Animal Models Of Glioma Research Articles

Application of Dendrimer-based Nanoparticles in Glioma Imaging.

Dendritic polymers or dendrimers present an alternate template for the development of nanoparticulate-based drug delivery and imaging systems. The smaller size (~7–12 nm) of dendrimers have the advantage over the other particles, because its smaller size can possibly improve tumor penetration and the inclusion of tumor specific drug release mechanisms. A Paramagnetic Chemical Exchange Saturation Transfer (PARACEST) MRI contrast agent, Eu-DOTA-Gly4 or a clinical relevant Gd-DOTA was conjugated on the surface of a G5 PAMAM dendrimer. To create a dual mode MRI-optical imaging nanoparticle, Dylight680 was also incorporated on the amines surface of a G5 dendrimer. The particle was detected with in vivo MRI in preclinical glioma animal model. Furthermore, noninvasive imaging results were validated with in vivo and ex-vivo optical imaging.

Arctigenin, a natural lignan compound, induces G0/G1 cell cycle arrest and apoptosis in human glioma cells.

The aim of the current study was to investigate the anticancer potential of arctigenin, a natural lignan compound, in malignant gliomas. The U87MG and T98G human glioma cell lines were treated with various concentrations of arctigenin for 48 h and the effects of arctigenin on the aggressive phenotypes of glioma cells were assessed. The results demonstrated that arctigenin dose-dependently inhibited the growth of U87MG and T98G cells, as determined using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and bromodeoxyuridine incorporation assays. Arctigenin exposure also induced a 60-75% reduction in colony formation compared with vehicle-treated control cells. However, arctigenin was not observed to affect the invasiveness of glioma cells. Arctigenin significantly increased the proportion of cells in the G0/G1 phase and reduced the number of cells in the S phase, as compared with the control group (P<0.05). Western blot analysis demonstrated that arctigenin increased the expression levels of p21, retinoblastoma and p53 proteins, and significantly decreased the expression levels of cyclin D1 and cyclin-dependent kinase 4 proteins. Additionally, arctigenin was able to induce apoptosis in glioma cells, coupled with increased expression levels of cleaved caspase-3 and the pro-apoptotic BCL2-associated X protein. Furthermore, arctigenin-induced apoptosis was significantly suppressed by the pretreatment of cells with Z-DEVD-FMK, a caspase-3 inhibitor. In conclusion, the results suggest that arctigenin is able to inhibit cell proliferation and may induce apoptosis and cell cycle arrest at the G0/G1 phase in glioma cells. These results warrant further investigation of the anticancer effects of arctigenin in animal models of gliomas.

Biased signalling is an essential feature of TLR4 in glioma cells

A distinct feature of the Toll-like receptor 4 (TLR4) is its ability to trigger both MyD88-dependent and MyD88-independent signalling, culminating in activation of pro-inflammatory NF-κB and/or the antiviral IRF3. Although TLR4 agonists (lipopolysaccharides; LPSs) derived from different bacterial species have different endotoxic activity, the impact of LPS chemotype on the downstream signalling is not fully understood. Notably, different TLR4 agonists exhibit anti-tumoural activity in animal models of glioma, but the underlying molecular mechanisms are largely unknown.Thus, we investigated the impact of LPS chemotype on the signalling events in the human glioma cell line U251. We found that LPS of Escherichia coli origin (LPSEC) leads to NF-κB-biased downstream signalling compared to Salmonella minnesota-derived LPS (LPSSM). Exposure of U251 cells to LPSEC resulted in faster nuclear translocation of the NF-κB subunit p65, higher NF-κB-activity and expression of its targets genes, and higher amount of secreted IL-6 compared to LPSSM. Using super-resolution microscopy we showed that the biased agonism of TLR4 in glioma cells is neither a result of differential regulation of receptor density nor of formation of higher order oligomers. Consistent with previous reports, LPSEC-mediated NF-κB activation led to significantly increased U251 proliferation, whereas LPSSM-induced IRF3 activity negatively influenced their invasiveness. Finally, treatment with methyl-β-cyclodextrin (MCD) selectively increased LPSSM-induced nuclear translocation of p65 and NF-κB activity without affecting IRF3.Our data may explain how TLR4 agonists differently affect glioma cell proliferation and migration.

