World Health Organization Grade II-IV Research Articles

Leptomeningeal Metastasis in Gliomas : Clinical Characteristics and Risk Factors.

Our objective is to analyze the occurrence, clinical course and risk factors for glioma patients with leptomeningeal metastasis (LM) according to different metastasis patterns and clinical variables. We retrospectively reviewed data from 376 World Health Organization (WHO) grade II-IV adult glioma patients who were treated in the National Cancer Center from 2001 to 2020. Patients who underwent surgery at other institutions, those without initial images or those with pathologically unconfirmed cases were excluded. LM was diagnosed based on magnetic resonance imaging (MRI) findings or cerebrospinal fluid (CSF) cytology. The metastasis pattern was categorized as nodular or linear according to the enhancement pattern. Tumor proximity to the CSF space was classified as involved or separated, whereas location of the tumor was dichotomized as midline, for tumors residing in the thalamus, basal ganglia and brainstem, or lateral, for tumors residing in the cerebral and cerebellar hemispheres. A total of 138 patients were enrolled in the study. A total of 44 patients (38%) were diagnosed with LM during a median follow-up of 9 months (range, 0-60). Among the clinical variables, tumor proximity to CSF space, the location of the tumor and the WHO grade were significant factors for LM development in univariate analysis. In multivariate analysis, the midline location of the tumor and WHO grade IV gliomas were the most significant factor for LM development. The hazard ratio was 2.624 for midline located gliomas (95% confidence interval [CI], 1.384-4.974; p=0.003) and 3.008 for WHO grade IV gliomas (95% CI, 1.379-6.561; p=0.006). Midline location and histological grading are an important factor for LM in glioma patients. The proximity to the CSF circulation pathway is also an important factor for WHO grade IV glioma LM. Patients carrying high risks should be followed up more thoroughly.

Effects of endoplasmic reticulum stress-mediated CREB3L1 on apoptosis of glioma cells.

The association between endoplasmic reticulum stress (ERS) and apoptosis has been extensively studied. Cyclic adenosine monophosphate responsive element binding protein 3 like 1 (CREB3L1) has an important role in the development of glioma. In the present study, the potential association between ERS-induced apoptosis and CREB3L1 and its clinical implications were investigated. From a total of 30 cases, brain gliomas with different pathological grades surgically resected at the Department of Neurosurgery of the Affiliated Hospital of Guizhou Medical University (Guiyang, China) between January 2018 and January 2019 were collected. The expression of CREB3L1 and ERS-related proteins in gliomas with different degrees of malignancy was detected by immunohistochemistry. Furthermore, U87-MG glioma cells were cultured in vitro and treated with different concentrations of ERS inducer thapsigargin (TG). The Cell Counting Kit-8 (CCK-8) assay was performed to detect changes in cell activity at different incubation times and drug concentrations. Cell apoptosis was detected by Annexin Ⅴ-FITC/propidium iodide double staining and the protein expression levels of CREB3L1 and ERS were detected by western blot analysis. Immunohistochemical analysis suggested that the expression levels of CREB3L1, glucose-regulated protein, 78 kDa (GRP78) and C/EBP-homologous protein (CHOP) in World Health Organization (WHO) grade I glioma were higher than those in WHO grade Ⅱ-Ⅳ (all P<0.01). The results of the CCK-8 assay suggested that the activity of U87-MG glioma cells was significantly decreased after treatment with TG (all P<0.05), and this effect was time- and drug concentration-dependent. Flow cytometric analysis indicated that the apoptotic rate of the cells was increased, which was significant when the concentration of TG was 0.1 µmol/l (P<0.01). Furthermore, the protein expression of CREB3L1, GRP78 and CHOP in glioma cells treated with TG was increased (P<0.05). However, the expression level of Bcl-2 decreased (P<0.05). In conclusion, ERS may reduce the cell proliferative activity and promote apoptosis through mediating CREB3L1 expression. CREB3L1 may be a novel potential target for glioma therapy.

Impact of awake mapping on overall survival and extent of resection in patients with adult diffuse gliomas within or near eloquent areas: a retrospective propensity score-matched analysis of awake craniotomy vs. general anesthesia

