Glioma Invasiveness Responds Variably to Irradiation in a Co-Culture Model

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nakamura, Jean L.; Haas-Kogan, Daphne A.; Department of Neurological Surgery, University of California-San Francisco, San Francisco, CA

2007-11-01

Purpose: We developed a co-culture system to quantitate the growth and invasion of human malignant gliomas into a background of confluent normal human astrocytes, then used this assay to assess independently the effects of irradiating both cell types on glioma invasion. Methods and Materials: Enhanced green fluorescent protein (EGFP)-labeled immortalized human astrocytes, human malignant glioma cells, or transformed human astrocytes were focally plated onto a confluent layer of normal human astrocytes, and the invasiveness of EGFP-labeled cells was scored after 96 h. To address the consequences of irradiation on glioma invasion, the invasiveness of irradiated glioma cell lines and irradiatedmore » astrocytic backgrounds was assessed. Fluorescence-activated cell sorting was used to quantitate the total number of EGFP-labeled cells. Results: Growth in the co-culture assay consistently reflected transformation states of the plated cells. Immortalized, but untransformed human astrocytes failed even to establish growth on confluent normal human astrocytes. In contrast, all malignant human glioma cell lines and transformed human astrocytes demonstrated various degrees of infiltration into the astrocytic bed. Irradiation failed to alter the invasiveness of U87, A172, and U373. A 1-Gy dose slightly reduced the invasiveness of U251 MG by 75% (p < 0.05 by one-way analysis of variance and post hoc Neuman-Keuls), without reducing total cell numbers. Independently irradiating the human astrocytic bed did not alter the invasiveness of nonirradiated U251, whereas the matrix metalloproteinase (MMP) inhibitor GM6001 reduced U251 invasiveness in the co-culture assay. Conclusions: Growth in the co-culture assay reflects the transformation status and provides a useful in vitro model for assessing invasiveness. Human glioma invasiveness in the co-culture model responds variably to single low-dose fractions. MMP activity promotes invasiveness in the co-culture model. Reduced invasiveness in irradiated U251 appears to be mediated by MMP-independent mechanisms.« less

Blood glutamate scavengers prolong the survival of rats and mice with brain-implanted gliomas.

PubMed

Ruban, Angela; Berkutzki, Tamara; Cooper, Itzik; Mohar, Boaz; Teichberg, Vivian I

2012-12-01

L-Glutamate (Glu) plays a crucial role in the growth of malignant gliomas. We have established the feasibility of accelerating a naturally occurring brain to-blood Glu efflux by decreasing blood Glu levels with intravenous oxaloacetate, the respective Glu co-substrate of the blood resident enzyme humane glutamate–oxaloacetate transaminase(hGOT). We wished to demonstrate that blood Glu scavenging provides neuroprotection in the case of glioma.We now describe the neuroprotective effects of blood Glu scavenging in a fatal condition such as brain-implanted C6 glioma in rats and brain-implanted human U87 MG glioma in nude mice. Rat (C-6) or human (U87) glioma cells were grafted stereotactically in the brain of rats or mice. After development of tumors, the animals were drinking oxaloacetate with or without injections of hGOT. In addition, mice were treated with combination treatment, which included drinking oxaloacetate with intracutaneous injections of hGOT and intraperitoneal injection of Temozolomide. Animals drinking oxaloacetate with or without injections of hGOT displayed a smaller tumor volume, reduced invasiveness and prolonged survival than control animals drinking saline. These effects were significantly enhanced by Temozolomide in mice, which increased survival by 237%. This is the first demonstration of blood Glu scavenging in brain cancer, and because of its safety, is likely to be of clinical significance for the future treatment of human gliomas. As we demonstrated, the blood glutamate scavenging treatment in combination with TMZ could be a good candidate or as an alternative treatment to the patients that do not respond to TMZ.

An endogenous and ectopic expression of metabotropic glutamate receptor 8 (mGluR8) inhibits proliferation and increases chemosensitivity of human neuroblastoma and glioma cells.

PubMed

Jantas, Danuta; Grygier, Beata; Gołda, Sławomir; Chwastek, Jakub; Zatorska, Justyna; Tertil, Magdalena

2018-06-06

The present study aimed to determine the role of metabotropic glutamate receptor 8 (mGluR8) in tumor biology. Using various molecular approaches (RNAi or GRM8 cDNA), cell clones with downregulated (human neuroblastoma SH-SY5Y and human glioma LN229) or overexpressed (human glioma U87-MG and LN18 cell lines) mGluR8 were generated. Next, comparative studies on cell proliferation and migration rates, induction of apoptosis and chemosensitivity were performed among these clones. The mGluR8-downregulated SH-SY5Y clones proliferated faster and were more resistant to cytotoxic action of staurosporine, doxorubicin, irinotecan and cisplatin when compared to control cells. Moreover, these clones were characterized by a lower activity of caspases, calpains and some kinases (GSK-3β, Akt and JNK). The mGluR8-downregulated LN229 clones migrated faster and were less prone to cell-damaging effect of staurosporine and irinotecan when compared with relevant control cells. In contrast, in GRM8-overexpressing U87-MG and LN18 clones, a decreased cell proliferation, increased apoptosis and elevated vulnerability to some cytotoxic agents were found. Altogether, our in vitro data for the first time evidenced a tumor suppressor and chemosensitizing role of mGluR8. Copyright © 2018 Elsevier B.V. All rights reserved.

Antitumor activity of (2E,5Z)-5-(2-hydroxybenzylidene)-2-((4-phenoxyphenyl)imino) thiazolidin-4-one, a novel microtubule-depolymerizing agent, in U87MG human glioblastoma cells and corresponding mouse xenograft model.

PubMed

Zhang, Qiu; Liu, Xiaojun; Li, Xiue; Li, Changlong; Zhou, Hongyu; Yan, Bing

2013-01-01

Glioblastoma is the most lethal brain cancer. In spite of intensive therapy, the prognosis of patients with glioblastoma is very poor. To discover novel therapeutic agents, we screened a combinatorial compound library containing 372 thiazolidinone compounds using U87MG human glioblastoma cells. (2E,5Z)-5-(2-hydroxybenzylidene)-2-((4-phenoxyphenyl)imino) thiazolidin-4-one (HBPT) was identified as the most potent anti-glioblastoma compound. HBPT inhibits U87MG human glioblastoma cell proliferation with an IC50 of 20 μM, which is almost 5-fold more potent than temozolomide (a widely used drug for treating malignant glioma in the clinic). Mechanistic investigation demonstrated that HBPT is a novel microtubule-depolymerizing agent, which arrests cancer cells at the G2/M phase of the cell cycle and induces cell apoptosis. In the mouse U87MG xenograft model, HBPT elicits a robust tumor inhibitory effect. More importantly, no obvious toxicity was observed for HBPT therapy in animal experiments. These findings indicate that HBPT has the potential to be developed as a novel agent for the treatment of glioblastoma. [Supplementary Tables: available only at http://dx.doi.org/10.1254/jphs.13064FP].

