Glioma Biopsies Research Articles

Diffuse infiltrative pontine glioma biopsy in children with neuronavigation, frameless procedure: A single center experience of 10 cases

IntroductionThis study presented pediatric DIPG ‘s biopsy with frameless Neuronavigation. Patients and methodsWe report our experience about 10 patients who had Diffuse Intrinsic Pontine Glioma between 2014 and 2018. All patients were biopsied with BrainLab Varioguide Neuronavigation®. We always used fusion between specific CT Scan and MRI to selected target, made planning and biopsies. All patients were included in BIOMEDE after scientific and ethic discussions. We always selected a trans-cerebellar trajectory and made same procedure (lot of biopsies at one level). All patients have MRI at J1 to verify site of biopsy and to eliminate complication. ResultsThe average age was 8.1 years. Symptoms were common with principally headaches and nystagmus. All biopsies were contributive for histopathological diagnosis and establish molecular profile for molecular study. We have no definitive morbidity and procedure duration was 93minutes in average. All MRI didn’t showed intracranial complication after procedure and showed great precision of biopsy compared with the selected target. DiscussionWe reviewed the literature and compare our results with series of DIPG biopsies using stereotactic frame or robotic assisted frameless. It was a safe, accuracy and easiness procedure. We always have histopathological and molecular result to proceed next step of treatment. This modality is an alternative possibility to biopsy very young patients with low morbidity.

Abstract 3950: Spatiotemporal analysis of gliomas: Dynamics of mesenchymal multicellular structures as novel target for tumor treatment

Abstract Glioblastoma multiforme (GBM) is the most frequent and lethal tumor of the central nervous system. GBM are characterized by diffuse invasion and cellular heterogeneity which challenges treatment efficacy. Tumors with mesenchymal properties display the most aggressive phenotype. However, the biological function and molecular mechanisms underlying GBM mesenchymal transformation remain unknown. Analysis of mouse and human malignant gliomas revealed the presence of organized multicellular structures formed by elongated and aligned cells. These structures resemble areas of GBM mesenchymal transformation that we named Oncostreams. We determined the molecular signature underlying oncostreams function by performing laser capture microdissection followed by transcriptomic analysis. We found that oncostreams overexpressed Col1a1, ACTA2, MMP9, MMP10 and ADAMTS2 genes, all of them associated with regulation of extracellular matrix organization, collagen catabolic process and cellular migration pathways. Functional network analysis indicated that Col1a1 was a primary regulator gene. To analyze whether these structures display migratory properties we used time lapse imagining in a 3D organotypic glioma model. Morphological and statistical analysis revealed that oncostreams displayed a collective motion pattern, organized as streams or flocks. This dynamic patterns participate in local invasion and function as tumoral highways to facilitate the spread of several cells as intra-. Further analysis showed that oncostreams presence correlates with increased collagen expression and decreased animal survival. Patient's glioma biopsies also evidenced that these areas of Col1a1 overexpression were present in high grade but no in low-grade gliomas. We corroborated by immune-hystochemistry that Col1a1 were overexpressed and co-localized within GFP positive tumoral cells. Further, to analyze the origin and role of Col1a1 in glioma malignancy we generated genetically engineered mouse glioma models (GEMM) with Col1a1 downregulation. Interestingly, Col1a1 was retained only surrounding the blood vessel but was completed absent within the tumor parenchyma. We demonstrated that Col1a1 downregulation ablates oncostreams structures, reversed the malignant phenotype resembling low grade glioma histopathology and prolonged animal survival. This study reveals that oncostreams are organized dynamic structures that regulate glioma growth and invasion. They are areas of mesenchymal transformation defined by a molecular signature associated to extracellular matrix proteins expressed as Col1a1. Oncostreams with high expression of Col1a1 within glioma cells represent a novel potential targets for future translational development. Disruption of oncostreams will provide a new avenue to treat GBM. Citation Format: Andrea Comba, Patrick Dunn, Anna E. Argento, Padma Kadiyala, Sebastien Motsch, Daniel Zamler, Alon Kahana, Phillips E. Kish, Maria G. Castro, Pedro R. Lowenstein. Spatiotemporal analysis of gliomas: Dynamics of mesenchymal multicellular structures as novel target for tumor treatment [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3950.

