Development of a Single-Cell Migration and Extravasation Platform through Selective Surface Modification.

Cell migration through three-dimensional (3D) tissue spaces is integral to many biological and pathological processes, including metastasis. Circulating tumor cells (CTCs) are phenotypically heterogeneous, and in vitro analysis of their extravasation behavior is often impeded by the inability to establish complex tissue-like extracellular matrix (ECM) environments and chemotactic gradients within microfluidic devices. We have developed a novel microfluidic strategy to manipulate surface properties of enclosed microchannels and create 3D ECM structures for real-time observation of individual migrating cells. The wettability of selective interconnected channels is controlled by a plasma pulse, enabling the incorporation of ECM exclusively within the transmigration regions. We applied this approach to collectively analyze CTC-endothelial adhesion, trans-endothelial migration, and subsequent motility of breast cancer cells (MDA-MB-231) through a 3D ECM under artificial gradients of SDF-1α. We observed migration velocities ranging from 5.12 to 12.8 µm/h, which closely correspond to single-cell migration in collagen blocks, but are significantly faster than the migration of cell aggregates. The compartmentalized microchannels separated by the porous ECM makes this in vitro assay versatile and suitable for a variety of applications such as inflammation studies, drug screening, and coculture interactions.

The role of regulatory T cells and microglia in glioblastoma-associated immunosuppression.

Cell-mediated suppression of anti-tumor immunity is multifactorial in patients with cancer, and recent studies have focused on several distinct cellular agents that are associated with this phenomenon. This review will focus on the potential role of regulatory T cells (Tregs) and microglia in the suppression of cellular immunity observed in patients with glioblastoma. We discuss the ontogeny, basic biology, evidence for activity, and potential clinical options for targeting Tregs and microglia as part of immunotherapy in affected patients.

Phase II trial of hypofractionated intensity-modulated radiation therapy combined with temozolomide and bevacizumab for patients with newly diagnosed glioblastoma.

Bevacizumab blocks the effects of VEGF and may allow for more aggressive radiotherapy schedules. We evaluated the efficacy and toxicity of hypofractionated intensity-modulated radiation therapy with concurrent and adjuvant temozolomide and bevacizumab in patients with newly diagnosed glioblastoma. Patients with newly diagnosed glioblastoma were treated with hypofractionated intensity modulated radiation therapy to the surgical cavity and residual tumor with a 1 cm margin (PTV1) to 60 Gy and to the T2 abnormality with a 1 cm margin (PTV2) to 30 Gy in 10 daily fractions over 2 weeks. Concurrent temozolomide (75 mg/m(2) daily) and bevacizumab (10 mg/kg) was administered followed by adjuvant temozolomide (200 mg/m(2)) on a standard 5/28 day cycle and bevacizumab (10 mg/kg) every 2 weeks for 6 months. Thirty newly diagnosed patients were treated on study. Median PTV1 volume was 131.1 cm(3) and the median PTV2 volume was 342.6 cm(3). Six-month progression-free survival (PFS) was 90 %, with median follow-up of 15.9 months. The median PFS was 14.3 months, with a median overall survival (OS) of 16.3 months. Grade 4 hematologic toxicity included neutropenia (10 %) and thrombocytopenia (17 %). Grades 3/4 non-hematologic toxicity included fatigue (13 %), wound dehiscence (7 %) and stroke, pulmonary embolism and nausea each in 1 patient. Presumed radiation necrosis with clinical decline was seen in 50 % of patients, two with autopsy documentation. The study was closed early to accrual due to this finding. This study demonstrated 90 % 6-month PFS and OS comparable to historic data in patients receiving standard treatment. Bevacizumab did not prevent radiation necrosis associated with this hypofractionated radiation regimen and large PTV volumes may have contributed to high rates of presumed radiation necrosis.

Characterization of distinct immunophenotypes across pediatric brain tumor types.

