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OBJECTIVE Glioblastoma (GBM) is a devasting primary brain tumor with less than a 5% 5-year survival. Treatment response assessment can be challenging because of inflammatory pseudoprogression that mimics true tumor progression clinically and on imaging. Developing additional noninvasive assays is critical. In this article, the authors review various biomarkers that could be used in developing liquid biopsies for GBM, along with strengths, limitations, and future applications. In addition, they present a potential liquid biopsy design based on the use of an extracellular vesicle–based liquid biopsy targeting nonneoplastic extracellular vesicles. METHODS The authors conducted a current literature review of liquid biopsy in GBM by searching the PubMed, Scopus, and Google Scholar databases. Articles were assessed for type of biomarker, isolation methodology, analytical techniques, and clinical relevance. RESULTS Recent work has shown that liquid biopsies of plasma, blood, and/or CSF hold promise as noninvasive clinical tools that can be used to diagnose recurrence, assess treatment response, and predict patient outcomes in GBM. Liquid biopsy in GBM has focused primarily on extracellular vesicles, cell-free tumor nucleic acids, and whole-cell isolates as focal biomarkers. GBM tumor signatures have been generated via analysis of tumor gene mutations, unique RNA expression, and metabolic and proteomic alterations. Liquid biopsies capture tumor heterogeneity, identifying alterations in GBM tumors that may be undetectable via surgical biopsy specimens. Finally, biomarker burden can be used to assess treatment response and recurrence in GBM. CONCLUSIONS Liquid biopsy offers a promising avenue for monitoring treatment response and recurrence in GBM without invasive procedures. Although additional steps must be taken to bring liquid biopsy into the clinic, proof-of-principle studies and isolation methodologies are promising. Ultimately, CSF and/or plasma-based liquid biopsy is likely to be a powerful tool in the neurosurgeon’s arsenal in the near future for the treatment and management of GBM patients.

INTRODUCTION Dendritic cells (DCs) are potent antigen presenting cells that can be exploited to initiate an adaptive anti-tumoral immune response. DC vaccine clinical trials for primary glioblastoma (GBM) have reported prolonged progression-free survival without any impact on overall survival (OS). We report a radiomics approach that identifies a subpopulation of patients with prolonged OS in clinical trial MC1272. METHODS Twenty adults with primary GBM undergoing standard-of-care therapy were enrolled in MC1272. Autologous DCs were pulsed with allogenic GBM cell lines to generate vaccines that were administered for up to twelve cycles. Standard brain imaging was obtained at the initiation of treatment and two months afterwards. An independent cohort of image-localized biopsies underwent RNA sequencing followed by cellular deconvolution to estimate T cell abundance. A machine learning model was trained to predict intratumoral T cell abundance from imaging features, and the model was applied to MC1272 patient imaging. RESULTS Voxelwise predictions of T cell abundance were generated for each patient’s pre- and post-treatment images. The difference in total intratumoral T cell abundance between imaging timepoints classified patients into increasing or decreasing T cell groups. Patients whose T cells increased had longer OS (median, 21 months) than those whose T cells decreased (median, 10 months; p=0.0035). The significance remained in a Cox proportional hazards analysis that adjusted for patient age and sex (p=0.011). CONCLUSIONS A spatially-resolved radiomics model detected that an intratumoral T cell influx after DC vaccine therapy was associated with prolonged OS. The “post-treatment” imaging in this study was obtained a mere two months after DC vaccine initiation, suggesting that our radiomics model can provide an early indication of treatment responsiveness and prognosis in these patients.

Digital, OBJECTIVE Recent studies have proposed resection of the T2 FLAIR hyperintensity beyond the T1 contrast enhancement (supramarginal resection [SMR]) for IDH–wild-type glioblastoma (GBM) to further improve patients’ overall survival (OS). GBMs have significant variability in tumor cell density, distribution, and infiltration. Advanced mathematical models based on patient-specific radiographic features have provided new insights into GBM growth kinetics on two important parameters of tumor aggressiveness: proliferation rate (ρ) and diffusion rate (D). The aim of this study was to investigate OS of patients with IDH–wild-type GBM who underwent SMR based on a mathematical model of cell distribution and infiltration profile (tumor invasiveness profile). METHODS Volumetric measurements were obtained from the selected regions of interest from pre- and postoperative MRI studies of included patients. The tumor invasiveness profile (proliferation/diffusion [ρ/D] ratio) was calculated using the following formula: ρ/D ratio = (4π/3)2/3 × (6.106/[VT21/1 − VT11/1])2, where VT2 and VT1 are the preoperative FLAIR and contrast-enhancing volumes, respectively. Patients were split into subgroups based on their tumor invasiveness profiles. In this analysis, tumors were classified as nodular, moderately diffuse, or highly diffuse. RESULTS A total of 101 patients were included. Tumors were classified as nodular (n = 34), moderately diffuse (n = 34), and highly diffuse (n = 33). On multivariate analysis, increasing SMR had a significant positive correlation with OS for moderately and highly diffuse tumors (HR 0.99, 95% CI 0.98–0.99; p = 0.02; and HR 0.98, 95% CI 0.96–0.99; p = 0.04, respectively). On threshold analysis, OS benefit was seen with SMR from 10% to 29%, 10% to 59%, and 30% to 90%, for nodular, moderately diffuse, and highly diffuse, respectively. CONCLUSIONS The impact of SMR on OS for patients with IDH–wild-type GBM is influenced by the degree of tumor invasiveness. The authors’ results show that increasing SMR is associated with increased OS in patients with moderate and highly diffuse IDH–wild-type GBMs. When grouping SMR into 10% intervals, this benefit was seen for all tumor subgroups, although for nodular tumors, the maximum beneficial SMR percentage was considerably lower than in moderate and highly diffuse tumors., Ciencias Médicas y de la Salud

Glioblastoma (GBM) is the most common primary brain tumor in adults an carries and carries a terrible prognosis. The current regiment of surgical resection, radiation, and chemotherapy has remained largely unchanged in recent years as new therapeutic approaches have struggled to demonstrate benefit. One of the most challenging hurdles to overcome in developing novel treatments is the profound immune suppression found in many GBM patients. This limits the utility of all manner of immunotherapeutic agents, which have revolutionized the treatment of a number of cancers in recent years, but have failed to show similar benefit in GBM therapy. Understanding the mechanisms of tumor-mediated immune suppression in GBM is critical to the development of effective novel therapies, and reversal of this effect may prove key to effective immunotherapy for GBM. In this review, we discuss the current understanding of tumor-mediated immune suppression in GBM in both the local tumor microenvironment and systemically. We also discuss the effects of current GBM therapy on the immune system. We specifically explore some of the downstream effectors of tumor-driven immune suppression, particularly myeloid-derived suppressor cells (MDSCs) and other immunosuppressive monocytes, and the manner by which GBM induces their formation, with particular attention to the role of GBM-derived extracellular vesicles (EVs). Lastly, we briefly review the current state of immunotherapy for GBM and discuss additional hurdles to overcome identification and implementation of effective therapeutic strategies.

A 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.