Awake craniotomy (AC) with intraoperative mapping is the best approach to preserve neurological function for glioma surgery in eloquent or near eloquent areas, but whether AC improves the extent of resection (EOR) and overall survival (OS) is controversial. This study aimed to compare the long-term clinical outcomes of glioma resection under AC with those under general anesthesia (GA). Data of 335 patients who underwent surgery with intraoperative magnetic resonance imaging for newly diagnosed gliomas of World Health Organization (WHO) grades II-IV between 2000 and 2013 were reviewed. EOR and OS were quantitatively compared between the AC and GA groups after 1:1 propensity score matching. The two groups were matched for age, preoperative Karnofsky performance status (KPS), tumor location, and pathology. After propensity score matching, 91 pairs were obtained. The median EOR was 96.1% (interquartile range [IQR] 7.3) and 97.4% (IQR 14.4) in the AC and GA groups, respectively (p = 0.31). Median KPS score 3months after surgery was 90 (IQR 20) in both groups (p = 0.384). The median survival times were 163.3months (95% confidence interval [CI] 77.9-248.7) and 143.5months (95% CI 94.4-192.7) in the AC and GA groups, respectively (p = 0.585). Even if the glioma was within or close to the eloquent area, AC was comparable with GA in terms of EOR and OS. In case of difficulties in randomizing patients with eloquent or near eloquent glioma, our propensity score-matched analysis provides retrospective evidence that AC can obtain EOR and OS equivalent to removing glioma under GA.

Amide proton transfer-weighted MRI in the clinical setting – correlation with dynamic susceptibility contrast perfusion in the post-treatment imaging of adult glioma patients at 3T

IntroductionWe investigated the correlation between amide proton transfer-weighted magnetic resonance imaging (APTw MRI) and dynamic susceptibility contrast (DSC) perfusion in order to assess the potential of APTw MRI as an alternative to DSC in adult brain tumor (glioma) imaging. MethodsAfter Ethical Committee approval, forty adult patients, treated for histopathologically confirmed glioma (World Health Organization (WHO) grade II-IV), were prospectively imaged at 3 Tesla (3 T) with DSC perfusion and a commercially available three-dimensional (3D) APTw sequence. Two consultant neuroradiologists independently performed region of interest (ROI) measurements on relative cerebral blood volume (rCBV) and APTw maps, co-registered with anatomical images. The correlation APTw MRI-DSC perfusion was assessed using Spearman's rank-order test. Inter-observer agreement was evaluated by the intraclass correlation coefficient (ICC) and Bland-Altman (BA) plots. ResultsA statistically significant moderately strong positive correlation was observed between maximum rCBV (rCBVmax) and maximum APTw (APTwmax) values (observer 1: r = 0.73; p < 0.01; observer 2: r = 0.62; p < 0.01). We found good inter-observer agreement for APTwmax (ICC = 0.82; 95% confidence interval (CI) 0.66–0.90), with somewhat broad outer 95% CI for the BA Limits of Agreement (LoA) (−1.6 to 1.9). ICC for APTwmax was higher than ICC for rCBVmax (ICC = 0.74; 95%; CI 0.50–0.86), but the difference was not statistically significant. ConclusionAPTwmax values correlate positively with rCBVmax in patients treated for brain glioma. APTw imaging is a reproducible technique, with some observer dependence. Results need to be confirmed by a larger population analysis. Implications for practiceAPTw MRI can be a useful addition to glioma follow-up imaging and a potential alternative to DSC perfusion, especially in patients where contrast agent is contraindicated.

Support vector machine based aphasia classification of transcranial magnetic stimulation language mapping in brain tumor patients

Repetitive TMS (rTMS) allows for non-invasive and transient disruption of local neuronal functioning. We used machine learning approaches to assess whether brain tumor patients can be accurately classified into aphasic and non-aphasic groups using their rTMS language mapping results as input features. Given that each tumor affects the subject-specific language networks differently, resulting in heterogenous rTMS functional mappings, we propose the use of machine learning strategies to classify potential patterns of rTMS language mapping results. We retrospectively included 90 patients with left perisylvian world health organization (WHO) grade II-IV gliomas that underwent presurgical navigated rTMS language mapping. Within our cohort, 29 of 90 (32.2%) patients suffered from at least mild aphasia as shown in the Aachen Aphasia Test based Berlin Aphasia Score (BAS). After spatial normalization to MNI 152 of all rTMS spots, we calculated the error rate (ER) in each stimulated cortical area (28 regions of interest, ROI) by automated anatomical labeling parcellation (AAL3) and IIT. We used a support vector machine (SVM) to classify significant areas in relation to aphasia. After feeding the ROIs into the SVM model, it revealed that in addition to age (w = 2.98), the ERs of the left supramarginal gyrus (w = 3.64), left inferior parietal gyrus (w = 2.28) and right pars triangularis (w = 1.34) contributed more than other features to the model. The model’s sensitivity was 86.2%, the specificity was 82.0%, the overall accuracy was 85.5% and the AUC was 89.3%. Our results demonstrate an increased vulnerability of right inferior pars triangularis to rTMS in aphasic patients due to left perisylvian gliomas. This finding points towards a functional relevant involvement of the right pars triangularis in response to aphasia. The tumor location feature, specified by calculating overlaps with white and grey matter atlases, did not affect the SVM model. The left supramarginal gyrus as a feature improved our SVM model the most. Additionally, our results could point towards a decreasing potential for neuroplasticity with age.