Intermittent Induction of HIF-1α Produces Lasting Effects on Malignant Progression Independent of Its Continued Expression

PubMed Central

Choi, Hyunsung; Gillespie, David L.; Berg, Shauna; Rice, Christopher; Couldwell, Sandrine; Gu, Jie; Colman, Howard; Jensen, Randy L.; Huang, L. Eric

2015-01-01

Dysregulation of hypoxia-inducible transcription factors HIF-1α and HIF-2α correlates with poor prognosis in human cancers; yet, divergent and sometimes opposing activities of these factors in cancer biology have been observed. Adding to this complexity is that HIF-1α apparently possesses tumor-suppressing activities, as indicated by the loss-of-function mutations or even homozygous deletion of HIF1A in certain human cancers. As a step towards understanding this complexity, we employed 8-week intermittent induction of a stable HIF-1α variant, HIF1α(PP), in various cancer cell lines and examined the effects on malignant progression in xenografts of immunocompromised mice in comparison to those of HIF2α(PP). Although 8-week treatment led to eventual loss of HIF1α(PP) expression, treated osteosarcoma U-2 OS cells acquired tumorigenicity in the subcutaneous tissue. Furthermore, the prior treatment resulted in widespread invasion of malignant glioma U-87 MG cells in the mouse brain and sustained growth of U-118 MG glioma cells. The lasting effects of HIF-1α on malignant progression are specific because neither HIF2α(PP) nor β-galactosidase yielded similar effects. By contrast, transient expression of HIF1α(PP) in U-87 MG cells or constitutive expression of HIF1α(PP) but not HIF2α(PP) in a patient-derived glioma sphere culture inhibited tumor growth and spread. Our results indicate that intermittent induction of HIF-1α produces lasting effects on malignant progression even at its own expense. PMID:25893706

Recombinant epidermal growth factor-like domain-1 from coagulation factor VII functionalized iron oxide nanoparticles for targeted glioma magnetic resonance imaging.

PubMed

Liu, Heng; Chen, Xiao; Xue, Wei; Chu, Chengchao; Liu, Yu; Tong, Haipeng; Du, Xuesong; Xie, Tian; Liu, Gang; Zhang, Weiguo

The highly infiltrative and invasive nature of glioma cells often leads to blurred tumor margins, resulting in incomplete tumor resection and tumor recurrence. Accurate detection and precise delineation of glioma help in preoperative delineation, surgical planning and survival prediction. In this study, recombinant epidermal growth factor-like domain-1, derived from human coagulation factor VII, was conjugated to iron oxide nanoparticles (IONPs) for targeted glioma magnetic resonance (MR) imaging. The synthesized EGF1-EGFP-IONPs exhibited excellent targeting ability toward tissue factor (TF)-positive U87MG cells and human umbilical vein endothelial cells in vitro, and demonstrated persistent and efficient MR contrast enhancement up to 12 h for preclinical glioma models with high targeting specificity in vivo. They hold great potential for clinical translation and developing targeted theranostics against brain glioma.

HULC long noncoding RNA silencing suppresses angiogenesis by regulating ESM-1 via the PI3K/Akt/mTOR signaling pathway in human gliomas.

PubMed

Zhu, Yu; Zhang, Xuebin; Qi, Lisha; Cai, Ying; Yang, Ping; Xuan, Geng; Jiang, Yuan

2016-03-22

Tumor angiogenesis plays a critical role in the tumor progression. Highly upregulated in liver cancer (HULC) is a long noncoding RNA (lncRNA) that acts as an oncogene in gliomas. We found that HULC, vascular endothelial growth factor (VEGF), and ESM-1 (endothelial cell specific molecule 1) expression and microvessel density were positively correlated with grade dependency in glioma patient tissues, and that HULC silencing suppressed angiogenesis by inhibiting glioma cells proliferation and invasion. This process induced anoikis and blocked the cell cycle at G1/S phase via the PI3K/Akt/mTOR signaling pathway, thus regulating the tumor-related genes involved in the above biological behavior in human glioma U87MG and U251 cells. However, these effects were reversed by ESM-1 overexpression, suggesting a mediating role of ESM-1 in the pro-angiogenesis effect of HULC. Our results define the mechanism of the pro-angiogenesis activity of HULC, which shows potential for application as a therapeutic target in glioma.

Quantitative evaluation of malignant gliomas damage induced by photoactivation of IR700 dye

NASA Astrophysics Data System (ADS)

Sakuma, Morito; Kita, Sayaka; Higuchi, Hideo

2016-01-01

The processes involved in malignant gliomas damage were quantitatively evaluated by microscopy. The near-infrared fluorescent dye IR700 that is conjugated to an anti-CD133 antibody (IR700-CD133) specifically targets malignant gliomas (U87MG) and stem cells (BT142) and is endocytosed into the cells. The gliomas are then photodamaged by the release of reactive oxygen species (ROS) and the heat induced by illumination of IR700 by a red laser, and the motility of the vesicles within these cells is altered as a result of cellular damage. To investigate these changes in motility, we developed a new method that measures fluctuations in the intensity of phase-contrast images obtained from small areas within cells. The intensity fluctuation in U87MG cells gradually decreased as cell damage progressed, whereas the fluctuation in BT142 cells increased. The endocytosed IR700 dye was co-localized in acidic organelles such as endosomes and lysosomes. The pH in U87MG cells, as monitored by a pH indicator, was decreased and then gradually increased by the illumination of IR700, while the pH in BT142 cells increased monotonically. In these experiments, the processes of cell damage were quantitatively evaluated according to the motility of vesicles and changes in pH.

Human cytomegalovirus inhibits apoptosis by regulating the activating transcription factor 5 signaling pathway in human malignant glioma cells

PubMed Central

WANG, TONGMEI; QIAN, DONGMENG; HU, MING; LI, LING; ZHANG, LI; CHEN, HAO; YANG, RUI; WANG, BIN

2014-01-01

The activating transcription factor 5 (ATF5), also termed ATFx, is a member of the ATF/cAMP response element-binding protein (CREB) family of basic zipper proteins. ATF5 is an anti-apoptotic protein that is highly expressed in malignant glioma and is essential for glioma cell survival. Accumulating evidence indicates that human malignant gliomas are universally infected with human cytomegalovirus (HCMV). Recent studies have shown that HCMV may be resistant to the induction of apoptosis by disrupting cellular pathways in glioblastoma. To investigate the potential anti-apoptotic function of HCMV in glioma, malignant U87 glioma cells were infected with HCMV. The present study showed that HCMV infection suppressed apoptosis in glioblastoma U87 cells by regulating the expression of ATF5. Furthermore, in glioblastoma U87 cells, HCMV infection induced cellular proliferation in parallel with an increase in the expression level of ATF5 and B-cell lymphoma/leukemia-2 to Bcl-2-associated X protein ratio. Loss of ATF5 function was achieved using a dominant-negative form of ATF5 in U87 cells, whereby cells appeared to grow marginally following HCMV infection when compared with the control. However, the anti-apoptotic ability was appeared to decline in the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay. These results indicate that ATF5 signaling pathways may be important in the anti-apoptotic activity of HCMV-infected glioblastoma cells; therefore, the anti-apoptotic molecular mechanisms of HCMV in human glioblastoma cells were investigated in the current study. Prevention of HCMV infection may present a potential and promising approach for the treatment of malignant gliomas. PMID:25120656

Melatonin inhibits proliferation and invasion via repression of miRNA-155 in glioma cells.