OBJECTIVE The authors’ goal was to use a multicenter, observational cohort study to determine whether supramarginal resection (SMR) of FLAIR-hyperintense tumor beyond the contrast-enhanced (CE) area influences the overall survival (OS) of patients with isocitrate dehydrogenase–wild-type (IDH-wt) glioblastoma after gross-total resection (GTR). METHODS The medical records of 888 patients aged ≥ 18 years who underwent resection of GBM between January 2011 and December 2017 were reviewed. Volumetric measurements of the CE tumor and surrounding FLAIR-hyperintense tumor were performed, clinical variables were obtained, and associations with OS were analyzed. RESULTS In total, 101 patients with newly diagnosed IDH-wt GBM who underwent GTR of the CE tumor met the inclusion criteria. In multivariate analysis, age ≥ 65 years (HR 1.97; 95% CI 1.01–2.56; p < 0.001) and contact with the lateral ventricles (HR 1.59; 95% CI 1.13–1.78; p = 0.025) were associated with shorter OS, but preoperative Karnofsky Performance Status ≥ 70 (HR 0.47; 95% CI 0.27–0.89; p = 0.006), MGMT promotor methylation (HR 0.63; 95% CI 0.52–0.99; p = 0.044), and increased percentage of SMR (HR 0.99; 95% CI 0.98–0.99; p = 0.02) were associated with longer OS. Finally, 20% SMR was the minimum percentage associated with beneficial OS (HR 0.56; 95% CI 0.35–0.89; p = 0.01), but > 60% SMR had no significant influence (HR 0.74; 95% CI 0.45–1.21; p = 0.234). CONCLUSIONS SMR is associated with improved OS in patients with IDH-wt GBM who undergo GTR of CE tumor. At least 20% SMR of the CE tumor was associated with beneficial OS, but greater than 60% SMR had no significant influence on OS.

Objectives Inhibitors to the checkpoint proteins cytotoxic T‐lymphocyte‐associated protein 4 (CTLA‐4) and programmed cell death protein 1 (PD‐1) are becoming widely used in cancer treatment. However, a lack of understanding of the patient response to treatment limits accurate identification of potential responders to immunotherapy. Methods In this study, we assessed the expression of PD‐1 and CTLA‐4 on 19 leucocyte populations in the peripheral blood of 74 cancer patients. A reference data set for PD‐1 and CTLA‐4 was established for 40 healthy volunteers to determine the normal expression patterns for these checkpoint proteins. Results Unsupervised hierarchical clustering found four immune profiles shared across the solid tumor types, while chronic lymphocytic leukaemia patients had an immune profile largely unique to them. Furthermore, we measured these leucocyte populations on an additional cohort of 16 cancer patients receiving the PD‐1 inhibitor pembrolizumab in order to identify differences between responders and non‐responders, as well as compared to healthy volunteers (n = 20). We observed that cancer patients had pre‐treatment PD‐1 and CTLA‐4 expression on their leucocyte populations at different levels compared to healthy volunteers and identified two leucocyte populations positive for CTLA‐4 that had not been previously described. We found higher levels of PD‐1+ CD3+ CD4− CD8− cells in patients with progressive disease and have identified it as a potential biomarker of response, as well as identifying other significant differences in phenotypes between responders and non‐responders. Conclusion These results are suggestive that categorisation of patients based on immune profiles may differentiate responders from non‐responders to immunotherapy for solid tumors., Here, we have developed a multiparameter flow cytometry assay that allowed us to measure checkpoint proteins CTLA‐4 and PD‐1 on 19 leucocyte populations in the peripheral blood. By looking at the data independent of disease, we used unsupervised hierarchical clustering of PD‐1 expression on patients with different types of cancers and identified five patient profiles. These profiles, or groups of immunologically similar patients, can be used for predicting responders vs non‐responders and applied towards decisions in choosing appropriate inhibitors.

Background Antibody drug conjugates (ADCs) targeting the epidermal growth factor receptor (EGFR), such as depatuxizumab mafodotin (Depatux-M), is a promising therapeutic strategy for glioblastoma (GBM) but recent clinical trials did not demonstrate a survival benefit. Understanding the mechanisms of failure for this promising strategy is critically important. Methods PDX models were employed to study efficacy of systemic vs intracranial delivery of Depatux-M. Immunofluorescence and MALDI-MSI were performed to detect drug levels in the brain. EGFR levels and compensatory pathways were studied using quantitative flow cytometry, Western blots, RNAseq, FISH, and phosphoproteomics. Results Systemic delivery of Depatux-M was highly effective in nine of 10 EGFR-amplified heterotopic PDXs with survival extending beyond one year in eight PDXs. Acquired resistance in two PDXs (GBM12 and GBM46) was driven by suppression of EGFR expression or emergence of a novel short-variant of EGFR lacking the epitope for the Depatux-M antibody. In contrast to the profound benefit observed in heterotopic tumors, only two of seven intrinsically sensitive PDXs were responsive to Depatux-M as intracranial tumors. Poor efficacy in orthotopic PDXs was associated with limited and heterogeneous distribution of Depatux-M into tumor tissues, and artificial disruption of the BBB or bypass of the BBB by direct intracranial injection of Depatux-M into orthotopic tumors markedly enhanced the efficacy of drug treatment. Conclusions Despite profound intrinsic sensitivity to Depatux-M, limited drug delivery into brain tumor may have been a key contributor to lack of efficacy in recently failed clinical trials.

BACKGROUND Glioblastoma is a fatal infiltrative primary brain tumor, and standard care includes maximal safe surgical resection followed by radiation and Temozolomide (TMZ). Therapy-resistant residual cells persist in a latent state a long time before inevitable recurrence. Conventional radiation and Temozolomide (TMZ) treatment cause oxidative stress and DNA damage resulting senescent-like state of cell-cycle arrest. However, increasing evidence demonstrates escaping senescence leads to tumor recurrence. Thus, the ablation of senescent tumor cells after chemoradiation may be an avenue to limit tumor recurrence. METHODS 100uM TMZ for 7days or 10-20Gy radiation (cesium gamma radiator) was used for senescence induction in human glioblastoma in vitro and confirmed by SA-Beta gal staining and PCR. Replication arrest assessed by automated quantification of cellular confluence (Thermo Scientific Series 8000 WJ Incubator). We evaluated the IC50 for several senolytics targeting multiple SCAPs, including Dasatinib, Quercetin, AMG-232, Fisetin, Onalespib, Navitoclax, and A1331852, and in senescent vs. proliferating cells. RESULTS Among the senolytic tested, the Bcl-XL inhibitors A1331852 and Navitoclax both shown senolytic effect by selectively killing radiated, senescent tumor cells at lower concentrations as compared to 0Gy treated non-senescent cells. Across 12 GBM cell lines, IC50 for senescent cells was 6–500 times lower than non-senescent GBM(p< 0.005). Such differential sensitivity to Bcl-XL inhibition after radiation has also observed by BCL-XL knockdown in radiated glioma. CONCLUSION These findings suggest the potential to harness radiation-induced biology to ablate surviving quiescent cells and demonstrate Bcl-XL dependency as a potential vulnerability of surviving tumor cells after exposure to chemoradiation.

Immunosuppression of unknown etiology is a hallmark feature of glioblastoma (GBM) and is characterized by decreased CD4 T cell counts and down regulation of MHC class II expression on peripheral blood monocytes in patients. This immunosuppression is a critical barrier to the successful development of immunotherapies for GBM. We recapitulated the immunosuppression observed in GBM patients in the C57BL/6 mouse and investigated the etiology of low CD4 T cell counts. We determined that thymic involution was a hallmark feature of immunosuppression in three distinct models of CNS cancer, including mice harboring GL261 glioma, B16 melanoma, and in a spontaneous model of Diffuse Intrinsic Pontine Glioma (DIPG). In addition to thymic involution, we determined that tumor growth in the brain induced significant splenic involution, reductions in peripheral T cells, reduced MHC class II expression on hematopoietic cells, and a modest increase in bone marrow resident CD4 T cells with a naïve phenotype. Using parabiosis we report that thymic involution, declines in peripheral T cell counts, and reduced MHC class II expression levels were mediated through circulating blood-derived factors. Conversely, T cell sequestration in the bone marrow was not governed through circulating factors. Serum isolated from glioma-bearing mice potently inhibited proliferation and functions of T cells both in vitro and in vivo. Interestingly, the factor responsible for immunosuppression in serum is nonsteroidal and of high molecular weight. Through further analysis of neurological disease models, we determined that the aforementioned immunosuppression was not unique to cancer itself, but rather occurs in response to CNS injury. Noncancerous acute neurological insults also induced significant thymic involution and rendered serum immunosuppressive. Both thymic involution and serum-derived immunosuppression were reversible upon clearance of brain insults. These findings demonstrate that CNS cancers cause multifaceted immunosuppression and pinpoint circulating factors as a target of intervention to restore immunity.

