Clinical Validation of Stimulated Raman Histology for Rapid Intraoperative Diagnosis of Central Nervous System Tumors.

Stimulated Raman histology (SRH) is an ex vivo optical imaging method that enables microscopic examination of fresh tissue intraoperatively. The conventional intraoperative method uses frozen section analysis, which is labor and time intensive, introduces artifacts that limit diagnostic accuracy, and consumes tissue. SRH imaging allows rapid microscopic imaging of fresh tissue, avoids tissue loss, and enables remote telepathology review. This improves access to expert neuropathology consultation in both low- and high-resource practices. We clinically validated SRH by performing a blinded, retrospective two-arm telepathology study to clinically validate SRH for telepathology at our institution. Using surgical specimens from 47 subjects, we generated a data set composed of 47 SRH images and 47 matched whole slide images (WSIs) of formalin-fixed, paraffin-embedded tissue stained with hematoxylin and eosin, with associated intraoperative clinicoradiologic information and structured diagnostic questions. We compared diagnostic concordance between WSI and SRH-rendered diagnoses. Also, we compared the 1-year median turnaround time (TAT) of intraoperative conventional neuropathology frozen sections with prospectively rendered SRH-telepathology TAT. All SRH images were of sufficient quality for diagnostic review. A review of SRH images showed high accuracy in distinguishing glial from nonglial tumors (96.5% SRH vs 98% WSIs) and predicting final diagnosis (85.9% SRH vs 93.1% WSIs). SRH-based diagnosis and WSI-permanent section diagnosis had high concordance (κ = 0.76). The median TAT for prospectively SRH-rendered diagnosis was 3.7 minutes, approximately 10-fold shorter than the median frozen section TAT (31 minutes). The SRH-imaging procedure did not affect ancillary studies. SRH generates diagnostic virtual histologic images with accuracy comparable to conventional hematoxylin and eosin-based methods in a rapid manner. Our study represents the largest and most rigorous clinical validation of SRH to date. It supports the feasibility of implementing SRH as a rapid method for intraoperative diagnosis complementary to conventional pathology laboratory methods.

GNA11 Mutation in an Intracranial Melanocytoma with Orbital Involvement and Nevus of Ota.

The prognostic value of mutations in G-protein genes GNAQ and GNA11 in patients with intracranial and orbital melanocytomas is unknown. The authors present a case of GNA11 mutation (GNA11Q209L) in a 32-year-old male suffering from a meningeal melanocytoma with orbital involvement and ipsilateral Nevus of Ota. The patient underwent gamma knife stereotactic radiosurgery without biopsy and later partial transcranial resection of the melanocytic tumor that was subject to immunohistochemical and molecular analysis. A 50-gene next-generation sequencing panel revealed a 626A>T mutation in the GNA11 gene. One year later, intracranial extension of the melanocytoma necessitated a ventriculoperitoneal shunt and immunotherapy. Future work is needed to determine how GNA11 mutations in melanocytomas influence prognosis and monitoring strategies.

Pediatric midline H3K27M-mutant tumor with disseminated leptomeningeal disease and glioneuronal features: case report and literature review.