Despite increasing evidence that antitumor immune control exists in the pediatric brain, these findings have yet to be exploited successfully in the clinic. A barrier to development of immunotherapeutic strategies in pediatric brain tumors is that the immunophenotype of these tumors' microenvironment has not been defined. To address this, the current study used multicolor FACS of disaggregated tumor to systematically characterize the frequency and phenotype of infiltrating immune cells in the most common pediatric brain tumor types. The initial study cohort consisted of 7 pilocytic astrocytoma (PA), 19 ependymoma (EPN), 5 glioblastoma (GBM), 6 medulloblastoma (MED), and 5 nontumor brain (NT) control samples obtained from epilepsy surgery. Immune cell types analyzed included both myeloid and T cell lineages and respective markers of activated or suppressed functional phenotypes. Immune parameters that distinguished each of the tumor types were identified. PA and EPN demonstrated significantly higher infiltrating myeloid and lymphoid cells compared with GBM, MED, or NT. Additionally, PA and EPN conveyed a comparatively activated/classically activated myeloid cell-skewed functional phenotype denoted in particular by HLA-DR and CD64 expression. In contrast, GBM and MED contained progressively fewer infiltrating leukocytes and more muted functional phenotypes similar to that of NT. These findings were recapitulated using whole tumor expression of corresponding immune marker genes in a large gene expression microarray cohort of pediatric brain tumors. The results of this cross-tumor comparative analysis demonstrate that different pediatric brain tumor types exhibit distinct immunophenotypes, implying that specific immunotherapeutic approaches may be most effective for each tumor type.

Development of the AANS/CNS joint Section on Tumors: challenges and opportunities.

Over the 30 years since the formation of the AANS/CNS Section on Tumors, the breadth and scope of our activities have continued to expand. An initial focus on education and collaboration between those members of the neurosurgical community dedicated to the care of patients with neurological tumors has broadened to include development of international relationships, participation in clinical trials and efforts to define standards of care and quality metrics. As we navigate the rapidly changing environment of health care, the Section on Tumors will occupy a central role in promoting advocacy, establishing collaboration, providing education, and supporting research for the community of neurosurgeons dedicated to the care of patients with neurological tumors. This article will provide an update and status report on the development of the Section on Tumors, followed by a brief discussion of the challenges and opportunities emerging at the onset of our fourth decade of service.

Increased immune gene expression and immune cell infiltration in high-grade astrocytoma distinguish long-term from short-term survivors.

Survival in the majority of high-grade astrocytoma (HGA) patients is very poor, with only a rare population of long-term survivors. A better understanding of the biological factors associated with long-term survival in HGA would aid development of more effective therapy and survival prediction. Factors associated with long-term survival have not been extensively studied using unbiased genome-wide expression analyses. In the current study, gene expression microarray profiles of HGA from long-term survivors were interrogated for discovery of survival-associated biological factors. Ontology analyses revealed that increased expression of immune function-related genes was the predominant biological factor that positively correlated with longer survival. A notable T cell signature was present within this prognostic immune gene set. Using immune cell-specific gene classifiers, both T cell-associated and myeloid linage-associated genes were shown to be enriched in HGA from long-term versus short-term survivors. Association of immune function and cell-specific genes with survival was confirmed independently in a larger publicly available glioblastoma gene expression microarray data set. Histology was used to validate the results of microarray analyses in a larger cohort of long-term survivors of HGA. Multivariate analyses demonstrated that increased immune cell infiltration was a significant independent variable contributing to longer survival, as was Karnofsky/Lansky performance score. These data provide evidence of a prognostic anti-tumor adaptive immune response and rationale for future development of immunotherapy in HGA.

Preoperative ethmoid artery ligation facilitates resection of large sub-frontal meningiomas.

Optimal vascular control during neurosurgical resection of large sub-frontal meningioma is hindered by limited early access to the ethmoidal arteries. Pre-operative ligation of the ethmoidal arteries 1) induces tumor necrosis simplifying resection and 2) minimizes blood loss and operative time. Early arterial ligation is an advantage of endoscopic approaches to transnasal resection of anterior skull base meningiomas that is not appreciated in open approaches with larger meningioma. Here we present a case of a colossal meningioma where minimally invasive pre-operative ligation of ethmoidal arteries prior to a traditional open surgical approach allowed for improved vascular control and decreased surgical time.

Prospective evaluation of health-related quality of life in patients with glioblastoma multiforme treated on a phase II trial of hypofractionated IMRT with temozolomide.

