criteria ...worldwide, it does not consider the substantial progress that has been made in the molecular classification of many brain tumours. Recent practice-changing clinical trials have defined a role for routine assessment of MGMT promoter methylation in glioblastomas in elderly people, and 1p and 19q codeletions in anaplastic oligodendroglial tumours. Moreover, large-scale molecular profiling approaches have identified new mutations in gliomas, affecting IDH1, IDH2, H3F3, ATRX, and CIC , which has allowed subclassification of gliomas into distinct molecular subgroups with characteristic features of age, localisation, and outcome. However, these molecular approaches cannot yet predict patients' benefit from therapeutic interventions. Similarly, transcriptome-based classification of medulloblastoma has delineated four variants that might now be candidate diseases in which to explore novel targeted agents.
Abstract
Background
Malignant astrocytic gliomas in children show a remarkable biological and clinical diversity. Small in-frame insertions or missense mutations in the epidermal growth factor ...receptor gene (EGFR) have recently been identified in a distinct subset of pediatric-type bithalamic gliomas with a unique DNA methylation pattern.
Methods
Here, we investigated an epigenetically homogeneous cohort of malignant gliomas (n = 58) distinct from other subtypes and enriched for pediatric cases and thalamic location, in comparison with this recently identified subtype of pediatric bithalamic gliomas.
Results
EGFR gene amplification was detected in 16/58 (27%) tumors, and missense mutations or small in-frame insertions in EGFR were found in 20/30 tumors with available sequencing data (67%; 5 of them co-occurring with EGFR amplification). Additionally, 8 of the 30 tumors (27%) harbored an H3.1 or H3.3 K27M mutation (6 of them with a concomitant EGFR alteration). All tumors tested showed loss of H3K27me3 staining, with evidence of overexpression of the EZH inhibitory protein (EZHIP) in the H3 wildtype cases. Although some tumors indeed showed a bithalamic growth pattern, a significant proportion of tumors occurred in the unilateral thalamus or in other (predominantly midline) locations.
Conclusions
Our findings present a distinct molecular class of pediatric-type malignant gliomas largely overlapping with the recently reported bithalamic gliomas characterized by EGFR alteration, but additionally showing a broader spectrum of EGFR alterations and tumor localization. Global H3K27me3 loss in this group appears to be mediated by either H3 K27 mutation or EZHIP overexpression. EGFR inhibition may represent a potential therapeutic strategy in these highly aggressive gliomas.
Cerebral gliomas of World Health Organization (WHO) grade II and III represent a major challenge in terms of histological classification and clinical management. Here, we asked whether large-scale ...genomic and transcriptomic profiling improves the definition of prognostically distinct entities. We performed microarray-based genome- and transcriptome-wide analyses of primary tumor samples from a prospective German Glioma Network cohort of 137 patients with cerebral gliomas, including 61 WHO grade II and 76 WHO grade III tumors. Integrative bioinformatic analyses were employed to define molecular subgroups, which were then related to histology, molecular biomarkers, including isocitrate dehydrogenase 1 or 2 (
IDH1/2
) mutation, 1p/19q co-deletion and telomerase reverse transcriptase (
TERT
) promoter mutations, and patient outcome. Genomic profiling identified five distinct glioma groups, including three
IDH1/2
mutant and two
IDH1/2
wild-type groups. Expression profiling revealed evidence for eight transcriptionally different groups (five
IDH1/2
mutant, three
IDH1/2
wild type), which were only partially linked to the genomic groups. Correlation of DNA-based molecular stratification with clinical outcome allowed to define three major prognostic groups with characteristic genomic aberrations. The best prognosis was found in patients with
IDH1/2
mutant and 1p/19q co-deleted tumors. Patients with
IDH1/2
wild-type gliomas and glioblastoma-like genomic alterations, including gain on chromosome arm 7q (+7q), loss on chromosome arm 10q (−10q),
TERT
promoter mutation and oncogene amplification, displayed the worst outcome. Intermediate survival was seen in patients with
IDH1/2
mutant, but 1p/19q intact, mostly astrocytic gliomas, and in patients with
IDH1/2
wild-type gliomas lacking the +7q/−10q genotype and
TERT
promoter mutation. This molecular subgrouping stratified patients into prognostically distinct groups better than histological classification. Addition of gene expression data to this genomic classifier did not further improve prognostic stratification. In summary, DNA-based molecular profiling of WHO grade II and III gliomas distinguishes biologically distinct tumor groups and provides prognostically relevant information beyond histological classification as well as
IDH1/2
mutation and 1p/19q co-deletion status.
