Tumor mutational burden and immune infiltrates in renal cell carcinoma and matched brain metastases.

BACKGROUND: Tumor mutational burden (TMB) and density of tumor-infiltrating lymphocytes (TIL) have been postulated as predictive biomarkers for immunotherapy. Therefore, we investigated the concordance of TMB and TIL of primary/extracranial renal cell carcinoma (RCC) specimens and matched brain metastases (BM). PATIENTS AND METHODS: Twenty specimens from 10 patients were retrieved from the Vienna Brain Metastasis Registry (6/10 primary tumor, 4/10 lung metastasis, 10/10 matched BM). TMB was assessed using the TruSight Oncology 500 gene panel with libraries sequenced on a NextSeq instrument. TIL subsets (CD3+, CD8+, CD45RO+, FOXP3+, PD-L1+) were investigated using immunohistochemistry (Ventana Benchmark Ultra system) and automated tissue analysis (Definiens software). RESULTS: No significant difference in TMB, CD3+, CD8+, CD45RO+, FOXP3+ or PD-L1+ expression was observed between extracranial and matched intracranial specimens (P > 0.05). Higher CD8+ TIL (P = 0.053) and CD45RO+ TIL (P = 0.030) densities in the primary tumor compared with the intracranial samples were observed in specimens collected after exposure to systemic treatment. Neither extracranial sample origin (lung metastasis versus primary RCC) nor extracranial disease status at BM diagnosis (progressive versus stable disease) were significantly associated with TMB or TIL densities in extracranial and intracranial samples (P > 0.05). No significant correlation was found between the median differences of TMB or TIL densities from extracranial to intracranial samples and BM-free survival. CONCLUSION: The comparable immunological microenvironment of extra- and intracranial tumor samples in our study underscores the immunological activation also in BM from RCC, and therefore, supports the development of immune modulatory treatments also in patients with brain metastatic RCC.

Lesion-Specific Language Network Alterations in Temporal Lobe Epilepsy.

BACKGROUND AND PURPOSE: Temporal lobe epilepsy, structural or nonlesional, may negatively affect language function. However, little is known about the lesion-specific influence on language networks. We hypothesized that different epileptogenic lesions are related to distinct alterations in the functional language connectome detected by fMRI. MATERIALS AND METHODS: One hundred one patients with epilepsy due to mesiotemporal sclerosis (21 left, 22 right), low-grade mesiotemporal tumors (12 left), or nonlesional temporal lobe epilepsy (22 left, 24 right) and 22 healthy subjects performed 3T task-based language fMRI. Task-based activation maps (laterality indices) and functional connectivity analysis (global and connectivity strengths between language areas) were correlated with language scores. RESULTS: Laterality indices based on fMRI activation maps failed to discriminate among patient groups. Functional connectivity analysis revealed the most extended language network alterations in left mesiotemporal sclerosis (involving the left temporal pole, left inferior frontal gyrus, and bilateral premotor areas). The other patient groups showed less extended but also predominantly ipsilesional network changes compared with healthy controls. Left-to-right hippocampal connectivity strength correlated positively with naming function (P = .01), and connectivity strength between the left Wernicke area and the left hippocampus was linked to verbal fluency scores (P = .01) across all groups. CONCLUSIONS: Different pathologies underlying temporal lobe epilepsy are related to distinct alterations of the functional language connectome visualized by fMRI functional connectivity analysis. Network analysis allows new insights into language organization and provides possible imaging biomarkers for language function. These imaging findings emphasize the importance of a personalized treatment strategy in patients with epilepsy.

Applied Precision Cancer Medicine in Neuro-Oncology.