BACKGROUND: H3K27M-mutant midline lesions were recently reclassified by the World Health Organization (WHO) as "diffuse midline glioma" (DMG) based entirely on their molecular signature. DMG is one of the most common and most lethal pediatric brain tumors; terminal progression is typically caused by local midbrain or brainstem progression, or secondary leptomeningeal dissemination. H3K27M mutations have also been infrequently associated with a histologically and prognostically diverse set of lesions, particularly spinal masses with early leptomeningeal spread. CASE PRESENTATION: A 15-year-old girl after 1 week of symptoms was found to have a T2/FLAIR-hyperintense and contrast-enhancing thalamic mass accompanied by leptomeningeal enhancement along the entire neuraxis. Initial infectious workup was negative, and intracranial biopsy was inconclusive. Spinal arachnoid biopsy revealed an H3K27M-mutant lesion with glioneuronal features, classified thereafter as DMG. She received craniospinal irradiation with a boost to the thalamic lesion. Imaging 1-month post-radiation demonstrated significant treatment response with residual enhancement at the conus. CONCLUSIONS: This case report describes the unique presentation of an H3K27M-mutant midline lesion with significant craniospinal leptomeningeal spread on admission and atypical glioneuronal histopathological markers. With such florid leptomeningeal disease, spinal dural biopsy should be considered earlier given its diagnostic yield in classifying the lesion as DMG. Consistent with similar prior reports, this lesion additionally demonstrated synaptophysin positivity-also potentially consistent with a diagnosis of diffuse leptomeningeal glioneuronal tumor (DLGNT). In atypical DMG cases, particularly with leptomeningeal spread, further consideration of clinical and histopathological context is necessary for accurate diagnosis and prognostication.

Carotid web: an occult mechanism of embolic stroke.

The carotid web is a proposed stroke mechanism that may underlie cryptogenic stroke, particularly in younger patients without vascular risk factors. The web appears as a shelf-like projection into the lumen of the proximal cervical internal carotid artery without evidence of calcification. It is pathologically defined as intimal fibromuscular dysplasia. Altered haemodynamics distal to the web cause flow stagnation and remote embolisation of fibrin-based clots. It is best demonstrated and diagnosed on CT angiography (CTA) of the neck because of its ability to resolve calcium and create multiplanar reconstructions. Although they can be readily visualised on CTA, carotid webs may be missed or misinterpreted because they do not typically cause haemodynamically significant stenosis and can mimic arterial dissection, non-calcified atherosclerotic plaque and intraluminal thrombus. Options for management include antiplatelet therapy, carotid endarterectomy and carotid artery stenting. Modern management strategies for cryptogenic stroke include long-term cardiac monitoring, further investigation for structural cardiac disease and a diagnostic workup for arterial hypercoagulability, however, these strategies are not likely to capture the possibility of a carotid web. Carotid webs should be suspected in a young patient presenting with recurrent unihemispheric strokes particularly when conventional vascular risk factors are not present.

Hemorrhagic stroke and anticoagulation in COVID-19.

BACKGROUND AND PURPOSE: Patients with the Coronavirus Disease of 2019 (COVID-19) are at increased risk for thrombotic events and mortality. Various anticoagulation regimens are now being considered for these patients. Anticoagulation is known to increase the risk for adverse bleeding events, of which intracranial hemorrhage (ICH) is one of the most feared. We present a retrospective study of 33 patients positive for COVID-19 with neuroimaging-documented ICH and examine anticoagulation use in this population. METHODS: Patients over the age of 18 with confirmed COVID-19 and radiographic evidence of ICH were included in this study. Evidence of hemorrhage was confirmed and categorized by a fellowship trained neuroradiologist. Electronic health records were analyzed for patient information including demographic data, medical history, hospital course, laboratory values, and medications. RESULTS: We identified 33 COVID-19 positive patients with ICH, mean age 61.6 years (range 37-83 years), 21.2% of whom were female. Parenchymal hemorrhages with mass effect and herniation occurred in 5 (15.2%) patients, with a 100% mortality rate. Of the remaining 28 patients with ICH, 7 (25%) had punctate hemorrhages, 17 (60.7%) had small- moderate size hemorrhages, and 4 (14.3%) had a large single site of hemorrhage without evidence of herniation. Almost all patients received either therapeutic dose anticoagulation (in 22 [66.7%] patients) or prophylactic dose (in 3 [9.1] patients) prior to ICH discovery. CONCLUSIONS: Anticoagulation therapy may be considered in patients with COVID-19 though the risk of ICH should be taken into account when developing a treatment regimen.

Somatic SLC35A2 variants in the brain are associated with intractable neocortical epilepsy.

