DNA methylation, transcriptome and genetic copy number signatures of diffuse cerebral WHO grade II/III gliomas resolve cancer heterogeneity and development.

BACKGROUND: Diffuse lower WHO grade II and III gliomas (LGG) are slowly progressing brain tumors, many of which eventually transform into a more aggressive type. LGG is characterized by widespread genetic and transcriptional heterogeneity, yet little is known about the heterogeneity of the DNA methylome, its function in tumor biology, coupling with the transcriptome and tumor microenvironment and its possible impact for tumor development. METHODS: We here present novel DNA methylation data of an LGG-cohort collected in the German Glioma Network containing about 85% isocitrate dehydrogenase (IDH) mutated tumors and performed a combined bioinformatics analysis using patient-matched genome and transcriptome data. RESULTS: Stratification of LGG based on gene expression and DNA-methylation provided four consensus subtypes. We characterized them in terms of genetic alterations, functional context, cellular composition, tumor microenvironment and their possible impact for treatment resistance and prognosis. Glioma with astrocytoma-resembling phenotypes constitute the largest fraction of nearly 60%. They revealed largest diversity and were divided into four expression and three methylation groups which only partly match each other thus reflecting largely decoupled expression and methylation patterns. We identified a novel G-protein coupled receptor and a cancer-related 'keratinization' methylation signature in in addition to the glioma-CpG island methylator phenotype (G-CIMP) signature. These different signatures overlap and combine in various ways giving rise to diverse methylation and expression patterns that shape the glioma phenotypes. The decrease of global methylation in astrocytoma-like LGG associates with higher WHO grade, age at diagnosis and inferior prognosis. We found analogies between astrocytoma-like LGG with grade IV IDH-wild type tumors regarding possible worsening of treatment resistance along a proneural-to-mesenchymal axis. Using gene signature-based inference we elucidated the impact of cellular composition of the tumors including immune cell bystanders such as macrophages. CONCLUSIONS: Genomic, epigenomic and transcriptomic factors act in concert but partly also in a decoupled fashion what underpins the need for integrative, multidimensional stratification of LGG by combining these data on gene and cellular levels to delineate mechanisms of gene (de-)regulation and to enable better patient stratification and individualization of treatment.

[Chronic subdural hematoma in the elderly]. / Das chronische Subduralhämatom im Alter.

Chronic subdural hematoma (cSDH) is a common condition, the frequency of which further increases due to an aging population and more frequent use of antithrombotic drugs. It leads to unspecific symptoms and neurological deficits and is usually treated surgically. Burr hole trepanation and twist drill craniostomy have become the therapeutic standards with craniotomy being rarely used for recurrent cases. Although recurrences are relatively common, in most cases a good outcome can be achieved even in the elderly; however, as cSDH is associated with other comorbidities, it is indicative of an increased morbidity and mortality. Controlled trials need to be carried out to determine whether pharmacological therapies can also be beneficial in addition to surgical treatment.

[Importance of preoperative and intraoperative imaging for operative strategies]. / Bedeutung der prä- und intraoperativen Bildgebung für die operative Strategie.

Recent advances in preoperative and postoperative imaging have an increasing influence on surgical decision-making and make more complex surgical interventions possible. This improves the possibilities for frequently occurring challenges and promoting improved functional and oncological outcome. This manuscript reviews the role of preoperative and intraoperative imaging in surgery. Various techniques are explained based on examples from hepatobiliary surgery and neurosurgery, in particular real-time procedures, such as the online use of augmented reality and in vivo fluorescence, as well as new and promising optical techniques including imaging of intrinsic signals and vibrational spectroscopy.

[Surgery for metastases, anatomical and ethical limits. Special aspect: oligometastases]. / Metastasenchirurgie - anatomische und ethische Grenzen. Besonderer Aspekt: Oligometastasierung.

