Care For Glioblastoma Patients Research Articles

IMMU-71. CRANIOENCEPHALIC FUNCTIONAL LYMPHOID UNITS IN GLIOBLASTOMA

Abstract The general accessibility of the brain to immune cells has been a subject of intense debate and reinterpretation within the last decade. While glioblastoma’s brain tumor ecosystem is typically considered immunosuppressed, we report the presence of a tumor-reactive immune cell niche within the cranial bone marrow (CB) situated adjacent to treatment-naive tumors. At time of initial diagnosis, glioblastoma patients underwent clinical and anatomical imaging, which revealed an unexpected co-morbid association between the CB and the tumor. Following this observation, we conducted an extensive experimental study including multiplex immunofluorescence, immunoprofiling, functional assays, and single-cell analysis, which identified an active lymphoid niche within the patient’s CB. CD8+ T cell effector cells residing in this niche exhibited acute and durable antitumor responses, and, compared to cells from the distal bone marrow, showed increased expression of the lymphoid egress marker S1PR1. Single-cell T cell receptor sequencing revealed a rich reservoir of non-exhausted tumor-shared CD8+ T cell clonotypes within the CB lymphoid niche, suggesting re-circulation between the proximal bone and the tumor tissue. These clonotypes were characterized by a high predicted tumor-reactivity, significantly surpassing levels found in blood and distal bone marrow. We discovered the entire developmental spectrum of CD8+ T cells in the CB, indicating that anatomical proximity may play a critical role in the process of early anti-tumor response. Correspondingly, cranial enhancement of CXCR4 radiolabel may serve as a surrogate marker, indicating focal association with improved progression-free survival. Our findings support the preservation and further exploitation of these cranioencephalic units to improve clinical care in glioblastoma patients.

Metabolism and signaling crosstalk in glioblastoma progression and therapy resistance.

Glioblastoma is the most common form of primary malignant brain tumor in adults and one of the most lethal human cancers, with high recurrence and therapy resistance. Glioblastoma cells display extensive genetic and cellular heterogeneity, which precludes a unique and common therapeutic approach. The standard of care in glioblastoma patients includes surgery followed by radiotherapy plus concomitant temozolomide. As in many other cancers, cell signaling is deeply affected due to mutations or alterations in the so-called molecular drivers. Moreover, glioblastoma cells undergo metabolic adaptations to meet the new demands in terms of energy and building blocks, with an increasing amount of evidence connecting metabolic transformation and cell signaling deregulation in this type of aggressive brain tumor. In this review, we summarize some of the most common alterations both in cell signaling and metabolism in glioblastoma, presenting an integrative discussion about how they contribute to therapy resistance. Furthermore, this review aims at providing a comprehensive overview of the state-of-the-art of therapeutic approaches and clinical trials exploiting signaling and metabolism in glioblastoma.

International practice variation in perioperative laboratory testing in glioblastoma patients\u2014a retrospective cohort study

PurposeAlthough standard-of-care has been defined for the treatment of glioblastoma patients, substantial practice variation exists in the day-to-day clinical management. This study aims to compare the use of laboratory tests in the perioperative care of glioblastoma patients between two tertiary academic centers—Brigham and Women’s Hospital (BWH), Boston, USA, and University Medical Center Utrecht (UMCU), Utrecht, the Netherlands.MethodsAll glioblastoma patients treated according to standard-of-care between 2005 and 2013 were included. We compared the number of blood drawings and laboratory tests performed during the 70-day perioperative period using a Poisson regression model, as well as the estimated laboratory costs per patient. Additionally, we compared the likelihood of an abnormal test result using a generalized linear mixed effects model.ResultsAfter correction for age, sex, IDH1 status, postoperative KPS score, length of stay, and survival status, the number of blood drawings and laboratory tests during the perioperative period were 3.7-fold (p < 0.001) and 4.7-fold (p < 0.001) higher, respectively, in BWH compared to UMCU patients. The estimated median laboratory costs per patient were 82 euros in UMCU and 256 euros in BWH. Furthermore, the likelihood of an abnormal test result was lower in BWH (odds ratio [OR] 0.75, p < 0.001), except when the prior test result was abnormal as well (OR 2.09, p < 0.001).ConclusionsOur results suggest a substantially lower clinical threshold for ordering laboratory tests in BWH compared to UMCU. Further investigating the clinical consequences of laboratory testing could identify over and underuse, decrease healthcare costs, and reduce unnecessary discomfort that patients are exposed to.