PubMed

Gu, Junyi; Lu, Zhongsheng; Ji, Chenghong; Chen, Yuchao; Liu, Yuzhao; Lei, Zhe; Wang, Longqiang; Zhang, Hong-Tao; Li, Xiangdong

2017-09-01

Melatonin, an indolamine mostly synthesized in the pineal gland, exerts the anti-cancer effect by various mechanisms in glioma cells. Our previous study showed that miR-155 promoted glioma cell proliferation and invasion. However, the question of whether melatonin may inhibit glioma by regulating miRNAs has not yet been addressed. In this study, we found that melatonin (100μM, 1μM and 1nM) significantly inhibited the expression of miR-155 in human glioma cell lines U87, U373 and U251. Especially, the lowest expression of miR-155 was detected in 1μM melatonin-treated glioma cells. Melatonin (1μM) inhibits cell proliferation of U87 by promoting cell apoptosis. Nevertheless, melatonin had no effect on cell cycle distribution of U87 cells. Moreover, U87 cells treated with 1μM melatonin presented significantly lower migration and invasion ability when compared with control cells. Importantly, melatonin inhibited c-MYB expression, and c-MYB knockdown reduced miR-155 expression and migration and invasion in U87 cells. Taken together, for the first time, our findings show that melatonin inhibits miR-155 expression and thereby represses glioma cell proliferation, migration and invasion, and suggest that melatonin may downregulate the expression of miR-155 via repression of c-MYB. This will provide a theoretical basis for revealing the anti-glioma mechanisms of melatonin. Copyright © 2017. Published by Elsevier Masson SAS.

Down-regulation of 14-3-3β exerts anti-cancer effects through inducing ER stress in human glioma U87 cells: Involvement of CHOP–Wnt pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cao, Lei; Lei, Hui; Chang, Ming-Ze

We previously identified 14-3-3β as a tumor-specific isoform of 14-3-3 protein in astrocytoma, but its functional role in glioma cells and underlying mechanisms are poorly understood. In the present study, we investigated the effects of 14-3-3β inhibition in human glioma U87 cells using specific targeted small interfering RNA (siRNA). The results showed that 14-3-3β is highly expressed in U87 cells but not in normal astrocyte SVGp12 cells. Knockdown of 14-3-3β by Si-14-3-3β transfection significantly decreased the cell viability but increased the LDH release in a time-dependent fashion in U87 cells, and these effects were accompanied with G0/G1 cell cycle arrestmore » and apoptosis. In addition, 14-3-3β knockdown induced ER stress in U87 cells, as evidenced by ER calcium release, increased expression of XBP1S mRNA and induction of ER related pro-apoptotic factors. Down-regulation of 14-3-3β significantly decreased the nuclear localization of β-catenin and inhibited Topflash activity, which was shown to be reversely correlated with CHOP. Furthermore, Si-CHOP and sFRP were used to inhibit CHOP and Wnt, respectively. The results showed that the anti-cancer effects of 14-3-3β knockdown in U87 cells were mediated by increased expression of CHOP and followed inhibition of Wnt/β-catenin pathway. In summary, the remarkable efficiency of 14-3-3β knockdown to induce apoptotic cell death in U87 cells may find therapeutic application for the treatment of glioma patients. - Highlights: • Knockdown of 14-3-3β leads to cytotoxicity in human glioma U87 cells. • Knockdown of 14-3-3β induces cell cycle arrest and apoptosis in U87 cells. • Knockdown of 14-3-3β results in ER stress in U87 cells. • Knockdown of 14-3-3β inhibits Wnt/β-catenin pathway via CHOP activation.« less

MAb 806 Enhances the Efficacy of Ionizing Radiation in Glioma Xenografts Expressing the de2-7 Epidermal Growth Factor Receptor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Johns, Terrance G.; McKay, Michael J.; Cvrljevic, Anna N.

2010-10-01

Purpose: Mutations of the epidermal growth factor receptor (EGFR) are common in glioma. The most frequent mutation, de2-7 EGFR/EGFRvIII, occurs in approximately 40% of high-grade gliomas and confers resistance to ionizing radiation (IR). We have previously shown that mAb 806, a novel EGFR-specific antibody, is able to inhibit the growth of U87MG.{Delta}2-7 glioma xenografts expressing the de2-7 EGFR and may have potential as a therapeutic. Methods and Materials: Nude mice bearing U87MG.{Delta}2-7 xenografts were treated with mAb 806 and/or IR. Comparison of tumor volumes, the effect of treatment on angiogenesis as determined by mean vessel density, and expression changes inmore » prosurvival protein pAkt between treatment groups were undertaken. Results: Treatment of mice bearing U87MG.{Delta}2-7 xenografts with mAb 806 and IR resulted in schedule-dependent radiosensitization. Maximal benefit was obtained when antibody treatment was given before irradiation, with the greatest inhibition of both tumor angiogenesis and tumor growth. Combination treatment mediated radiosensitization by selectively blocking the phosphorylation of the prosurvival protein Akt at serine 473, a process that is independent of DNA-dependent protein kinase catalytic subunit. Conclusions: Our results provide a rationale for the use of mAb 806 in combination with IR for the treatment of glioma and potentially other solid tumors bearing the de2-7 EGFR.« less

Methylation Status of the RIZ1 Gene Promoter in Human Glioma Tissues and Cell Lines.