Global activation of oncogenic pathways underlies therapy resistance in diffuse midline glioma

Diffuse midline gliomas (DMGs) are aggressive pediatric brain tumors with dismal prognosis due to therapy-resistant tumor growth and invasion. We performed the first integrated histologic/genomic/proteomic analysis of 21 foci from three pontine DMG cases with supratentorial dissemination. Histone H3.3-K27M was the driver mutation, usually at high variant allele fraction due to recurrent chromosome 1q copy number gain, in combination with germline variants in ATM, FANCM and MYCN genes. Both previously reported and novel recurrent copy number variations and somatic pathogenic mutations in chromatin remodeling, DNA damage response and PI3K/MAPK growth pathways were variably detected, either in multiple or isolated foci. Proteomic analysis showed global upregulation of histone H3, lack of H3-K27 trimethylation, and further impairment of polycomb repressive complex 2 by ASXL1 downregulation. Activation of oncogenic pathways resulted from combined upregulation of N-MYC, SOX2, p65/p50 NF-κB and STAT3 transcription factors, EGFR, FGFR2, PDGFRα/β receptor tyrosine kinases, and downregulation of PHLPP1/2, PTEN and p16/INK4A tumor suppressors. Upregulation of SMAD4, PAI-1, CD44, and c-SRC in multiple foci most likely contributed to invasiveness. This integrated comprehensive analysis revealed a complex spatiotemporal evolution in diffuse intrisic pontine glioma, recommending pontine and cerebellar biopsies for accurate populational genetic characterization, and delineated common signaling pathways and potential therapeutic targets. It also revealed an unsuspected activation of a multitude of oncogenic pathways, including cancer cell reprogramming, explaining the resistance of DMG to current therapies.

Spectrum of radiological presentations of Intracranial Glioma among the Indian population

Background: Gliomas comprises the group of most common primary tumour of central nervous system. The current study was undertaken to evaluate the various usual and unusual radiological presentations among the patients of glioma among the Indian population.
 Aims and Objective: The aim of the current study was to observe the various radiological presentations of glioma occurring among the subjects. It was also intended to correlate the radiological and histopathological grading in glioma among subjects.
 Materials and Methods: The current study was a prospective observational study carried out among seventy-five patients admitted in a tertiary care hospital in Eastern India with provisional diagnosis of supratentorial glioma and operative biopsy confirmed to be Glioma.
 Result: The most common grade of tumour encountered was grade IV tumour. Of the clinical features, 70.66 % patients had hypodense lesion, 25.33% patients had iso dense and hypodense (mixed density) lesion and 3 patients had hyperdense lesion on CT Brain. On MRI in T1 Weighted images, 81.33% patients had hypointense lesion lesion on MRI T1 image. On T2 Weighted Images, 76% patients had hyperintense lesion., 77.33% demonstrated heterogenous enhancement. Ring enhancement was seen in 9 cases Minimal to no enhancement was seen in 8 cases. MRS showed maximum cases had Choline peak with altered Choline: creatinine ratio and decreased NAA peaks.
 Conclusion: Good imaging interpretations is crucial for planning of surgical excision and adjuvant radiotherapy. This review has been put to solve the basic problem in interpreting the Ct Scan and MRI of Glioma.

Applications of cerebrospinal fluid circulating tumor DNA in the diagnosis of gliomas.