BACKGROUND Glioblastoma (GBM) is an infiltrative, uniformly fatal brain tumor, treated with surgery, radiation, and Temozolomide (TMZ). Chemoradiation induces a senescent-like phenotype, which contributes to disease recurrence. We recently found that, radiated GBM cells can be eliminated by inhibition of BCL-XL. However, it remains unknown whether p53 is involved in this process and whether the elimination of senescent cells by BCL-XL inhibition could be augmented by MDM2 inhibition, a negative p53 regulator. METHODS p53-mutant (GBM6/GBM123) and p53-WT (GBM39/GBM76) human GBM cells were treated with 5Gy or TMZ following 48 hours of MDM2 inhibitor (AMG232) or vehicle treatment and maintained for seven days to establish a senescent-like phenotype. We evaluated the IC50 for BCL-XL inhibitor (A1331852) in radiated vs. non-radiated cells with or without MDM2 inhibitor pre-treatment. RESULTS MDM2-inhibitor treatment prior to radiation increased the expression of p21 and lead to increased cell death when combined with BCL-XL inhibition in p53-WT GBM cells. IC50 of BCL-XL inhibitor after prior MDM2 pretreatment and radiation in GBM76 was 4.5□M compared to 33.5□M, 18.1□M, and 32.3□M in vehicle without radiation, vehicle with radiation, and MDM2 inhibitor-alone treatment groups, respectively(p=0.0036). The IC50 of BCL-XL inhibitor without MDM2-inhibition in non-irradiated and radiated, as well as pre-administration of MDM2 inhibitor in non-irradiated and radiated GBM39 cells was 5618nM, 16117nM, 3926nM and 276.5nM, respectively(p=0.0003). Conversely, MDM2 co-inhibition with BCL-XL did not lead to increased rates of cell death in p53-mutant cell(p>0.05). CONCLUSION We previously identified that BCL-XL inhibition promotes cell death in senescent GBM cells. We build upon that work, demonstrating the increased rates of cell death can be augmented by MDM2 inhibition, but only in p53-WT cells. These findings highlight a novel therapeutic target for treating latent GBM tumors prior to recurrence, with the additional of MDM2 inhibition greatly increasing the efficacy of BCL-XL targeting agents.

Glioblastoma (GBM) is the most common and fatal primary brain tumor in adults. The development of novel therapies is critical, as little has changed regarding the standard of care in nearly two decades. Immunotherapy holds much promise, as treatments including chimeric antigen receptor (CAR) T cells and immune checkpoint blockade inhibitors have transformed the treatment of a number of cancers in recent years. However, GBM patients exhibit profound immunosuppression, limiting the efficacy of these therapies. Understanding the mechanisms of GBM-mediated immunosuppression is critical to overcoming this barrier. GBM-derived extracellular vesicles (EVs) have been shown to mediate the induction of immunosuppressive monocytes, which may point to a mechanism of immunosuppression. EVs make initial contact with target cells through interactions between heparan sulfate proteoglycans, and soluble heparin has been shown to inhibit these interactions in some models. We demonstrate that soluble heparin inhibits the binding of GBM-derived EVs to monocytes in a dose-dependent manner, and that heparin treatment reduces the induction of immunosuppressive monocytes upon in vitro conditioning of monocytes with GBM-derived EVs (p< 0.01). Further, we demonstrate that heparin treated EV-conditioned monocytes are functionally less immunosuppressive than untreated EV-conditioned monocytes as measured by T cell proliferation in co-culture studies (p< 0.05). Taken together, these findings underscore the import of tumor-derived EVs in immunosuppression in GBM, and demonstrate the feasibility of targeting EV-monocyte interactions in treating GBM-mediated immunosuppression.

Background Patterns of recurrence and survival with different surgical and radiotherapy (RT) techniques were evaluated to guide RT target volumes for patients with temporal lobe glioma. Methods and Materials This retrospective cohort study included patients with World Health Organization grades II to IV temporal lobe glioma treated with either partial (PTL) or complete temporal lobectomy (CTL) followed by RT covering both the parenchymal and dural resection bed (whole-cavity radiotherapy [WCRT]) or the parenchymal resection bed only (partial-cavity radiotherapy [PCRT]). Patterns of recurrence, progression-free survival (PFS) and overall survival (OS) were evaluated. Results Fifty-one patients were included and 84.3% of patients had high-grade glioma (HGG). CTL and PTL were performed for 11 (21.6%) and 40 (78.4%) patients, respectively. Median RT dose was 60 Gy (range, 40-76 Gy). There were 82.4% and 17.6% of patients who received WCRT and PCRT, respectively. Median follow-up time was 18.4 months (range, 4-161 months). Forty-six patients (90.2%) experienced disease recurrence, most commonly at the parenchymal resection bed (76.5%). No patients experienced an isolated dural recurrence. The median PFS and OS for the PCRT and WCRT cohorts were 8.6 vs 10.8 months (P = .979) and 19.9 vs 18.6 months (P = .859), respectively. PCRT was associated with a lower RT dose to the brainstem, optic, and ocular structures, hippocampus, and pituitary. Conclusion We identified no isolated dural recurrence and similar PFS and OS regardless of postoperative RT volume, whereas PCRT was associated with dose reduction to critical structures. Omission of dural RT may be considered a reasonable alternative approach. Further validation with larger comparative studies is warranted.

Several studies have linked codeletion of chromosome arms 1p/19q in low-grade gliomas (LGG) with positive response to treatment and longer progression-free survival. Hence, predicting 1p/19q status is crucial for effective treatment planning of LGG. In this study, we predict the 1p/19q status from MR images using convolutional neural networks (CNN), which could be a non-invasive alternative to surgical biopsy and histopathological analysis. Our method consists of three main steps: image registration, tumor segmentation, and classification of 1p/19q status using CNN. We included a total of 159 LGG with 3 image slices each who had biopsy-proven 1p/19q status (57 non-deleted and 102 codeleted) and preoperative postcontrast-T1 (T1C) and T2 images. We divided our data into training, validation, and test sets. The training data was balanced for equal class probability and was then augmented with iterations of random translational shift, rotation, and horizontal and vertical flips to increase the size of the training set. We shuffled and augmented the training data to counter overfitting in each epoch. Finally, we evaluated several configurations of a multi-scale CNN architecture until training and validation accuracies became consistent. The results of the best performing configuration on the unseen test set were 93.3% (sensitivity), 82.22% (specificity), and 87.7% (accuracy). Multi-scale CNN with their self-learning capability provides promising results for predicting 1p/19q status non-invasively based on T1C and T2 images. Predicting 1p/19q status non-invasively from MR images would allow selecting effective treatment strategies for LGG patients without the need for surgical biopsy.

