Parney, Ian F.

OBJECTIVE The objective of this study was to isolate extracellular vesicles (EVs) from plasma in a cohort of patients with traumatic brain injury (TBI) and analyze their contents for novel biomarkers that could prove useful for rapid diagnosis and classification of brain injury during initial evaluation. METHODS Plasma EVs were isolated by serial ultracentrifugation from patients with TBI (n = 15) and healthy controls (n = 5). Samples were obtained from the TRACK-TBI biorepository (2010–present). Size and concentration were determined by nanoparticle tracking. Glial fibrillary acidic protein (GFAP) concentration was determined in EV protein. EV RNA was isolated and deep sequencing of short noncoding RNA was performed. RESULTS Plasma EVs are physically similar but contained approximately 10 times more GFAP in TBI patients with altered consciousness than patients and controls with normal consciousness. Eleven highly differentially expressed microRNAs (miRNAs) were identified between these groups. Genes targeted by these miRNAs are highly associated with biologically relevant cellular pathways, including organismal injury, cellular development, and organismal development. Multiple additional coding and noncoding RNA species with potential biomarker utility were identified. CONCLUSIONS Isolating plasma EVs in patients with TBI is feasible. Increased GFAP concentration—a validated plasma TBI marker—in EVs from TBI patients with altered consciousness, along with differential expression of multiple miRNAs targeting TBI-relevant pathways, suggests that EVs may be a useful source of TBI biomarkers. Additional evaluation in larger patient cohorts is indicated.

Abstract PURPOSE We investigated the long term impact of SRS and WBRT in two large prospective phase III trials. METHODS Patients with 1–4 BMs +/- resection were randomized to SRS or WBRT. Cognitive deterioration was a drop of >1 standard deviation from baseline in >2/6 cognitive measures (CM). Quality of life (QOL) scores were scored 0–100 point scale. CM and QOL scores were modeled using baseline adjusted Linear Mixed Models (LMM) with uncorrelated random intercept for subject and random slopes for time. Differences in trend over time between groups and the effect of >2 cognitive scores with >2 SD change from baseline were assessed. RESULTS 88 patients were included with median follow up of 24 months. We observed decreasing CM over time (SRS: 4/6; WBRT: 5/6). Mean CM was significantly higher in SRS for Total recall and Delayed Recall at 3, 6, 9, 12 months. More patients in WBRT arm declined 1 SD in >1 and >2 CM at the 3, 6, 9, and 12 months. A 1 SD decline in >3 CM at 1 year was 21% SRS vs 47% WBRT (p=0.02). SRS had fewer patients with a 2 SD decline in >1 CM at every time point. SRS had fewer patients with a 2 SD decline at >2 and >3 CM. WBRT had lower QOL at 3 months, but switched to SRS having lower QOL at 24 months for PWB, EWB, FWB, FactG, BR, and FactBR (p<0.05). A 2 SD decline in cognition decreased mean FWB by 6.4 units (95% CI: -11, -1.75; p=0.007) and decreased QOL by 5.1 units (95% CI: -7.7, -2.5; p<0.001). CONCLUSIONS We report the first pooled prospective study demonstrating the long term outcomes of patients with BMs after cranial radiation. WBRT was associated with worse cognitive outcomes. Impaired cognition is associated with worse QOL.

2072 Background: Human glioma stem cells and xenograft lines are common translational models in neuro-oncology but it has not been established if they are genetically and phenotypically comparable. This study aimed to determine if human glioma xenografts and stem cell-derived tumors had similar genotypes. Methods: Matched glioma stem cell cultures and subcutaneous xenograft lines were generated from four human glioblastoma specimens (BT114, BT116, BT120, BT132). Comparison was made between subcutaneous stem cell-derived tumors (flank) and xenografts established in nude mice. Copy number variation (CNV) and gene expression microarray studies were performed. Results: Various differences in copy number and gene expression were seen. Observed CNVs included regions within EGFR, myc, and p16 (INK). For example, EGFR copy number was two fold higher in xenografts vs. stem cell-derived tumor in one line (BT114). This difference was corroborated by western blot. Other differences included a heat shock protein homolog (DNAJA4), tetraspanin 13, and a p53 family target gene (ISG20L1). Two lines (BT114, BT116) had a greater than two fold increase in DNAJA4 expression in xenografts vs. stem cell-derived tumors (p = 0.04, 0.01). Two cell lines (BT116, BT120) had a two to eight fold increase in tetraspanin 13 expression in xenografts (p = 0.02, 0.05). However, neither copy number nor gene expression variations were consistent across all cell lines. Conclusions: Xenografts and glioma stem cell-derived tumors established from the same patient specimens have distinct genotypes. Further work is needed to establish if these differences are random or represent characteristic changes selected by different in vitro or in vivo pressures. However, these variations raise questions regarding which model is ideal for studying glioma biology, and which ones best replicate glioma characteristics in human patients.