OBJECTIVE: Somatic variants are a recognized cause of epilepsy-associated focal malformations of cortical development (MCD). We hypothesized that somatic variants may underlie a wider range of focal epilepsy, including nonlesional focal epilepsy (NLFE). Through genetic analysis of brain tissue, we evaluated the role of somatic variation in focal epilepsy with and without MCD. METHODS: We identified somatic variants through high-depth exome and ultra-high-depth candidate gene sequencing of DNA from epilepsy surgery specimens and leukocytes from 18 individuals with NLFE and 38 with focal MCD. RESULTS: We observed somatic variants in 5 cases in SLC35A2, a gene associated with glycosylation defects and rare X-linked epileptic encephalopathies. Nonsynonymous variants in SLC35A2 were detected in resected brain, and absent from leukocytes, in 3 of 18 individuals (17%) with NLFE, 1 female and 2 males, with variant allele frequencies (VAFs) in brain-derived DNA of 2 to 14%. Pathologic evaluation revealed focal cortical dysplasia type Ia (FCD1a) in 2 of the 3 NLFE cases. In the MCD cohort, nonsynonymous variants in SCL35A2 were detected in the brains of 2 males with intractable epilepsy, developmental delay, and magnetic resonance imaging suggesting FCD, with VAFs of 19 to 53%; Evidence for FCD was not observed in either brain tissue specimen. INTERPRETATION: We report somatic variants in SLC35A2 as an explanation for a substantial fraction of NLFE, a largely unexplained condition, as well as focal MCD, previously shown to result from somatic mutation but until now only in PI3K-AKT-mTOR pathway genes. Collectively, our findings suggest a larger role than previously recognized for glycosylation defects in the intractable epilepsies. Ann Neurol 2018.

MR imaging phenotype correlates with extent of genome-wide copy number abundance in IDH mutant gliomas.

PURPOSE: There is variability in survival within IDH mutant gliomas determined by chromosomal events. Copy number variation (CNV) abundance associated with survival in low-grade and IDH mutant astrocytoma has been reported. Our purpose was to correlate the extent of genome-wide CNV abundance in IDH mutant astrocytomas with MRI features. METHODS: Presurgical MRI and CNV plots derived from Illumina 850k EPIC DNA methylation arrays of 18 cases of WHO grade II-IV IDH mutant astrocytomas were reviewed. IDH mutant astrocytomas were divided into CNV stable group (CNV-S) with ≤ 3 chromosomal gains or losses and lack of focal gene amplifications and CNV unstable group (CNV-U) with > 3 large chromosomal gains/losses and/or focal amplifications. The associations between MR features, relative cerebral blood volume (rCBV), CNV abundance, and time to progression were assessed. Tumor rCBV estimates were obtained using DSC T2* perfusion analysis. RESULTS: There were nine (50%) CNV-S and nine (50%) CNV-U IDH mutant astrocytomas. CNV-U tumors showed larger mean tumor size (P = 0.004) and maximum diameter on FLAIR (P = 0.004) and also demonstrated significantly higher median rCBV than CNV-S tumors (2.62 vs 0.78, P = 0.019). CNV-U tumors tended to have shorter time to progression although without statistical significance (P = 0.393). CONCLUSIONS: Larger size/diameter and higher rCBVs were seen associated CNV-U astrocytomas, suggesting a correlation of aggressive imaging phenotype with unstable and aggressive genotype in IDH mutant astrocytomas.

Hypoxia-Mediated Mechanisms Associated with Antiangiogenic Treatment Resistance in Glioblastomas.