P12.03 A Delphi survey to inform core ‘red flag’ symptoms for an electronic patient reported outcome system in glioblastoma follow up

Abstract BACKGROUND Glioblastoma is a common and aggressive primary malignant brain tumour in adults associated with a poor prognosis and considerable symptom burden. Clinical review and serial neuroimaging remain the primary monitoring tools to assess for disease status. However, the evidence base for the existing surveillance imaging schedule is yet to be established. New models of follow up care are needed to demonstrate better patient outcomes in this patient cohort. There are indications that patient-reported outcome measures (PROMs) can contribute to improved survival and maintaining an optimal quality of life in other oncology populations. MATERIALS AND METHODS There are no standardised PROMs for the priority symptoms in glioblastoma patients, which can be used for evaluation in clinical care as a surrogate marker for disease progression. A broad set of symptoms were therefore initially identified in a targeted literature search and were further refined via Delphi methodology, with the aim to obtain consensus amongst a small expert panel involved in the care of glioblastoma patients. A three round Delphi email survey was conducted. Consensus was defined as 70% agreement. RESULTS Symptom and function constructs were assessed for relevance, relationship to disease and treatment, sensitivity to change, psychometric properties and patient acceptability. Consensus was reached on the red flag symptoms and symptom clusters to monitor, as well as the minimum severity thresholds needed to trigger an alert on an electronic symptom reporting system, which has been developed to allow patient self-reporting of symptoms during treatment. CONCLUSION The red flag symptoms to monitor, along with the symptom severity thresholds, informed an app-based symptom reporting system, which is currently being piloted in a feasibility study exploring triggered imaging using ePROMs. This has the potential to inform future clinical practice through development of patient reported biomarkers.

Automatic Prediction of MGMT Status in Glioblastoma via Deep Learning-Based MR Image Analysis.

Methylation of the O6-methylguanine methyltransferase (MGMT) gene promoter is correlated with the effectiveness of the current standard of care in glioblastoma patients. In this study, a deep learning pipeline is designed for automatic prediction of MGMT status in 87 glioblastoma patients with contrast-enhanced T1W images and 66 with fluid-attenuated inversion recovery(FLAIR) images. The end-to-end pipeline completes both tumor segmentation and status classification. The better tumor segmentation performance comes from FLAIR images (Dice score, 0.897 ± 0.007) compared to contrast-enhanced T1WI (Dice score, 0.828 ± 0.108), and the better status prediction is also from the FLAIR images (accuracy, 0.827 ± 0.056; recall, 0.852 ± 0.080; precision, 0.821 ± 0.022; and F1 score, 0.836 ± 0.072). This proposed pipeline not only saves the time in tumor annotation and avoids interrater variability in glioma segmentation but also achieves good prediction of MGMT methylation status. It would help find molecular biomarkers from routine medical images and further facilitate treatment planning.

Causes of death in glioblastoma: insights from the SEER database.

Glioblastoma (GB) and its variants portend a poor prognosis. The predominant cause of death (COD) is related to the cancer diagnosis, but a significant subset is related to other causes. As GB is a systemic disease requiring systemic treatment, focus regarding all COD provides a comprehensive illustration of the disease. The SEER-18 was queried for patients with cranial GB and its variants. Age, gender, race, marital status, tumor characteristics, treatment details, and follow-up data were acquired. The patients were classified into group A (death attributed to this cancer diagnosis) or group B (death attributed to causes other than this cancer diagnosis). From 1973 to 2013, 36,632 deaths (94%) constituted group A, and 2,324 deaths (5.9%) constituted group B. The latter significantly exhibited lower proportions of age <60, Caucasians, married status, frontal/brain stem/ventricle tumor locations, and receipt of radiation. From logistic regression, age >60, male gender, race, not married, tumor location, and no radiation were significant independent predictors for group B. The top known CODs in group B are diseases of heart, pneumonia and influenza, cerebrovascular diseases, accidents and adverse effects, and infections. CODs not attributed to GB remains a significant subset of all CODs. Many of these, particularly diseases of heart, are frequent comorbidities. Moreover, infection-related CODs after GB diagnosis appear more salient compared to CODs in the general population. Consideration of these CODs, and vigilant treatment aimed at these CODs, may improve overall care for GB patients.

Survival effects of a strategy favoring second-line multimodal treatment compared to supportive care in glioblastoma patients at first progression.