The 2016 World Health Organization (WHO) Classification of Tumors of the Central Nervous System (CNS) was revised to include molecular biomarkers as diagnostic criteria. However, conventional biopsies of gliomas were spatially and temporally limited. This study aimed to determine whether circulating tumor DNA (ctDNA) from cerebrospinal fluid (CSF) could provide more comprehensive diagnostic information to gliomas. Combined with clinical data, we analyzed gene alterations from CSF and tumor tissues of newly diagnosed patients, and detected mutations of ctDNA in recurrent patients. We simultaneously analyzed mutations of ctDNA in different glioma subtypes, and in lower-grade gliomas (LrGG) versus glioblastoma multiforme (GBM). CSF ctDNA mutations had high concordance rates with tumor DNA (tDNA). CSF ctDNA mutations of PTEN and TP53 were commonly detected in recurrent gliomas patients. IDH mutation was detected in most of CSF ctDNA derived from IDH-mutant diffuse astrocytomas, while CSF ctDNA mutations of RB1 and EGFR were found in IDH-wild-type GBM. IDH mutation was detected in LrGG, whereas Rb1 mutation was more commonly detected in GBM. CSF ctDNA detection can be an alternative method as liquid biopsy in gliomas.

Biopsy in diffuse pontine gliomas: expert neurosurgeon opinion-a survey from the SIOPE brain tumor group.

The prognosis of diffuse intrinsic pontine glioma (DIPG) is poor. The role of biopsy in DIPG remains controversial since the diagnosis may be established with imaging alone. Recent advances in understanding molecular biology and targeting of brain tumors have created a renewed interest in biopsy for DIPG. The Neurosurgery Working Group (NWG) of the SIOP-Europe Brain Tumor Group (BTG) undertook a survey among international pediatric neurosurgeons to define their current perceptions and practice regarding DIPG biopsy. The NWG developed a 20-question survey which was emailed to neurosurgeons in the International Society for Pediatric Neurosurgery (ISPN). The questionnaire included questions on diagnosis, indications, and techniques for biopsy, clinical trials, and healthcare infrastructure. The survey was sent to 202 neurosurgeons and 73 (36%) responded. Consensus of > 75% agreement was reached for 12/20 questions, which included (1) radiological diagnosis of DIPG is sufficient outside a trial, (2) clinical trial-based DIPG biopsy is justified if molecular targets are investigated and may be used for treatment, and (3) morbidity/mortality data must be collected to define the risk:benefit ratio. The remaining 8/20 questions proved controversial and failed to reach consensus. Routine DIPG biopsy continues to be debated. Most neurosurgeons agreed that DIPG biopsy within a clinical trial should be supported, with the aims of defining the procedure risks, improving understanding of tumor biology, and evaluating new treatment targets. Careful family counseling and consent remain important.

Liquid Biopsy in Gliomas- A Review.

Are we witnessing the end of the biopsy as we know it? Is this the start of a revolution in cancer diagnostics and treatment where analysis of somatic mutations present in the blood, CSF, or urine followed by targeted therapy replaces the traditional surgery followed by chemo-radiation? Since 2016, molecular markers are an integral part of the 'glioma' treatment decision-making process- diagnostic, prognostic, and therapeutic. A lot of these somatic mutations that identify and prognosticate tumors are also detected in the adjoining bio-fluids in serum or CSF- the sampling of which is known as liquid biopsy. The objective of this study is to review the advancement of scientific techniques that now allows the investigation of these bio-fluids, to diagnose, prognosticate and treat gliomas. This review article is an exhaustive review of the literature that summarises the role of the three main liquid biopsy modalities- Circulating Tumor Cells, Cell-free Tumor DNA and Exosomes in the detection of known diagnostic and prognostic markers in gliomas. The current review highlights the strengths and weaknesses of the diffrerent modalities in use, and their potential use in the clinical setting. Liquid biopsies hold tremendous potential in the diagnosis and management of gliomas in the future.

Terceiroventriculostomia e biópsia endoscópica em glioma de baixo grau:relato de caso

Terceiroventriculostomia e biópsia endoscópica em glioma de baixo grau:relato de caso

Biomarker concordance between molecular stereotactic biopsy and open surgical specimens in gliomas.