The surgical resection of metastases is nowadays feasible in selected patients with multifocal metastatic disease due to the implementation of interdisciplinary multimodal therapeutic options. Anatomical limitations do not seem to represent obstacles which cannot be overcome because of the development of new surgical techniques. The cornerstone of the selection of patients is the correct staging diagnosis achieved through modern diagnostic tools; however, surgery alone does not always offer acceptable survival and recurrence-free rates. Furthermore, in every complex surgical procedure there is the risk of morbidity and mortality; therefore, parameters such as alternative therapeutic modalities, the individual situation of the patient and tumor biology have to be considered in order to make the correct selection of patients. This is one of the major future challenges and should never be driven by unfounded hopes and expectations of the patients. The same principle also applies for brain metastases, which represent the most common brain tumors. Approximately 70 % of patients with brain metastases have 1-3 lesions (oligometastases). Treatment is now individualized and the goal of therapy has shifted towards long-term survival (≥ 24 months) and improved quality of life. Under this aspect surgery is one of the important treatment options, particularly in patients with a single metastasis or oligometastases. Furthermore, approximately 20 % of patients who have recurrent brain metastases, successfully undergo a complete resection of tumors and with a Karnofsky performance status (KPS) score > 70 show a long-term survival of ≥ 24 months.

NDRG1 prognosticates the natural course of disease in WHO grade II glioma.

There is a lack of relevant prognostic and predictive factors in neurooncology besides mutation of isocitrate dehydrogenase 1, codeletion of 1p/19q and promoter hypermethylation of O (6) -methylguanine-DNA-methyltransferase. More importantly, there is limited translation of these factors into clinical practice. The cancer genome atlas data and also clinical correlative analyses suggest a pivotal role for the epidermal growth factor receptor /protein kinase B/mammalian target of rapamycin (mTOR) pathway in both biology and the clinical course of gliomas. However, attempts to stratify gliomas by activating alterations in this pathway have failed thus far. The tumors of 40 patients with WHO grade II gliomas without immediate postoperative genotoxic treatment and known progression and survival status at a median follow-up of 12.2 years were analyzed for expression of the mTOR complex 2 downstream target N-myc downstream regulated gene (NDRG)1 using immunohistochemistry. Baseline characteristics for NDRG1 absent/low versus moderate/high patients were similar. Time to reintervention was significantly longer in the NDRG1 group (P = 0.026). NDRG1 may become a novel biomarker to guide the decision which WHO°II glioma patients may be followed without postsurgical intervention and which patients should receive genotoxic treatment early on. Validation of this hypothesis will be possible with the observational arm of the RTOG 9802 and the pretreatment step of the EORTC 22033/26032 trials.

Gross total but not incomplete resection of glioblastoma prolongs survival in the era of radiochemotherapy.

BACKGROUND: This prospective multicenter study assessed the prognostic influence of the extent of resection when compared with biopsy only in a contemporary patient population with newly diagnosed glioblastoma. PATIENTS AND METHODS: Histology, O(6)-methylguanine-DNA methyltransferase (MGMT) promoter methylation status, and clinical data were centrally analyzed. Survival analyses were carried out with the Kaplan-Meier method. Prognostic factors were assessed with proportional hazard models. RESULTS: Of 345 patients, 273 underwent open tumor resection and 72 biopsies; 125 patients had gross total resections (GTRs) and 148, incomplete resections. Surgery-related morbidity was lower after biopsy (1.4% versus 12.1%, P = 0.007). 64.3% of patients received radiotherapy and chemotherapy (RT plus CT), 20.0% RT alone, 4.3% CT alone, and 11.3% best supportive care as an initial treatment. Patients ≤60 years with a Karnofsky performance score (KPS) of ≥90 were more likely to receive RT plus CT (P < 0.01). Median overall survival (OS) (progression free survival; PFS) ranged from 33.2 months (15 months) for patients with MGMT-methylated tumors after GTR and RT plus CT to 3.0 months (2.4 months) for biopsied patients receiving supportive care only. Favorable prognostic factors in multivariate analyses for OS were age ≤60 years [hazard ratio (HR) = 0.52; P < 0.001], preoperative KPS of ≥80 (HR = 0.55; P < 0.001), GTR (HR = 0.60; P = 0.003), MGMT promoter methylation (HR = 0.44; P < 0.001), and RT plus CT (HR = 0.18, P < 0.001); patients undergoing incomplete resection did not better than those receiving biopsy only (HR = 0.85; P = 0.31). CONCLUSIONS: The value of incomplete resection remains questionable. If GTR cannot be safely achieved, biopsy only might be used as an alternative surgical strategy.

Association of transcobalamin c. 776C>G with overall survival in patients with primary central nervous system lymphoma.