Suppression of the Eag1 potassium channel sensitizes glioblastoma cells to injury caused by temozolomide

Glioblastoma multiforme (GBM) is the most aggressive type of human primary brain tumor. The standard treatment protocol includes radiotherapy in combination with temozolomide (TMZ). Despite advances in GBM treatment, the survival time of patients diagnosed with glioma is 14.5 months. Regarding tumor biology, various types of cancer cell overexpress the ether à go-go 1 (Eag1) potassium channel. Therefore, the present study examined the role of Eag1 in the cell damage caused by TMZ on the U87MG glioblastoma cell line. Eag1 was inhibited using a channel blocker (astemizole) or silenced by a short-hairpin RNA expression vector (pKv10.1-3). pKv10.1-3 (0.2 µg) improved the Eag1 silencing caused by 250 µM TMZ, as determined by reverse transcription-quantitative polymerase chain reaction and immunocytochemistry. Additionally, inhibiting Eag1 with the vector or astemizole (5 µM) reduced glioblastoma cell viability and sensitized cells to TMZ. Cell viability decreased by 63% for pKv10.1-3 + TMZ compared with 34% for TMZ alone, and by 77% for astemizole + TMZ compared with 46% for TMZ alone, as determined by MTT assay. In addition, both the vector and astemizole increased the apoptosis rate of glioblastoma cells triggered by TMZ, as determined by an Annexin V apoptosis assay. Collectively, the current data reveal that Eag1 has a role in the damage caused to glioblastoma by TMZ. Furthermore, suppression of this channel can improve the action of TMZ on U87MG glioblastoma cells. Thus, silencing Eag1 is a promising strategy to improve GBM treatment and merits additional studies in animal models of glioma.

Abstract 1670: Radiosensitization of glioma cells by the ketone body β-hydroxybutyrate is associated with enhanced cell cycle arrest in the G2/M phase

Abstract Glioblastoma multiforme (GBM) is an aggressive primary brain tumor with a 5 year survival rate of 25% in children and less than 10% in adults. Improvement in the prognosis of GBM patients requires the development of new therapeutic approaches. One emerging strategy is to target aberrant cell metabolism, a trait shared by virtually all tumor cells. The ketogenic diet (KD), a high fat, low carbohydrate and protein metabolic therapy has been shown to prolong survival in animal glioma models, and when used in conjunction with radiation cured 9 of 11 mice of their implanted tumors. We have also shown that the KD alters hypoxia, angiogenesis, and other hallmarks of glioma progression. To elucidate the underlying mechanisms through which ketones exert their effects on glioma, we are doing analyses of the effect of β-hydroxybutyrate (βHB), the most prevalent ketone body synthesized during ketosis, on glioma cells in vitro. We found that βHB both alone and in conjunction with radiation significantly inhibits proliferation of human and mouse glioma cells and human glioma stem cells (GSC). Alterations in passage through the cell cycle may affect proliferation, and cells are also more sensitive to radiation in the G2/M cell cycle phase. We therefore analyzed the effect of βHB on cell cycle distribution of cells treated with βHB and/or radiation. An analysis of cell cycle status by flow cytometry demonstrated that treating the GSC line L0 with 5mM βHB in combination with 4 Gy of radiation significantly increased the number of cells in G2/M cell cycle arrest. In GL261-Luc2 mouse glioma cells, 5mM βHB alone significantly enhanced G2/M cell cycle arrest, which could lead to radiosensitization. Currently we are analyzing proteins involved in cell cycle progression and apoptosis to better understand βHB mediated changes in growth and radioresistance. In summary, these data provide insight to the radiosensitization and anti-proliferative mechanisms of βHB and may hold implications for the use of the KD in the treatment of GBM. Citation Format: Helena B. Silva-Nichols, Alex P. Rossi, Eric C. Woolf, Marshall J. Fairres, Loic P. Deleyrolle, Brent A. Reynolds, Adrienne C. Scheck. Radiosensitization of glioma cells by the ketone body β-hydroxybutyrate is associated with enhanced cell cycle arrest in the G2/M phase. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1670.