Glioblastomas (GBMs) are malignant tumors characterized by their vascularity and invasive capabilities. Antiangiogenic therapy (AAT) is a treatment option that targets GBM-associated vasculature to mitigate the growth of GBMs. However, AAT demonstrates transient effects because many patients eventually develop resistance to this treatment. Several recent studies attempt to explain the molecular and biochemical basis of resistance to AAT in GBM patients. Experimental investigations suggest that the induction of extensive intratumoral hypoxia plays a key role in GBM escape from AAT. In this review, we examine AAT resistance in GBMs, with an emphasis on six potential hypoxia-mediated mechanisms: enhanced invasion and migration, including increased expression of matrix metalloproteinases and activation of the c-MET tyrosine kinase pathway; shifts in cellular metabolism, including up-regulation of hypoxia inducible factor-1α's downstream processes and the Warburg effect; induction of autophagy; augmentation of GBM stem cell self-renewal; possible implications of GBM-endothelial cell transdifferentiation; and vasoformative responses, including vasculogenesis, alternative angiogenic pathways, and vascular mimicry. Juxtaposing recent studies on well-established resistance pathways with that of emerging mechanisms highlights the overall complexity of GBM treatment resistance while also providing direction for further investigation.

Multifaceted C-X-C Chemokine Receptor 4 (CXCR4) Inhibition Interferes with Anti-Vascular Endothelial Growth Factor Therapy-Induced Glioma Dissemination.

Resistance to antiangiogenic therapy in glioblastoma (GBM) patients may involve hypoxia-induced expression of C-X-C motif chemokine receptor 4 (CXCR4) on invading tumor cells, macrophage/microglial cells (MGCs), and glioma stem cells (GSCs). We determined whether antagonizing CXCR4 with POL5551 disrupts anti-vascular endothelial growth factor (VEGF) therapy-induced glioma growth and dissemination. Mice bearing orthotopic CT-2A or GL261 gliomas received POL5551 and/or anti-VEGF antibody B20-4.1.1. Brain tissue was analyzed for tumor volume, invasiveness, hypoxia, vascular density, proliferation, apoptosis, GSCs, and MGCs. Glioma cells were evaluated for CXCR4 expression and polymorphism and POL5551's effects on CXCR4 ligand binding, cell viability, and migration. No CXCR4 mutations were identified. POL5551 inhibited CXCR4 binding to its ligand, stromal cell-derived factor-1α, and reduced hypoxia- and stromal cell-derived factor-1α-mediated migration dose-dependently but minimally affected cell viability. In vivo, B20-4.1.1 increased hypoxic foci and invasiveness, as seen in GBM patients receiving anti-VEGF therapy. Combination of POL5551 and B20-4.1.1 reduced both glioma invasiveness by 16% to 39% and vascular density compared to B20-4.1.1 alone in both glioma models. Reduced populations of GSCs and MGCs were also seen in CT-2A tumors. POL5551 concentrations, evaluated by mass spectrometry, were higher in tumors than in neighboring brain tissues, likely accounting for the results. Inhibition of CXCR4-regulated tumoral, stem cell, and immune mechanisms by adjunctive CXCR4 antagonists may help overcome antiangiogenic therapy resistance, benefiting GBM patients.

Endothelium-Independent Primitive Myxoid Vascularization Creates Invertebrate-Like Channels to Maintain Blood Supply in Optic Gliomas.

Optic gliomas are brain tumors characterized by slow growth, progressive loss of vision, and limited therapeutic options. Optic gliomas contain various amounts of myxoid matrix, which can represent most of the tumor mass. We sought to investigate biological function and protein structure of the myxoid matrix in optic gliomas to identify novel therapeutic targets. We reviewed histological features and clinical imaging properties, analyzed vasculature by immunohistochemistry and electron microscopy, and performed liquid chromatography-mass spectrometry on optic gliomas, which varied in the amount of myxoid matrix. We found that although subtypes of optic gliomas are indistinguishable on imaging, the microvascular network of pilomyxoid astrocytoma, a subtype of optic glioma with abundant myxoid matrix, is characterized by the presence of endothelium-free channels in the myxoid matrix. These tumors show normal perfusion by clinical imaging and lack histological evidence of hemorrhage organization or thrombosis. The myxoid matrix is composed predominantly of the proteoglycan versican and its linking protein, a vertebrate hyaluronan and proteoglycan link protein 1. We propose that pediatric optic gliomas can maintain blood supply without endothelial cells by using invertebrate-like channels, which we termed primitive myxoid vascularization. Enzymatic targeting of the proteoglycan versican/hyaluronan and proteoglycan link protein 1 rich myxoid matrix, which is in direct contact with circulating blood, can provide novel therapeutic avenues for optic gliomas of childhood.