Brain tumours that are refractory to treatment have a poor prognosis and constitute a major challenge in offering effective treatment strategies. By targeting molecular alterations, precision cancer medicine may be a viable option for the treatment of brain tumours. In this retrospective analysis of our PCM platform, we describe the molecular profiling of primary brain tumours from 50 patients. Tumour samples of the patients were examined by a 161-gene next-generation sequencing panel, immunohistochemistry, and fluorescence in situ hybridization (FISH). We identified 103 molecular aberrations in 36 (72%) of the 50 patients. The predominant mutations were TP53 (14.6%), IDH1 (9.7%) and PIK3CA (6.8%). No mutations were detected in 14 (28%) of the 50 patients. IHC demonstrated frequent overexpression of EGFR and mTOR, in 38 (76%) and 35 (70%) patients, respectively. Overexpression of PDGFRa and PDGFRb were less common and detected in 16 and four patients, respectively. For 35 patients a targeted therapy was recommended. In our database, the majority of patients displayed mutations, against which targeted therapy could be offered. Based on our observations, PCM may be a feasible novel treatment approach in neuro-oncology.

Brain metastases as first manifestation of advanced cancer: exploratory analysis of 459 patients at a tertiary care center.

Symptomatic brain metastases (BM) are a frequent and late complication in cancer patients. However, a subgroup of cancer patients presents with BM as the first symptom of metastatic cancer. Here we aimed to analyze the clinical course and prognostic factors of this particular BM patient population. Patients presenting with newly diagnosed BM without a history of metastatic cancer were identified from the Vienna Brain Metastasis Registry. Clinical characteristics and overall survival were retrieved by chart review. 459/2419 (19.0%) BM patients presented with BM as first symptom of advanced cancer. In 374/459 (81.5%) patients, an extracranial primary tumor, most commonly lung cancer, could be identified within 3 months after BM diagnosis. In 85/459 (18.5%) patients no extracranial primary tumor could be identified despite comprehensive diagnostic workup within the first 3 months after diagnosis of BM. Survival of patients with identified extracranial tumor differed only numerically from patients with cancer of unknown primary (CUP), however patients receiving targeted therapy after molecular workup showed significantly enhanced survival (20 months vs. 7 months; p = 0.003; log rank test). The GPA score showed a statistically significant association with median overall survival times in the CUP BM patients (class I: 46 months; class II: 7 months; class III: 4 months; class IV: 2 months; p < 0.001; log rank test). The GPA score has a strong prognostic value in patients with CUP BM and may be useful for patient stratification in the clinical setting. Comprehensive diagnostic workup including advanced imaging techniques and molecular tissue analyses appears to benefit patients by directing specific molecular targeted therapies.

Combination of isocitrate dehydrogenase 1 (IDH1) mutation and podoplanin expression in brain tumors identifies patients at high or low risk of venous thromboembolism.

Essentials Risk stratification for venous thromboembolism (VTE) in patients with brain tumors is challenging. Patients with IDH1 wildtype and high podoplanin expression have a 6-month VTE risk of 18.2%. Patients with IDH1 mutation and no podoplanin expression have a 6-month VTE risk of 0%. IDH1 mutation and podoplanin overexpression in primary brain tumors appear to be exclusive. SUMMARY: Background Venous thromboembolism (VTE) is a frequent complication in primary brain tumor patients. Independent studies revealed that podoplanin expression in brain tumors is associated with increased VTE risk, whereas the isocitrate dehydrogenase 1 (IDH1) mutation is associated with very low VTE risk. Objectives To investigate the interrelation between intratumoral podoplanin expression and IDH1 mutation, and their mutual impact on VTE development. Patients/Methods In a prospective cohort study, intratumoral IDH1 R132H mutation and podoplanin were determined in brain tumor specimens (mainly glioma) by immunohistochemistry. The primary endpoint of the study was symptomatic VTE during a 2-year follow-up. Results All brain tumors that expressed podoplanin to a medium-high extent showed also an IDH1 wild-type status. A score based on IDH1 status and podoplanin expression levels allowed prediction of the risk of VTE. Patients with wild-type IDH1 brain tumors and high podoplanin expression had a significantly increased VTE risk compared with those with mutant IDH1 tumors and no podoplanin expression (6-month risk 18.2% vs. 0%). Conclusions IDH1 mutation and podoplanin overexpression seem to be exclusive. Although brain tumor patients with IDH1 mutation are at very low risk of VTE, the risk of VTE in patients with IDH1 wild-type tumors is strongly linked to podoplanin expression levels.