656 - Hydrolyzed rutin in an animal model of human glioma

656 - Hydrolyzed rutin in an animal model of human glioma

Fatty acid oxidation is required for the respiration and proliferation of malignant glioma cells.

Background.Glioma is the most common form of primary malignant brain tumor in adults, with approximately 4 cases per 100 000 people each year. Gliomas, like many tumors, are thought to primarily metabolize glucose for energy production; however, the reliance upon glycolysis has recently been called into question. In this study, we aimed to identify the metabolic fuel requirements of human glioma cells.Methods.We used database searches and tissue culture resources to evaluate genotype and protein expression, tracked oxygen consumption rates to study metabolic responses to various substrates, performed histochemical techniques and fluorescence-activated cell sorting-based mitotic profiling to study cellular proliferation rates, and employed an animal model of malignant glioma to evaluate a new therapeutic intervention.Results.We observed the presence of enzymes required for fatty acid oxidation within human glioma tissues. In addition, we demonstrated that this metabolic pathway is a major contributor to aerobic respiration in primary-cultured cells isolated from human glioma and grown under serum-free conditions. Moreover, inhibiting fatty acid oxidation reduces proliferative activity in these primary-cultured cells and prolongs survival in a syngeneic mouse model of malignant glioma.Conclusions.Fatty acid oxidation enzymes are present and active within glioma tissues. Targeting this metabolic pathway reduces energy production and cellular proliferation in glioma cells. The drug etomoxir may provide therapeutic benefit to patients with malignant glioma. In addition, the expression of fatty acid oxidation enzymes may provide prognostic indicators for clinical practice.

678. Neural Stem Cell-Based Gene Therapy for Malignant Brain Tumor by Doubl-Stranded Adeno-Associated Viral Vectors

Despite intensive efforts exploring gene therapeutic approaches for human cancers, the clinical results in treating of malignant brain tumors are still poor. We demonstrated that the combined therapeutic strategy by using an adenoviral vector carrying the thymidine kinase (suicidal) gene and an adeno-associated viral vector carrying the angiostatin gene achieved better therapeutic results. However, the malignant brain tumor could not be completely eradicated. Therefore, novel vectors and strategies are urgently needed. Neural stem cells (NSCs) may be used either for cell replacement or for gene delivery vehicles in neurodegenerative diseases. The expression of therapeutic proteins by NSCs can be enhanced by viral-mediated gene transfer, though the effects of several common recombinant viruses on primary progenitor cell populations have not been widely tested. The goal of this study is to identify and develop a safe and effective NSCs-based gene therapy protocol to treat malignant glioma. We examined what seral type of the novel double-stranded recombinant adeno-associated viral (AAV) vector can efficiently deliver anti-angiogenic genes and suicidal gene into the rat NSCs. Meanwhile, we investigated the gene product delivered by the genetically engineered NSCs in the adult normal brain. Then, we injected genetically engineered NSCs into different sites of the animal model of human malignant glioma. Flowcytometry data showed that AAV2 had higher transduction efficiency in neural stem cells. The NSCs were engineered to express Decorin and Angiostatin by AAV2, and then injected into the hemisphere contralateral to tumor cell implantation. We demonstrated that genetically-modified NSCs have the capacity to migrate into brain tumors. The therapeutic efficacy of genetically-modified NSCs is remaining explored.

Combination of vatalanib and a 20-HETE synthesis inhibitor results in decreased tumor growth in an animal model of human glioma.