Loss of histone H3K27me3 identifies a subset of meningiomas with increased risk of recurrence.

Epigenetic patterns on the level of DNA methylation have already been shown to separate clinically relevant subgroups of meningiomas. We here set out to identify potential prognostic implications of epigenetic modification on the level of histones with focus on H3K27 trimethylation (H3K27me3). H3K27me3 was assessed by immunohistochemistry on 232 meningiomas from 232 patients. In 194 cases, trimethylation was detected in tumor cells. In 25 cases, staining was limited to vessels while all tumor cells were negative. Finally, 13 cases yielded equivocal staining patterns. Reduced abundance of H3K27me3 in cases with staining limited to vessels was confirmed by mass spectrometry on a subset of cases. Lack of staining for H3K27me3 in all tumor cells was significantly associated with more rapid progression (p = 0.009). In line, H3K27me3-negative cases were associated with a DNA methylation pattern of the more aggressive types among the recently introduced DNA methylation groups. Also, NF2 and SUFU mutations were enriched among cases with complete lack of H3K27me3 staining in tumor cells (p < 0.0001 and p = 0.029, respectively). H3K27me3 staining pattern added significant prognostic insight into WHO grade II cases and in the compound subset of WHO grade I and II cases (p = 0.04 and p = 0.007, respectively). However, it did not further stratify within WHO grade III cases. Collectively, these data indicate that epigenetic modifications beyond DNA methylation are involved in the aggressiveness of meningioma. It also suggests that H3K27me3 immunohistochemistry might be a useful adjunct in meningioma diagnostics, particularly for cases with WHO grade II histology or at the borderline between WHO grade I and II.

Clonal selection drives genetic divergence of metastatic medulloblastoma.

Medulloblastoma, the most common malignant paediatric brain tumour, arises in the cerebellum and disseminates through the cerebrospinal fluid in the leptomeningeal space to coat the brain and spinal cord. Dissemination, a marker of poor prognosis, is found in up to 40% of children at diagnosis and in most children at the time of recurrence. Affected children therefore are treated with radiation to the entire developing brain and spinal cord, followed by high-dose chemotherapy, with the ensuing deleterious effects on the developing nervous system. The mechanisms of dissemination through the cerebrospinal fluid are poorly studied, and medulloblastoma metastases have been assumed to be biologically similar to the primary tumour. Here we show that in both mouse and human medulloblastoma, the metastases from an individual are extremely similar to each other but are divergent from the matched primary tumour. Clonal genetic events in the metastases can be demonstrated in a restricted subclone of the primary tumour, suggesting that only rare cells within the primary tumour have the ability to metastasize. Failure to account for the bicompartmental nature of metastatic medulloblastoma could be a major barrier to the development of effective targeted therapies.

Unique findings of subependymal giant cell astrocytoma within cortical tubers in patients with tuberous sclerosis complex: a histopathological evaluation.