The physiological phosphorylation of tau is critically changed in fetal brains of individuals with Down syndrome.

AIMS: Down syndrome (DS) is a common cause of mental retardation accompanied by cognitive impairment. Comprehensive studies suggested a link between development and ageing, as nearly all individuals with DS develop Alzheimer disease (AD)-like pathology. However, there is still a paucity of data on tau in early DS to support this notion. METHODS: Using morphometric immunohistochemistry we compared tau phosphorylation in normal brains and in brains of individuals with DS from early development until early postnatal life. RESULTS: We observed in DS a critical loss of physiological phosphorylation of tau. Rhombencephalic structures showed prominent differences between controls and DS using antibodies AT8 (Ser-202/Thr-205) and AT180 (Thr-231). In contrast, in the subiculum only a small portion of controls deviated from DS using antibodies AT100 (Thr-212/Ser-214) and AT270 (Thr-181). With exception of the subiculum, phosphorylation-independent tau did not differ between groups, as confirmed by immunostaining for the HT-7 antibody (epitope between 159 and 163 of the human tau) as well. DISCUSSION: Our observations suggest functional tau disturbance in DS brains during development, rather than axonal loss. This supports the role of tau as a further important player in the pathophysiology of cognitive impairment in DS and related AD.

Dysregulation of the (immuno)proteasome pathway in malformations of cortical development.

BACKGROUND: The proteasome is a multisubunit enzyme complex involved in protein degradation, which is essential for many cellular processes. During inflammation, the constitutive subunits are replaced by their inducible counterparts, resulting in the formation of the immunoproteasome. METHODS: We investigated the expression pattern of constitutive (ß1, ß5) and immunoproteasome (ß1i, ß5i) subunits using immunohistochemistry in malformations of cortical development (MCD; focal cortical dysplasia (FCD) IIa and b, cortical tubers from patients with tuberous sclerosis complex (TSC), and mild MCD (mMCD)). Glial cells in culture were used to elucidate the mechanisms regulating immunoproteasome subunit expression. RESULTS: Increased expression was observed in both FCD II and TSC; ß1, ß1i, ß5, and ß5i were detected (within cytosol and nucleus) in dysmorphic neurons, balloon/giant cells, and reactive astrocytes. Glial and neuronal nuclear expression positively correlated with seizure frequency. Positive correlation was also observed between the glial expression of constitutive and immunoproteasome subunits and IL-1ß. Accordingly, the proteasome subunit expression was modulated by IL-1ß in human astrocytes in vitro. Expression of both constitutive and immunoproteasome subunits in FCD II-derived astroglial cultures was negatively regulated by treatment with the immunomodulatory drug rapamycin (inhibitor of the mammalian target of rapamycin (mTOR) pathway, which is activated in both TSC and FCD II). CONCLUSIONS: These observations support the dysregulation of the proteasome system in both FCD and TSC and provide new insights on the mechanism of regulation the (immuno)proteasome in astrocytes and the molecular links between inflammation, mTOR activation, and epilepsy.

Promoter-Specific Hypomethylation Correlates with IL-1ß Overexpression in Tuberous Sclerosis Complex (TSC).

In tuberous sclerosis complex (TSC), overexpression of numerous genes associated with inflammation has been observed. Among different proinflammatory cytokines, interleukin-1ß (IL-1ß) has been shown to be significantly involved in epileptogenesis and maintenance of seizures. Recent evidence indicates that IL-1ß gene expression can be regulated by DNA methylation of its promoter. In the present study, we hypothesized that hypomethylation in the promoter region of the IL-1ß gene may underlie its overexpression observed in TSC brain tissue. Bisulfite sequencing was used to study the methylation status of the promoter region of the IL-1ß gene in TSC and control samples. We identified hypomethylation in the promoter region of the IL-1ß gene in TSC samples. IL-1ß is overexpressed in tubers, and gene expression is correlated with promoter hypomethylation at CpG and non-CpG sites. Our results provide the first evidence of epigenetic modulation of the IL-1ß signaling in TSC. Thus, strategies that target epigenetic alterations could offer new therapeutic avenues to control the persistent activation of interleukin-1ß-mediated inflammatory signaling in TSC brain.