BackgroundDue to the hypervascular nature of glioblastoma (GBM), antiangiogenic treatments, such as vatalanib, have been added as an adjuvant to control angiogenesis and tumor growth. However, evidence of progressive tumor growth and resistance to antiangiogenic treatment has been observed. To counter the unwanted effect of vatalanib on GBM growth, we have added a new agent known as N-hydroxy-N′-(4-butyl-2 methylphenyl)formamidine (HET0016), which is a selective inhibitor of 20-hydroxyeicosatetraenoic acid (20-HETE) synthesis. The aims of the studies were to determine 1) whether the addition of HET0016 can attenuate the unwanted effect of vatalanib on tumor growth and 2) whether the treatment schedule would have a crucial impact on controlling GBM.MethodsU251 human glioma cells (4×105) were implanted orthotopically. Two different treatment schedules were investigated. Treatment starting on day 8 (8–21 days treatment) of the tumor implantation was to mimic treatment following detection of tumor, where tumor would have hypoxic microenvironment and well-developed neovascularization. Drug treatment starting on the same day of tumor implantation (0–21 days treatment) was to mimic cases following radiation therapy or surgery. There were four different treatment groups: vehicle, vatalanib (oral treatment 50 mg/kg/d), HET0016 (intraperitoneal treatment 10 mg/kg/d), and combined (vatalanib and HET0016). Following scheduled treatments, all animals underwent magnetic resonance imaging on day 22, followed by euthanasia. Brain specimens were equally divided for immunohistochemistry and protein array analysis.ResultsOur results demonstrated a trend that HET0016, alone or in combination with vatalanib, is capable of controlling the tumor growth compared with that of vatalanib alone, indicating attenuation of the unwanted effect of vatalanib. When both vatalanib and HET0016 were administered together on the day of the tumor implantation (0–21 days treatment), tumor volume, tumor blood volume, permeability, extravascular and extracellular space volume, tumor cell proliferation, and cell migration were decreased compared with that of the vehicle-treated group.ConclusionHET0016 is capable of controlling tumor growth and migration, but these effects are dependent on the timing of drug administration. The addition of HET0016 to vatalanib may attenuate the unwanted effect of vatalanib.

ATPS-77THE KETONE BODY ß-HYDROXYBUTYRATE RADIOSENSITIZES GLIOBLASTOMA MULTIFORME STEM CELLS

Glioblastoma multiforme (GBM) is a malignant brain tumor that typically recurs following standard treatment with surgery, radiation and chemotherapy. A rare population of radio-chemoresistant glioma stem cells (GSCs) is implicated in tumor repopulation and progression following treatment. Improvement in the prognosis of these patients requires the development of new therapeutic approaches. One emerging strategy is to target aberrant cell metabolism, a trait shared by virtually all tumor cells. The ketogenic diet (KD), a high fat, low carbohydrate and protein metabolic therapy has demonstrated anti-tumor effects alone, and in combination with radiation. The KD prolongs survival in animal glioma models, and when used in conjunction with radiation 9 of 11 mice were cured of their implanted tumors. We are now extending this work to determine if the ketone body, β-hydroxybutyrate (βHB) radiosensitizes human GSCs in vitro. The effects of βHB on growth were examined at various βHB concentrations in two GSC lines, L0 and L1. Both 5mM and 10mM βHB significantly reduced GSC growth. When coupled with 4Gy of radiation, 5mM βHB treatment significantly reduced L1 neurosphere formation and frequency, more than either βHB treatment or radiation alone. In L0, combination treatment significantly reduced neurosphere formation and frequency, more than radiation alone. To explore underlying mechanisms we are investigating the effect of βHB on DNA damage and repair in GSCs. Additionally, we are determining the effect of βHB on hypoxia within GSC neurospheres by analyzing expression of hypoxia inducible factor 1 alpha and the hypoxic marker carbonic anhydrase 9. We previously showed that the KD reduced the expression of these genes in a mouse glioma model. Taken altogether, these data suggest that βHB may sensitize GBMs to radiation by inhibiting growth of both glioma cells and GSCs, which may have implications for its use to treat tumor recurrence.