To compare 1p/19q codeletion, MGMT promoter methylation, and IDH mutation status in stereotactic biopsy and open craniotomy specimens. The latest WHO classification of gliomas requires assessment of the expression of molecular markers. Samples can be obtained for molecular assays via open craniotomy or molecular stereotactic biopsy (MSB). However, there is uncertainty as to whether MSB is representative of the entire tumour, and therefore how reliable it is for treatment planning. We examined 11 patients diagnosed with brain tumours suspicious of glioma who underwent open craniotomy after stereotactic biopsy and in whom multiple biomarkers were assessed in both sets of samples by methylation-specific multiplex ligation-dependent probe amplification. Institutional Review Board ethical approval was granted (KB 694/2018). The initial histopathological grade as determined by stereotactic biopsy was the same as in the samples obtained by open surgery. Further, the marker profile used here was valid in both high- and low-grade gliomas. MSB is a reliable way to obtain material for precision medicine approaches.

Reduced expression of proteolipid protein 2 increases ER stress-induced apoptosis and autophagy in glioblastoma.

Proteolipid protein 2 (PLP2) is an integral ion channel membrane protein of the endoplasmic reticulum. The protein has been shown to be highly expressed in many cancer types, but its importance in glioma progression is poorly understood. Using publicly available datasets (Rembrandt, TCGA and CGGA), we found that the expression of PLP2 was significantly higher in high‐grade gliomas than in low‐grade gliomas. We confirmed these results at the protein level through IHC staining of high‐grade (n = 56) and low‐grade glioma biopsies (n = 16). Kaplan‐Meier analysis demonstrated that increased PLP2 expression was associated with poorer patient survival. In functional experiments, siRNA and shRNA PLP2 knockdown induced ER stress and increased apoptosis and autophagy in U87 and U251 glioma cell lines. Inhibition of autophagy with chloroquine augmented apoptotic cell death in U87‐ and U251‐siPLP2 cells. Finally, intracranial xenografts derived from U87‐ and U251‐shPLP2 cells revealed that loss of PLP2 reduced glioma growth in vivo. Our results therefore indicate that increased PLP2 expression promotes GBM growth and that PLP2 represents a potential future therapeutic target.

Genomic analysis of primary and recurrent gliomas reveals clinical outcome related molecular features

Tremendous efforts have been made to explore biomarkers for classification and grading on gliomas. The goal of this study was to identify more molecular features that are associated with clinical outcomes by comparing the genomic profiles of primary and recurrent gliomas and determine potential recurrence leading factors that are significantly enriched in relapse tumors. Hybrid capture based next generation sequencing (NGS) analysis was performed on 64 primary and 17 recurrent glioma biopsies. Copy number variation (CNV) was more frequent in recurrent tumors and CDKN2A/B loss was significantly enriched. In addition, overall mutations in cell cycle pathway are more common in relapse tumors. The patterns of gene sets, including IDH1/TERT and IDH1/TP53 exhibited significant difference between the groups. Survival analysis uncovered the worse disease-free survival (DFS) and overall survival (OS) associated with altered copy number and excessive activation of CELL CYCLE pathway. High Tumor Mutation Burden (TMB) was also a biomarker with great potential for poor prognosis. The assessment of genomic characteristics in primary versus recurrent gliomas aids the discovery of potential predictive biomarkers. The prognostic value of TMB in gliomas was raised for the first time.