Data on the survival effects of supportive care compared to second-line multimodal treatment for glioblastoma progression are scarce. Thus, the authors assessed survival in two population-based, similar cohorts from two European university hospitals with different treatment strategies at first progression. The authors retrospectively identified patients with newly diagnosed glioblastoma treated at two neurooncological centers. After diagnosis, patients from both centers received identical treatments, but at tumor progression each center used a different approach. In the majority of cases, at center A (Greece), supportive care or a single therapeutic modality was offered at progression, whereas center B (Germany) provided multimodal second-line therapy. The main outcome measure was survival after progression (SaP). The influence of the treatment strategy on SaP was assessed by multivariate analysis. One hundred three patients from center A and 156 from center B were included. Tumor progression was observed in 86 patients (center A) and 136 patients (center B). At center A, 53 patients (72.6%) received supportive care alone, while at center B, 91 patients (80.5%) received second-line treatment. Progression-free survival at both centers was similar (9.4 months [center A] vs 9.0 months [center B]; p = 0.97), but SaP was significantly improved in the patients treated with multimodal second-line therapy at center B (7 months, 95% CI 5.3-8.7 months) compared to those treated with supportive care or a single therapeutic modality at center A (4.5 months, 95% CI 3.5-5.5 months; p = 0.003). In the multivariate analysis, the treatment center was an independent prognostic factor for overall survival (HR 1.59, 95% CI 0.17-2.15; p = 0.002). Treatment strategy favoring multimodal second-line treatment over minimal treatment or supportive care at glioblastoma progression is associated with significantly better overall survival.

Abstract 4220: Heterogeneous distribution of prognostic protein markers in glioblastoma

Abstract Background: Glioblastoma (GBM) is the most common and lethal primary malignant brain tumor in adults. The heterogeneity of the disease leads to significant variability in response to standard therapy (surgery plus concurrent radiation and temozolomide). An accurate and reliable predictor of patient prognosis represents an unmet need to improve the care of GBM patients. While protein markers are an effective readout of cellular function, proteomics has rarely been utilized in GBM prognostic marker discovery. Experimental Procedures: GBM patients were prospectively recruited (Ohio Brain Tumor Study) and proteomics discovery using liquid chromatography mass spectrometry analysis (LC MS/MS) was performed in a discovery set of 27 patients including 13 short-term survivors (&amp;lt; 9 months, STS) and 14 long-term survivors (&amp;gt;= 18 months, LTS). Statistically significant proteins were evaluated in two independent datasets, including in18 samples micro-dissected from multiple tumor areas of 6 GBM patients. Results: Proteomics discovery identified 11,941 peptides in 2,495 unique proteins, with 172 proteins exhibiting significant dysregulation when comparing STS and LTS. Proteins involved in glycolysis/TCA cycle were up-regulated in STS compared to LTS by examination of individual targets as well as upon application of a novel protein and peptide pathway enrichment analysis. Validation of these dysregulated proteins and other protein markers from the literature in our first validation set (18 samples micro-dissected from n=6 patients) demonstrated that they were very unevenly distributed throughout individual patients' tumors. Several proteins overcame the heterogeneous nature of the tumors and were both prognostic markers differentially expressed between LTS and STS, as well as potential drug targets owing to their even distribution throughout the tumor. Protein abundance of significant marker proteins were verified in a second independent validation set using Western immunoblots. Conclusion: The current study verified the importance of metabolism in GBM pathology and demonstrated the heterogeneous nature of GBMs at the protein level. Citation Format: Lindsay C. Stetson, Quinn T. Ostrom, Peter Liao, Andrew E. Sloan, Mark R. Chance, Jill S. Barnholtz-Sloan. Heterogeneous distribution of prognostic protein markers in glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4220.

Antisecretory Factor-Mediated Inhibition of Cell Volume Dynamics Produces Antitumor Activity in Glioblastoma.