To report health-related quality of life (HRQOL) in glioblastoma (GBM) patients treated on a phase II trial of hypofractionated intensity-modulated radiotherapy (hypo-IMRT) with temozolomide (TMZ). GBM patients received postoperative hypo-IMRT to 60 Gy in 10 fractions with TMZ. HRQOL was assessed using the EORTC quality of life questionnaire core-30 and the EORTC brain cancer module, performed at baseline, RT completion, 1 mo post-RT, and every 3 mos thereafter. Changes from baseline were calculated for each specific HRQOL scale. A ≥ 10 point change in any HRQOL scale from the mean baseline score was significant. 24 patients were treated. Compliance with HRQOL assessments at baseline, RT completion, and 1, 3, 6, 9, and 12 mos post-RT was 100, 96, 92, 79, 70, 68 and 53 %, respectively. Up to 12 mos post-RT, no significant changes were seen in global health status, physical functioning, role functioning, emotional functioning, fatigue, nausea, vision, headache or seizure. Significant improvement was seen in insomnia, future uncertainty, motor dysfunction and drowsiness. Significant worsening was observed in cognitive functioning, social functioning, appetite loss and communication deficit. 60 Gy hypo-IMRT in 6-Gy fractions with TMZ does not appear to negatively impact overall HRQOL.

Frontoethmoid osteoma with pneumocephalus: options for surgical management.

Pneumocephalus is an exceedingly rare complication associated with neurological deficit in cases of frontoethmoid osteoma. The overarching management strategy for affected patients remains undefined. We describe the case of a 61-year-old female patient presenting with frontoethmoid osteoma manifesting as profound intraparenchymal pneumocephalus and associated neurological deficit, treated through a minimally invasive combined surgical strategy involving image-guided burr hole decompression of the pneumocephalus followed by transnasal endoscopic removal of the tumor. Using this approach, the patient rapidly recovered full neurologic function. We review the existing literature and, given the likely intraparenchymal location of pneumocephalus associated with these lesions with the potential of rapid clinical deterioration, recommend aggressive surgical management. Although these lesions can be removed from a purely endoscopic approach, we recommend burr-hole decompression of the pneumocephalus as an adjunct to ensure prompt resolution of the neurologic symptoms.

Immune gene and cell enrichment is associated with a good prognosis in ependymoma.

Approximately 50% of children with ependymoma will suffer from tumor recurrences that will ultimately lead to death. Development of more effective therapies and patient stratification in ependymoma mandates better prognostication. In this study, tumor gene expression microarray profiles from pediatric ependymoma clinical samples were subject to ontological analyses to identify outcome-associated biological factors. Histology was subsequently used to evaluate the results of ontological analyses. Ontology analyses revealed that genes associated with nonrecurrent ependymoma were predominantly immune function-related. Additionally, increased expression of immune-related genes was correlated with longer time to progression in recurrent ependymoma. Of those genes associated with both the nonrecurrent phenotype and that positively correlated with time to progression, 95% were associated with immune function. Histological analysis of a subset of these immune function genes revealed that their expression was restricted to a subpopulation of tumor-infiltrating cells. Analysis of tumor-infiltrating immune cells showed increased infiltration of CD4(+) T cells in the nonrecurrent ependymomas. No genomic sequences for SV40, BK, JC, or Merkel polyomaviruses were found in nonrecurrent ependymoma. This study reveals that up-regulation of immune function genes is the predominant ontology associated with a good prognosis in ependymoma and it provides preliminary evidence of a beneficial host proinflammatory and/or Ag-specific immune response.

Intracortical electroencephalography in acute brain injury.

OBJECTIVE: Continuous electroencephalography (EEG) is used in patients with neurological injury to detect electrographic seizures and clinically important changes in brain function. Scalp EEG has poor spatial resolution, is often contaminated by artifact, and frequently demonstrates activity that is suspicious for but not diagnostic of ictal activity. We hypothesized that bedside placement of an intracortical multicontact electrode would allow for improved monitoring of cortical potentials in critically ill neurological patients. METHODS: Sixteen individuals with brain injury, requiring invasive neuromonitoring, underwent implantation of an eight-contact minidepth electrode. RESULTS: Intracortical EEG (ICE) was successfully performed and compared with scalp EEG in 14 of these 16 individuals. ICE provided considerable improvement in signal-to-noise ratio compared with surface EEG, demonstrating clinically important findings in 12 of 14 patients (86%) including electrographic seizures (n = 10) and acute changes related to secondary neurological injury (n = 2, 1 ischemia, 1 hemorrhage). In patients with electrographic seizures detected by ICE, scalp EEG demonstrated no concurrent ictal activity in six, nonictal-appearing rhythmic delta in two, and intermittently correlated ictal activity in two. In two patients with secondary neurological complications, ICE demonstrated prominent attenuation 2 to 6 hours before changes in other neuromonitoring modalities and more than 8 hours before the onset of clinical deterioration. INTERPRETATION: ICE can provide high-fidelity intracranial EEG in an intensive care unit setting, can detect ictal discharges not readily apparent on scalp EEG, and can identify early changes in brain activity caused by secondary neurological complications. We predict that ICE will facilitate the development of EEG-based alarm systems and lead to prevention of secondary neuronal injury.