P13.08 1H-MRS metabolomics as a predictor of overall survival in glioma patients

Abstract BACKGROUND We assess the efficacy of the metabolomic profile from glioma biopsies, in providing estimates of postsurgical Overall Survival in glioma patients. MATERIAL AND METHODS Tumor biopsies from 46 patients bearing gliomas, obtained neurosurgically in the period 1992–1998, were analyzed by high resolution 1H magnetic resonance spectroscopy (HR- 1H MRS), following retrospectively individual postsurgical Overall Survival up to 720 weeks. RESULTS The Overall Survival profile could be resolved in three groups; Short (shorter than 52 weeks, n=19), Intermediate (between 53 and 364 weeks, n=19) or Long (longer than 365 weeks, n=8), respectively. Classical histopathological analysis assigned WHO grades II-IV to every biopsy but notably, some patients with low grade glioma depicted unexpectedly Short Overall Survival, while some patients with high grade glioma, presented unpredictably Long Overall Survival. To explore reasons underlying this behavior, we analyzed HR- 1H MRS spectra from acid extracts of these biopsies, to identify the metabolite patterns underlying OS predictions. Poor prognosis was found in biopsies with higher contents of alanine, acetate, glutamate, total choline, phosphorylcholine and glycine, while more favorable prognosis was achieved in biopsies with larger contents of total creatine, glycerol-phosphorylcholine and myo-inositol. We implemented then a multivariate analysis approach to identify hierarchically the influence of these metabolomic biomarkers on OS predictions, using Classification Regression Trees (CRTs). Metabolomic CRTs grew up to 3 branches and split into 8 nodes, predicting correctly the outcome of 94.7% of the patients in the Short Overall Survival group, 78.9 % of the patients in the Intermediate Overall Survival group, and 75% of the patients in the Long Overall Survival group, respectively. CONCLUSION Present results suggest that metabolic profiling by HR-1H MRS provides more accurate Overall Survival estimates of glioma patients than classical histopathological grading, thus allowing to implement more accurate therapeutic decisions.

P13.05 Chemical Biopsy as an Alternative Sampling Method in Neurooncology

Abstract BACKGROUND Histopathology is a gold standard for diagnostics of brain tumors but it is time consuming, demands removal of the part of the tissue what makes it highly invasive. That is why it seems to be relevant to develop new diagnostic methods addressing the abovementioned drawbacks. Ideal tool should be easy to use, as low invasive as possible and of high clinical value. The aim of the study was to assess the usefulness of chemical biopsy based on solid phase microextraction as a sampling method for lipidomic analysis of brain tumors. MATERIAL AND METHODS The experiment was carried out on 38 brain tumors. Sampling was performed ex vivo, directly after removal of brain tumors. SPME fibers (coated with biocompatible extraction phase, length and diameter of the coating 7mm and 200um, respectively) were inserted into the sample without prior preparation of the tissue. Desorption of lipids was performed in isopropanol-methanol mixture. Analysis of extracts was carried out using liquid chromatography coupled to high resolution mass spectrometry. RESULTS Wide range of tentative lipid species was obtained in the studies. Two histologically different types of brain tumors: gliomas and meningiomas were subjected to the chemical biopsy. The results showed very good separation between the tumor types in unsupervised chemometric tests. The lipid species responsible for this separation were previously reported in the literature as cancer biomarkers. With regards to malignancy grading of gliomas the separation was notable but not as clear as between the tumor types, what is probably related to high heterogeneity of this cancerous tissue and subjectivity of histopathological analysis which were a referring point in methods comparison. Very good agreement was noted for changes in lipidome obtained by chemical biopsy and histological subtypes of gliomas. However, the most promising results showed impact of IDH1 R132H mutation on lipidome of gliomas. CONCLUSION Chemical biopsy seems to be useful method in lipidomics studies and for differentiation of brain tumors according their histological origin and malignancy. It also enables to find a relation between IDH1 mutation and lipidomic profile. ACKNOWLEDGMENTS The work was supported by the grant 2015/18/M/ST4/00059 funded by National Science Centre, Poland. The authors want to acknowledge Supelco/MilliporeSigma for the SPME probes, Thermo Fisher Scientific for the access to Q-Exactive Focus mass spectrometer, Anchem for the technical support and Shim-Pol for the access to LCMS-8060 mass spectrometer and technical support.