Interstitial fluid pressure (IFP) presents a barrier to drug uptake in solid tumors, including the aggressive primary brain tumor glioblastoma (GBM). It remains unclear how fluid dynamics impacts tumor progression and can be targeted therapeutically. To address this issue, a novel telemetry-based approach was developed to measure changes in IFP during progression of GBM xenografts. Antisecretory factor (AF) is an endogenous protein that displays antisecretory effects in animals and patients. Here, endogenous induction of AF protein or exogenous administration of AF peptide reduced IFP and increased drug uptake in GBM xenografts. AF inhibited cell volume regulation of GBM cells, an effect that was phenocopied in vitro by the sodium-potassium-chloride cotransporter 1 (SLC12A2/NKCC1) inhibitor bumetanide. As a result, AF induced apoptosis and increased survival in GBM models. In vitro, the ability of AF to reduce GBM cell proliferation was phenocopied by bumetanide and NKCC1 knockdown. Next, AF's ability to sensitize GBM cells to the alkylating agent temozolomide, standard of care in GBM patients, was evaluated. Importantly, combination of AF induction and temozolomide treatment blocked regrowth in GBM xenografts. Thus, AF-mediated inhibition of cell volume regulation represents a novel strategy to increase drug uptake and improve outcome in GBM. Mol Cancer Res; 16(5); 777-90. ©2018 AACR.

End-of-life care for glioblastoma multiforme patients at an academic institution.

202 Background: End of life care for glioblastoma patients has not been studied in the United States. We studied 100 patients with GBM to examine use of advance directives (AD), symptom assessments, hospitalizations, chemotherapy use, and hospice use. Methods: A single center retrospective cohort study of adult patients actively treated for GBM at Johns Hopkins Hospital from 2009-2014. 100 patients were randomly chosen from the group of 317 GBM patients. Information included documentation of AD, code status, hospitalizations, chemotherapy use, code status and symptom assessments. A secondary analysis of all JHH GBM patients who received hospice care at Gilchrist Hospice during this time period was analyzed using the electronic medical record and hospice records to evaluate date of referral to hospice, length of stay in hospice and location of death. Results: Of 100 patients, 76 were referred to hospice. Only 40% of individuals had documentation of code status and only 17% had any documentation of ADs in the outpatient chart. Formal symptom, spiritual, or psychosocial assessments were seldom documented in the outpatient record. Only 17% used chemotherapy in their last month of life. 37% were hospitalized in the last month of life for an average of 9 days. Of the Gilchrist Hospice patients, the median length of stay in hospice was 21 days and 64% of these patients died in their residence with hospice services. Conclusions: More formalized palliative care utilization and structure could improve end of life care for GBM patients, especially in the use of ADs; formal symptom, spiritual, and psychosocial assessments; and earlier use of hospice to prevent end of life hospitalizations.

HCP-12IMPROVING THE EFFICIENCY OF MOLECULAR TESTING FOR EXPEDITED BRAIN TUMOR PATIENT MANAGEMENT AND CLINICAL TRIAL ENROLLMENT

BACKGROUND: O6-methylguanine methyltransferase (MGMT) methylation status is prognostic in glioblastoma. It is a critical datum in optimizing care in elderly glioblastoma patients. Further, MGMT methylation status is increasingly required for clinical trial enrollment (e.g. NCT02336165, NCT02287428). As such, maximizing efficiency of molecular tumor testing is critical in order to provide the best care for glioblastoma patients. METHODS: Using the longitudinal electronic medical record and pathology sample tracking systems, we determined the turn around time (TAT) from tumor resection to MGMT methylation status reporting, along with various intermediate steps along the way. We performed interdepartmental process mapping to identify opportunities for process change, followed by 2 process interventions with follow-up tracking of TAT with statistical process control testing. RESULTS: The baseline average interval from craniotomy to MGMT methylation result was 25.1 days for the 30 weeks prior to intervention, which included ∼5 days from craniotomy to pathological diagnosis, and ∼18 days from pathological diagnosis to MGMT result. Two tests of change were implemented, including a switch from batched to daily DNA extraction and MGMT methylation testing, and implementation of a new procedure for pathology sample inventory within the molecular pathology laboratory. The median time from craniotomy to MGMT methylation status reporting was reduced to from 25.1 days to 17 days after the first intervention, and to 15 days after the second intervention, with significant TAT reduction. CONCLUSIONS: Through clinical process improvement strategies and interdepartmental collaboration, we significantly reduced the TAT from craniotomy to MGMT methylation status reporting by an average of 10 days (40%). This allows for improved patient care by accelerating the delivery of potentially treatment-altering information as well as supplying earlier eligibility information for clinical trial enrollment. We continue to study additional opportunities for intervention, as this work highlights the possibility for neuro-oncology care improvement through process improvement strategies.