BACKGROUND: Chemotherapy for primary central nervous system lymphoma (PCNSL) is based on methotrexate (MTX), which interferes with both nucleic acid synthesis and methionine metabolism. We have reported previously that genetic variants with influence on methionine metabolism are associated with MTX side effects, that is, the occurrence of white matter lesions as a sign of MTX neurotoxicity. Here, we investigated whether such variants are associated with MTX efficacy in terms of overall survival in MTX-treated PCNSL patients. METHODS: We analysed seven genetic variants influencing methionine metabolism in 68 PCNSL patients treated with systemic and facultative intraventricular MTX-based polychemotherapy (Bonn protocol). RESULTS: Median age at diagnosis was 59 years (range: 28-77), 32 patients were female. Younger age (Wald=8.9; P=0.003) and the wild-type C (CC) allele of the genotype transcobalamin c (Tc2). 776C>G (Wald=6.7; P=0.010) were associated with longer overall survival in a multivariate COX regression analysis. CONCLUSION: This observation suggests that the missense variant Tc2. 776C>G influences both neurotoxicity and efficacy of MTX in the Bonn PCNSL protocol.

The prognostic value of IDH mutations and MGMT promoter status in secondary high-grade gliomas.

Reports about the prognostic value of IDH mutations and the promoter region of the O6-Methyl-guanyl-methyl-transferase gene in secondary high-grade gliomas (sHGG) are few in number. We investigated the prognostic value of IDH mutations and methylation of the promoter region of the MGMT gene in 99 patients with sHGG and analyzed the clinical course of those tumors. Patients with sHGG were screened for IDH mutations by direct sequencing, and, for promoter status of MGMT gene, by the methylation-specific polymerase chain reaction. A total of 48 of 99 patients (48.5 %) had secondary anaplastic gliomas (Group 1), while 51 patients had secondary glioblastomas (Group 2). The median survival time after malignant progression of all patients with sHGG and with an IDH mutation was 4 years, which is significantly longer than in patients with wild-type IDH (1.2 years, p = 0.009). Patients' survival was not significantly influenced by the tumors' MGMT promoter status, both in Group 1- 9.7 years vs. 6.1 years, methylated vs. unmethylated promoter (p = 0.330)-as well as in Group 2-1.5 years vs. 1.6 years, methylated versus unmethylated promoter (p = 0.829). In our population, the IDH mutation status was not associated with increased PFS or median survival time in sGBM patients. However, patients with secondary anaplastic glioma and IDH mutation had a significantly improved outcome. In addition, IDH mutations are a more powerful prognostic marker concerning both PFS and MS than the MGMT promoter status in those patients.

The prostate stem cell antigen represents a novel glioma-associated antigen.

Gliomas of WHO grades III-IV are malignant brain tumors mostly resistant to conventional therapies. Therefore, novel strategies for the treatment of gliomas are warranted. Although immunotherapy is gaining increased attention for the treatment of malignant gliomas and in particular of glioblastoma multiforme (GBM), this approach requires the identification of appropriate antigens. Our aim was to investigate the expression of the prostate stem cell antigen (PSCA), a highly N-glycosylated phosphatidylinositol (GPI)-anchored cell surface protein, in gliomas of different WHO grades in order to evaluate its potential as a diagnostic marker and as a target for immunotherapy. Tumor specimens and controls were assessed by quantitative RT-PCR, Western blotting and immunohistochemistry. The samples investigated in the study consisted of 210 human glial tumors, among which 31 were oligodendrogliomas, 9 ependymomas and 170 were astrocytomas (including 134 glioblastomas). PSCA was absent in normal brain tissue, but was detected in WHO grade III-IV gliomas. Weak PSCA protein expression was also recognized in some WHO grade I and WHO grade II tumors. The difference between WHO grade I-II tumors and WHO grade III-IV tumors was statistically significant (p<0.001). Our results suggest that increased PSCA expression levels are linked to gliomas of WHO grades III and IV, and may represent a suitable additional target for immunotherapy of gliomas.

Rosette-forming glioneuronal tumor of the fourth ventricle in an elderly patient.