Prognostic significance of Ki67 proliferation index, HIF1 alpha index and microvascular density in patients with non-small cell lung cancer brain metastases.

BACKGROUND: Survival upon diagnosis of brain metastases (BM) in patients with non-small cell lung cancer (NSCLC) is highly variable and established prognostic scores do not include tissue-based parameters. METHODS: Patients who underwent neurosurgical resection as first-line therapy for newly diagnosed NSCLC BM were included. Microvascular density (MVD), Ki67 tumor cell proliferation index and hypoxia-inducible factor 1 alpha (HIF-1 alpha) index were determined by immunohistochemistry. RESULTS: NSCLC BM specimens from 230 patients (151 male, 79 female; median age 56 years; 199 nonsquamous histology) and 53/230 (23.0%) matched primary tumor samples were available. Adjuvant whole-brain radiation therapy (WBRT) was given to 153/230 (66.5%) patients after neurosurgical resection. MVD and HIF-1 alpha indices were significantly higher in BM than in matched primary tumors. In patients treated with adjuvant WBRT, low BM HIF-1 alpha expression was associated with favorable overall survival (OS), while among patients not treated with adjuvant WBRT, BM HIF-1 alpha expression did not correlate with OS. Low diagnosis-specific graded prognostic assessment score (DS-GPA), low Ki67 index, high MVD, low HIF-1 alpha index and administration of adjuvant WBRT were independently associated with favorable OS. Incorporation of tissue-based parameters into the commonly used DS-GPA allowed refined discrimination of prognostic subgroups. CONCLUSION: Ki67 index, MVD and HIF-1 alpha index have promising prognostic value in BM and should be validated in further studies.

Non-invasive assessment of intratumoral vascularity using arterial spin labeling: A comparison to susceptibility-weighted imaging for the differentiation of primary cerebral lymphoma and glioblastoma.

Using conventional MRI methods, the differentiation of primary cerebral lymphomas (PCNSL) and other primary brain tumors, such as glioblastomas, is difficult due to overlapping imaging characteristics. This study was designed to discriminate tumor entities using normalized vascular intratumoral signal intensity values (nVITS) obtained from pulsed arterial spin labeling (PASL), combined with intratumoral susceptibility signals (ITSS) from susceptibility-weighted imaging (SWI). Thirty consecutive patients with glioblastoma (n=22) and PCNSL (n=8), histologically classified according to the WHO brain tumor classification, were included. MRIs were acquired on a 3T scanner, and included PASL and SWI sequences. nVITS was defined by the signal intensity ratio between the tumor and the contralateral normal brain tissue, as obtained by PASL images. ITSS was determined as intratumoral low signal intensity structures detected on SWI sequences and were divided into four different grades. Potential differences in the nVITS and ITSS between glioblastomas and PCNSLs were revealed using statistical testing. To determine sensitivity, specificity, and diagnostic accuracy, as well as an optimum cut-off value for the differentiation of PCNSL and glioblastoma, a receiver operating characteristic analysis was used. We found that nVITS (p=0.011) and ITSS (p=0.001) values were significantly higher in glioblastoma than in PCNSL. The optimal cut-off value for nVITS was 1.41 and 1.5 for ITSS, with a sensitivity, specificity, and accuracy of more than 95%. These findings indicate that nVITS values have a comparable diagnostic accuracy to ITSS values in differentiating glioblastoma and PCNSL, offering a completely non-invasive and fast assessment of tumoral vascularity in a clinical setting.

Arterial spin-labeling assessment of normalized vascular intratumoral signal intensity as a predictor of histologic grade of astrocytic neoplasms.