ATPS-83REPURPOSING MEBENDAZOLE AS A REPLACEMENT FOR VINCRISTINE FOR THE TREATMENT OF BRAIN TUMORS

The microtubule inhibitor vincristine is currently used to treat a number of brain tumors, including newly diagnosed diffuse low-grade and anaplastic glioma. Vincristine however does not penetrate well into brain tumor tissue and moreover, it displays dose-limiting toxicities, including peripheral neuropathy. Mebendazole, an anti-helminthic with an excellent safety profile, has recently been shown to display strong therapeutic efficacy in animal models of both glioma and medulloblastoma (Bai R-Y et al, 2011 and Bai R-Y et al, 2014). In addition, appropriate formulations of mebendazole yield therapeutically effective concentrations in the brain (Bai R-Y et al, 2015). Mebendazole has been shown to inhibit microtubule formation, but it is not known whether its potency against tumor cells is mediated by this microtubule inhibitory effect. To investigate this, we examined the effects of mebendazole on GL261 glioblastoma cell viability, metaphase arrest and microtubule polymerization and found that the effective concentrations to inhibit these functions are very similar. In addition, using mebendazole as a seed for the NCI COMPARE program revealed that the top-scoring drugs were highly enriched in microtubule-targeting drugs. Taken together, these results indicate that the cell toxicity of mebendazole indeed is caused by inhibiting microtubule formation. We also have compared the therapeutic efficacies of mebendazole and vincristine against GL261 orthotopic tumors at their respective maximum tolerated doses (respectively 100 mg/kg/day and 1 mg/kg/week). We found that mebendazole showed a 61% increase in animal survival time, whereas vincristine failed to show any efficacy. However, we did observe significant neuropathy (as measured by sensory allodynia) induced by mebendazole treatment, similar to that caused by vincristine. In conclusion, our results strongly support the clinical use of mebendazole as a replacement for vincristine for the treatment of brain tumors.

2-Hydoxyglutarate: D/Riving Pathology in gLiomaS.

Common pathways and mechanisms can be found in both cancers and inborn errors of metabolism. 2-Hydroxyglutarate (2-HG) acidurias and isocitrate dehydrogenase (IDH) 1/2 mutant tumors are examples of this phenomenon. 2-HG can exist in two chiral forms, D(R)-2-HG and L(S)-2-HG, which are elevated in D- and L-acidurias, respectively. D-2-HG was subsequently discovered to be synthesized in IDH 1/2 mutant tumors including ∼70% of intermediate-grade gliomas and secondary glioblastomas (GBM). Recent studies have revealed that L-2-HG is generated in hypoxia in IDH wild-type tumors. Both 2-HG enantiomers have similar structures as α-ketoglutarate (α-KG) and can competitively inhibit α-KG-dependent enzymes. This inhibition modulates numerous cellular processes, including histone and DNA methylation, and can ultimately impact oncogenesis. D-2-HG can be detected in vivo in glioma patients and animal models using advanced imaging modalities. Finally, pharmacologic inhibitors of mutant IDH 1/2 attenuate the production of D-2-HG and show great promise as therapeutic agents.

Abstract 240: The ketogenic diet alters the expression of microRNAs that play key roles in tumor development