PubMed

Zhang, Chenran; Meng, Wei; Wang, Jiajia; Lu, Yicheng; Hu, Guohan; Hu, Liuhua; Ma, Jie

2017-08-01

Retinoblastoma protein-interacting zinc-finger gene 1 (RIZ1), a strong tumor suppressor, is silenced in many human cancers. Our previous studies showed that RIZ1 expression was negatively correlated with the grade of glioma and was a key predictor of patient survival. Therefore, RIZ1 could be a potential tumor suppressor during glioma pathogenesis, although the mechanism underlying RIZ1 gene inactivation in gliomas is unknown. We investigated the methylation status of the RIZ1 promoter in human glioma tissues and four glioblastoma (GBM) cell lines, and verified the effect of the methyltransferase inhibitor 5-aza-2-deoxycytidine (5-aza-CdR) on RIZ1 transcription and cell proliferation. Methylation-specific PCR (MSP) was performed to determine RIZ1 promoter methylation in human glioma specimens. The correlation between RIZ1 hypermethylation in tumors and clinicopathological features also was analyzed. 5-Aza-CdR treatment was used to reactivate gene expression silenced by hypermethylation in the U87 glioblastoma cell line, and real-time PCR was then used to measure RIZ1 expression. The ability of 5-aza-CdR to inhibit the proliferation of glioma cell lines whose RIZ1 promoters were hypermethylated was measured by bromodeoxyuridine (BrdU) incorporation. Among 51 human glioma specimens, RIZ1 promoter methylation was detected in 23 cases. Clinicopathological evaluation suggested that RIZ1 hypermethylation was negatively associated with tumor grade and patient age (P < 0.05). Hypermethylation of the RIZ1 promoter was detected in the U87 and U251 cell lines. RIZ1 mRNA expression in U87 cells was upregulated after treatment with 5-aza-Cdr, which correlated with inhibition of cell proliferation in a time- and concentration-dependent manner. Promoter hypermethylation may play an important role in the epigenetic silencing of RIZ1 expression in human glioma tissues and GBM cell lines.

High expression of B7-H6 in human glioma tissues promotes tumor progression.

PubMed

Jiang, Tianwei; Wu, Wei; Zhang, Huasheng; Zhang, Xiangsheng; Zhang, Dingding; Wang, Qiang; Huang, Lei; Wang, Ye; Hang, Chunhua

2017-06-06

B7-H6, a new member of B7-family ligand, also known as NCR3LG1, plays an important role in NK cells mediated immune responses. Many studies have shown that it is highly expressed in various human cancers, and its expression levels are significantly associated with cancer patients' clinicopathological parameters and postoperative prognoses. But, still the exact role of B7-H6 expression in human glioma remains elusive. In the present study, we have characterized the B7-H6 expression in the human glioma tissues as well as glioma cell lines, U87 and U251. We observed that B7-H6 was highly expressed in the human glioma tissues, and its expression was significantly associated with cancer progression. By using the RNA interference technology, we successfully ablated B7-H6 expression in human glioma cell lines to further study its contribution towards various biological features of this malignancy. Our study identified that the B7-H6 knockdown in U87 and U251 glioma cells significantly suppressed cell proliferation, migration, invasion, and enhanced apoptosis along with induction of cell cycle arrest. It thus suggested that B7-H6 play an important role in the regulation of the biological behavior of these glioma cells. However, the detailed mechanism of B7-H6 mediated regulation of glioma cancer cell transformation and its prognostic value merits further investigation.

Silencing of cZNF292 circular RNA suppresses human glioma tube formation via the Wnt/β-catenin signaling pathway.

PubMed

Yang, Ping; Qiu, Zhijun; Jiang, Yuan; Dong, Lei; Yang, Wensheng; Gu, Chao; Li, Guang; Zhu, Yu

2016-09-27

CircRNA is a novel type of RNA molecule formed by a covalently closed loop which have no 5'-3' polarity and possess no polyA tail and relatively stable due to the cyclic structure. Therefore, they may serve as potential targets and diagnosis biomarkers for tumor therapy. cZNF292 is an important circular oncogenic RNA and plays a critical role in the progression of tube formation. This study is aimed at exploring the role of cZNF292 in human glioma tube formation and its potential mechanism of action. We found that cZNF292 silencing suppresses tube formation by inhibiting glioma cell proliferation and cell cycle progression. Cell cycle progression in human glioma U87MG and U251 cells was halted at S/G2/M phase via the Wnt/β-catenin signaling pathway and related genes such as PRR11, Cyclin A, p-CDK2, VEGFR-1/2, p-VEGFR-1/2 and EGFR. The results suggest that cZNF292 silencing plays an important role in the tube formation process and has potential for application as a therapeutic target and biomarker in glioma.

Chloroquine activates the p53 pathway and induces apoptosis in human glioma cells

PubMed Central

Kim, Ella L.; Wüstenberg, Robin; Rübsam, Anne; Schmitz-Salue, Christoph; Warnecke, Gabriele; Bücker, Eva-Maria; Pettkus, Nadine; Speidel, Daniel; Rohde, Veit; Schulz-Schaeffer, Walter; Deppert, Wolfgang; Giese, Alf

2010-01-01

Glioblastoma is the most common malignant brain tumor in adults. The currently available treatments offer only a palliative survival advantage and the need for effective treatments remains an urgent priority. Activation of the p53 growth suppression/apoptotic pathway is one of the promising strategies in targeting glioma cells. We show that the quinoline derivative chloroquine activates the p53 pathway and suppresses growth of glioma cells in vitro and in vivo in an orthotopic (U87MG) human glioblastoma mouse model. Induction of apoptosis is one of the mechanisms underlying the effects of chloroquine on suppressing glioma cell growth and viability. siRNA-mediated downregulation of p53 in wild-type but not mutant p53 glioblastoma cells substantially impaired chloroquine-induced apoptosis. In addition to its p53-activating effects, chloroquine may also inhibit glioma cell growth via p53-independent mechanisms. Our results clarify the mechanistic basis underlying the antineoplastic effect of chloroquine and reveal its therapeutic potential as an adjunct to glioma chemotherapy. PMID:20308316

Suppression of HIV-1 Infectivity by Human Glioma Cells

PubMed Central

Hoque, Sheikh Ariful; Tanaka, Atsushi; Islam, Salequl; Ahsan, Gias Uddin; Jinno-Oue, Atsushi

2016-01-01

Abstract HIV-1 infection to the central nervous system (CNS) is very common in AIDS patients. The predominant cell types infected in the brain are monocytes and macrophages, which are surrounded by several HIV-1–resistant cell types, such as astrocytes, oligodendrocytes, neurons, and microvascular cells. The effect of these HIV-1–resistant cells on HIV-1 infection is largely unknown. In this study, we examined the stability of HIV-1 cultured with several human glioblastoma cell lines, for example, NP-2, U87MG, T98G, and A172, to determine whether these HIV-1–resistant brain cells could enhance or suppress HIV-1 infection and thus modulate HIV-1 infection in the CNS. The HIV-1 titer was determined using the MAGIC-5A indicator cell line as well as naturally occurring CD4+ T cells. We found that the stability of HIV-1 incubated with NP-2 or U87MG cells at 37°C was significantly shorter (half-life, 2.5–4 h) compared to that of HIV-1 incubated with T98G or A172 cells or in culture medium without cells (half-life, 8–18 h). The spent culture media (SCM) of NP-2 and U87MG cells had the ability to suppress both R5- and X4-HIV-1 infection by inhibiting HIV-1 attachment to target cells. This inhibitory effect was eliminated by the treatment of the SCM with chondroitinase ABC but not heparinase, suggesting that the inhibitory factor(s) secreted by NP-2 and U87MG cells was chiefly mediated by chondroitin sulfate (CS) or CS-like moiety. Thus, this study reveals that some but not all glioma cells secrete inhibitory molecules to HIV-1 infection that may contribute in lowering HIV-1 infection in the CNS in vivo. PMID:26650729

Concomitant administration of radiation with eribulin improves the survival of mice harboring intracerebral glioblastoma.