BACKGROUND AND PURPOSE: Pulsed arterial spin-labeling is a noninvasive MR imaging perfusion method performed with the use of water in the arterial blood as an endogenous contrast agent. The purpose of this study was to determine the inversion time with the largest difference in normalized intratumoral signal intensity between high-grade and low-grade astrocytomas. MATERIALS AND METHODS: Thirty-three patients with gliomas, histologically classified as low-grade (n = 7) or high-grade astrocytomas (n = 26) according to the World Health Organization brain tumor classification, were included. A 3T MR scanner was used to perform pulsed arterial spin-labeling measurements at 8 different inversion times (370 ms, 614 ms, 864 ms, 1114 ms, 1364 ms, 1614 ms, 1864 ms, and 2114 ms). Normalized intratumoral signal intensity was calculated, which was defined by the signal intensity ratio of the tumor and the contralateral normal brain tissue for all fixed inversion times. A 3-way mixed ANOVA was used to reveal potential differences in the normalized vascular intratumoral signal intensity between high-grade and low-grade astrocytomas. RESULTS: The difference in normalized vascular intratumoral signal intensity between high-grade and low-grade astrocytomas obtained the most statistically significant results at 370 ms (P = .003, other P values ranged from .012-.955). CONCLUSIONS: The inversion time by which to differentiate high-grade and low-grade astrocytomas by use of normalized vascular intratumoral signal intensity was 370 ms in our study. The normalized vascular intratumoral signal intensity values at this inversion time mainly reflect the labeled intra-arterial blood bolus and therefore could be referred to as normalized vascular intratumoral signal intensity. Our data indicate that the use of normalized vascular intratumoral signal intensity values allows differentiation between low-grade and high-grade astrocytomas and thus may serve as a new, noninvasive marker for astrocytoma grading.

Relative survival of patients with non-malignant central nervous system tumours: a descriptive study by the Austrian Brain Tumour Registry.

BACKGROUND: Unlike malignant primary central nervous system (CNS) tumours outcome data on non-malignant CNS tumours are scarce. For patients diagnosed from 1996 to 2002 5-year relative survival of only 85.0% has been reported. We investigated this rate in a contemporary patient cohort to update information on survival. METHODS: We followed a cohort of 3983 cases within the Austrian Brain Tumour Registry. All patients were newly diagnosed from 2005 to 2010 with a histologically confirmed non-malignant CNS tumour. Vital status, cause of death, and population life tables were obtained by 31 December 2011 to calculate relative survival. RESULTS: Overall 5-year relative survival was 96.1% (95% CI 95.1-97.1%), being significantly lower in tumours of borderline (90.2%, 87.2-92.7%) than benign behaviour (97.4%, 96.3-98.3%). Benign tumour survival ranged from 86.8 for neurofibroma to 99.7% for Schwannoma; for borderline tumours survival rates varied from 83.2 for haemangiopericytoma to 98.4% for myxopapillary ependymoma. Cause of death was directly attributed to the CNS tumour in 39.6%, followed by other cancer (20.4%) and cardiovascular disease (15.8%). CONCLUSION: The overall excess mortality in patients with non-malignant CNS tumours is 5.5%, indicating a significant improvement in survival over the last decade. Still, the remaining adverse impact on survival underpins the importance of systematic registration of these tumours.

[Rapid frozen sections in neuropathology]. / Der Schnellschnitt in der Neuropathologie.

Neuropathological evaluation of frozen sections requires a) special expertise in neuropathological specimen assessment and neurooncology as well as b) a trustful and open communication culture with the neurosurgeons. In addition to frozen sections, cytological examinations of smear and touch preparations as supporting methods are available to reach a correct diagnosis: these additional methods should therefore be performed whenever possible. Besides evaluation of biopsy specimens, appraisal of resection specimens and resection margin controls are of high clinical relevance. In the case of diffusely infiltrating central nervous system (CNS) neoplasms, in particular gliomas, resection margin control is often not feasible in contrast to other types of solid tumor.

Kynurenic Acid Metabolism in Various Types of Brain Pathology in HIV-1 Infected Patients.