Differentiation of recurrent diffuse glioma from treatment-induced change using amide proton transfer imaging: incremental value to diffusion and perfusion parameters.

To evaluate the incremental value of amide proton transfer (APT) imaging to diffusion tensor imaging (DTI), dynamic susceptibility contrast (DSC) imaging, and dynamic contrast-enhanced (DCE) imaging in differentiating recurrent diffuse gliomas (World Health Organization grade II-IV) from treatment-induced change after concurrent chemoradiotherapy or radiotherapy. This study included 36 patients (25 patients with recurrent gliomas and 11 with treatment-induced changes) with post-treatment gliomas. The mean values of apparent diffusion coefficient (ADC), fractional anisotropy (FA), normalized cerebral blood volume (nCBV), normalized cerebral blood flow, volume transfer constant, rate transfer coefficient, extravascular extracellular volume fraction, plasma volume fraction, and APT asymmetry index were assessed. Independent quantitative parameters were investigated to predict recurrent glioma using multivariable logistic regression. The incremental value of APT signal to other parameters was assessed by the increase of the area under the curve, net reclassification index, and integrated discrimination improvement. Univariable analysis showed that lower ADC (p = 0.018), higher FA (p = 0.031), higher nCBV (p = 0.021), and higher APT signal (p = 0.009) were associated with recurrent gliomas. In multivariable logistic regression, the diagnostic performance of the model with ADC, FA, and nCBV significantly increased when APT signal was added, with areas under the curve of 0.87 and 0.92, respectively (net reclassification index of 0.77 and integrated discrimination improvement of 0.13). APT imaging may be a useful imaging biomarker that adds value to DTI, DCE, and DSC parameters for distinguishing between recurrent gliomas and treatment-induced changes.

Factors associated with health-related quality of life in patients with glioma: impact of symptoms and implications for rehabilitation.

The factors associated with health-related quality of life in patients with glioma remain unclear; particularly, the impact of symptoms on quality of life has not been studied comprehensively. This study aims to document the quality of life of patients with glioma and clarify the impact of symptoms. In this cross-sectional study, participants were recruited from patients at The University of Tokyo Hospital and from patients who were registered at the Japan Brain Tumor Alliance. We included adult patients with World Health Organization grade II-IV glioma and excluded those with disturbances of consciousness or aphasia. We used the European Organization for Research and Treatment of Cancer QLQ-C30 and BN20 to evaluate quality of life and the symptoms. Multiple regression analyses were performed to investigate the impact of symptoms on European Organization for Research and Treatment of Cancer global health status and QLQ-C30 social functioning. In addition, we performed univariate subgroup analyses classified by World Health Organization grade and history of chemotherapy. This study included 76 patients. Seven symptoms occurred in more than 50% of the patients: fatigue, future uncertainty, drowsiness, communication deficit, financial difficulties, motor dysfunction and weakness of legs. Multiple regression analyses showed that insomnia affected their global health status, and appetite loss, financial difficulties and motor dysfunction were significantly related to their social functioning. In subgroup analysis, the number of symptom subscales that were significantly related to global health status and social functioning was larger in World Health Organization grade II patients compared with grade III/IV patients. In addition to neurological deficits, symptoms were associated with poor quality of life in patients with glioma. This study provided the basis on further investigation of usefulness of symptom evaluation on quality of life improvement.

Evaluating angioarchitectural characteristics of glial and metastatic brain tumors with conventional magnetic resonance imaging

Primary and metastatic brain tumors can overlap in traditional imaging features detected on preoperative conventional magnetic resonance imaging (MRI). The research objective was to determine whether morphological vascular characteristics present in routine preoperative imaging using traditional MRI sequences are predictive of primary versus metastatic brain tumors; secondarily to determine association of conventional and vascular-related imaging parameters with intraoperative blood loss, pathological invasion, and World Health Organization (WHO) tumor grade. A retrospective review analyzed 100 consecutive intracranial tumor surgeries, 50 WHO grade II-IV gliomas and 50 intracranial metastases. Two blinded expert readers independently evaluated preoperative MRIs, obtained via standard morphological imaging sequences, for adjacent or intra-tumoral arterial aneurysm, peritumoral venous ectasia, prominence, or engorgement (“aberrant peritumoral vessels”), and prominent intra-tumoral flow voids. Multivariate analysis was performed to develop models predictive of glioma and glioblastoma (GBM). Aberrant peritumoral vessels and prominent intra-tumoral flow voids were statistically significant predictors of glioma in univariate analyses (p = 0.048, p = 0.001, respectively) and when combined in multivariate analysis (OR = 5.23, p = 0.001), particularly for GBM (OR = 9.08, p < 0.001). Multivariate modeling identified prominent intra-tumoral flow voids and FLAIR invasion as the strongest combined predictors of gliomas and GBM. Aberrant peritumoral vessels and larger tumor volume predicted higher intraoperative blood loss in all analyses. No vascular-related parameters predicted pathological invasion on multivariate analysis. Aberrant peritumoral vessels and prominent intra-tumoral flow voids were predictive of gliomas, specifically GBM. These vascular characteristics, evaluated on routine clinical preoperative MRI imaging, may aid in distinguishinggliomafrom brainmetastases andmay predict intraoperative blood loss.