Early and progressive colonization of the healthy brain is one hallmark of diffuse gliomas, including glioblastomas. We recently discovered ultralong (>10 to hundreds of microns) membrane protrusions ...tumor microtubes (TMs) extended by glioma cells. TMs have been associated with the capacity of glioma cells to effectively invade the brain and proliferate. Moreover, TMs are also used by some tumor cells to interconnect to one large, resistant multicellular network. Here, we performed a correlative gene-expression microarray and
imaging analysis, and identified novel molecular candidates for TM formation and function. Interestingly, these genes were previously linked to normal CNS development. One of the genes scoring highest in tests related to the outgrowth of TMs was
(
), which was highly expressed in a fraction of TMs in mice and patients. Ttyh1 was confirmed to be a potent regulator of normal TM morphology and of TM-mediated tumor-cell invasion and proliferation. Glioma cells with one or two TMs were mainly responsible for effective brain colonization, and Ttyh1 downregulation particularly affected this cellular subtype, resulting in reduced tumor progression and prolonged survival of mice. The remaining Ttyh1-deficient tumor cells, however, had more interconnecting TMs, which were associated with increased radioresistance in those small tumors. These findings imply a cellular and molecular heterogeneity in gliomas regarding formation and function of distinct TM subtypes, with multiple parallels to neuronal development, and suggest that Ttyh1 might be a promising target to specifically reduce TM-associated brain colonization by glioma cells in patients.
In this report, we identify tweety-homolog 1 (Ttyh1), a membrane protein linked to neuronal development, as a potent driver of tumor microtube (TM)-mediated brain colonization by glioma cells. Targeting of Ttyh1 effectively inhibited the formation of invasive TMs and glioma growth, but increased network formation by intercellular TMs, suggesting a functional and molecular heterogeneity of the recently discovered TMs with potential implications for future TM-targeting strategies.
The standard of care for anaplastic gliomas is surgery followed by radiotherapy. The NOA-04 phase III trial compared efficacy and safety of radiotherapy followed by chemotherapy at progression with ...the reverse sequence in patients with newly diagnosed anaplastic gliomas.
Patients (N = 318) were randomly assigned 2:1:1 (A:B1:B2) to receive conventional radiotherapy (arm A); procarbazine, lomustine (CCNU), and vincristine (PCV; arm B1); or temozolomide (arm B2) at diagnosis. At occurrence of unacceptable toxicity or disease progression, patients in arm A were treated with PCV or temozolomide (1:1 random assignment), whereas patients in arms B1 or B2 received radiotherapy. The primary end point was time to treatment failure (TTF), defined as progression after radiotherapy and one chemotherapy in either sequence.
Patient characteristics in the intention-to-treat population (n = 274) were balanced between arms. All histologic diagnoses were centrally confirmed. Median TTF (hazard ratio HR = 1.2; 95% CI, 0.8 to 1.8), progression-free survival (PFS; HR = 1.0; 95% CI, 0.7 to 1.3, and overall survival (HR = 1.2; 95% CI, 0.8 to 1.9) were similar for arms A and B1/B2. Extent of resection was an important prognosticator. Anaplastic oligodendrogliomas and oligoastrocytomas share the same, better prognosis than anaplastic astrocytomas. Hypermethylation of the O(6)-methylguanine DNA-methyltransferase (MGMT) promoter (HR = 0.59; 95% CI, 0.36 to 1.0), mutations of the isocitrate dehydrogenase (IDH1) gene (HR = 0.48; 95% CI, 0.29 to 0.77), and oligodendroglial histology (HR = 0.33; 95% CI, 0.2 to 0.55) reduced the risk of progression. Hypermethylation of the MGMT promoter was associated with prolonged PFS in the chemotherapy and radiotherapy arm.
Initial radiotherapy or chemotherapy achieved comparable results in patients with anaplastic gliomas. IDH1 mutations are a novel positive prognostic factor in anaplastic gliomas, with a favorable impact stronger than that of 1p/19q codeletion or MGMT promoter methylation.