PubMed

Miki, Shunichiro; Imamichi, Shoji; Fujimori, Hiroaki; Tomiyama, Arata; Fujimoto, Kenji; Satomi, Kaishi; Matsushita, Yuko; Matsuzaki, Sanae; Takahashi, Masamichi; Ishikawa, Eiichi; Yamamoto, Tetsuya; Matsumura, Akira; Mukasa, Akitake; Nishikawa, Ryo; Masutomi, Kenkichi; Narita, Yoshitaka; Masutani, Mitsuko; Ichimura, Koichi

2018-05-14

Glioblastoma is the most common and devastating type of malignant brain tumor. We recently found that eribulin suppresses glioma growth in vitro and in vivo and that eribulin is efficiently transferred into mouse brain tumors at a high concentration. Eribulin is a non-taxane microtubule inhibitor approved for breast cancer and liposarcoma. Cells arrested in M-phase by chemotherapeutic agents such as microtubule inhibitors are highly sensitive to radiation-induced DNA damage. Several recent case reports demonstrated the clinical benefits of eribulin combined with radiation therapy for metastatic brain tumors. In this study, we investigated the efficacy of a combined eribulin and radiation treatment on human glioblastoma cells. The glioblastoma cell lines U87MG, U251MG, U118MG, and SJ28 cells, a patient-derived sphere culture cell line, were used to determine the radiosensitizing effect of eribulin using western blotting, flow cytometry, and clonogenic assay. Subcutaneous and intracerebral glioma xenografts were generated in mice to assess the efficacy of the combined treatment. The combination of eribulin and radiation enhanced DNA damage in vitro. The clonogenic assay of U87MG demonstrated the radiosensitizing effect of eribulin. The concomitant eribulin and radiation treatment significantly prolonged the survival of mice harboring intracerebral glioma xenografts compared with eribulin or radiation alone (p<0.0001). In addition, maintenance administration of eribulin after the concomitant treatment further controlled brain tumor growth. Aberrant microvasculature was decreased in these tumors. Concomitant treatment with eribulin and radiation followed by maintenance administration of eribulin may serve as a novel therapeutic strategy for glioblastomas. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Osthole suppresses the proliferation and accelerates the apoptosis of human glioma cells via the upregulation of microRNA-16 and downregulation of MMP-9.

PubMed

Lin, Kai; Gao, Zhiyu; Shang, Bin; Sui, Shaohua; Fu, Qiang

2015-09-01

Osthole (7-methoxy-8-isoamyl alkenyl coumarin) has been reported to exhibit marked anticancer effects on several types of cancer. The expression levels of matrix metalloproteinase-9 (MMP-9) are closely associated with the pathogenesis of glioma. Furthermore, it is reported that the upregulation of microRNA‑16 (miR‑16) by the MMP‑9 signaling pathway can restrain the proliferation of cancer cells. To examine whether osthole increases the anticancer effect on human glioma cells in the present study, the common glioma cell line, U87, was treated with osthole at concentrations of 0, 50, 100 and 200 µΜ. The effects of osthole on cell viability were determined using a 3‑(4,5‑dimethylthiazol‑2‑thiazolyl)‑2,5‑diphenyl‑tetrazolium bromide assay. The rate of cellular apoptosis was analyzed by measuring the activity of caspase‑3 and using flow cytometry. The expression of MMP‑9 was determined using gelatin zymography assays and the expression of miR‑16 was determined using reverse transcription‑quantitative polymerase chain reaction. The results demonstrated that osthole significantly suppressed the proliferation and accelerated the apoptosis of the U87 cells. Furthermore, increased expression levels of miR‑16 and reduced protein expression levels of MMP‑9 were found in the U87 cells. In addition, miR‑16 was found to regulate the expression of MMP‑9 in the U87 cells through transfection of miR‑16 precursor and anti‑miR‑16 into the U87 cells. In conclusion, these observations indicated that osthole suppressed the proliferation and accelerated the apoptosis of human glioma cells through upregulation of the expression of miR‑16 and downregulation of the expression of MMP-9.

Application of Albumin-embedded Magnetic Nanoheaters for Release of Etoposide in Integrated Chemotherapy and Hyperthermia of U87-MG Glioma Cells.

PubMed

Babincová, Melánia; Vrbovská, Hana; Sourivong, Paul; Babinec, Peter; Durdík, Štefan

2018-05-01

Malignant gliomas remain refractory to several therapeutic approaches and the requirement for novel treatment modalities is critical to combat this disease. Etoposide is a topoisomerase-II inhibitor, which promotes DNA damage and apoptosis of cancer cells. In this study, we prepared albumin with embedded magnetic nanoparticles and etoposide for in vitro evaluation of combined hyperthermia and chemotherapy. Magnetic nanoparticles were prepared by a modified co-precipitation method in the presence of human serum albumin and etoposide. A cellular proliferation assay was used to determine the effects of these nanostructures on the viability of U87 glioma cells in an alternating magnetic field. The in vitro experiments showed that cell viability decreased to 59.4% after heat treatment alone and to 53.8% on that with free etoposide, while combined treatment resulted in 7.8% cell viability. Integrating hyperthermia and chemotherapy using albumin co-embedded magnetic nanoheaters and etoposide may represent a promising therapeutic option for glioblastoma. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Autophagy and Oxidative Stress in Gliomas with IDH1 Mutations

PubMed Central

Gilbert, Misty R.; Liu, Yinxing; Neltner, Janna; Pu, Hong; Morris, Andrew; Sunkara, Manjula; Pittman, Thomas; Kyprianou, Natasha; Horbinski, Craig

2013-01-01

IDH1 mutations in gliomas associate with longer survival. Prooxidant and antiproliferative effects of IDH1 mutations and its D-2-hydroxyglutarate (2-HG) product have been described in vitro, but inconsistently observed. It is also unclear whether overexpression of mutant IDH1 in wild-type cells accurately phenocopies the effects of endogenous IDH1-mutations on tumor apoptosis and autophagy. Herein we investigated the effects of 2-HG and mutant IDH1 overexpression on proliferation, apoptosis, oxidative stress, and autophagy in IDH1 wild-type glioma cells, and compared those results with patient-derived tumors. 2-HG reduced viability and proliferation of U87MG and LN18 cells, triggered apoptosis in LN18 cells, and autophagy in U87MG cells. In vitro studies and flank xenografts of U87MG cells overexpressing R132H IDH1 exhibited increased oxidative stress, including increases of both manganese superoxide dismutase (MnSOD) and p62. Patient-derived IDH1-mutant tumors showed no significant differences in apoptosis or autophagy, but showed p62 accumulation and actually trended toward reduced MnSOD expression. These data indicate that mutant IDH1 and 2-HG can induce oxidative stress, autophagy, and apoptosis, but these effects vary greatly according to cell type. PMID:24150401

A dual function fusion protein of Herpes simplex virus type 1 thymidine kinase and firefly luciferase for noninvasive in vivo imaging of gene therapy in malignant glioma.