Background This study retrospectively analyzed outcomes for patients undergoing gamma knife radiosurgery (GKR) for uveal melanoma (UM) and intraocular metastases. Methods Patients who underwent GKR for UM or intraocular metastases between 1/1/1990 and 6/1/2015 at Mayo Clinic, Rochester, MN, USA, were retrospectively analyzed. Results Eleven patients (11 eyes) had UM while seven patients (7 eyes) had intraocular metastases. Patients with UM were followed for a median of 19.74 ± 10.4 months. Visual acuity (VA) logMAR 0.30 ± 0.53 (Snellen 20/40) versus 0.40 ± 0.97 (Snellen 20/50), tumor thickness (5.30 ± 2.17 vs. 3.60 ± 2.32 mm), were not significantly different between preoperative and postoperative measurements, respectively. Nine percent (1/11) patients required enucleation. Subsequently, no patients experienced metastases. Patients with intraocular metastases were followed for a median of 6.03 ± 6.32 months. They did not have significant changes in VA (logMAR 0.30 ± 0.59 vs. 0.30 ± 1.57; Snellen 20/40 vs. 20/40) or tumor thickness (3.50 ± 1.36 vs. 1.30 ± 0.76 mm) postoperatively. Fourteen percent (1/7 patients) required enucleation. Complications experienced by patients with UM include radiation retinopathy (2/11), papillopathy (1/11), cystoid macular edema (1/11), vitreomacular traction (1/11), exudative retinal detachment (1/11). Patients with metastases had treatment complicated by recurrence (2/7). Dose to the margin, maximum dose of radiation, and clinical target volume did not correlate with post-procedural VA, risk of enucleation, or death in patients with either UM or patients with intraocular metastases. Conclusions Visual outcomes were satisfactory for patients undergoing GKR without significant morbidity and without significant risk of enucleation or metastases.

Purpose Whole brain radiation therapy (WBRT) remains a commonly used cancer treatment, although controversy exists regarding the optimal dose/fractionation to optimize intracranial tumor control and minimize resultant cognitive deficits. Methods and Materials NCCTG N107C [Alliance]/CEC.3 randomized 194 patients with brain metastases to either stereotactic radiosurgery alone or WBRT after surgical resection. Among the 92 patients receiving WBRT, sites predetermined the dose/fractionation that would be used for all patients treated at that site (either 30 Gy in 10 fractions or 37.5 Gy in 15 fractions). Analyses were performed using Kaplan-Meier estimates, log rank tests, and Fisher’s exact tests. Results Among 92 patients treated with surgical resection and adjuvant WBRT, 49 were treated with 30 Gy in 10 fractions (53%), and 43 were treated with 37.5 Gy in 15 fractions (47%). Baseline characteristics, including cognitive testing, were well balanced between groups with the exception of primary tumor type (lung cancer histology was more frequent with protracted WBRT: 72% vs 45%, P = .01), and 93% of patients completed the full course of WBRT. A more protracted WBRT dose regimen (37.5 Gy in 15 fractions) did not significantly affect time to cognitive failure (hazard ratio [HR], 0.9; 95% confidence interval [CI], 0.6-1.39; P = .66), surgical bed control (HR, 0.52 [95% CI, 0.22-1.25], P = .14), intracranial tumor control (HR, 0.56 [95% CI, 0.28-1.12], P = .09), or overall survival (HR, 0.72 [95% CI, 0.45-1.16], P = .18). Although there was no reported radionecrosis, there is a statistically significant increase in the risk of at least 1 grade ≥3 adverse event with 37.5 Gy in 15 fractions versus 30 Gy in 10 fractions (54% vs 31%, respectively, P = .03). Conclusions This post hoc analysis does not demonstrate that protracted WBRT courses reduce the risk of cognitive deficit, improve tumor control in the hypoxic surgical cavity, or otherwise improve the therapeutic ratio. Adverse events were significantly higher with the lengthened course of WBRT. For patients with brain metastases where WBRT is recommended, shorter course hypofractionated regimens remain the current standard of care.

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.

Central nervous system hemangioblastoma (CNS HGB) is a rare neoplasm. Sporadic and von-Hippel Lindau (VHL)-associated forms exist. Predicting tumor growth is difficult, making the indication for intervention challenging. A retrospective analysis of consecutive, neurosurgically managed CNS HGB at Mayo Clinic spanning 1988–2018 was performed to clarify their characteristics. 117 sporadic and 67 VHL HGBs were reviewed. 9 sporadic lesions (8 cases) and 80 VHL lesions (35 cases) had tumor size data at ≥4 follow-up time-points beyond 1 year. The median follow-up was 8.0 years. No statistical difference existed between sporadic and VHL lesions regarding growth speed or growth latency (period of time without growth). Symptomatic lesions showed faster growth than asymptomatic lesions (2.79 vs 0.59 cm3/month, p=0.005). Solid and cystic portions often showed different growth patterns in cases harboring both (66.7%). Cyst growth was faster than solid portions (1.22 vs 1.05cm3/month, p=0.009) and growth latency was longer in the latter (15.2 vs 18.5%, p=0.25). Lesions’ growth were classified as linear (7), exponential (49), cubic (regressed to cubic function curve, 7), saltatory (19) and no-growth (7) patterns. Growth latency often existed in growing lesions (27-out-of-64 linear/exponential/cubic lesions, 57.8%), being average 8.3% of follow-up time. Contrarily, although saltatory lesions grew slower than exponential lesions (p=0.03), their overall growth speed was comparable to linear/cubic lesions (0.67cm3/month), and 9-out-of-19 showed growth surges at their ends. Growth speed at each follow-up roughly depended on the tumor size in cubic regression (R2=0.52). However, whereas total 18/821 time-point lesions showed rapid growth (≥1cm3/m, 2.2%), relatively small lesions (≤2cm3) also occasionally showed rapid growth (5/687, 0.73%). Not all large (>5cm3) lesions grew rapidly (8/62, 12.9%). These results revealed while symptomatic/large/cystic lesions tend to grow more rapidly, HGB behavior is difficult to predict, which stresses the importance of careful follow-up.

Patients with glioblastoma (GBM) have a median survival of 15 months despite aggressive treatment. Immunotherapies such as dendritic cell (DC) vaccines have modest clinical efficacy in small clinical trials. Treatment-related pseudo-progression confounds outcome assessment by MRI, particularly in patients receiving immunotherapy. Thus, there is a need for additional non-invasive methods to monitor treatment response. Extracellular vesicles (EVs), especially plasma exosomes, contain tumor-specific microRNA (miRNA) cargo that could serve as a liquid biopsy to distinguish true progression from treatment-related pseudo-progression. Plasma exosomes were isolated by serial density gradient ultracentrifugation from 20 newly diagnosed GBM patients enrolled in a clinical trial of allogeneic tumor lysate-pulsed autologous DC vaccination. Short non-coding RNA sequencing and bioinformatics analysis was performed for each patient at three time points (TP): pre-vaccine (TP1), post-initial vaccine (TP2), and at end of treatment (TP3). miRNA expression analysis revealed a total of 14 upregulated and 12 downregulated miRNAs across time points (p-value < 0.05, |logFC| >1), few of which have been previously reported to be differentially expressed in GBM. Interestingly, patients’ miRNA profile expression differed more at the beginning of treatment (e.g. TP1-vs-TP2) and at subsequent time points (e.g. TP2-vs-TP3). Ingenuity Pathway Analysis is in progress to identify pathways associated with immunotherapy treatment response in malignant gliomas. In conclusion, miRNA sequencing from GBM patients’ plasma exosomes enrolled in our DC clinical trial shows marked differential miRNA expression between time points. These results suggest that as patients progress through treatment, consistent differences in plasma exosomal miRNA expression profile can be identified that could be utilized as predictors of treatment response. Thus, plasma EVs may serve as a robust platform to monitor treatment outcome.