J. Neurochem. (2012) 122, 444–455.
Temozolomide (TMZ) is an alkylating chemotherapeutic agent that prolongs the survival of patients with glioblastoma. Clinical benefit is more prominent in patients ...with methylation of the O6‐methyl‐guanine DNA methyltransferase (MGMT) promoter. However, all patients eventually suffer from tumor progression because their tumors become resistant to TMZ. Here, we modeled acquired TMZ resistance in glioma cells in vitro to identify underlying molecular mechanisms. To this end, the glioma cell lines LNT‐229, LN‐308, and LN‐18 were exposed repetitively to increasing concentrations of TMZ to induce a stable resistant phenotype (R) defined by clonogenic survival assays. The molecular mechanisms mediating acquired resistance were assessed by immunoblot, PCR, and flow cytometry. Rescue experiments were performed with siRNA‐mediated candidate gene silencing. We found in LN‐18 cells constitutively expressing MGMT a strong up‐regulation of MGMT levels in TMZ‐resistant cells. TMZ resistance in the MGMT‐negative cell lines LNT‐229 and LN‐308 was not associated with de novo expression of MGMT. Instead, we found a down‐regulation of several DNA mismatch‐repair proteins in resistant LNT‐229 cells. A TMZ‐resistant phenotype was also achieved by silencing selected DNA mismatch repair proteins in parental LNT‐229 cells. No obvious mechanism of resistance was identified in the third cell line, LN‐308, except for reduced methylation of LINE‐1 repetitive elements. In conclusion, we demonstrate that different molecular mechanisms may contribute to the development of acquired TMZ resistance in glioma cells, indicating the need to develop distinct strategies to overcome resistance.
Identification of molecular mechanisms leading to temozolomide resistanceAcquired resistance to temozolomide is an important challenge in the setting of therapy failure in glioblastoma patients. Here, we identify distinct mechanisms by which glioma cells become resistant to the anti‐tumor effects of temozolomide. These findings highlight the importance of different therapy‐induced molecular changes, which might be exploited for novel therapeutic approaches in the future.
Somatic mutations in the
IDH1
gene encoding cytosolic NADP+-dependent isocitrate dehydrogenase have been shown in the majority of astrocytomas, oligodendrogliomas and oligoastrocytomas of WHO grades ...II and III.
IDH2
encoding mitochondrial NADP+-dependent isocitrate dehydrogenase is also mutated in these tumors, albeit at much lower frequencies. Preliminary data suggest an importance of
IDH1
mutation for prognosis showing that patients with anaplastic astrocytomas, oligodendrogliomas and oligoastrocytomas harboring
IDH1
mutations seem to fare much better than patients without this mutation in their tumors. To determine mutation types and their frequencies, we examined 1,010 diffuse gliomas. We detected 716
IDH1
mutations and 31
IDH2
mutations. We found 165
IDH1
(72.7%) and 2
IDH2
mutations (0.9%) in 227 diffuse astrocytomas WHO grade II, 146
IDH1
(64.0%) and 2
IDH2
mutations (0.9%) in 228 anaplastic astrocytomas WHO grade III, 105
IDH1
(82.0%) and 6
IDH2
mutations (4.7%) in 128 oligodendrogliomas WHO grade II, 121
IDH1
(69.5%) and 9
IDH2
mutations (5.2%) in 174 anaplastic oligodendrogliomas WHO grade III, 62
IDH1
(81.6%) and 1
IDH2
mutations (1.3%) in 76 oligoastrocytomas WHO grade II and 117
IDH1
(66.1%) and 11
IDH2
mutations (6.2%) in 177 anaplastic oligoastrocytomas WHO grade III. We report on an inverse association of
IDH1
and
IDH2
mutations in these gliomas and a non-random distribution of the mutation types within the tumor entities.
IDH1
mutations of the R132C type are strongly associated with astrocytoma, while
IDH2
mutations predominantly occur in oligodendroglial tumors. In addition, patients with anaplastic glioma harboring
IDH1
mutations were on average 6 years younger than those without these alterations.
Astrocytoma and oligodendroglioma are histologically and genetically well-defined entities. The majority of astrocytomas harbor concurrent
TP53
and
ATRX
mutations, while most oligodendrogliomas carry ...the 1p/19q co-deletion. Both entities share high frequencies of
IDH
mutations. In contrast, oligoastrocytomas (OA) appear less clearly defined and, therefore, there is an ongoing debate whether these tumors indeed constitute an entity or whether they represent a mixed bag containing both astrocytomas and oligodendrogliomas. We investigated 43 OA diagnosed in different institutions employing histology, immunohistochemistry and in situ hybridization addressing surrogates for the molecular genetic markers
IDH1
R132H,
TP53
,
ATRX
and 1p/19q loss. In all but one OA the combination of nuclear p53 accumulation and ATRX loss was mutually exclusive with 1p/19q co-deletion. In 31/43 OA, only alterations typical for oligodendroglioma were observed, while in 11/43 OA, only indicators for mutations typical for astrocytomas were detected. A single case exhibited a distinct pattern, nuclear expression of p53, ATRX loss,
IDH1
mutation and partial 1p/19q loss. However, this was the only patient undergoing radiotherapy prior to surgery, possibly contributing to the acquisition of this uncommon combination. In OA with oligodendroglioma typical alterations, the portions corresponding to astrocytic part were determined as reactive, while in OA with astrocytoma typical alterations the portions corresponding to oligodendroglial differentiation were neoplastic. These data provide strong evidence against the existence of an independent OA entity.
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