PubMed

Söling, Ariane; Theiss, Christian; Jungmichel, Stephanie; Rainov, Nikolai G

2004-08-04

BACKGROUND: Suicide gene therapy employing the prodrug activating system Herpes simplex virus type 1 thymidine kinase (HSV-TK)/ ganciclovir (GCV) has proven to be effective in killing experimental brain tumors. In contrast, glioma patients treated with HSV-TK/ GCV did not show significant treatment benefit, most likely due to insufficient transgene delivery to tumor cells. Therefore, this study aimed at developing a strategy for real-time noninvasive in vivo monitoring of the activity of a therapeutic gene in brain tumor cells. METHODS: The HSV-TK gene was fused to the firefly luciferase (Luc) gene and the fusion construct HSV-TK-Luc was expressed in U87MG human malignant glioma cells. Nude mice with subcutaneous gliomas stably expressing HSV-TK-Luc were subjected to GCV treatment and tumor response to therapy was monitored in vivo by serial bioluminescence imaging. Bioluminescent signals over time were compared with tumor volumes determined by caliper. RESULTS: Transient and stable expression of the HSV-TK-Luc fusion protein in U87MG glioma cells demonstrated close correlation of both enzyme activities. Serial optical imaging of tumor bearing mice detected in all cases GCV induced death of tumor cells expressing the fusion protein and proved that bioluminescence can be reliably used for repetitive and noninvasive quantification of HSV-TK/ GCV mediated cell kill in vivo. CONCLUSION: This approach may represent a valuable tool for the in vivo evaluation of gene therapy strategies for treatment of malignant disease.

Expression of complement membrane regulators membrane cofactor protein (CD46), decay accelerating factor (CD55), and protectin (CD59) in human malignant gliomas.

PubMed Central

Mäenpää, A.; Junnikkala, S.; Hakulinen, J.; Timonen, T.; Meri, S.

1996-01-01

Gliomas are malignant brain tumors, which, despite recent progress in surgical and radiological treatment, still have a poor prognosis. Since gliomas apparently resist immunological clearance mechanisms, we became interested in examining bow gliomas resist killing by the human complement system. The resistance of human cells to complement-mediated damage is, in large part, mediated by specific inhibitors of complement:membrane cofactor protein (CD46), decay-accelerating factor (CD55), and protectin (CD59). In the present study we examined the expression of complement regulators in 14 human glioma tumors and in 7 glioma cell lines (U251, U87, HS683, U373, U138, U118, and H2). Protectin was found to be strongly expressed by all glioma tumors and cell lines. Northern blotting analysis demonstrated the typical pattern of four to five protectin mRNAs in the glioma cells. Except for blood vessels, the expression of decay-accelerating factor was weak or absent in the tumors in situ, whereas in the cell lines its expression varied, ranging from negative to intermediate. Membrane cofactor protein was moderately expressed by all the cell lines but only weakly in the tumors. Cell-killing experiments demonstrated that the glioma cell lines were exceptionally resistant to C-mediated lysis. Five of the seven cell lines (U373, HS683, U118, U138, and H2) resisted complement lysis under conditions where most other cell lines were sensitive to killing. Neutralization experiments using specific monoclonal antibodies indicated that protectin was functionally the most important complement regulator in the glioma cells. The killing of the U87 and U251 cells could be significantly increased by a blocking anti-protectin monoclonal antibody, whereas for the other cell lines only moderate or no response was observed. The H2 cell line resisted killing by all antibodies and by complement. These results show that protectin is the most important complement regulator on human glioma cells. The exceptional complement resistance of some glioma cell lines suggests that they may utilize other, hitherto less well characterized, mechanisms to resist complement killing. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 6 Figure 7 PMID:8644856

Inhibition of glioblastoma cell invasion by hsa-miR-145-5p and hsa-miR-31-5p co-overexpression in human mesenchymal stem cells.

PubMed

Kurogi, Ryota; Nakamizo, Akira; Suzuki, Satoshi O; Mizoguchi, Masahiro; Yoshimoto, Koji; Amano, Toshiyuki; Amemiya, Takeo; Takagishi, So; Iihara, Koji

2018-03-09

OBJECTIVE Human bone marrow-derived mesenchymal stem cells (hMSCs) show tropism for brain tumors and may be a useful vehicle for drug or gene delivery to malignant gliomas. Recently, some microRNAs (miRNAs) have been shown to suppress the invasiveness of malignant gliomas. METHODS To test their potential to become vehicles for the delivery of miRNA to malignant gliomas, hMSCs were engineered so that hMSC secretion of miRNAs that inhibit glioma cell invasion was enabled without altering the hMSC tropism for glioma cells. RESULTS In coculture, hMSCs cotransfected with hsa-miR-145-5p and -31-5p miRNAs showed markedly reduced invasion by U87 glioma cells in a contact-dependent manner both in vitro and ex vivo, with invasion of hMSCs cotransfected with these 2 miRNAs by the U87 cells reduced to 60.7% compared with control cells. According to a Matrigel invasion assay, the tropism of the hMSCs for U87 cells was not affected. In glioma cell lines U251 and LN229, hMSCs exhibited tropism in vivo, and invasion of hMSCs cotransfected with hsa-miR-145-5p and -31-5p was also significantly less than that of control cells. When U87 cells were coimplanted into the striatum of organotypic rat brain slices with hMSCs cotransfected with hsa-miR-145 and -31-5p, the relative invasive area decreased by 37.1%; interestingly, these U87 cells showed a change to a rounded morphology that was apparent at the invasion front. Whole-genome microarray analysis of the expression levels of 58,341 genes revealed that the co-overexpression of hsa-miR-145-5p and -31-5p downregulated FSCN1 expression in U87 cells. CONCLUSIONS This study demonstrates that miRNA overexpression in hMSCs can alter the function of glioma cells via contact-dependent transfer. Co-overexpression of multiple miRNAs may be a useful and novel therapeutic strategy. The study results suggest that hMSCs can be applied as a delivery vehicle for miRNAs.

Preclinical Pharmacological Evaluation of Letrozole as a Novel Treatment for Gliomas

PubMed Central

Dave, Nimita; Chow, Lionel M.L.; Gudelsky, Gary A.; LaSance, Kathleen; Qi, Xiaoyang; Desai, Pankaj B.