Expression and clinical significance of calumenin in brain gliomas

Objective To detect the expression of calumenin (CALU) in human brain glioma samples, to explore its effect on the patient′s outcome, and to analyze the related biological functions of the genes co-expressed with CALU. Methods A retrospective analysis was conducted on CALU sequencing expression and clinical data of 288 glioma patients with positive CALU expression in the mRNAseq_325 database of Chinese Glioma Genome Atlas (CGGA). We analyzed the effects of CALU expression levels on survival in different types of glioma patients. The multivariate Cox regression method was used to determine the relevant factors affecting the survival of glioma patients. Pearson correlation analysis was employed to screen the related genes co-expressed with CALU and their biological functions were explored using the online bioinformatic analysis tools. Results Among the 288 patients, the expression of CALU was 4.74±0.68, 5.49±1.12 and 6.17±0.94 in World Health Organization (WHO) grade Ⅱ, Ⅲ, and Ⅳ patients respectively, and 6.13±1.17 and 5.08±0.76 in isocitrate dehydrogenase l (IDH1) wild-type and IDH1 mutant gliomas respectively, and 5.79±0.07 and 4.71±0.07 in gliomas with 1p/19q non-codeletion and 1p/19q co-deletion. The comparative differences in the above indicators were statistically significant (all P 0.05). The median survival of CALU patients with high and low expression levels (144 cases in each) was 13.0(1-132)months and 82.5(2-149)months, respectively, which had statistically significant difference (P<0.01). In patients with WHO grade Ⅱ-Ⅳ gliomas, the high expression of CALU mainly reduced the survival time of grade Ⅲ and Ⅳ patients (both P<0.01). The multivariate Cox regression analysis showed that CALU expression was an independent factor influencing the survival of glioma patients (RR=2.193, 95%CI: 1.507-3.192, P<0.01). In the CGGA database, there were 325 genes that were positively correlated with CALU expression. The biological functions of those genes were mainly signal transduction, cell adhesion and cellular metabolism, and were mainly involved in endoplasmic reticulum protein processing, adhesion and cancer pathways. Conclusion CALU is significantly related to the malignancy of brain glioma and may serve as an independent prognostic factor in high grade glioma patients, which thus provides a potential therapeutic target in the future research. Key words: Glioma; Gene expression; Prognosis; Calumenin

Quantitative Dynamic Oxygen 17 MRI at 7.0 T for the Cerebral Oxygen Metabolism in Glioma.

Background Altered metabolism is a characteristic of cancer. Because of a shift in glucose metabolism from oxidative phosphorylation to lactate production for energy generation, malignant tumors are characterized by increased glycolysis followed by lactic acid fermentation, even in the presence of abundant oxygen (the Warburg effect). Purpose To quantitatively investigate dynamic oxygen 17 (17O) MRI in healthy participants and participants with untreated glioma to understand altered cerebral oxygen metabolism in glioma. Materials and Methods In this prospective study conducted from September 2016 to June 2018, individuals with newly diagnosed previously untreated glioma (World Health Organization grade II-IV) and healthy volunteers were included. Dynamic 17O MRI was performed with a 7.0-T whole-body system. 17O2 gas inhalation enabled dynamic measurement of the cerebral metabolic rate of oxygen (CMRO2) consumption. In healthy volunteers and participants with glioma, CMRO2 values in gray matter and white matter volumes were compared by using Wilcoxon signed rank tests. In participants with glioma, the tumor volume and tumor subcompartments were compared with normal-appearing gray matter and white matter by using Friedman test followed by Holm-Sidak post hoc tests. Results Ten participants (mean age, 42 years ± 18 [standard deviation]; nine men) with glioma and three healthy volunteers (mean age, 44 years ± 21; all men) were evaluated. CMRO2 was higher in normal-appearing gray matter compared with white matter in both participants with glioma (2.36 μmol/g/min ± 0.22 vs 0.75 μmol/g/min ± 0.10, respectively) and healthy volunteers (2.38 μmol/g/min ± 0.15 vs 0.63 μmol/g/min ± 0.05, respectively) (P < .001 and P = .03, respectively). In the tumor region, CMRO2 was reduced (high-grade tumor CMRO2, 0.23 μmol/g/min ± 0.07; low-grade tumor CMRO2, 0.39 μmol/g/min ± 0.16; overall CMRO2, 0.34 μmol/g/min ± 0.16) compared with normal-appearing gray matter (P < .001) and normal-appearing white matter (P < .001) in accordance with the Warburg theorem. Conclusion Dynamic oxygen 17 MRI method at 7.0 T as a direct metabolic imaging technique in glioma enabled quantitative visualization of the Warburg effect. A general reduction in oxidative glycolysis was observed in accordance with the Warburg theorem. © RSNA, 2020 Online supplemental material is available for this article. See also the editorial by Rapalino in this issue.