Abstract Glioblastoma multiforme (GBM) is the most common, and most aggressive, primary brain tumour in adults. Despite advances in surgical techniques and improvements in radiotherapy and chemotherapy protocols, the prognosis for patients with these tumors is extremely poor, with a median survival of 14 months in patients receiving maximal aggressive treatment. Although several targeted therapies have been tried in GBM, these have proved to be ineffective, with no targeted agent showing an improvement in overall survival. These dismal facts highlight the need for new therapeutic approaches. One strategy is to take advantage of the metabolic dysregulation of tumor cells and their high dependence on glucose for survival using dietary measures. This approach would target the tumor as a whole and help to overcome the problems associated with tumour heterogeneity. One such approach is the ketogenic diet, (KD). This high fat, low carbohydrate diet has been used clinically for the management of refractory paediatric epilepsy and has been shown to extend survival in a number of animal models of glioma. Furthermore, studies using the GL261-luc2 syngeneic intracranial model of malignant glioma also highlighted the ability of the KD to potentiate the effects of chemotherapy and radiotherapy. However, the mechanisms underlying these responses are poorly understood. MicroRNAs (miRs) are small noncoding single-stranded RNAs that are capable of altering mRNA expression of target genes and hence have crucial roles in a diverse array of biological processes. To investigate changes in the expression of miRs during the KD, we profiled brain tumors from mice fed either a KD or standard diet for differential miR expression using the miScript miRNA PCR Array Mouse Brain Cancer (Qiagen Ltd., Manchester, UK). We observed that miRs with an oncogenic potential were generally downregulated whereas those with a tumor suppressor function were upregulated in tumors from mice fed KD compared to tumors from mice fed SD. Upregulation of miR 296-5p is of particular interest, since its target, Pin 1, a Ser/Thr-Pro cis/trans isomerase, is a regulator of cell metabolism through its interaction with glycolytic enzymes for the Warburg effect. Furthermore, miR 296-5p has been implicated in DNA damaging chemotherapy resistance in breast and lung cancer cell lines. Taken together our results suggest that the KD could inhibit tumor growth and contribute to therapy sensitivity through differential expression of miRs. Citation Format: Julia Pazmandi, Kevin S. O'Neill, Adrienne C. Scheck, Peter W. Szlosarek, Eric C. Woolf, Kenneth S. Brooks, Nelofer Syed. The ketogenic diet alters the expression of microRNAs that play key roles in tumor development. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 240. doi:10.1158/1538-7445.AM2015-240

Image guided drug release from pH-sensitive Ion channel-functionalized stealth liposomes into an in vivo glioblastoma model

Liposomal drug delivery vehicles are promising nanomedicine tools for bringing cytotoxic drugs to cancerous tissues selectively. However, the triggered cargo release from liposomes in response to a target-specific stimulus has remained elusive. We report on functionalizing stealth-liposomes with an engineered ion channel and using these liposomes in vivo for releasing an imaging agent into a cerebral glioma rodent model. If the ambient pH drops below a threshold value, the channel generates temporary pores on the liposomes, thus allowing leakage of the intraluminal medicines. By using magnetic resonance spectroscopy and imaging, we show that engineered liposomes can detect the mildly acidic pH of the tumor microenvironment with 0.2 pH unit precision and they release their content into C6 glioma tumors selectively, in vivo. A drug delivery system with this level of sensitivity and selectivity to environmental stimuli may well serve as an optimal tool for environmentally-triggered and image-guided drug release. From the Clinical EditorCancer remains a leading cause of mortality worldwide. With advances in science, delivery systems of anti-cancer drugs have also become sophisticated. In this article, the authors designed and characterized functionalized liposomal vehicles, which would release the drug payload in a highly sensitive manner in response to a change in pH environment in an animal glioma model. The novel data would enable better future designs of drug delivery systems.

Enhancement in blood-tumor barrier permeability and delivery of liposomal doxorubicin using focused ultrasound and microbubbles: evaluation during tumor progression in a rat glioma model