Kynurenic acid, an intermediate metabolite of L-kynurenine, is a competitive antagonist of inotropic excitatory amino acid (EAA) receptors as well as a non competitive antagonist of 7 alpha nicotine cholinergic receptors and its involvement in memory deficit and cognition impairment has been suggested. Alterations of kynurenic acid metabolism in the brain after HIV-1 (human immunodeficiency virus type-1) infection have been demonstrated. The present study evaluates the biosynthetic machinery of kynurenic acid e.g. the content of L-kynurenine and kynurenic acid, as well as the activity of enzymes synthesizing kynurenic acid, kynurenine aminotransferase I (KAT I) and kynurenine aminotransferase II (KAT II) in the frontal cortex and cerebellum of HIV-1 infected patients in relation to different types of pathology classified as follows: HIV in brain (HIV); opportunistic infection (OPP); infarction of brain (INF); malignant lymphoma of brain (LY); and glial dystrophy (GD) and of control (CO) subjects. Of all investigated pathologies the most frequent was OPP (65%), followed by HIV (26%), LY, INF, and GD (each 22%, respectively). Further, 68% of HIV-1 patients had bronchopneumonia, the highest incidence of which, at 60%, was seen in the OPP and LY group. Kynurenic acid was increased significantly in the frontal cortex of LY (392% of CO, P < 0.001), HIV (231% of CO, P < 0.01) and GD (193% of CO, P < 0.05), as well as in the cerebellum of GD (261% of CO, P < 0.01). A significant increase of L-kynurenine was observed in the frontal cortex of LY (385% of CO, P < 0.001) and INF (206% of CO, P < 0.01), and in the cerebellum of GD, LY, OPP and HIV (between 177% and 147% of CO). The KAT I activity increased significantly in the frontal cortex of all pathological subgroups, ie OPP = 420% > INF > LY > HIV > GD = 192% of CO. In the cerebellum, too, all pathological subgroups showed marked increase of KAT I activity (OPP = 320% > LY, HIV > GD > INF = 176% of CO). On contrary, the activity of KAT II was moderately, but significantly, higher in the frontal cortex of INF and OPP; in the cerebellum of HIV, OPP and LY it was comparable to the control, while mildly reduced in INF and GD. Interestingly, normal subjects with the diagnosis of bronchopneumonia were characterized by high kynurenic acid metabolism in the brain, too. Correlation analyses between kynurenine parameters revealed association between high ratio KAT I/KAT II and increased kynurenic acid level and lower L-kynurenine in the frontal cortex and cerebellum of HIV and LY subgroups. The present study revealed a different pattern of alteration of kynurenic acid metabolism in frontal cortex and cerebellum among investigated pathological subgroups of HIV-1 infected patients. Interestingly, a marked enhancement of kynurenic acid metabolism in the brain has been found with occurrence of bronchopneumonia. This finding indicates a notable association between impaired conditions of oxygen availability and enhancement of kynurenic acid formation in the human brain. These observation(s) might have an impact on the understanding of pathological processes in the brain after HIV-1 infection involving the development of neuropsychiatric and neurological symptoms, including memory and cognition impairment.

FISH-based detection of 1p 19q codeletion in oligodendroglial tumors: procedures and protocols for neuropathological practice - a publication under the auspices of the Research Committee of the European Confederation of Neuropathological Societies (Euro-CNS).

The codeletion of chromosomal arms 1p 19q is a characteristic and early genetic event in oligodendroglial tumors, that is associated with a better prognosis and enhanced response to therapy. Over the last years, the increasing clinical demand to determine the 1p 19q status has led to the implementation of its testing in many neuropathology laboratories. Several different methods for 1p 19q testing are available: PCR-based loss of heterozygosity analysis, multiplex ligation-dependent probe amplification, array comparative genomic hybridization, and fluorescence in situ hybridization (FISH). Herein, we focus and critically discuss the latter method because a detailed description of procedures and protocols for FISH-based 1p 19q testing in practice is lacking. We present a practical approach to the FISH-based assessment of the 1p 19q status in oligodendroglial tumors, from commonly used locus-specific probes and technical protocols to the neuropathological interpretation of results. Thereby, we aim to facilitate the implementation of FISH-based 1p 19q testing for clinical purposes in standard neuropathology laboratories without special focus on brain tumor research.