18F-FET PET Imaging in Differentiating Glioma Progression from Treatment-Related Changes: A Single-Center Experience.

In glioma patients, differentiation between tumor progression (TP) and treatment-related changes (TRCs) remains challenging. Difficulties in classifying imaging alterations may result in a delay or an unnecessary discontinuation of treatment. PET using O-(2-18F-fluoroethyl)-l-tyrosine (18F-FET) has been shown to be a useful tool for detecting TP and TRCs. Methods: We retrospectively evaluated 127 consecutive patients with World Health Organization grade II-IV glioma who underwent 18F-FET PET imaging to distinguish between TP and TRCs. 18F-FET PET findings were verified by neuropathology (40 patients) or clinicoradiologic follow-up (87 patients). Maximum tumor-to-brain ratios (TBRmax) of 18F-FET uptake and the slope of the time-activity curves (20-50 min after injection) were determined. The diagnostic accuracy of 18F-FET PET parameters was evaluated by receiver-operating-characteristic analysis and χ2 testing. The prognostic value of 18F-FET PET was estimated using the Kaplan-Meier method. Results: TP was diagnosed in 94 patients (74%) and TRCs in 33 (26%). For differentiating TP from TRCs, receiver-operating-characteristic analysis yielded an optimal 18F-FET TBRmax cutoff of 1.95 (sensitivity, 70%; specificity, 71%; accuracy, 70%; area under the curve, 0.75 ± 0.05). The highest accuracy was achieved by a combination of TBRmax and slope (sensitivity, 86%; specificity, 67%; accuracy, 81%). However, accuracy was poorer when tumors harbored isocitrate dehydrogenase (IDH) mutations (91% in IDH-wild-type tumors, 67% in IDH-mutant tumors, P < 0.001). 18F-FET PET results correlated with overall survival (P < 0.001). Conclusion: In our neurooncology department, the diagnostic performance of 18F-FET PET was convincing but slightly inferior to that of previous reports.

Ex-vivo analysis of quantitative 5-ALA fluorescence intensity in diffusely infiltrating gliomas using a handheld spectroscopic probe: Correlation with histopathology, proliferation and microvascular density.

Intraoperative semiquantitative classification of different visible 5-aminolevulinic acid (5-ALA) fluorescence levels by the neurosurgeon is subjective. Recently, handheld spectroscopic probes were introduced enabling quantitative analysis of 5-ALA induced fluorescence intensity (FI). The aim of this ex-vivo study was to correlate the FI in gliomas of different grades with histopathology, proliferation and microvasular density (MVD). Patients with suspected World Health Organization (WHO) grade II-IV gliomas were included and tissue samples from different visible fluorescence levels (strong, vague or none) were intraoperatively collected. After resection, the FI of each sample was investigated ex-vivo by a handheld spectroscopic probe. The FI values were correlated with visible fluorescence, histopathology (WHO grade, quality of tissue, histopathological parameters of anaplasia), proliferation (MIB-1) and MVD. Altogether, 143 tumor samples with strong (n = 61), vague (n = 21) and no fluorescence (n = 61) were collected in 68 patients. We found significantly different median FI values between all three visible fluorescence levels. Moreover, the median FI value was significantly higher in WHO grade III/IV samples and compact tumor tissue compared to WHO grade II samples and infiltrated tumor tissue. Further, significant differences in median FI values were observed in specific histopathological parameters of anaplasia (mitotic rate, cell density, nuclear pleomorphism and microvascular proliferation) in multivariable analysis. Finally, a significant correlation between the proliferation rate and FI, but not between MVD and FI was noted. Our data indicate that handheld spectroscopic probes are capable of visualizing intratumoral glioma heterogeneity by objective assessment of fluorescence and may thus optimize future glioma surgery.