OBJECTIVES Dogma suggests that for brain tumors, regions of enhancement on T1-weighted gadolinium contrast enhanced (T1Gd) magnetic resonance imaging (MRI) correlate with intravenously delivered drug distribution as enhancement indicates a compromised blood-brain barrier (BBB). However, poor response to intravenous therapies highlights the importance of the diffuse disease beyond enhancing regions. This study investigated whether imaging features can provide an accurate prediction of drug distribution. METHODS Eight brain tumor patients (7 gliomas and 1 metastatic adenocarcinoma) were included in this Phase 0 trial. Presurgery T1-weighted, T1Gd, T2-weighted gadolinium contrast enhanced (T2Gd), and T2-weighted Fluid Attenuated Inversion Recovery (T2FLAIR) MRIs were acquired. All images underwent bias correction using the N4 algorithm, standardization of intensities, and registration. Prior to incision, patients received both an antibiotic cefazolin (6% BBB penetrance) and levetiracetam (80% BBB penetrance), an anti-seizure drug. Subsequently, multiple blood samples and image-guided biopsies were taken and analyzed for drug concentration using liquid chromatography mass spectrometry. Biopsy drug levels are reported as Brain-Plasma Ratio (BPR), the ratio of biopsy concentration relative to plasma concentration. Mean image intensity was extracted from an 8x8 mm window surrounding each biopsy location. Regression analysis was performed to determine which combination of image types were linearly predictive of BPR for both drugs. Correlations were also analyzed according to the biopsy location radiographic appearance. RESULTS Regression analysis revealed that T2Gd intensity was linearly predictive of cefazolin BPR and FLAIR intensity was linearly predictive of levetiracetam BPR (p=0.009 and 0.041, respectively). Grouping samples according the the radiographic appearance revealed that levetiracetam BPR had a similar pattern of values to that of FLAIR intensities and cefazolin BPR had a similar pattern to T2, further supporting the regression analysis results. CONCLUSIONS Local concentrations of drug may be related to T2-weighted signals (T2Gd and T2FLAIR) rather than the gadolinium distribution on T1Gd images.

Glioblastoma is the most common adult primary brain tumor and carries a dismal prognosis. Radiation is a standard first-line therapy, typically deployed following maximal safe surgical debulking, when possible, in combination with cytotoxic chemotherapy. For other systemic cancers, standard of care is being transformed by immunotherapies, including checkpoint-blocking antibodies targeting CTLA-4 and PD-1/PD-L1, with potential for long-term remission. Ongoing studies are evaluating the role of immunotherapies for GBM. Despite dramatic responses in some cases, randomized trials to date have not met primary outcomes. Challenges have been attributed in part to the immunologically "cold" nature of glioblastoma relative to other malignancies successfully treated with immunotherapy. Radiation may serve as a mechanism to improve tumor immunogenicity. In this review, we critically evaluate current evidence regarding radiation as a synergistic facilitator of immunotherapies through modulation of both the innate and adaptive immune milieu. Although current preclinical data encourage efforts to harness synergistic biology between radiation and immunotherapy, several practical and scientific challenges remain. Moreover, insights from radiation biology may unveil additional novel opportunities to help mobilize immunity against GBM.

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 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.

Stereotactic radiosurgery (SRS) is increasingly utilized to treat the resection cavity following resection of brain metastases and recent randomized trials have confirmed postoperative SRS as a standard of care. Postoperative SRS for resected brain metastases improves local control compared to observation, while also preserving neurocognitive function in comparison to whole brain radiation therapy (WBRT). However, even with surgery and SRS, rates of local recurrence at 1 year may be as high as 40%, especially for larger cavities, and there is also a known risk of leptomeningeal disease after surgery. Additional treatment strategies are needed to improve control while maintaining or decreasing the toxicity profile associated with treatment. Preoperative SRS is discussed here as one such approach. Preoperative SRS allows for contouring of an intact metastasis, as opposed to an irregularly shaped surgical cavity in the post-op setting. Delivering SRS prior to surgery may also allow for a "sterilizing" effect, with the potential to increase tumor control by decreasing intra-operative seeding of viable tumor cells beyond the treated cavity, and decreasing risk of leptomeningeal disease. Because there is no need to treat brain surrounding tumor in the preoperative setting, and since the majority of the high dose volume can then be resected at surgery, the rate of symptomatic radiation necrosis may also be reduced with preoperative SRS. In this mini review, we explore the potential benefits and risks of preoperative vs. postoperative SRS for brain metastases as well as the existing literature to date, including published outcomes with preoperative SRS.

Recent studies have highlighted the importance of using molecular biomarkers (IDH, 1p19q, TERT) to group gliomas that have similar clinical behavior, response to therapy, and outcome. An emerging hypothesis is that glioma specific genetic and/or molecular alterations manifest as specific observable changes in MR anatomical imaging. Morphologic and texture features, originating from anatomical MRI, have been investigated as imaging biomarkers to predict MGMT and glioma group status. These texture or morphologic based approaches pose several challenges including requirements for several preprocessing steps such as intensity standardization, skull stripping, and tumor segmentation. Deep learning is an important evolving technology in many different fields, including anatomic imaging, and can be used to empirically identify important features in a variety of modalities, including MRI. Importantly deep learning precludes the need for extensive pre-processing. We describe a convolutional neural network, evaluating resnet50, vgg16, inception and xception neural network architectures, that can predict IDH, 1p19q and TERT status utilizing conventional T2 weighted MRI imaging with intensity normalization and nonuniform intensity normalization (N4) bias corrections. The dataset consisted of 432 images (340 for training and 92 for validation) from patients published by Eckel-Passow et al New England Journal of Medicine (2015). The system achieved a weighted f1 score of 0.901, 0.937 and 0.924 for IDH, 1p19q and TERT prediction on the test dataset, respectively. IDH status was misclassified in 9 out of 92 patients, while 1p19q and TERT status was misclassified in 6 and 7 patients respectively. Our results demonstrate the potential of deep learning architectures applied to conventional MRI to predict molecular glioma groups.

Gliomas including glioblastoma (GBM) are the most common malignant brain tumors. Glioma extracellular vesicles (EVs), especially plasma exosomes, have biological effects such as mediating immunosuppression and contain signature tumor-specific cargo that could serve as liquid biopsies. Increasing interest in molecular biomarkers to determine patient prognosis in GBM has suggested that EV miRNA-based signatures may be able to predict progression-free and overall survival, differentiate normal donors from GBM patients, and distinguish true progression from treatment-related pseudo-progression. We have established a simple two-step technique, using density gradient ultracentrifugation (DGU), to isolate plasma exosomes from glioma patients and normal donors. One-step DGU (90 minutes/24,000 RPM) efficiently isolated exosomes from plasma, while a second DGU (16 hours/24,000 RPM) efficiently concentrated exosomes for subsequent cargo analysis, including miRNA signature analysis. Purification of total RNA, including miRNA, was performed on plasma exosomes from normal donors (n=8) and GBM patients (n=8) using the miRNeasy kit (Qiagen). Next generation short non-coding RNA sequencing samples was performed by Illumina HiSeq 2000 and revealed many differentially expressed miRNAs in GBM patients with high fold change/low false discovery rates compared to normal donor plasma exosomes. In order to test the diagnostic accuracy of the proposed technique for the differentiation of GBM patients and normal donors, ROC analysis was performed based on the top 30 differentially expressed miRNA samples. The area under the ROC curve (AUC; a figure of merit to determine the optimal miRNA signature) was 0.968. In addition, multiple novel miRNAs and other short non-coding RNA species (Y-RNA, piRNA, snoRNA) were found with some differential expression. In conclusion, miRNA sequencing from plasma exosomes shows marked differential miRNA expression between healthy donors and GBM patients. These findings as well as additional differentially expressed short non-coding RNA species suggest plasma EVs may serve as a robust platform to develop GBM liquid biopsies.