Effective drug delivery to brain tumors is often challenging because of the heterogeneous permeability of the ‘blood tumor barrier’ (BTB) along with other factors such as increased interstitial pressure and drug efflux pumps. Focused ultrasound (FUS) combined with microbubbles can enhance the permeability of the BTB in brain tumors, as well as the blood–brain barrier in the surrounding tissue. In this study, dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) was used to characterize the FUS-induced permeability changes of the BTB in a rat glioma model at different times after implantation. 9L gliosarcoma cells were implanted in both hemispheres in male rats. At day 9, 14, or 17 days after implantation, FUS-induced BTB disruption using 690 kHz ultrasound and definity microbubbles was performed in one tumor in each animal. Before FUS, liposomal doxorubicin was administered at a dose of 5.67 mg kg−1. This chemotherapy agent was previously shown to improve survival in animal glioma models. The transfer coefficient Ktrans describing extravasation of the MRI contrast agent Gd-DTPA was measured via DCE-MRI before and after sonication. We found that tumor doxorubicin concentrations increased monotonically (823 ± 600, 1817 ± 732 and 2432 ± 448 ng g−1) in the control tumors at 9, 14 and 17 d. With FUS-induced BTB disruption, the doxorubicin concentrations were enhanced significantly (P < 0.05, P < 0.01, and P < 0.0001 at days 9, 14, and 17, respectively) and were greater than the control tumors by a factor of two or more (2222 ± 784, 3687 ± 796 and 5658 ± 821 ng g−1) regardless of the stage of tumor growth. The transfer coefficient Ktrans was significantly (P < 0.05) enhanced compared to control tumors only at day 9 but not at day 14 or 17. These results suggest that FUS-induced enhancements in tumor drug delivery are relatively consistent over time, at least in this tumor model. These results are encouraging for the use of large drug carriers, as they suggest that even large/late-stage tumors can benefit from FUS-induced drug enhancement. Corresponding enhancements in Ktrans were found to be variable in large/late-stage tumors and not significantly different than controls, perhaps reflecting the size mismatch between the liposomal drug (~100 nm) and Gd-DTPA (molecular weight: 938 Da; hydrodynamic diameter: ≃2 nm). It may be necessary to use a larger MRI contrast agent to effectively evaluate the sonication-induced enhanced permeabilization in large/late-stage tumors when a large drug carrier such as a liposome is used.

The ketogenic diet for the treatment of malignant glioma

Advances in our understanding of glioma biology has led to an increase in targeted therapies in preclinical and clinical trials; however, cellular heterogeneity often precludes the targeted molecules from being found on all glioma cells, thus reducing the efficacy of these treatments. In contrast, one trait shared by virtually all tumor cells is altered (dysregulated) metabolism. Tumor cells have an increased reliance on glucose, suggesting that treatments affecting cellular metabolism may be an effective method to improve current therapies. Indeed, metabolism has been a focus of cancer research in the last few years, as many pathways long associated with tumor growth have been found to intersect metabolic pathways in the cell. The ketogenic diet (high fat, low carbohydrate and protein), caloric restriction, and fasting all cause a metabolic change, specifically, a reduction in blood glucose and an increase in blood ketones. We, and others, have demonstrated that these metabolic changes improve survival in animal models of malignant gliomas and can potentiate the anti-tumor effect of chemotherapies and radiation treatment. In this review we discuss the use of metabolic alteration for the treatment of malignant brain tumors.

TM-15 * GLUTAMINE BASED PET IMAGING FACILITATES ENHANCED METABOLIC DETECTION OF GLIOMAS IN VIVO

Glutamine is the most abundant plasma amino acid and many cancers show altered glutamine metabolism. We evaluated glutamine uptake and metabolism in gliomas using PET imaging and biochemical approaches. We demonstrate that glutamine is a key TCA cycle anaplerotic substrate and is metabolized to generate 2-HG in IDH1-mutant gliomas. PET imaging with18F-labeled glutamine (18F-FGln) showed high uptake in gliomas in vivo but low background uptake in the surrounding brain in RCAS-PDGF/PTEN null and IDH1-mutant glioma animal models, facilitating clear tumor delineation in contrast to that seen with 18F-FDG. We did not observe 18F-FGln uptake in animals with neuroinflammation or animals with a disrupted BBB. Further, 18F-FGln uptake was specifically reduced on chemo/radiation therapy. Finally, 18F-FGln showed high avidity in human glioma with low uptake in the surrounding brain. These data suggest that 18F-FGln is specifically taken up by gliomas, can be used to assess the metabolic state of gliomas in vivo and may serve as a valuable tool in the clinical management of gliomas.