Intracortical EEG for the detection of vasospasm in patients with poor-grade subarachnoid hemorrhage.

BACKGROUND: To study the feasibility of utilizing intracortical electroencephalography (ICE) including quantitative EEG (qEEG) analysis for the detection of vasospasm in five consecutive poor-grade SAH patients. METHODS: Intracortical electroencephalography (ICE) was obtained via a single miniature parenchymal 8-contact depth electrode placed at the bedside. Quantitative EEG parameters, calculated on surface EEG and ICE, included alpha/delta ratio (ADR), mean amplitude, suppression percent, and total power. Percent changes between averaged values over 4-6 h of baseline EEG and EEG prior to angiography were calculated. The entire continuous qEEG recording for each patient was then reviewed to determine optimal automated alarm criteria. RESULTS: ICE ADR was the most accurate for predicting angiographic vasospasm (5/5). ICE ADR decreased between baseline and follow-up by 42% (from 0.56 ± 0.07 to 0.32 ± 0.03) for those with vasospasm (N = 3) compared to 17% (0.62 ± 0.06 to 0.51 ± 0.03) for those without (N = 2). A sustained decrease in the ICE ADR from baseline (>25% for ≥ 4 h) occurred in all three patients with angiographically confirmed vasospasm and not in the two without; this decline occurred 1-3 days prior to angiographic confirmation. CONCLUSIONS: Intracortical EEG is promising for detecting ischemia from vasospasm in poor-grade SAH patients, may be superior to scalp EEG, and allow automated detection, particularly using the ADR. Larger studies are needed to better define the effectiveness of this approach.

Intracranial multimodal monitoring for acute brain injury: a single institution review of current practices.

BACKGROUND: Critical care management of patients with severe acute brain injury has undergone tremendous advances. Neurosurgeons and neurointensivists have a large armamentarium of invasive monitoring devices available to help detect secondary brain injury and guide therapy. No consensus exists regarding patient specific selection of monitoring devices, the placement of devices in relation to injured brain tissue, or the preferred insertion technique. Here we review our experience in a consecutive series of acutely brain injured patients who underwent multimodality monitoring. METHODS: Sixty-one patients admitted to the Neurological Intensive Care Unit underwent multimodality intracranial monitoring between January 2005 and October 2008. Patient demographics, hospital length of stay, types of monitoring devices and modalities monitored, insertion techniques, device placement location relative to injury, and complications are reported. RESULTS: Monitored modalities included brain tissue oxygen (PbtO(2)) in 97% (N = 59), microdialysis (MD) in 79% (N = 48), intracranial electroencephalography in 31% (N = 19), brain temperature in 18% (N = 11), and cerebral blood flow in 11% (N = 7). On average, monitoring started within 2 days (0-8) of admission and was continued for 7 days (1-17). The majority of probes (56%; N = 35) were placed into patients with focal brain injuries, while in 43% N = 26 the injury was diffuse. Among those with focal injury, probe placement was categorized as peri-lesional in 46% (N = 16), and within a clot or infarct in 17% (N = 6). The most frequent complication of multimodality brain monitoring was device malfunction or dislodgement (43%; N = 26). Rates of hematoma and infection were 3 and 5%, respectively. Average NICU length of stay was 17 days (3-48) and 26% (N = 16) of patients were dead at discharge. CONCLUSIONS: Collaboration among institutions is necessary to establish practice guidelines for the choice and placement of multimodal monitors. Further advancement in device technology is needed to improve insertion techniques, inter-device compatibility, and device durability. Multimodality data needs to be analyzed to determine the preferable device location.

Dynamics of central and peripheral immunomodulation in a murine glioma model.