Patient derived xenograft (PDX) models have shown great utility for pre-clinical and translational studies for a range of malignancies. We have established a comprehensive, publically available collection of 95 high-grade glioma flank PDX models. Viable PDX were derived from Glioblastoma, IDH-wildtype (n=91), Glioblastoma, IDH-mutant (n=2), Diffuse Midline Glioma, H3 K27M-mutant (n=1), and Anaplastic Oligodendroglioma (n=1) and include both primary (n=60) and recurrent (n=35) tumors. Comprehensive molecular characterization of PDX is ongoing and, to date, has included whole exome sequencing (WES, n=82), RNA-sequencing (n=40), and genome-wide methylation profiling (n=78) that included MGMT promoter, with data available in cBioPortal. PDX reflected the genetic characteristics of glioblastoma, with the majority harboring TERT promoter mutations, chromosomal gain +7, loss -10 and homozygous deletion of CKDN2A/B. EGFR alterations were frequent (~40%) and included amplification, point mutation, EGFRvIII, and other splice variants. Other common alterations, including amplifications of MET, CDK4, CDK6, MDM2, MDM4 and mutation of TP53, PTEN, NF1, RB1, PIK3CA, PIK3R1 were present at similar frequencies reported by TCGA, with the exception of PDGFRA alterations that were underrepresented in PDX. RNA-sequencing showed representation of each of the glioblastoma gene expression subtypes. To assess preservation of genetic features during xeongrafting, we performed WES on 20 matched patient tumors. The vast majority of genetic driver alterations were shared between patient and PDX, including EGFR, EGFRvIII, CDK4, MDM2, and MET amplifications. In 3 PDX, subclonal selection events were observed, including amplifications of MYCN, CDK6 and an EGFR splice variant, as well as selection against PDGFRA amplification. Overall, our large panel of PDX models reflects the genetic heterogeneity of glioblastoma and largely preserves the genetic features of the primary patient tumors. The PDX National Resource is a powerful tool for neuro-oncology research, with all PDX models and genomic data openly available (http://www.mayo.edu/research/labs/translational-neuro-oncology/mayo-clinic-brain-tumor-patient-derived-xenograft-national-resource).

Background Attenuated measles virus (MV) strains are promising agents currently being tested against solid tumors or hematologic malignancies in ongoing phase I and II clinical trials; factors determining oncolytic virotherapy success remain poorly understood, however. Methods We performed RNA sequencing and gene set enrichment analysis to identify pathways differentially activated in MV-resistant (n = 3) and -permissive (n = 2) tumors derived from resected human glioblastoma (GBM) specimens and propagated as xenografts (PDX). Using a unique gene signature we identified, we generated a diagonal linear discriminant analysis (DLDA) classification algorithm to predict MV responders and nonresponders, which was validated in additional randomly selected GBM and ovarian cancer PDX and 10 GBM patients treated with MV in a phase I trial. GBM PDX lines were also treated with the US Food and Drug Administration-approved JAK inhibitor, ruxolitinib, for 48 hours prior to MV infection and virus production, STAT1/3 signaling and interferon stimulated gene expression was assessed. All statistical tests were two-sided. Results Constitutive interferon pathway activation, as reflected in the DLDA algorithm, was identified as the key determinant for MV replication, independent of virus receptor expression, in MV-permissive and -resistant GBM PDXs. Using these lines as the training data for the DLDA algorithm, we confirmed the accuracy of our algorithm in predicting MV response in randomly selected GBM PDX ovarian cancer PDXs. Using the DLDA prediction algorithm, we demonstrate that virus replication in patient tumors is inversely correlated with expression of this resistance gene signature (ρ = -0.717, P = .03). In vitro inhibition of the interferon response pathway with the JAK inhibitor ruxolitinib was able to overcome resistance and increase virus production (1000-fold, P = .03) in GBM PDX lines. Conclusions These findings document a key mechanism of tumor resistance to oncolytic MV therapy and describe for the first time the development of a prediction algorithm to preselect for oncolytic treatment or combinatorial strategies.

Treatment-related changes can be difficult to differentiate from progressive glioma using MRI with contrast (CE). The purpose of this study is to compare the sensitivity and specificity of 18F-DOPA-PET and MRI in patients with recurrent glioma. Thirteen patients with MRI findings suspicious for recurrent glioma were prospectively enrolled and underwent 18F-DOPA-PET and MRI for neurosurgical planning. Stereotactic biopsies were obtained from regions of concordant and discordant PET and MRI CE, all within regions of T2/FLAIR signal hyperintensity. The sensitivity and specificity of 18F-DOPA-PET and CE were calculated based on histopathologic analysis. Receiver operating characteristic curve analysis revealed optimal tumor to normal (T/N) and SUVmax thresholds. In the 37 specimens obtained, 51% exhibited MRI contrast enhancement (M+) and 78% demonstrated 18F-DOPA-PET avidity (P+). Imaging characteristics included M−P− in 16%, M−P+ in 32%, M+P+ in 46% and M+P− in 5%. Histopathologic review of biopsies revealed grade II components in 16%, grade III in 43%, grade IV in 30% and no tumor in 11%. MRI CE sensitivity for recurrent tumor was 52% and specificity was 50%. PET sensitivity for tumor was 82% and specificity was 50%. A T/N threshold > 2.0 altered sensitivity to 76% and specificity to 100% and SUVmax > 1.36 improved sensitivity and specificity to 94 and 75%, respectively. 18F-DOPA-PET can provide increased sensitivity and specificity compared with MRI CE for visualizing the spatial distribution of recurrent gliomas. Future studies will incorporate 18F-DOPA-PET into re-irradiation target volume delineation for RT planning.

Temozolomide (TMZ) was used for the treatment of glioblastoma (GBM) for over a decade, but its treatment benefits are limited by acquired resistance, a process that remains incompletely understood. Here we report that an enhancer, located between the promoters of marker of proliferation Ki67 (MKI67) and O6-methylguanine-DNA-methyltransferase (MGMT) genes, is activated in TMZ-resistant patient-derived xenograft (PDX) lines and recurrent tumor samples. Activation of the enhancer correlates with increased MGMT expression, a major known mechanism for TMZ resistance. We show that forced activation of the enhancer in cell lines with low MGMT expression results in elevated MGMT expression. Deletion of this enhancer in cell lines with high MGMT expression leads to a dramatic reduction of MGMT and a lesser extent of Ki67 expression, increased TMZ sensitivity, and impaired proliferation. Together, these studies uncover a mechanism that regulates MGMT expression, confers TMZ resistance, and potentially regulates tumor proliferation., Temozolomide (TMZ) resistance in glioblastomas (GBM) is associated with increased MGMT expression. Here, the authors identify an enhancer between the promoters of MKI67 and MGMT, that when activated drives MGMT expression despite MGMT promoter methylation to confer TMZ resistance in GBM.