2015-01-01

We present data that letrozole, an extensively used aromatase inhibitor in the treatment of estrogen receptor-positive breast tumors in postmenopausal women, may be potentially used in the treatment of glioblastomas. First, we measured the in vitro cytotoxicity of letrozole and aromatase (CYP19A1) expression and activity in human LN229, T98G, U373MG, U251MG, and U87MG, and rat C6 glioma cell lines. Estrogen receptor (ER)positive MCF-7 and ER-negative MDA-MB-231 cells served as controls. Cytotoxicity was determined employing the MTT assay, and aromatase activity using an immunoassay that measures the conversion of testosterone to estrogen. Second, in vivo activity of letrozole was assessed in Sprague-Dawley rats orthotopically implanted with C6 gliomas. The changes in tumor volume with letrozole treatment (4 mg/kg/day) were assessed employing μPET/CT imaging, employing [18F]-fluorodeoxyglucose (F18-FDG) as the radiotracer. Brain tissues were collected for histologic evaluations. All glioma cell lines included here expressed CYP19A1 and letrozole exerted considerable cytotoxicity and decrease in aromatase activity against these cells (IC50, 0.1–3.5 μmol/L). Imaging analysis employing F18-FDG μPET/CT demonstrated a marked reduction of active tumor volume (>75%) after 8 days of letrozole treatment. Immunohistochemical analysis revealed marked reduction in aromatase expression in tumoral regions of the brain after letrozole treatment. Thus, employing multifaceted tools, we demonstrate that aromatase may be a novel target for the treatment of gliomas and that letrozole, an FDA-approved drug with an outstanding record of safety may be repurposed for the treatment of such primary brain tumors, which currently have few therapeutic options. PMID:25695958

Preclinical pharmacological evaluation of letrozole as a novel treatment for gliomas.

PubMed

Dave, Nimita; Chow, Lionel M L; Gudelsky, Gary A; LaSance, Kathleen; Qi, Xiaoyang; Desai, Pankaj B

2015-04-01

We present data that letrozole, an extensively used aromatase inhibitor in the treatment of estrogen receptor-positive breast tumors in postmenopausal women, may be potentially used in the treatment of glioblastomas. First, we measured the in vitro cytotoxicity of letrozole and aromatase (CYP19A1) expression and activity in human LN229, T98G, U373MG, U251MG, and U87MG, and rat C6 glioma cell lines. Estrogen receptor (ER)-positive MCF-7 and ER-negative MDA-MB-231 cells served as controls. Cytotoxicity was determined employing the MTT assay, and aromatase activity using an immunoassay that measures the conversion of testosterone to estrogen. Second, in vivo activity of letrozole was assessed in Sprague-Dawley rats orthotopically implanted with C6 gliomas. The changes in tumor volume with letrozole treatment (4 mg/kg/day) were assessed employing μPET/CT imaging, employing [(18)F]-fluorodeoxyglucose (F18-FDG) as the radiotracer. Brain tissues were collected for histologic evaluations. All glioma cell lines included here expressed CYP19A1 and letrozole exerted considerable cytotoxicity and decrease in aromatase activity against these cells (IC50, 0.1-3.5 μmol/L). Imaging analysis employing F18-FDG μPET/CT demonstrated a marked reduction of active tumor volume (>75%) after 8 days of letrozole treatment. Immunohistochemical analysis revealed marked reduction in aromatase expression in tumoral regions of the brain after letrozole treatment. Thus, employing multifaceted tools, we demonstrate that aromatase may be a novel target for the treatment of gliomas and that letrozole, an FDA-approved drug with an outstanding record of safety may be repurposed for the treatment of such primary brain tumors, which currently have few therapeutic options. ©2015 American Association for Cancer Research.

JS-K, a glutathione S-transferase-activated nitric oxide donor with antineoplastic activity in malignant gliomas

PubMed Central

Weyerbrock, Astrid; Osterberg, Nadja; Psarras, Nikolaos; Baumer, Brunhilde; Kogias, Evangelos; Werres, Anna; Bette, Stefanie; Saavedra, Joseph E.; Keefer, Larry K.; Papazoglou, Anna

2011-01-01

Background Glutathione S-transferases (GSTs) control multidrug-resistance and are upregulated in many cancers including malignant gliomas. The diazeniumdiolate JS-K generates nitric oxide (NO) on enzymatic activation by glutathione and GST, showing promising NO-based anticancer efficacy. Objective To evaluate the role of NO-based antitumor therapy with JS-K in U87 gliomas in vitro and in vivo. Methods U87 glioma cells and primary glioblastoma cell lines were exposed to JS-K and a variety of inhibitors to study cell death by necrosis, apoptosis and other mechanisms. GST-expression was evaluated by immunocytochemistry, PCR and Western blot and NO release from JS-K using a NO assay. The growth-inhibitory effect of JS-K was studied in a U87 xenograft model in vivo. Results Dose-dependent inhibition of cell proliferation was observed in human U87 glioma cells and primary glioblastoma cells in vitro. Cell death was partially induced by caspase-dependent apoptosis which could be blocked by Z-VAD-FMK and Q-VD-OPH. GST-inhibition by sulfasalazine, cGMP inhibition by ODQ and MEK 1/2 inhibition by UO126 attenuated the antiproliferative effect of JS-K, suggesting the involvement of various intracellular death signalling pathways. Response to JS-K correlated with mRNA and protein expression of GST and the amount of NO released by the glioma cells. Growth of U87 xenografts was significantly reduced, with immunohistochemical evidence for increased necrosis, apoptosis and reduced proliferation. Conclusion Our data for the first time show the potent antiproliferative effect of JS-K in gliomas in vitro and in vivo. These findings warrant further investigation of this novel NO-releasing prodrug in gliomas. PMID:21849924

JS-K, a glutathione S-transferase-activated nitric oxide donor with antineoplastic activity in malignant gliomas.

PubMed

Weyerbrock, Astrid; Osterberg, Nadja; Psarras, Nikolaos; Baumer, Brunhilde; Kogias, Evangelos; Werres, Anna; Bette, Stefanie; Saavedra, Joseph E; Keefer, Larry K; Papazoglou, Anna

2012-02-01

Glutathione S-transferases (GSTs) control multidrug resistance and are upregulated in many cancers, including malignant gliomas. The diazeniumdiolate JS-K generates nitric oxide (NO) on enzymatic activation by glutathione and GST, showing promising NO-based anticancer efficacy. To evaluate the role of NO-based antitumor therapy with JS-K in U87 gliomas in vitro and in vivo. U87 glioma cells and primary glioblastoma cell lines were exposed to JS-K and a variety of inhibitors to study cell death by necrosis, apoptosis, and other mechanisms. GST expression was evaluated by immunocytochemistry, polymerase chain reaction, and Western blot, and NO release from JS-K was studied with a NO assay. The growth-inhibitory effect of JS-K was studied in a U87 xenograft model in vivo. Dose-dependent inhibition of cell proliferation was observed in human U87 glioma cells and primary glioblastoma cells in vitro. Cell death was partially induced by caspase-dependent apoptosis, which could be blocked by Z-VAD-FMK and Q-VD-OPH. Inhibition of GST by sulfasalazine, cGMP inhibition by ODQ, and MEK1/2 inhibition by UO126 attenuated the antiproliferative effect of JS-K, suggesting the involvement of various intracellular death signaling pathways. Response to JS-K correlated with mRNA and protein expression of GST and the amount of NO released by the glioma cells. Growth of U87 xenografts was reduced significantly, with immunohistochemical evidence for increased necrosis and apoptosis and reduced proliferation. Our data show for the first time the potent antiproliferative effect of JS-K in gliomas in vitro and in vivo. These findings warrant further investigation of this novel NO-releasing prodrug in gliomas.