P14.88 Comparison of SNaPshot PCR and digital droplet PCR for the identification of TERT promoter in non-diagnostic biopsies from glioma patients

Abstract BACKGROUND We previously showed that TERT promotor (pTERT) mutations can be detected in a high proportion of non-diagnostic biopsies from glioma patients using SNaPshot PCR. The aim of the present study was to assess the diagnostic value of digital droplet PCR (ddPCR) in this setting. MATERIAL AND METHODS pTERT mutations were retrospectively assessed using SNaPshot PCR and ddPCR in the non-diagnostic biopsies of 14 adult patients who underwent an initial non-diagnostic needle biopsy that led to perform another biopsy that finally demonstrated a pTERT-mutant glioma. RESULTS Median age at diagnosis was 69 years and the median interval between the initial and final biopsy was 22 days. The initial biopsy consisted of unspecific infiltrating glial cells (n=10), hemorrhage, necrosis and/or fibrosis (n=4). Using SNaPshot PCR, the final biopsy found an IDH-WT pTERT-mutant glioma in 12 cases (glioblastoma n=12), an IDH-mutant pTERT-mutant oligodendroglioma in one case and an IDH-WT pTERT-WT glioblastoma in one case. Retrospectively, a pTERT mutation was identified by both ddPCR and SNaPshot in the non-diagnostic biopsies in 85% (11 /13) of pTERT mutant-gliomas. No mutation was detected in the non-diagnostic biopsies of IDH-WT pTERT-WT gliomas using either technique. Overall, a pTERT mutation were detected in 79% (11/14) of non-diagnostic biopsies. CONCLUSION Digital Droplet PCR, as SNaPshot PCR, enables the identification of pTERT mutations in a high rate of non-diagnostic biopsies from glioma patients. The sensitivity of both techniques seems similar in this setting.

Quantitative proteomics reveals reduction of endocytic machinery components in gliomas

BackgroundGliomas are the most frequent and aggressive malignancies of the central nervous system. Decades of molecular analyses have demonstrated that gliomas accumulate genetic alterations that culminate in enhanced activity of receptor tyrosine kinases and downstream mediators. While the genetic alterations, like gene amplification or loss, have been well characterized, little information exists about changes in the proteome of gliomas of different grades. MethodsWe performed unbiased quantitative proteomics of human glioma biopsies by mass spectrometry followed by bioinformatic analysis. FindingsVarious pathways were found to be up- or downregulated. In particular, endocytosis as pathway was affected by a vast and concomitant reduction of multiple machinery components involved in initiation, formation, and scission of endocytic carriers. Both clathrin-dependent and -independent endocytosis were changed, since not only clathrin, AP-2 adaptins, and endophilins were downregulated, but also dynamin that is shared by both pathways. The reduction of endocytic machinery components caused increased receptor cell surface levels, a prominent phenotype of defective endocytosis. Analysis of additional biopsies revealed that depletion of endocytic machinery components was a common trait of various glioma grades and subclasses. InterpretationWe propose that impaired endocytosis creates a selective advantage in glioma tumor progression due to prolonged receptor tyrosine kinase signaling from the cell surface. FundThis work was supported by Grants 316030-164105 (to P. Jenö), 31003A-162643 (to M. Spiess) and PP00P3-176974 (to G. Hutter) from the Swiss National Science Foundation. Further funding was received by the Department of Surgery from the University Hospital Basel.

Assessment of Overall Survival in Glioma Patients as Predicted by Metabolomic Criteria.