Glioblastoma (GBM) is the most common brain tumor and is nearly universally fatal within five years despite aggressive treatment. Tissue histopathology is required for definitive diagnosis. Imaging alone is inadequate to distinguish tumor progression from treatment-related inflammatory pseudoprogression, particularly following immunotherapy. Therefore, there is a need for non-invasive diagnostic alternatives. GBM’s release extracellular vesicles (EV’s) including exosomes (

Glioblastoma is the most common primary tumor of the brain and has few long-term survivors. The local and systemic immunosuppressive environment created by glioblastoma allows it to evade immunosurveillance. Myeloid-derived suppressor cells (MDSCs) are a critical component of this immunosuppression. Understanding mechanisms of MDSC formation and function are key to developing effective immunotherapies. In this study, we developed a novel model to reliably generate human MDSCs from healthy-donor CD14+ monocytes by culture in human glioma-conditioned media. Monocytic MDSC frequency was assessed by flow cytometry and confocal microscopy. The resulting MDSCs robustly inhibited T cell proliferation. A cytokine array identified multiple components of the GCM potentially contributing to MDSC generation, including Monocyte Chemoattractive Protein-1, interleukin-6, interleukin-8, and Macrophage Migration Inhibitory Factor (MIF). Of these, Macrophage Migration Inhibitory Factor is a particularly attractive therapeutic target as sulforaphane, a naturally occurring MIF inhibitor derived from broccoli sprouts, has excellent oral bioavailability. Sulforaphane inhibits the transformation of normal monocytes to MDSCs by glioma-conditioned media in vitro at pharmacologically relevant concentrations that are non-toxic to normal leukocytes. This is associated with a corresponding increase in mature dendritic cells. Interestingly, sulforaphane treatment had similar pro-inflammatory effects on normal monocytes in fresh media but specifically increased immature dendritic cells. Thus, we have used a simple in vitro model system to identify a novel contributor to glioblastoma immunosuppression for which a natural inhibitor exists that increases mature dendritic cell development at the expense of myeloid-derived suppressor cells when normal monocytes are exposed to glioma conditioned media.

BACKGROUND AND PURPOSE: In 2016, the World Health Organization revised the brain tumor classification, making IDH mutation and 1p/19q codeletion the defining features of oligodendroglioma. To determine whether imaging characteristics previously associated with oligodendroglial tumors are still applicable, we evaluated the MR imaging features of genetically defined oligodendrogliomas. MATERIALSANDMETHODS: One hundred forty-eight adult patients with untreated World Health Organization grade II and III infiltrating gliomas with histologic oligodendroglial morphology, known 1p/19q status, and at least 1 preoperative MR imaging were retrospectively identified. The association of 1p/19q codeletion with tumor imaging characteristics and ADC values was evaluated. RESULTS: Ninety of 148 (61%) patients had 1p/19q codeleted tumors, corresponding to genetically defined oligodendroglioma, and 58/148 (39%) did not show 1p/19q codeletion, corresponding to astrocytic tumors. Eighty-three of 90 (92%) genetically defined oligodendrogliomas had noncircumscribed borders, compared with 26/58 (45%) non-1p/19q codeleted tumors with at least partial histologic oligodendroglial morphology (P < .0001). Eighty-nine of 90 (99%) oligodendrogliomas were heterogeneous on T1- and/or T2-weighted imaging. In patients with available ADC values, a lower mean ADC value predicted 1p/19q codeletion (P = .0005). CONCLUSIONS: Imaging characteristics of World Health Organization 2016 genetically defined oligodendrogliomas differ from the previously considered characteristics of morphologically defined oligodendrogliomas. We found that genetically defined oligodendrogliomas were commonly poorly circumscribed and were almost always heterogeneous in signal intensity.

Glioblastoma (GBM) is among the most invasive and lethal of cancers, frequently infiltrating surrounding healthy tissue and giving rise to rapid recurrence. It is therefore critical to establish experimental model systems and develop therapeutic approaches that enhance anti-tumor immunity. In the current study, we have employed a newly developed murine glioma model to assess the efficacy of a novel picornavirus vaccination approach for the treatment of established tumors. The GL261-Quad system is a variation of the GL261 syngeneic glioma that has been engineered to expresses model T cell epitopes including OVA257–264. MRI revealed that both GL261 and GL261-Quad tumors display characteristic features of human gliomas such as heterogeneous gadolinium leakage and larger T2 weighted volumes. Analysis of brain-infiltrating immune cells demonstrated that GL261-Quad gliomas generate detectable CD8+ T cell responses toward the tumor-specific Kb:OVA257–264 antigen. Enhancing this response via a single intracranial or peripheral vaccination with picornavirus expressing the OVA257–264 antigen increased anti-tumor CD8+ T cells infiltrating the brain, attenuated progression of established tumors, and extended survival of treated mice. Importantly, the efficacy of the picornavirus vaccination is dependent on functional cytotoxic activity of CD8+ T cells, as the beneficial response was completely abrogated in mice lacking perforin expression. Therefore, we have developed a novel system for evaluating mechanisms of anti-tumor immunity in vivo, incorporating the GL261-Quad model, 3D volumetric MRI, and picornavirus vaccination to enhance tumor-specific cytotoxic CD8+ T cell responses and track their effectiveness at eradicating established gliomas in vivo.

BACKGROUND: Dendritic cell (DC) vaccines for glioblastoma (GBM) are promising but significant conceptual shortcomings may have limited their clinical efficacy. First, most trials have not employed optimal DC culture techniques resulting in large numbers of immature (immunosuppressive) DC's. Second, most have used autologous tumor lysate. While highly personalized, this limits vaccine availability and precludes antigen-specific response testing. Finally, GBM-mediated immunosuppression has been largely unaddressed. We have developed a novel DC vaccine protocol that addresses these issues. METHODS: The ability of standard DC culture techniques to produce mature (CD83+) DC's from healthy donors' and GBM patients' CD14+ monocytes was assessed. An optimized technique using GBM patients' monocytes was developed. A library of human Good Manufacturing Practices (GMP) GBM cell lines was established using a novel platelet lysate supplement. Immature/mature glioneuronal markers and tumor-associated antigens (TAA's) were assessed by western blot. Finally, multicolor FACS immunophenotyping of 120 leukocyte surface markers in 5mL whole blood was established. RESULTS: Standard culture techniques produce >90% mature (CD83+) DC's from healthy donor monocytes but < 60% CD83+ DC's from GBM patients' monocytes (P90% mature DC from GBM patients. Seven GMP-compatible GBM cell lines have been developed with clonigenic karyotypic abnormalities, immature (nestin) and some mature (GFAP, olig2, β-tubulin) glioneuronal marker expression, and expression of multiple TAA's. Our 120 marker panel produces comprehensive, reproducible immunophenotyping. CONCLUSION: We have developed a DC vaccination strategy that optimizes DC maturity, utilizes renewable GBM cell lines with established TAA expression profiles, and allows comprehensive immunophenotyping to identify responding/non-responding patient populations. Based on these studies, we have initiated a phase I clinical trial (MC1272; IND#15233) of autologous DC / allogeneic tumor lysate in newly diagnosed adult GBM patients that enables multiple vaccinations in most GBM patients with extensive immunophenotyping and TAA-specific response testing.