Advances in the management of glioblastoma: the role of temozolomide and MGMT testing

Glioblastoma (GB) is one of the most lethal forms of cancer, with an invasive growth pattern that requires the use of adjuvant therapies, including chemotherapy and radiation, to prolong survival. Temozolomide (TMZ) is an oral chemotherapy with a limited side effect profile that has become the standard of care in GB treatment. While TMZ has made an impact on survival, tumor recurrence and TMZ resistance remain major challenges. Molecular markers, such as O6-methylguanine-DNA methyltransferase methylation status, can be helpful in predicting tumor response to TMZ, and therefore guides clinical decision making. This review will discuss the epidemiology and possible genetic underpinnings of GB, how TMZ became the standard of care for GB patients, the pharmacology of TMZ, the practical aspects of using TMZ in clinic, and how molecular diagnostics – particularly the use of O6-methylguanine-DNA methyltransferase status – affect clinical management.

Patterns of care and survival of glioblastoma patients: a comparative study between 2004 and 2008 in Lyon, France

Patterns of care and survival of glioblastoma patients: a comparative study between 2004 and 2008 in Lyon, France

Abstract 2821: The PKC-iota inhibitor ICA-1 overcomes chemoresistance and induces cell death in glioblastoma multiforme

Abstract Glioblastoma multiforme is a very aggressive brain cancer that is refractory to most chemotherapeutics. The standard of care for glioblastoma patients is the administration of temozolomide following surgery. However, temozolomide provides an approximately 20% enhanced survival rate. Therefore, it is imperative to develop other novel chemotherapeutics to better enhance the survival rate and induce tumor regression in patients. A potential candidate for a pre-clinical glioblastoma study is the novel chemotherapeutic, ICA-1. ICA-1 is a PKC-iota inhibitor which specifically inhibits PKC-iota and not its homologue, PKC-zeta. Results demonstrated that ICA-1 inhibited cell proliferation rates in a range of T98G glioblastoma concentrations in vitro. Also, results showed that low density T98G cultures which have a high proliferation status and high S-phase promotion concomitantly demonstrated resistance to ICA-1 treatment. However, high density T98G cultures which have a lower proliferation status and low S-phase promotion are concurrently chemosensitive toward ICA-1. At 48h post ICA-1 treatment, the majority of cells in the high density cultures were in G0/G1 with a rise of cells in this phase of the cell cycle shown by day 5 post-treatment. Conversely, between 48h and day 5, 30-44% of cells from the low density cultures were in S phase irrespective of ICA-1 therapy. Furthermore, by day 5 post-treatment, ICA-1 induced significant cell death in T98G cells only at higher ranges (i.e., 20-40 microM of ICA-1). Interestingly, 40 microM ICA-1 caused a significant cell death rate of 45% when 7,500 cells were seeded into a 24-well plate on Day 0 followed by ICA-1 exposure on the next day. Forty microM ICA-1 induced 15% cell death when 3,750 cells were seeded. There was a stepwise increase in the amount of ICA-1 required to induce cell death as the cell concentration increased (i.e., high density T98G cultures required elevated levels of ICA-1). Therefore, the data suggested that the G0/G1 phase glioblastoma cells (i.e., the higher cell concentrations) were more sensitive to 40 microM ICA-1 therapy. Whereas, the S phase glioblastoma cells were less sensitive to the ICA-1 therapy. Future animal models may show that the more aggressive the tumor becomes, the greater the potential for the effectiveness of ICA-1 (i.e., greater chemosensitivity). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2821. doi:1538-7445.AM2012-2821

Abstract 3321: The benefits from combined temozolomide and sunitinib in medulloblastoma cells: The antitumor effect, inhibition of invasiveness, and blocking of cancer stem cells