Objective: We assess the efficacy of the metabolomic profile from glioma biopsies in providing estimates of postsurgical Overall Survival in glioma patients.Methods: Tumor biopsies from 46 patients bearing gliomas, obtained neurosurgically in the period 1992–1998, were analyzed by high resolution 1H magnetic resonance spectroscopy (HR- 1H MRS), following retrospectively individual postsurgical Overall Survival up to 720 weeks.Results: The Overall Survival profile could be resolved in three groups; Short (shorter than 52 weeks, n = 19), Intermediate (between 53 and 364 weeks, n = 19) or Long (longer than 365 weeks, n = 8), respectively. Classical histopathological analysis assigned WHO grades II–IV to every biopsy but notably, some patients with low grade glioma depicted unexpectedly Short Overall Survival, while some patients with high grade glioma, presented unpredictably Long Overall Survival. To explore the reasons underlying these different responses, we analyzed HR-1H MRS spectra from acid extracts of the same biopsies, to characterize the metabolite patterns associated to OS predictions. Poor prognosis was found in biopsies with higher contents of alanine, acetate, glutamate, total choline, phosphorylcholine, and glycine, while more favorable prognosis was achieved in biopsies with larger contents of total creatine, glycerol-phosphorylcholine, and myo-inositol. We then implemented a multivariate analysis to identify hierarchically the influence of metabolomic biomarkers on OS predictions, using a Classification Regression Tree (CRT) approach. The CRT based in metabolomic biomarkers grew up to three branches and split into eight nodes, predicting correctly the outcome of 94.7% of the patients in the Short Overall Survival group, 78.9% of the patients in the Intermediate Overall Survival group, and 75% of the patients in the Long Overall Survival group, respectively.Conclusion: Present results indicate that metabolic profiling by HR-1H MRS improves the Overall Survival predictions derived exclusively from classical histopathological gradings, thus favoring more precise therapeutic decisions.

Biopsy and Ablation of H3K27 Glioma Using Skull-Mounted Smartframe Device: Technical Case Report

Clearpoint Smartframe is a magnetic resonance imaging-compatible stereotactic system often used to perform magnetic resonance imaging-guided biopsies. The system is typically mounted on the scalp through screws that pierce the skin and penetrate the outer table of the cranium. However, the frame can also be configured to be mounted directly onto the skull. Here, we describe the clinical context well suited for a skull-mount Clearpoint Smartframe surgery. The patient suffered from a subcentimeter right parafourth ventricular contrast-enhancing lesion with hydrocephalus and underwent a right suboccipital needle biopsy followed by an occipital ventriculoperitoneal shunt. Although the hydrocephalus resolved with the ventriculoperitoneal shunt, the biopsy sample proved nondiagnostic. The patient underwent a second procedure during which the Clearpoint Smartframe was mounted onto the skull through space dissected free during the previous surgery. Diagnostic biopsy (H3K27 glioma) was performed followed by stereotactic laser ablation of the lesion. We describe a case in which the skull mount Clearpoint Smartframe was used to biopsy and ablate a midline H3K27 glioma.

3131 ONCOSTREAMS: NOVEL DYNAMICS PATHOLOGICAL MULTICELLULAR STRUCTURES INVOLVED IN GLIOBLATOMA GROWTH AND INVASION

OBJECTIVES/SPECIFIC AIMS: Oncostreams represent a novel growth pattern of GBM. In this study we uncovered the cellular and molecular mechanism that regulates the oncostreams function in GBM growth and invasion. METHODS/STUDY POPULATION: We studied oncostreams organization and function using genetically engineered mouse gliomas models (GEMM), mouse primary patient derived GBM model and human glioma biopsies. We evaluated the molecular landscape of oncostreams by laser capture microdissection (LCM) followed by RNA-Sequencing and bioinformatics analysis. RESULTS/ANTICIPATED RESULTS: Oncostreams are multicellular structures of 10-20 cells wide and 2-400 μm long. They are distributed throughout the tumors in mouse and human GBM. Oncostreams are heterogeneous structures positive for GFAP, Nestin, Olig2 and Iba1 cells and negative for Neurofilament. Using GEMM we found a negative correlation between oncostream density and animal survival. Moreover, examination of patient’s glioma biopsies evidenced that oncostreams are present in high grade but no in low grade gliomas. This suggests that oncostreams may play a role in tumor malignancy. Our data also indicated that oncostreams aid local invasion of normal brain. Transcriptome analysis of oncostreams revealed 43 differentially expressed (DE) genes. Functional enrichment analysis of DE genes showed that “collagen catabolic processes”, “positive regulation of cell migration”, and “extracellular matrix organization” were the most over-represented GO biological process. Network analysis indicated that Col1a1, ACTA2, MMP9 and MMP10 are primary target genes. These genes were also overexpressed in more malignant tumors (WT-IDH) compared to the less malignant (IDH1- R132H) tumors. Confocal time lapse imagining of 3D tumor slices demonstrated that oncostreams display a collective motion pattern within gliomas that has not been seen before. DISCUSSION/SIGNIFICANCE OF IMPACT: In summary, oncostreams are anatomically and molecularly distinctive, regulate glioma growth and invasion, display collective motion and are regulated by the extracellular matrix. We propose oncostreams as novel pathological markers valuable for diagnosis, prognosis and designing therapeutics for GBM patients.