The role of Gamma Knife stereotactic radiosurgery (GK-SRS) as a treatment option for patients with primary or recurrent high grade gliomas (HGG) is not well defined. To better understand clinical outcomes with GK-SRS we reviewed the charts of 94 consecutive patients treated for primary (n = 10) or recurrent (n = 84) WHO grade III/IV astrocytomas (n = 91), oligodendrogliomas (n = 2), or mixed (n = 1) lesions. Treatment prior to GK-SRS included External beam radiotherapy (EBRT) (median dose, 60Gy) in 90% and chemotherapy in 87%. Overall survival (OS) was estimated using the Kaplan-Meier method. Univariate and multivariate analyses (MVA) were performed using Cox proportional hazards regression models incorporating age, Karnofsky performance status (KPS) prior to GK-SRS (100 vs. 18mos), and tumor volume (≤12,000, >12,000). Median age at treatment was 49.6yrs (range, 18.4-77.2). KPS was ≥ 70 for all patients. Median follow-up was 0.9 years (range, 0.15-21.5). Tumor grade at diagnosis was II (10), III (18), or IV (66), with several progressions of II-to-III (9), II-to-IV (1), and III-to-IV (4) at time of GK-SRS. Median treatment volume was 12,800 mm3 (range, 484-139,000) with a median prescription dose of 15 Gy (50% isodose; range, 8-24). Median survival was 4.8 years for grade III (initial grade III: 3.9yrs; grade II-to-III progression: 5.6yrs) and 1.8 years for grade IV (initial grade IV 1.8yrs; grade II/III-to-IV progression: 4.2yrs). Predictors of OS on MVA include age at GK-SRS treatment per year (p = 0.0014, HR 1.033, 95% CI 1.012-1.053), grade IV at GK-SRS (p = 0.0078, HR 2.218, 95% CI 1.233-3.990), and interval EBRT to GK-SRS >18 months (p = 0.0011, HR 2.909, 95% CI 1.531-5.528). Our results show favorable survival outcomes with the use of GK-SRS compared to historic controls. Further prospective investigation is warranted.

OBJECTIVE:: To assess the survival and role of adjuvant chemotherapy in adult medulloblastoma. METHODS:: We reviewed outcomes of 66 patients (aged 18 y or more; median age, 33 y) with medulloblastoma. Forty-four (67%) patients had M0 disease, 9 had M1-M4, and 13 had MX. Thirty-one patients each for whom risk stratification was available were classified as high risk or standard risk. Fifty-six patients had histologic results: classic histology was the most common [n=46 (84%)], followed by desmoplastic (n=9), and large cell/anaplastic (n=1). Overall survival (OS) and progression-free survival (PFS) were estimated with Kaplan-Meier curves and log-rank tests. Cox regression analysis was used to compare recurrences. RESULTS:: Median follow-up was 6.7 years. The estimated 5-year OS and PFS were 74% and 59%, respectively. High-risk versus standard-risk classification was ass ociated with worse OS (61% vs. 86%; P=0.03) and recurrence (hazard ratio, 2.56; P=0.05) and a tren d for worse PFS (49% vs. 69%; P=0.13). Gross total resection was associated with improved OS (P=0.03) and a trend toward improved PFS (P=0.09). No chemotherapy benefit could be demonstrated for the group as a whole. For high-risk patients with classic histology (n=25), chemotherapy was associated with a trend for improvement in 5-year PFS from 36% to 71% (P=0.10) and in 5-year OS from 49% to 100% (P=0.08). CONCLUSIONS:: In adult patients with medulloblastoma, the extent of resection and risk classification predicts the outcome. These results suggest a chemotherapy benefit for high-risk patients with classic histology. PMID: 23111362 [PubMed - as supplied by publisher]

MRI is currently the gold standard for image-guided neurosurgical resection or biopsy and target volume delineation in radiation therapy (RT) planning of brain tumors. MRI findings typically include a heterogeneous area of contrast enhancement (CE) of solid tumor surrounded by a large area of vasogenic edema. However, anaplastic tumors can present with non–contrast enhancement (NCE) of lesions on MRI, which makes surgical planning for biopsy or resection and RT delineation of high-grade tumor volumes challenging. Dependence on CE from MRI alone can lead to inaccurate grading of the tumor, insufficient resection of highly aggressive disease, and undertreatment in areas of aggressive disease. Reliance on the T2 signal abnormality, which includes a heterogeneous mixture of tumor and normal brain, can lead to unnecessary RT of normal brain. PET provides visual information about biological processes and shows promise for improving tumor delineation accuracy for RT and surgical planning of gliomas, which would positively impact accurate assessment of prognosis and improved treatment efficacy. In contrast to the high glucose metabolism in normal brain from 18-fluoro-deoxyglucose (18F-FDG) PET, amino acid PET tracers, such as 3,4-dihydroxy-6-[18F] fluoro-L-phenylalanine (18F-DOPA), demonstrate high uptake in tumor tissue and low uptake in normal brain tissue.1–3 Amino acid transport is generally increased in malignant transformation due to the flux of the amino acid to the tissue, the intrinsic activity of the amino acid transporter, and the rate of the intracellular amino acid metabolism.4,5 Limited studies evaluating the sensitivity of 18F-DOPA indicated that although 18F-FDG PET demonstrated a higher absolute standard uptake value (SUV) than 18F-DOPA, the sensitivity for detection of low- or high-grade tumors was 96% for 18F-DOPA vs 61% for 18F-FDG.2,6 The objectives of this study were to compare pathology findings from biopsies taken in concordant and discordant regions of T1-weighted CE and 18F-DOPA uptake, to determine correlations of high-grade and low-grade pathology components with 18F-DOPA PET SUV, and from these analyses to develop 18F-DOPA PET thresholds to guide clinical decision making and RT target delineation.

High-grade gliomas, such as glioblastomas (GBMs), are very aggressive, invasive brain tumors with low patient survival rates. The recent identification of distinct glioma tumor subtypes offers the potential for understanding disease pathogenesis, responses to treatment and identification of molecular targets for personalized cancer therapies. However, the key alterations that drive tumorigenesis within each subtype are still poorly understood. Although aberrant NF-κB activity has been implicated in glioma, the roles of specific members of this protein family in tumorigenesis and pathogenesis have not been elucidated. In this study, we show that the NF-κB protein RelB is expressed in a particularly aggressive mesenchymal subtype of glioma, and loss of RelB significantly attenuated glioma cell survival, motility and invasion. We find that RelB promotes the expression of mesenchymal genes including YKL-40, a marker of the MES glioma subtype. Additionally, RelB regulates expression of Olig2, a regulator of cancer stem cell proliferation and a candidate marker for the cell of origin in glioma. Furthermore, loss of RelB in glioma cells significantly diminished tumor growth in orthotopic mouse xenografts. The relevance of our studies for human disease was confirmed by analysis of a human GBM genome database, which revealed that high RelB expression strongly correlates with rapid tumor progression and poor patient survival rates. Thus, our findings demonstrate that RelB is an oncogenic driver of mesenchymal glioma tumor growth and invasion, highlighting the therapeutic potential of inhibiting the noncanonical NF-κB (RelB-mediated) pathway to treat these deadly tumors.

Background Phosphaturic mesenchymal tumor, mixed connective tissue variant (PMTMCT) is a rare tumor typically occurring in soft tissues and bone, causing oncogenic (tumor-induced) osteomalacia (TIO) through secretion of the phosphaturic hormone, fibroblast growth factor-23 (FGF-23). Rare tumors identical to PMTMCT occur without known TIO. Intracranial localization of PMTMCT is extremely rare, with only two cases reported in the literature. We present a very unusual case of a patient with an intracranial PMTMCT that presented with neurologic changes without osteomalacia. Case description A 67-year-old woman presented with progressive incontinence, apathy, and abulia after having undergone a total knee replacement 1 month earlier. Imaging disclosed a large left frontal anterior fossa mass. She underwent uncomplicated surgical resection of this tumor. Surprisingly, histopathology suggested PMTMCT. Reverse transcription polymerase chain reaction (RT-PCR) assay demonstrating FGF-23 expression in the tumor confirmed the diagnosis. Serum FGF-23 levels postoperatively were normal and she had no clinical or laboratory evidence of osteomalacia or phosphaturia. Conclusion This report should serve to alert clinicians to the possibility that PMTMCT can be included in the differential diagnosis of intracranial masses even in the absence of tumor-induced osteomalacia.