2025 Background: Although adjuvant radiation has demonstrated a clear survival benefit in anaplastic astrocytomas (AA), the role of concurrent temozolomide (TMZ) remains controversial. Methods: All adult patients diagnosed with AA from 2001 to 2011 and treated with standard doses of adjuvant radiation were identified retrospectively using the neuro-oncology database at the Mayo Clinic (Rochester, MN). Clinical data was extracted from the electronic medical record. IDH mutation status was also determined by obtaining either IDH1R132H immunostain on existing unstained slides or paraffin block sections, or by sequencing. Cumulative survival probabilities were estimated using the Kaplan-Meier method. Univariable and multivariable Cox proportional hazards regression models were fit to compare patients who did/did not receive TMZ. Results: 218 patients were identified that met inclusion criteria. 34 patients were excluded due to missing data on adjuvant chemotherapy. 146 patients had received adjuvant TMZ while 38 had not. Of these, IDH mutation status was determined on 124 patients who received TMZ and 33 of those who had not. The median duration of follow up was 36.4 months. On univariable analysis, adjuvant TMZ demonstrated a trend towards improved median survival from 35.2 to 53.1 months (p=0.06). As an independent variable, patients with IDH mutations had longer median survivals (111.7 months) when compared to IDH wild-type patients (23.3 months) (p<0.001). On multivariable analysis, five-year-survival was significantly impacted by receipt of adjuvant TMZ (HR = 0.56, p=0.03) and IDH mutation status remained a significant prognostic factor (HR = 0.19, p < 0.001) (see Table). Conclusions: IDH mutation strongly predicts a favorable outcome in patients with AA. Secondarily, concurrent TMZ is also associated with improved survival in patients with AA who are receiving adjuvant radiotherapy. [Table: see text]

Abstract Background Immunosuppression in glioblastoma (GBM) is an obstacle to effective immunotherapy. GBM-derived immunosuppressive monocytes are central to this. Programmed cell death ligand 1 (PD-L1) is an immune checkpoint molecule, expressed by GBM cells and GBM extracellular vesicles (EVs). We sought to determine the role of EV-associated PD-L1 in the formation of immunosuppressive monocytes. Methods Monocytes collected from healthy donors were conditioned with GBM-derived EVs to induce the formation of immunosuppressive monocytes, which were quantified via flow cytometry. Donor-matched T cells were subsequently co-cultured with EV-conditioned monocytes in order to assess effects on T-cell proliferation. PD-L1 constitutive overexpression or short hairpin RNA–mediated knockdown was used to determined the role of altered PD-L1 expression. Results GBM EVs interact with both T cells and monocytes but do not directly inhibit T-cell activation. However, GBM EVs induce immunosuppressive monocytes, including myeloid-derived suppressor cells (MDSCs) and nonclassical monocytes (NCMs). MDSCs and NCMs inhibit T-cell proliferation in vitro and are found within GBM in situ. EV PD-L1 expression induces NCMs but not MDSCs, and does not affect EV-conditioned monocytes T-cell inhibition. Conclusion These findings indicate that GBM EV-mediated immunosuppression occurs through induction of immunosuppressive monocytes rather than direct T-cell inhibition and that, while PD-L1 expression is important for the induction of specific immunosuppressive monocyte populations, immunosuppressive signaling mechanisms through EVs are complex and not limited to PD-L1.

AbstractA recent meeting was held on March 22, 2019, among the FDA, clinical scientists, pharmaceutical and biotech companies, clinical trials cooperative groups, and patient advocacy groups to discuss challenges and potential solutions for increasing development of therapeutics for central nervous system metastases. A key issue identified at this meeting was the need for consistent tumor measurement for reliable tumor response assessment, including the first step of standardized image acquisition with an MRI protocol that could be implemented in multicenter studies aimed at testing new therapeutics. This document builds upon previous consensus recommendations for a standardized brain tumor imaging protocol (BTIP) in high-grade gliomas and defines a protocol for brain metastases (BTIP-BM) that addresses unique challenges associated with assessment of CNS metastases. The “minimum standard” recommended pulse sequences include: (i) parameter matched pre- and post-contrast inversion recovery (IR)–prepared, isotropic 3D T1-weighted gradient echo (IR-GRE); (ii) axial 2D T2-weighted turbo spin echo acquired after injection of gadolinium-based contrast agent and before post-contrast 3D T1-weighted images; (iii) axial 2D or 3D T2-weighted fluid attenuated inversion recovery; (iv) axial 2D, 3-directional diffusion-weighted images; and (v) post-contrast 2D T1-weighted spin echo images for increased lesion conspicuity. Recommended sequence parameters are provided for both 1.5T and 3T MR systems. An “ideal” protocol is also provided, which replaces IR-GRE with 3D TSE T1-weighted imaging pre- and post-gadolinium, and is best performed at 3T, for which dynamic susceptibility contrast perfusion is included. Recommended perfusion parameters are given.