Rosette-forming glioneuronal tumor of the fourth ventricle (RGNT) is a rare condition, which previously has been described predominantly in middle-aged patients. There is limited experience with this kind of tumor in the elderly. Clinical, neuroimaging, and histological features of an example in a 70-year-old male who presented initially with vertigo are detailed and compared with published cases. Neuroimaging studies demonstrated a 4-cm cystic lesion in posterior fossa containing a 1-cm contrast-enhancing nodule on its lateral margin. The lesion was confined to the fourth ventricle and initially thought to be a hemangioblastoma until angiography clarified the minimal tumor vascularization. Gross total resection was achieved. Pathological examination showed a rosette-forming low grade tumor with a cell proliferation rate of 2% being consistent with RGNT. The postoperative course was uneventful and clinical symptoms resolved completely. There was no tumor recurrence after 2 years follow-up. We confirm that the rare and only recently characterized tumor entity of RGNT can also be found in elderly patients; furthermore, it can be associated with a benign course. The main differential diagnosis of RGNT resulting from CNS-imaging modalities in elderly patients are pilocytic astrocytoma and hemangioblastoma of the posterior fossa, which after metastasis are the most common primary adult intra-axial posterior fossa tumors. Therefore, a subtle preoperative radiological diagnosis is warranted and surgery should be performed by experienced hands to avoid neurological deterioration.

Complementary therapy use in patients with glioma: an observational study.

OBJECTIVE: Despite novel multimodal therapeutic approaches, the vast majority of glial tumors are not curable. Patients may search for complementary therapies in order to contribute to the fight against their disease or to relieve symptoms induced by their brain tumor. The extent of the use of complementary or alternative therapies, the patients' rationale behind it, and the cost of complementary therapy for gliomas are not known. We used a questionnaire and the database of the German Glioma Network to evaluate these questions. METHODS: A total of 621 questionnaires were available for evaluation from patients with glial tumors of WHO grades II to grade IV. The patients were recruited from 6 neuro-oncologic centers in Germany. Complementary therapy was defined as methods or compounds not used in routine clinical practice and not scientifically evaluated. RESULTS: Forty percent of the responding patients reported the use of complementary therapies. Significant differences between the group of complementary therapy users and nonusers were seen with respect to age (younger > older), gender (female > male), and education (high education level > low education level). The motivation for complementary therapy use was not driven by unsatisfactory clinical care by the neuro-oncologists, but by the wish to add something beneficial to the standard of care. CONCLUSIONS: In clinical practice, patients' use of complementary therapies may be largely overseen and underestimated. The major motivation is not distrust in conventional therapies. Neuro-oncologists should be aware of this phenomenon and encourage an open but critical dialogue with their patients.

A MDR1 (ABCB1) gene single nucleotide polymorphism predicts outcome of temozolomide treatment in glioblastoma patients.

BACKGROUND: Some patients with glioblastoma multiform do not respond to temozolomide even though they have aberrant promoter methylation of the DNA repair enzyme O(6)-methylguanine methyltransferase (MGMT). This suggests that additional factors hamper temozolomide cytotoxicity. We aimed to confirm first that temozolomide is a target for the multidrug resistance transporter MDR1/ABCB1 and second to investigate whether genetic variants of the MDR1 gene are associated with the survival of glioblastoma patients treated with temozolomide. MATERIALS AND METHODS: Temozolomide-mediated cytotoxicity was determined by the colorimetric methyl-thiazol-tetrazolium assay in MDR-expressing and MDR-nonexpressing cell lines. Genotypes of three single nucleotide polymorphisms (SNPs) of the MDR1 gene (C1236T, G2677T, and C3435T), MDR1 mRNA expression levels, and the MGMT promoter methylation status were analyzed in 112 glioblastoma patients who had been treated either by surgery plus radiotherapy alone or by additional temozolomide chemotherapy. RESULTS: In vitro analysis revealed that temozolomide-mediated cytotoxicity is dependent on MDR1 expression. Multivariate analysis of MDR1 genotypes showed that the C/C variant of the exon12 C1236T SNP is predictive for survival of patients treated with temozolomide. This effect was independent of the MGMT methylation status. Patients with the C/C genotype had a 2-year overall survival of 37% compared with 8% and 10% for patients with C/T and T/T genotypes, respectively (P=0.02). No influence was seen in the group of patients with radiotherapy only. CONCLUSION: The genotype of the MDR1 exon12 C1236T SNP is a novel independent predictive factor for outcome of temozolomide treatment in glioblastoma patients.

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.

Integration of intraoperative 3D-ultrasound in a commercial navigation system.