Phenolic compounds oleuropein and hydroxytyrosol exert differential effects on glioma development via antioxidant defense systems

Oxidative stress is involved in many of the stages of tumorigenesis, and the administration of exogenous antioxidants seems to modulate them. Thus, it has been described that oleuropein and hydroxytyrosol exert important anti-cancer activities. We analyse in vivo the anti-tumor properties of both phenolic compounds in an animal model of glioma, and their effects on oxidative stress, enzymatic and non-enzymatic antioxidant defence systems and on several biochemical biomarkers. Hydroxytyrosol, but not oleuropein nor the mixture of both compounds, inhibits tumor growth through mechanisms that involve enzymatic and non-enzymatic antioxidant defences, as demonstrated by a decrease in lipid peroxidation and protein oxidation levels. Furthermore, hydroxytyrosol maintains the non-enzymatic antioxidants as in healthy animals, and positively modifies the enzymatic antioxidants. However, hydroxytyrosol probably acts not as an antioxidant, but through other mechanisms that only indirectly modify the redox status. Finally, these compounds yield few adverse effects related to changes in hepatic enzymes.

Abstract 4196: An RNA aptamer-based approach for human glioma treatment

Abstract Gliomas are the most common primary central nervous system tumors with a dismal prognosis. Despite recent advances in surgery, radiotherapy, and chemotherapy, current treatment regimens have a modest survival benefit. Due to a combination of its complex phenotype and organ-specific clinical manifestations, efforts to refine glioma treatment with targeted therapies have largely been frustrated. Hence, finding specific ligands capable of detecting and measuring the altered pattern of gene expression and to discriminate between different tumor phenotypes, is a strategic and plausible objective for the diagnosis and therapy of glioma. To date, antibody-based approaches have been developed for in vivo applications but, in most cases, they show toxicity in vivo and do not reach adequate sensitivity. Thanks to their unique characteristics (low size, good affinity for the target, no immunogenicity, chemical structures that can be easily modified to improve their in vivo applications), single-stranded nucleic acid ligand molecules, named aptamers, represent a valid alternative to antibodies for in vivo targeted recognition as therapeutics or delivery agents for nanoparticles, small interfering RNAs, chemotherapeutic cargos and molecular imaging probes. By applying cell-SELEX on cancer cells we have generated and characterized different nuclease resistant 2′fluoro-pyrimidines RNA aptamers as high affinity ligands and inhibitors of human receptor tyrosine kinases (RTKs) with a crucial role in glioma, including EGFR, EGFRVIII, EphB3, Axl and PDGFRβ. All the aptamers are able to efficiently and specifically inhibit tumor growth in cell based assays and in animal models of human glioma. Further, some of these aptamers strongly cooperates in inducing inhibition of tumor growth thus providing the basis for further development of antitumor combination therapies. Citation Format: Simona Camorani, Carla L. Esposito, Silvia Catuogno, Paola Amero, Anna Rienzo, Gennaro De Vita, Gerolama Condorelli, Vittorio de Franciscis, Laura Cerchia. An RNA aptamer-based approach for human glioma treatment. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4196. doi:10.1158/1538-7445.AM2014-4196

CPEB1 modulates differentiation of glioma stem cells via downregulation of HES1 and SIRT1 expression.

Glioma stemness has been recognized as the most important reason for glioma relapse and drug resistance. Differentiation of glioma stem cells (GSCs) has been implicated as a novel approach to target recurrent glioma. However, the detailed molecular mechanism involved in the differentiation of GSCs has not yet been elucidated. This study identified CPEB1 as the key modulator that induces the differentiation of GSCs at the post-transcriptional level. Gain and loss of function experiments showed that CPEB1 expression reduced sphere formation ability and the expression of stemness markers such as Nestin and Notch. To elucidate the detailed molecular mechanism underlying the action of CPEB1, we investigated the interacting ribonome of the CPEB1 complex using a Ribonomics approach. CPEB1 specifically suppressed the translation of HES1 and SIRT1 by interacting with a cytoplasmic polyadenylation element. The expression profile of CPEB1 negatively correlated with overall survival in glioma patients. Overexpression of CPEB1 decreased the number of GSCs in an orthotopically implanted glioma animal model. These results suggest that CPEB1-mediated translational control is essential for the differentiation of GSCs and provides novel therapeutic concepts for differentiation therapy.