BACKGROUND: Immunosuppression by gliomas contributes to tumor progression and treatment resistance. It is not known when immunosuppression occurs during tumor development but it likely involves cross-talk among tumor cells, tumor-associated macrophages and microglia (TAMs), and peripheral as well as tumor-infiltrating lymphocytes (TILs). RESULTS: We have performed a kinetic study of this immunomodulation, assessing the dynamics of immune infiltration and function, within the central nervous system (CNS) and peripherally. PDGF-driven murine glioma cells were injected into the white matter of 13 mice. Four mice were sacrificed 13 days post-injection (dpi), four mice at 26 dpi, and five mice at 40 dpi. Using multiparameter flow cytometry, splenic T cells were assessed for FoxP3 expression to identify regulatory T cells (Tregs) and production of IFN-gamma and IL-10 after stimulation with PMA/ionomycin; within the CNS, CD4+ TILs were quantified, and TAMs were quantified and assessed for TNF-alpha and IL-10 production after stimulation with LPS. Peripheral changes associated with tumor development were noted prior to effects within the CNS. The percentage of FoxP3+ regulatory T cells (Tregs) increased by day 26, with elevated frequencies throughout the duration of the study. This early increase in Tregs was paralleled by an increase in IL-10 production from Tregs. At the final time points examined (tumor morbidity or 40 dpi), there was an increase in the frequency of TAMs with decreased capacity to secrete TNF-alpha. An increase in TIL frequency was also observed at these final time points. CONCLUSION: These data provide insight into the kinetics of the immunosuppressive state associated with tumor growth in a murine model of human gliomas. Functional impairment of TAMs occurs relatively late in the course of GBM tumor growth, potentially providing a window of opportunity for therapeutic strategies directed towards preventing their functional impairment.

Rapid recurrence and malignant transformation of pilocytic astrocytoma in adult patients.

Pilocytic astrocytoma is a slow-growing, circumscribed glioma that most frequently occurs within the pediatric population. In general, surgical resection for pilocytic astrocytoma is thought to be curative with tumor recurrence or malignant transformation being relatively rare. However, there have been very few studies specifically looking at the prognosis for adult patients diagnosed with pilocytic astrocytoma. To evaluate the frequency of recurrence and malignant transformation of pilocytic astrocytoma in adults, we performed a retrospective analysis of all adult patients who underwent surgical resection for this tumor at our institution over a period of 10 years. In our cohort of 20 patients, there were 6 (30%) recurrences with four patients requiring repeat surgery due to symptomatic progression. Relatively rapid recurrences were noted with the median time to recurrence being 16.5 months. All recurrences occurred within 4 years of initial surgery while patients requiring repeat surgery presented within 17 months of initial surgery. Based on this study estimated rates of freedom from recurrence (FFR) at 12 and 24 months after initial surgery are 94 +/- 5% and 76 +/- 10%, respectively. A high rate of malignant transformation was observed in the patients that underwent repeat surgery with 75% (3/4) progressing to anaplastic astrocytoma on pathological examination. This study provides further evidence that the clinical course of a subset of adult patients with pilocytic astrocytoma will not be benign. The potential for rapid tumor recurrence and malignant transformation necessitates careful post-operative follow-up for adult patients with this tumor.

Targeted genomic CRISPR-Cas9 screen identifies MAP4K4 as essential for glioblastoma invasion.

Among high-grade brain tumors, glioblastoma is particularly difficult to treat, in part due to its highly infiltrative nature which contributes to the malignant phenotype and high mortality in patients. In order to better understand the signaling pathways underlying glioblastoma invasion, we performed the first large-scale CRISPR-Cas9 loss of function screen specifically designed to identify genes that facilitate cell invasion. We tested 4,574 genes predicted to be involved in trafficking and motility. Using a transwell invasion assay, we discovered 33 genes essential for invasion. Of the 11 genes we selected for secondary testing using a wound healing assay, 6 demonstrated a significant decrease in migration. The strongest regulator of invasion was mitogen-activated protein kinase 4 (MAP4K4). Targeting of MAP4K4 with single guide RNAs or a MAP4K4 inhibitor reduced migration and invasion in vitro. This effect was consistent across three additional patient derived glioblastoma cell lines. Analysis of epithelial-mesenchymal transition markers in U138 cells with lack or inhibition of MAP4K4 demonstrated protein expression consistent with a non-invasive state. Importantly, MAP4K4 inhibition limited migration in a subset of human glioma organotypic slice cultures. Our results identify MAP4K4 as a novel potential therapeutic target to limit glioblastoma invasion.

Optimization of postoperative computerized tomographic imaging in patients with the implanted responsive neurostimulation system.

The Responsive Neurostimulation System is currently under investigation as a neurosurgical option for medically refractory epilepsy. The device produces significant metallic artifact on conventional axial CT scans, resulting in limited diagnostic imaging options for implanted patients. We have developed a strategy to overcome this technical difficulty utilizing optimized patient positioning, thin-slice image acquisition, and postprocessing image reconstruction with commercially available software. Significant improvements were noted in the severity of device-related metallic streak artifact on reconstructed axial images. In addition, thin-section data were successfully used to generate detailed three-dimensional reconstructions, providing for improved visualization of the stimulator and the intracranial position of attached electrodes.