Validation of vessel size imaging (VSI) in high-grade human gliomas using magnetic resonance imaging, image-guided biopsies, and quantitative immunohistochemistry

To evaluate the association between a vessel size index (VSIMRI) derived from dynamic susceptibility contrast (DSC) perfusion imaging using a custom spin-and-gradient echo echoplanar imaging (SAGE-EPI) sequence and quantitative estimates of vessel morphometry based on immunohistochemistry from image-guided biopsy samples. The current study evaluated both relative cerebral blood volume (rCBV) and VSIMRI in eleven patients with high-grade glioma (7 WHO grade III and 4 WHO grade IV). Following 26 MRI-guided glioma biopsies in these 11 patients, we evaluated tissue morphometry, including vessel density and average radius, using an automated procedure based on the endothelial cell marker CD31 to highlight tumor vasculature. Measures of rCBV and VSIMRI were then compared to histological measures. We demonstrate good agreement between VSI measured by MRI and histology; VSIMRI = 13.67 μm and VSIHistology = 12.60 μm, with slight overestimation of VSIMRI in grade III patients compared to histology. rCBV showed a moderate but significant correlation with vessel density (r = 0.42, p = 0.03), and a correlation was also observed between VSIMRI and VSIHistology (r = 0.49, p = 0.01). The current study supports the hypothesis that vessel size measures using MRI accurately reflect vessel caliber within high-grade gliomas, while traditional measures of rCBV are correlated with vessel density and not vessel caliber.

Cystathionine as a marker for 1p/19q codeleted gliomas by in vivo magnetic resonance spectroscopy.

Codeletion of chromosome arms 1p and 19q (1p/19q codeletion) highly benefits diagnosis and prognosis in gliomas. In this study, we investigated the effect of 1p/19q codeletion on cancer cell metabolism and evaluated possible metabolic targets for tailored therapies. We combined in vivo 1H (proton) magnetic resonance spectroscopy (MRS) measurements in human gliomas with the analysis of a series of standard amino acids by liquid chromatography-mass spectroscopy (LC-MS) in human glioma biopsies. Sixty-five subjects with low-grade glioma were included in the study: 31 underwent the MRI/MRS examination, 47 brain tumor tissue samples were analyzed with LC-MS, and 33 samples were analyzed for gene expression with quantitative PCR. Additionally, we performed metabolic tracer experiments in cell models with 1p deletion. We report the first in vivo detection of cystathionine by MRS in 1p/19q codeleted gliomas. Selective accumulation of cystathionine was observed in codeleted gliomas in vivo, in brain tissue samples, as well as in cells harboring heterozygous deletions for serine- and cystathionine-pathway genes located on 1p: phosphoglycerate dehydrogenase (PHGDH) and cystathionine gamma-lyase (CTH). Quantitative PCR analyses showed 40-50% lower expression of both PHGDH and CTH in 1p/19q codeleted gliomas compared with their non-codeleted counterparts. Our results provide strong evidence of a selective vulnerability of codeleted gliomas to serine and glutathione depletion and point to cystathionine as a possible noninvasive marker of treatment response.