STUDY AIMS: The purpose of this study was the integration of three-dimensional ultrasound data into a neuronavigation system, in order to allow a guided intraoperative resection control during neurosurgical interventions. MATERIAL AND METHODS: A system for iterative neuronavigation based on 3D-ultrasound (US) has been developed. The main components of the system are the ultrasound device Voluson 730 (GE Healthcare) with a 5 - 9 MHz probe, the navigation system VectorVision2 (BrainLAB AG) and a standard PC with Windows XP. The ultrasound data are transferred via DICOM from the ultrasound device to an external computer, where they are processed with a C++ program for representation in the neuronavigation coordinate system. The data transfer between the navigation system and the external computer is performed via the VVLink interface from BrainLAB. The feasibility test of the system was performed with an ultrasound phantom RMI 403GS (Gammex-RMI GmbH). RESULTS: The error of homologous points mapping from US datasets to a CT dataset in the neuronavigation system was determined to be 1.9 +/- 0.97 mm. The maximum time required to technically integrate the ultrasound data into the navigation system was 1.5 min. CONCLUSIONS: The developed system allows 3D-ultrasound based navigation to be carried out with a commercially available navigation system. The functionality of this system has been proven by technical tests. Recording and integration of the ultrasound data can be repeated at any time during surgery and can be used to update anatomical data and consequently for resection control. Another application is the intraoperative adaptation of preoperative datasets (MRI or CT) in order to compensate for "brain shift" during neurosurgical operations.

Outcomes of surgical resection of large solitary hemangioblastomas of the craniocervical junction with limitations in preoperative angiographic intervention: report of three cases.

INTRODUCTION: Hemangioblastomas of the central nervous system may occur sporadically, or in association with von Hippel-Lindau (vHL) disease. The treatment of large solitary hemangioblastomas of the posterior cranial fossa mandates a combination of angiographic intervention and surgery. However, large tumors may derive their vascularity from major cerebellar vessels, which can make their embolization hazardous. AIM: To describe the surgical outcomes of three cases of large hemangioblastomas with compression of the medulla oblongata, where the potential for preoperative embolization was extremely limited. CASES: Three patients (all males; 68, 36 and 38 years) presented with a history of chronic headache and caudal cranial nerve deficiencies. Diagnostic imaging showed large vascular lesions (4 x 3, 4 x 5 and 5 x 5 cm) at the craniocervical junction, compressing the brainstem. There were no concomitant findings associated with vHL disease. TREATMENT: Staged treatment was administered. Preoperative embolization was attempted at first. One patient (68 yrs) showed a PICA occlusion and associated cerebellar infarction after embolization; embolization was deemed hazardous in the other two. In the second phase, the lesions were removed via a midline suboccipital approach with resection of the arch of altas. Complete removal was possible in all three cases. POSTOPERATIVE COURSE AND FOLLOW-UP: The caudal cranial nerve deficiencies deteriorated soon after surgery in all three patients. A tracheotomy was required in two patients, which was removed uneventfully during the rehabilitation phase. Ventriculo-peritoneal shunts were implanted in two patients. MRI follow-up three (1 case) and four years (2 cases) after surgery showed no relapse. The Karnofsky Index scores were 80, 70 and 90 in the three patients aged 68, 36 and 38, respectively. CONCLUSION: Total microneurosurgical removal of large hemangioblastomas at the craniocervical junction with limited preoperative embolization (associated with morbidity) should be seriously considered. Although the early outcome is not encouraging, the long-term outcomes seem favorable.

Primary aspergillosis of the sphenoid sinus with pituitary invasion - a rare differential diagnosis of sellar lesions.

Aspergillosis belongs to the group of mycotic diseases of paranasal sinuses. The invasive forms, and particularly the fulminant forms, are potentially fatal. Isolated aspergillosis of the sphenoid sinus or the clivus is a difficult diagnosis, since the often misleading clinical manifestations of this rare disease develop late. These patients become apparent by neurological signs such as cavernous sinus syndrome, pseudotumor of the pituitary or the orbit. Diagnosis is often made intra-operatively or on histological examination. We report a case of invasive aspergillosis uniquely involving the sellar area revealed by clinical features suggesting a pseudotumor of the pituitary. Although such lesions are almost always seen in immune suppressed subjects, in our case, the patient was immune competent and had no past history of sinusitis.The question of whether, and when to perform limited or extensive surgery remains an issue for discussion, owing to the rarity of this disease honed by lack of experience. It depends on several factors: the kind of disease, the immunity, the subtype of invasive fungal sinusitis and the degree of tissue invasion.