C-Met Expression Is a Useful Marker for Prognosis Prediction in IDH-Mutant Lower-Grade Gliomas and IDH-Wildtype Glioblastomas

c-Met has been shown to be associated with tumor growth in several human cancers. This study aims to evaluate the correlation between the c-Met expression and histopathologic/clinical characteristics. A total of 153 patients with histologically defined World Health Organization grade II-IV diffuse astrocytic and oligodendroglial tumors were analyzed. For each histopathologic diagnosis, the number of cases and positive rate of c-Met expression are as follows: oligodendroglioma, IDH-mutant, and 1p19q codeletion (OD): 16 cases, 6.3%; anaplastic oligodendroglioma, IDH-mutant, and 1p19q codeletion (AO): 11 cases, 36.4%; diffuse astrocytoma (DA), IDH-mutant: 21 cases, 28.6%; anaplastic astrocytoma (AA), IDH- mutant: 15 cases, 20%; glioblastoma, IDH-mutant: 2, 100%, DA, IDH-wildtype: 9 cases, 33.3%; AA, IDH-wildtype: 20 cases, 30.0%; and glioblastoma, IDH-wildtype: 59 cases, 52.5%. c-Met expression was correlated with progression-free survival in oligodendroglial tumors and glioblastoma, IDH-wildtype. Furthermore, it was correlated with overall survival in AO, oligodendroglial tumors, DA, IDH-mutant, DA, IDH-wildtype, and glioblastoma, IDH-wildtype, and tend to be correlated with overall survival in IDH-mutant lower-grade astrocytic tumors. c-Met expression was revealed to be a useful marker for prognosis prediction in IDH-mutant lower-grade gliomas and glioblastoma, IDH-wildtype, representing a new independent prognostic marker that can be easily measured.

ACTB and SDHA Are Suitable Endogenous Reference Genes for Gene Expression Studies in Human Astrocytomas Using Quantitative RT-PCR.

Background:Quantitative real-time reverse-transcription polymerase chain reaction is frequently used as research tool in experimental oncology. There are some studies of valid endogenous control genes in the field of human glioma research, which, however, only focus on the comparison between normal brain with tumor tissue and malignant transformation toward secondary glioblastomas. Aim of this study was to validate a more general reference gene also suitable for pre- and posttreatment analysis and other evaluations (eg, primary vs secondary glioblastoma).Methods:This quantitative polymerase chain reaction analysis was performed to test a panel of the 6 most suitable reference genes from other studies representing different physiological pathways (ACTB, GAPDH, POLR2A, RPL13A, SDHA, and TBP) in all common glioma groups, namely: diffuse astrocytoma World Health Organization II, anaplastic astrocytoma World Health Organization III, secondary glioblastoma World Health Organization IV with and without chemotherapy, primary glioblastoma, recurrent glioblastoma, and gliomas before and after radiation. Expression stability was tested during the longitudinal course of the disease in 8 single patients.Results:Evaluation of the expression levels of the 6 target genes showed that ACTB, GAPDH, and RPL13A show higher expression compared to SDHA, POLR2A, and TBP. ACTB, GAPDH, and RPL13A showed different expression levels between astrozytoma grade II and primary glioblastoma. Except for this difference, the candidate genes were not differentially expressed between primary and secondary glioblastomas and between the World Health Organization tumor grades. Furthermore, they remained stable before and after radiotherapy and/or chemotherapy. Therefore, they are adequate references for glioblastoma gene expression studies. The comparison of all tested genes resulted in SDHA and ACTB as most stable reference genes determined by the NormFinder software. Our data revealed lowest intragroup variation in the SDHA, highest in the RPL13A gene.Conclusions:All tested genes may be recommended as universal reference genes for data normalization in gene expression studies under different treatment regimens both in primary glioblastomas and astrocytomas of different grades (World Health Organization grades II-IV), respectively. In summary, ACTB and SDHA exhibited the best stability values and showed the lowest intergroup expression variability.

Mutant IDH1 and thrombosis in gliomas.