Intratumoral heterogeneity of endogenous tumor cell invasive behavior in human glioblastoma.

Intratumoral genetic heterogeneity is a widely accepted characteristic of human cancer, including the most common primary malignant brain tumor, glioblastoma. However, the variability in biological behaviors amongst cells within individual tumors is not well described. Invasion into unaffected brain parenchyma is one such behavior, and a leading mechanism of tumor recurrence unaddressed by the current therapeutic armamentarium. Further, providing insight into variability of tumor cell migration within individual tumors may inform discovery of novel anti-invasive therapeutics. In this study, ex vivo organotypic slice cultures from EGFR-wild type and EGFR-amplified patient tumors were treated with the EGFR inhibitor gefitinib to evaluate potential sub-population restricted intratumoral drug-specific responses. High-resolution time-lapse microscopy and quantitative path tracking demonstrated migration of individual cells are punctuated by intermittent bursts of movement. Elevation of population aggregate mean speeds were driven by subpopulations of cells exhibiting frequent high-amplitude bursts, enriched within EGFR-amplified tumors. Treatment with gefitinib specifically targeted high-burst cell subpopulations only in EGFR-amplified tumors, decreasing bursting frequency and amplitude. We provide evidence of intratumoral subpopulations of cells with enhanced migratory behavior in human glioblastoma, selectively targeted via EGFR inhibition. These data justify use of direct human tumor slice cultures to investigate patient-specific therapies designed to limit tumor invasion.

Expansile, enhancing cervical cord lesion with an associated syrinx secondary to demyelination. Case report and review of the literature.

The authors describe the case of a patient with an enhancing, intramedullary cervical spinal cord lesion and associated syrinx. Biopsy sampling of the cervical lesion was performed, and the histological findings were consistent with a demyelinating process supporting the diagnosis of multiple sclerosis (MS). Syrinx formation associated with demyelinating disease has only been described in isolated cases, almost exclusively in Japanese patients with MS. A 22-year-old woman of Caribbean descent presented with a subacute, progressive myelopathy including symptoms of pain and weakness in all extremities, bladder incontinence, and the inability to ambulate. Magnetic resonance imaging of the brain and spinal cord demonstrated an enlarged cervical cord with enhancement and central cavitation consistent with a syrinx. The patient underwent a C3-7 laminoplasty and placement of a dural graft for cord decompression as well as fenestration of the central syrinx. Biopsy sampling of the lesion was performed, and the histopathological analysis, in conjunction with subsequent laboratory and diagnostic testing, supported the diagnosis of demyelinating disease. After treatment with a course of high-dose dexamethasone and inpatient rehabilitation therapy, the patient demonstrated significant clinical improvement. Spinal cord involvement is not uncommon in patients with demyelinating disease; however, enhancing lesions associated with extensive tissue loss and syrinx formation have rarely been reported. For the consulting neurological surgeon, demyelinating disease should be included in the differential diagnosis of such lesions given the level of complexity and risk to the patient associated with open biopsy of the spinal cord.

A Human Glioblastoma Organotypic Slice Culture Model for Study of Tumor Cell Migration and Patient-specific Effects of Anti-Invasive Drugs.

Glioblastoma (GBM) continues to carry an extremely poor clinical prognosis despite surgical, chemotherapeutic, and radiation therapy. Progressive tumor invasion into surrounding brain parenchyma represents an enduring therapeutic challenge. To develop anti-migration therapies for GBM, model systems that provide a physiologically relevant background for controlled experimentation are essential. Here, we present a protocol for generating slice cultures from human GBM tissue obtained during surgical resection. These cultures allow for ex vivo experimentation without passaging through animal xenografts or single cell cultures. Further, we describe the use of time-lapse laser scanning confocal microscopy in conjunction with cell tracking to quantitatively study the migratory behavior of tumor cells and associated response to therapeutics. Slices are reproducibly generated within 90 min of surgical tissue acquisition. Retrovirally-mediated fluorescent cell labeling, confocal imaging, and tumor cell migration analyses are subsequently completed within two weeks of culture. We have successfully used these slice cultures to uncover genetic factors associated with increased migratory behavior in human GBM. Further, we have validated the model's ability to detect patient-specific variation in response to anti-migration therapies. Moving forward, human GBM slice cultures are an attractive platform for rapid ex vivo assessment of tumor sensitivity to therapeutic agents, in order to advance personalized neuro-oncologic therapy.