INTRODUCTION: Tuberous sclerosis is associated with three central nervous system pathologies: cortical/subcortical tubers, subependymal nodules (SENs), and subependymal giant cell astrocytomas (SEGAs). Tubers are associated with epilepsy, which is often medication-resistant and often leads to resective surgery. Recently, mammalian target of rapamycin inhibitors (mTORi) have been shown to be effective reducing seizure burden in some patients with tuberous sclerosis complex (TSC)-related refractory epilepsy. mTORi have also been shown to be an alternative for surgery treating SEGAs. We describe several cases of resected tubers that contained SEGA tissue without an intraventricular SEGA. METHODS: After institutional review board (IRB) protocol approval, we retrospectively reviewed the surgical-pathological data for all TSC patients who underwent cortical resections for treatment of refractory epilepsy at NYU Langone Medical Center and Tel Aviv Medical Center between 2003 and 2013. Data included demographics, epilepsy type, MRI characteristics, epilepsy outcome, and histopathological staining. RESULTS: We reviewed cortical resections from 75 patients with complete pathological studies. In three patients, cortical lesions demonstrated histopathological findings consistent with a SEGA within the resected tuber tissue, with no intraventricular SEGA. All lesions were cortically based and none had any intraventricular extension. No patient had been treated before surgery with an mTORi. Two of the three patients remain Engel grade I-II. All lesions stained positive for glial fibrillary acidic protein (GFAP), synaptophysin, and neuronal nuclear antigen (NeuN). CONCLUSION: This is the first description of cortical tubers harboring SEGA tissue. This observation though preliminary may suggest a subgroup of patients with intractable epilepsy in whom mTORi may be considered before surgical intervention.

Tumoral Presentation of Homonymous Hemianopia and Prosopagnosia in Cerebral Amyloid Angiopathy-Related Inflammation.

While cerebral amyloid angiopathy is a common cause of lobar hemorrhage, rarely it may be associated with an inflammatory response, thought to be incited by amyloid deposits. We report a 73-year-old woman with an extensive cancer history who presented with tumor-like lesions and symptoms of homonymous hemianopia and prosopagnosia. Found to have cerebral amyloid angiopathy-related inflammation proven by brain biopsy, she was treated successfully with immunosuppression.

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.

Utility of MRI versus tumor markers for post-treatment surveillance of marker-positive CNS germ cell tumors.

Patients with marker-positive central nervous system (CNS) germ cell tumors are typically monitored for tumor recurrence with both tumor markers (AFP and b-hCG) and MRI. We hypothesize that the recurrence of these tumors will always be accompanied by an elevation in tumor markers, and that surveillance MRI may not be necessary. We retrospectively identified 28 patients with CNS germ cell tumors treated at our institution that presented with an elevated serum or cerebrospinal fluid (CSF) tumor marker at the time of diagnosis. We then identified those who had a tumor recurrence after having been in remission and whether each recurrence was detected via MRI changes, elevated tumor markers, or both. Four patients suffered a tumor recurrence. Only one patient had simultaneously elevated tumor markers and MRI evidence of recurrence. Two patients had evidence of recurrence on MRI without corresponding elevations in serum or CSF tumor markers. One patient had abnormal tumor markers with no evidence of recurrence on MRI until 6 months later. We conclude that in patients with marker-positive CNS germ cell tumors who achieve complete remission, continued surveillance imaging in addition to measurement of tumor markers is indicated to detect recurrences.

Medulloblastoma subgroups remain stable across primary and metastatic compartments.

Medulloblastoma comprises four distinct molecular variants with distinct genetics, transcriptomes, and outcomes. Subgroup affiliation has been previously shown to remain stable at the time of recurrence, which likely reflects their distinct cells of origin. However, a therapeutically relevant question that remains unanswered is subgroup stability in the metastatic compartment. We assembled a cohort of 12-paired primary-metastatic tumors collected in the MAGIC consortium, and established their molecular subgroup affiliation by performing integrative gene expression and DNA methylation analysis. Frozen tissues were collected and profiled using Affymetrix gene expression arrays and Illumina methylation arrays. Class prediction and hierarchical clustering were performed using existing published datasets. Our molecular analysis, using consensus integrative genomic data, establishes the unequivocal maintenance of molecular subgroup affiliation in metastatic medulloblastoma. We further validated these findings by interrogating a non-overlapping cohort of 19 pairs of primary-metastatic tumors from the Burdenko Neurosurgical Institute using an orthogonal technique of immunohistochemical staining. This investigation represents the largest reported primary-metastatic paired cohort profiled to date and provides a unique opportunity to evaluate subgroup-specific molecular aberrations within the metastatic compartment. Our findings further support the hypothesis that medulloblastoma subgroups arise from distinct cells of origin, which are carried forward from ontogeny to oncology.