Mutant isocitrate dehydrogenase 1 (IDH1) is common in gliomas, and produces D-2-hydroxyglutarate (D-2-HG). The full effects of IDH1 mutations on glioma biology and tumor microenvironment are unknown. We analyzed a discovery cohort of 169 World Health Organization (WHO) grade II-IV gliomas, followed by a validation cohort of 148 cases, for IDH1 mutations, intratumoral microthrombi, and venous thromboemboli (VTE). 430 gliomas from The Cancer Genome Atlas were analyzed for mRNAs associated with coagulation, and 95 gliomas in a tissue microarray were assessed for tissue factor (TF) protein. In vitro and in vivo assays evaluated platelet aggregation and clotting time in the presence of mutant IDH1 or D-2-HG. VTE occurred in 26-30% of patients with wild-type IDH1 gliomas, but not in patients with mutant IDH1 gliomas (0%). IDH1 mutation status was the most powerful predictive marker for VTE, independent of variables such as GBM diagnosis and prolonged hospital stay. Microthrombi were far less common within mutant IDH1 gliomas regardless of WHO grade (85-90% in wild-type versus 2-6% in mutant), and were an independent predictor of IDH1 wild-type status. Among all 35 coagulation-associated genes, F3 mRNA, encoding TF, showed the strongest inverse relationship with IDH1 mutations. Mutant IDH1 gliomas had F3 gene promoter hypermethylation, with lower TF protein expression. D-2-HG rapidly inhibited platelet aggregation and blood clotting via a novel calcium-dependent, methylation-independent mechanism. Mutant IDH1 glioma engraftment in mice significantly prolonged bleeding time. Our data suggest that mutant IDH1 has potent antithrombotic activity within gliomas and throughout the peripheral circulation. These findings have implications for the pathologic evaluation of gliomas, the effect of altered isocitrate metabolism on tumor microenvironment, and risk assessment of glioma patients for VTE.

Epileptic Activity Increases Cerebral Amino Acid Transport Assessed by 18F-Fluoroethyl-l-Tyrosine Amino Acid PET: A Potential Brain Tumor Mimic.

Ten patients with 11 episodes of serial seizures or status epilepticus, who underwent MRI and 18F-FET PET, were studied. The main diagnosis was glioma World Health Organization grade II-IV (n = 8); 2 patients suffered from nonneoplastic diseases. Immunohistochemical assessment of LAT1/LAT2/CD98 amino acid transporters was performed in seizure-affected cortex (n = 2) and compared with glioma tissues (n = 3). All patients exhibited increased seizure-associated strict gyral 18F-FET uptake, which was reversible in follow-up studies or negative shortly before and without any histologic or clinical signs of tumor recurrence. 18F-FET uptake corresponded to structural MRI changes, compatible with cortical vasogenic and cytotoxic edema, partial contrast enhancement, and hyperperfusion. Patients with prolonged postictal symptoms lasting up to 8 wk displayed intensive and widespread (≥ 1 lobe) cortical 18F-FET uptake. LAT1/LAT2/CD98 was strongly expressed in neurons and endothelium of seizure-affected brains and less in reactive astrocytosis. Seizure activity, in particular status epilepticus, increases cerebral amino acid transport with a strict gyral 18F-FET uptake pattern. Such periictal pseudoprogression represents a potential pitfall of 18F-FET PET and may mimic brain tumor. Our data also indicate a seizure-induced upregulation of neuronal, endothelial, and less astroglial LAT1/LAT2/CD98 amino acid transporter expression.

Symptom resolution in infiltrating WHO grade II-IV glioma patients undergoing surgical resection

Past studies of morbidity in patients with infiltrating gliomas have focused on the impact of surgery on quality of life. Surprisingly, little attention has been given to the rate at which the presenting symptoms improve after surgery, even though this is often the patient’s first concern. This study is an initial effort to provide useful information about symptom resolution and factors predicting persistence of symptoms in glioma patients who undergo surgery. We conducted a retrospective analysis on patients who underwent surgery for World Health Organization (WHO) grade II-IV astrocytoma/oligodendroglioma/oligoastrocytoma at our institution. All patients were seen 2–4months postoperatively, and asked about the persistence of symptoms they experienced preoperatively. Symptoms reported in clinic were assessed against symptoms reported prior to surgery. Our study includes 56 consecutive patients undergoing surgery for gliomas. Of patients who experienced symptoms initially, headache resolved in 18/27 postoperatively, weakness resolved in 8/14 postoperatively, altered mental status resolved in 8/12 postoperatively, vision problems resolved in 7/11 postoperatively, nausea resolved in 5/7 postoperatively, and ataxia resolved in 4/5 postoperatively. Headache was more likely to resolve in patients with frontal or temporal tumors (p=0.02). Preoperative Karnofsky Performance Scale (KPS) of 70 or less was associated with longer postsurgical hospital stay (p<0.01). Younger patients were more likely to experience a resolution of altered mental status (p=0.04). Our analysis provides data regarding the rate at which surgery alleviates patient symptoms and considers variables predicting likelihood of symptom resolution. Some patients will experience symptom resolution following resection of WHO grade II-IV gliomas in the months following surgery.

Phosphohistone H3 (pHH3) is a prognostic and epithelial to mesenchymal transition marker in diffuse gliomas.