Abstract Purpose: Glioblastoma is the most frequent and lethal primary brain tumor. Development of novel therapies relies on the availability of relevant preclinical models. We have established a panel of 96 glioblastoma patient-derived xenografts (PDX) and undertaken its genomic and phenotypic characterization. Experimental Design: PDXs were established from glioblastoma, IDH-wildtype (n = 93), glioblastoma, IDH-mutant (n = 2), diffuse midline glioma, H3 K27M-mutant (n = 1), and both primary (n = 60) and recurrent (n = 34) tumors. Tumor growth rates, histopathology, and treatment response were characterized. Integrated molecular profiling was performed by whole-exome sequencing (WES, n = 83), RNA-sequencing (n = 68), and genome-wide methylation profiling (n = 76). WES data from 24 patient tumors was compared with derivative models. Results: PDXs recapitulate many key phenotypic and molecular features of patient tumors. Orthotopic PDXs show characteristic tumor morphology and invasion patterns, but largely lack microvascular proliferation and necrosis. PDXs capture common and rare molecular drivers, including alterations of TERT, EGFR, PTEN, TP53, BRAF, and IDH1, most at frequencies comparable with human glioblastoma. However, PDGFRA amplification was absent. RNA-sequencing and genome-wide methylation profiling demonstrated broad representation of glioblastoma molecular subtypes. MGMT promoter methylation correlated with increased survival in response to temozolomide. WES of 24 matched patient tumors showed preservation of most genetic driver alterations, including EGFR amplification. However, in four patient–PDX pairs, driver alterations were gained or lost on engraftment, consistent with clonal selection. Conclusions: Our PDX panel captures the molecular heterogeneity of glioblastoma and recapitulates many salient genetic and phenotypic features. All models and genomic data are openly available to investigators.

Abstract INTRODUCTION Central nervous system hemangioblastoma (CNS HGB) is a rare neoplasm, which predominantly arise in the posterior fossa and spinal cord. The etiology is divided into sporadic and von-Hippel Lindau (VHL) disease. The difference in clinical picture of these 2 types of HGB and differentiation of treatment have not been extensively unraveled yet. METHODS Retrospective analysis of consecutive, neurosurgically managed CNS HGB at Mayo Clinic, 1988–2018. RESULTS 117 sporadic and 67 VHL HGBs were treated by Mayo Clinic. No significant difference in sex was observed. Compared with sporadic cases, VHL cases were younger (51.8 vs 36.0 years old, p&lt;0.0001), had more frequent family history (0.0 vs 41.5 %, p&lt;0.0001), and higher frequency of germline alteration (0.0 vs 84.2 %, p&lt;0.0001). Regarding imaging findings, VHL cases had multiple lesions at presentation more frequently (3.4 vs 82.1 %, p&lt;0.0001), it was more common for sporadic lesions to contain cysts (72.2 vs 51.0 %, p=0.0004), the solid portion rate in the entire lesion was larger in VHL lesions (60.2 vs 69.5 %, p=0.02), and the volume was larger in sporadic cases (15.1 vs 6.6 cc, p&lt;0.0001). Regarding treatment, 131 and 123 surgeries were performed for sporadic and VHL cases, respectively, among which the indication of surgery was preventative in 8.4 and 47.3 %, respectively (p&lt;0.0001). VHL cases had higher number of treatments per case in the follow-up (1.3 vs 2.1, p&lt;0.0001). Recurrence-free survival of sporadic cases was significantly longer than that of VHL cases (p=0.007) and overall survival was longer in sporadic cases than VHL, but not significant (p=0.07). CONCLUSION Clinical presentation and tumor appearance on imaging are highly dependent on the etiology. Differences in clinical manifestations require further study, but may reflect contrasting tumor biology that are tied to genetic differences.