Prognostic marker analysis in pediatric intracranial ependymomas.

Histologic grading methods dependent upon H&E staining review have not been shown to reliably predict survival in children with intracranial ependymomas due to the subjectivity of the analytical methods. We hypothesized that the immunohistochemical detection of MIB-1, Tenascin C, CD34, VEGF, and CA IX may represent objective markers of post-operative survival (Progression Free and Overall Survival; PFS, OS) in these patients. Intracranial ependymomas from patients aged 22 years or less were studied. The original histologic grade was recorded, H&E sections were reviewed for vascular proliferation status, and immunohistochemistry was used to determine MIB-1, Tenascin C, CD34, VEGF, and CA IX status. Based upon the World Health Organization (WHO) grading system, 3 Grade I, 18 Grade II and 9 Grade III ependymomas were studied. Median follow-up time was 9.0 years; median PFS was, 6.1 years. Original WHO grade did not correlate with PFS or OS. Peri-necrotic CA IX localization correlated with PFS (Log rank = 0.0181) and OS (Log rank p = 0.0015). All patients with a CA IX ≤ 5 % total area localization were alive at last follow-up. Perinecrotic CA IX staining was also associated with vascular proliferation (p = 0.006), though not with VEGF expression score. MIB-1 labeling index (LI) correlated with OS (HR 1.06, 95 % CI 1.01, 1.12) and PFS (HR 1.08, 95 % CI 1.02, 1.14). MIB-1 LI and perinecrotic CA IX individually correlated with PFS. The effect of perinecrotic CA IX remained when grade was added to a Cox model predicting PFS. Immunodetection of CA IX and MIB-1 expression are predictive biomarkers for survival in children with posterior fossa ependymomas. These markers represent objective indicators of survival that supplement H&E grading alone.

CCR7 signalling as an essential regulator of CNS infiltration in T-cell leukaemia.

T-cell acute lymphoblastic leukaemia (T-ALL) is a blood malignancy afflicting mainly children and adolescents. T-ALL patients present at diagnosis with increased white cell counts and hepatosplenomegaly, and are at an increased risk of central nervous system (CNS) relapse. For that reason, T-ALL patients usually receive cranial irradiation in addition to intensified intrathecal chemotherapy. The marked increase in survival is thought to be worth the considerable side-effects associated with this therapy. Such complications include secondary tumours, neurocognitive deficits, endocrine disorders and growth impairment. Little is known about the mechanism of leukaemic cell infiltration of the CNS, despite its clinical importance. Here we show, using T-ALL animal modelling and gene-expression profiling, that the chemokine receptor CCR7 (ref. 5) is the essential adhesion signal required for the targeting of leukaemic T-cells into the CNS. Ccr7 gene expression is controlled by the activity of the T-ALL oncogene Notch1 and is expressed in human tumours carrying Notch1-activating mutations. Silencing of either CCR7 or its chemokine ligand CCL19 (ref. 6) in an animal model of T-ALL specifically inhibits CNS infiltration. Furthermore, murine CNS-targeting by human T-ALL cells depends on their ability to express CCR7. These studies identify a single chemokine-receptor interaction as a CNS 'entry' signal, and open the way for future pharmacological targeting. Targeted inhibition of CNS involvement in T-ALL could potentially decrease the intensity of CNS-targeted therapy, thus reducing its associated short- and long-term complications.