Abstract Objectives: Medulloblastoma is the most common malignant and metastatic pediatric brain tumor in children. Angiogenesis inhibitors, such as sunitinib, represent a promising strategy to improve glioblastoma (GBM) tumor response. Temozolomide (TMZ) also became the standard of care for GBM patients by removing alkyl groups from the O6-position of guanine. But, there is little evidence about migration and invasion study on medulloblastoma cells treated with sunitinib or TMZ. In this study, we identified the effects of sunitinib or TMZ only, and the combination of them on migration and invasiveness in medulloblastoma cell line (DAOY). Also the characteristics of cancer stem cells were investigated. Methods and Results: The cell proliferation rate showed TMZ (1000µM) 70%, sunitinib (62.5µM) 60%, and the combination of TMZ and sunitinib (1000/62.5µM) 35% at 72hr after treatment with the drugs in DAOY cells. Treatment with sunitinib in addition to TMZ in the cells demonstrated more G2/M arrest than cells treated respective drugs at 24hr exposure of the drugs from cell cycle analysis. Through Rhodamine phalloidin staining, we also observed change of cell size and F-actin alterations in all cells treated TMZ, sunitinib only or the combination of the reagents, in contrast to cells treated DMSO as a control. And, in invasion assay using Matrigel chamber, the cells treated with combination of drugs were inhibited invasiveness the most when compared with control, sunitinib, and TMZ only at 72hr after exposure to the drugs. Moreover, when the cells were treated with sunitinib and TMZ simultaneously, migration ability from wound-healing assay represented higher than in cells treated TMZ or sunitinib only during exposure of 24hr. We investigated properties of cancer stem cells as well as characteristics of migration and invasion in cells treated the drugs analyzing the expression of protein during exposure of 24, 48, 72hr by western blotting. As a result, we identified the decreased levels of the protein such as p-PDGFR-β, p-VEGFR1, integrin-β1, MMP-9, CD44, Rac1 as migration and invasion markers, nestin, CD133, Oct3/4, Sox2 as cancer stem cell markers and O6-methylguanine methyltransferase(MGMT). Conclusions: Taken together, the combination therapy functions as very effective therapeutic tool in terms of, not only inhibititor of proliferation, migration and invasiveness, but also cancer stem cells in medulloblastoma cell line. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3321. doi:10.1158/1538-7445.AM2011-3321

Abstract C176: Combining oncolytic herpes simplex virus and chemotherapy: G47 and temozolomide synergize in killing glioma stem cells

Abstract Malignant gliomas, such as glioblastoma, are amongst the most common primary brain tumors in adults. Along with debulking surgery and post-operative radiation, the alkylating agent temozolomide (TMZ) is the standard of care for glioblastoma patients. However, the benefits of TMZ are correlated with tumors having transcriptional silencing of the O6-methylguanine methyltransferase (MGMT) gene, which occurs in approximately half of tumors. Unfortunately, regardless of MGMT status, recurrence or progression are inevitable, necessitating novel therapeutic approaches. Glioma stem cells (GSCs) have been implicated as a potential perpetrator of glioma recurrence. We have established human GSCs from surgically resected glioblastomas that vary in MGMT expression and TMZ sensitivity. Five human GSC lines were examined, two of which were MGMT-positive (confirmed by western blotting). MGMT-negative GSCs were more sensitive to TMZ (IC50&amp;lt;100 µM) than their counterparts grown in serum-containing media (representative of the bulk tumor) or established glioma cell lines such as U87 or U373. MGMT-positive GSCs were extremely resistant to TMZ (IC50&amp;gt;1000 µM). When MGMT was inactivated by O6-Benzylguanine (O6-BG), GSC sensitivity to TMZ approximately doubled. Oncolytic viruses destroy tumor cells by different mechanisms from conventional therapeutics, including TMZ, and therefore should interact positively. We have previously reported that clinically tested oncolytic herpes simplex virus (oHSV) G207 combined with TMZ synergistically kills established glioma cell lines. However, G207 does not replicate in GSCs. Therefore, we examined a newer generation oHSV, G47, which was effective at killing GSCs regardless of MGMT status. The interaction between TMZ and G47 on GSC killing was assessed by Chou-Talalay analysis. For MGMT-negative GSCs, the combination was synergistic, while it was antagonistic for MGMT-positive GSCs. When MGMT was inactivated with O6-BG, the interaction became synergistic for MGMT-positive GSCs. Interestingly, viral replication, entry, or spread was not affected by combination treatment. For those combinations that were synergistic, there was a large increase in H2AX, indicative of DNA double-strand breaks. Finally, the effect of combination therapy was assessed in two in vivo GSC models: GBM8 (MGMT-negative) and BT74 (MGMT-positive). Intracerebral tumors were established in athymic mice and treated with chemotherapy and/or intracerebral injection of G47 on the second day of chemotherapy. TMZ + G47 was more effective than either agent alone in extending survival in mice bearing GBM8 tumors. In the BT74 model, TMZ + O6-BG + G47 was more effective than TMZ+O6-BG or G47 alone. These studies demonstrate that GSCs can be both sensitive and resistant to TMZ, correlating with their MGMT status. All GSCs were sensitive to killing by G47 and this was synergistically enhanced by TMZ or TMZ+O6-BG, depending upon the MGMT status. Importantly, the combination was very efficacious in vivo in both models, suggesting that this combination strategy be tested clinically. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):C176.