The World Health Organization (WHO) grading of gliomas stratifies tumors by histology. However, the aggressiveness of tumors in each grade still shows great heterogeneity. Phosphohistone H3 (pHH3) has been reported as an accurate marker of cells within the mitotic phase of the cell cycle in many kinds of cancers. To evaluate the role of pHH3 in predicting patient outcome and to annotate the functions of pHH3 in WHO grade II-IV gliomas, we analyzed the expression pattern of pHH3 and pHH3 associated genes by IHC and mRNA expression profiling. Phosphohistone H3, mRNA enrichment of histone H3 and associated gene signature all showed prognostic value in adult diffuse gliomas. Gene set enrichment analysis suggested that the expression of pHH3 had positive correlation with both epithelial to mesenchymal transition and immune response. These findings suggest that subgroups of diffuse gliomas defined by pHH3 and pHH3 signatures possess distinctive prognostic and biological characteristics.

TMIC-13MUTANTIDH1/2SUPPRESSES LOCAL AND SYSTEMIC THROMBOSIS IN GLIOMA PATIENTS

Mutations in isocitrate dehydrogenase 1 and 2 (mutIDH1/2) are common in gliomas, resulting in enzymes that produce D-2-hydroxyglutarate (D-2-HG). The effects of mutIDH1/2 on the microenvironment, including thrombosis, are not known. In discovery and validation cohorts of World Health Organization (WHO) grade II-IV gliomas from two institutions (total N = 317), microthrombi were present in 85-90% of wild-type gliomas but in only 2-6% of mutant gliomas (P < 0.0001), and their absence was an independent predictor of mutIDH1/2. Systemic hypercoagulation occurred in 25-30% of patients with wild-type IDH1/2 gliomas, but in 0% of patients with mutIDH1/2 gliomas (P < 0.0001). The prognostic strength of microthrombi was comparable to necrosis and microvascular proliferation, the current WHO criteria for a diagnosis of glioblastoma. In silico analysis of 428 gliomas from The Cancer Genome Atlas (TCGA) revealed that, in the coagulome, the mRNA with the strongest inverse relationship to mutIDH1/2 was F3, encoding tissue factor procoagulant (P = 1.06E-52). MutIDH1/2 gliomas in the TCGA also had significantly elevated F3 promoter methylation (P < 0.0001). In a separate tissue microarray containing 95 gliomas, mutIDH1/2 tumors showed dramatically reduced tissue factor protein expression (P < 0.0001). Additionally, D-2-HG inhibited human platelet aggregation by blocking intracellular calcium accumulation. In human in vitro and mouse in vivo models, mutIDH1 and D-2-HG lowered free serum calcium by up to 40% and markedly impaired coagulation (P < 0.0001). In sum, mutIDH1/2 enzymes have potent antithrombotic activity within gliomas and systemically. These findings have profound implications for the pathologic evaluation of gliomas, the effect of altered isocitrate metabolism on tumor microenvironment and calcium homeostasis, and the pre-and postoperative management of these patients.

PCSM-05ANTIEPILEPTIC DRUG TREATMENT WITH BUCCAL CLONAZEPAM AND INTRANASAL MIDAZOLAM IN THE END OF LIFE PHASE OF GLIOMA PATIENTS: A FEASIBILITY STUDY

BACKGROUND: A rapid deterioration in neurological functioning may hamper the oral administration of antiepileptic drugs (AEDs) in the end of life (EOL) phase of glioma patients. We aimed to assess the feasibility of treatment with non-oral AEDs in an out-of-hospital setting. METHODS: We included patients with a histopathological diagnosis of World Health Organization (WHO) grade II-IV glioma and a history of epilepsy, in whom further antitumor treatment was considered to be no longer meaningful. When swallowing difficulties developed, the patient's caregiver started to administer prophylactic AED treatment with buccal clonazepam. Acute seizures were treated with intranasal midazolam. We evaluated the adherence to the study medication, seizure prevalence as well as caregiver's satisfaction. RESULTS: Thirty-four patients were approached, of which 25 gave consent to participate and 23 had died at the end of the study. Fourteen of 19 patients (73.7%) who had developed swallowing difficulties used the study medication. Prophylactic buccal clonazepam was used by 13 patients, of which 8 patients remained seizure free until death. Six patients received treatment with intranasal midazolam at least once. In all patients seizure control could be reached. None of the patients had to be transferred to the hospital because of recurrent seizures. All caregivers reported satisfaction with the use of the study medication. CONCLUSIONS: It is feasible to treat glioma patients with seizures in the EOL phase with a combination of buccal clonazepam and intranasal midazolam. Additionally, it seems to provide an important level of comfort among caregivers to be able to manage seizures at home.