Antitumor effects of cannabidiol, a nonpsychoactive cannabinoid, on human glioma cell lines.

PubMed

Massi, Paola; Vaccani, Angelo; Ceruti, Stefania; Colombo, Arianna; Abbracchio, Maria P; Parolaro, Daniela

2004-03-01

Recently, cannabinoids (CBs) have been shown to possess antitumor properties. Because the psychoactivity of cannabinoid compounds limits their medicinal usage, we undertook the present study to evaluate the in vitro antiproliferative ability of cannabidiol (CBD), a nonpsychoactive cannabinoid compound, on U87 and U373 human glioma cell lines. The addition of CBD to the culture medium led to a dramatic drop of mitochondrial oxidative metabolism [3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H tetrazolium bromide test] and viability in glioma cells, in a concentration-dependent manner that was already evident 24 h after CBD exposure, with an apparent IC(50) of 25 microM. The antiproliferative effect of CBD was partially prevented by the CB2 receptor antagonist N-[(1S)-endo-1,3,3-trimethylbicyclo[2,2,1]heptan-2-yl]-5-(4-chloro-3-methylphenyl)-1-(4-methylbenzyl)-pyrazole-3-carboxamide (SR144528; SR2) and alpha-tocopherol. By contrast, the CB1 cannabinoid receptor antagonist N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboximide hydrochloride (SR141716; SR1), capsazepine (vanilloid receptor antagonist), the inhibitors of ceramide generation, or pertussis toxin did not counteract CBD effects. We also show, for the first time, that the antiproliferative effect of CBD was correlated to induction of apoptosis, as determined by cytofluorimetric analysis and single-strand DNA staining, which was not reverted by cannabinoid antagonists. Finally, CBD, administered s.c. to nude mice at the dose of 0.5 mg/mouse, significantly inhibited the growth of subcutaneously implanted U87 human glioma cells. In conclusion, the nonpsychoactive CBD was able to produce a significant antitumor activity both in vitro and in vivo, thus suggesting a possible application of CBD as an antineoplastic agent.

Effect of doublecortin on self-renewal and differentiation in brain tumor stem cells

PubMed Central

Santra, Manoranjan; Santra, Sutapa; Buller, Ben; Santra, Kastuv; Nallani, Ankita; Chopp, Michael

2011-01-01

Analysis of Affymetrix Probe data from glioma patient samples in conjuction with patient Kaplan-Meier Survival Plot indicate that expression of a glioma suppressor gene doublecortin (DCX) favors glioma patient survival. From neurosphere formation in culture, Time-Lapse Microscopy video recording and tumor xenograft, we show that DCX synthesis significantly reduces self-renewal of brain tumor stem cells (BTSCs) in human primary glioma (YU-PG, HF66) cells from surgically-removed human glioma specimens and U87 cells in vitro and in vivo. Time-Lapse Microscopic video recording revealed that double transfection of YU-PG, HF66 and U87 cells with DCX and neurabin II caused incomplete cell cycle with failure of cytokinesis, i.e. endomitosis by dividing into three daughter cells from one mother BTSC. Activation of c-jun NH2-terminal kinase 1 (JNK1) after simvastatin (10nM) treatment of DCX+neurabin II+ BTSCs from YU-PG, HF66 and U87 cells induced terminal differentiation into neuron-like cells. TUNEL staining data demonstrated that JNK1 activation also induced apoptosis only in double transfected BTSCs with DCX and neurabin II, but not in single transfected BTSCs from YU-PG, HF66 and U87 cells. Western blot analysis showed that procaspase-3 was induced after DCX transfection and activated after simvastatin treatment in YU-PG, HF66 and U87 BTSCs. Sequential immunoprecipitation and Western blot data revealed that DCX synthesis blocked protein phosphatase-1 (PP1)/caspase-3 protein-protein interaction and increased PP1-DCX interaction. These data demonstrate that DCX synthesis induces apoptosis in BTSCs via a novel JNK1/neurabin II/DCX/PP1/caspase-3 pathway. PMID:21477071

Effect of doublecortin on self-renewal and differentiation in brain tumor stem cells.

PubMed

Santra, Manoranjan; Santra, Sutapa; Buller, Ben; Santra, Kastuv; Nallani, Ankita; Chopp, Michael

2011-07-01

Analysis of microarray probe data from glioma patient samples, in conjunction with patient Kaplan-Meier survival plots, indicates that expression of a glioma suppressor gene doublecortin (DCX) favors glioma patient survival. From neurosphere formation in culture, time-lapse microscopic video recording, and tumor xenograft, we show that DCX synthesis significantly reduces self-renewal of brain tumor stem cells (BTSC) in human primary glioma (YU-PG, HF66) cells from surgically removed human glioma specimens and U87 cells in vitro and in vivo. Time-lapse microscopic video recording revealed that double transfection of YU-PG, HF66, and U87 cells with DCX and neurabin II caused incomplete cell cycle with failure of cytokinesis, that is, endomitosis by dividing into three daughter cells from one mother BTSC. Activation of c-jun NH2-terminal kinase 1 (JNK1) after simvastatin (10 nM) treatment of DCX(+) neurabin II(+) BTSC from YU-PG, HF66, and U87 cells induced terminal differentiation into neuron-like cells. dUTP nick end labeling data indicated that JNK1 activation also induced apoptosis only in double transfected BTSC with DCX and neurabin II, but not in single transfected BTSC from YU-PG, HF66, and U87 cells. Western blot analysis showed that procaspase-3 was induced after DCX transfection and activated after simvastatin treatment in YU-PG, HF66, and U87 BTSC. Sequential immunoprecipitation and Western blot data revealed that DCX synthesis blocked protein phosphatase-1 (PP1)/caspase-3 protein-protein interaction and increased PP1-DCX interaction. These data show that DCX synthesis induces apoptosis in BTSC through a novel JNK1/neurabin II/DCX/PP1/caspase-3 pathway. © 2011 Japanese Cancer Association.

Anti-proliferative and anti-migration effects of Polish propolis combined with Hypericum perforatum L. on glioblastoma multiforme cell line U87MG.

PubMed

Borawska, Maria H; Naliwajko, Sylwia K; Moskwa, Justyna; Markiewicz-Żukowska, Renata; Puścion-Jakubik, Anna; Soroczyńska, Jolanta

2016-09-20

Propolis and Hypericum perforatum L. are natural products which contain many active compounds and have numerous beneficial effects, including an antitumor effect. Gliobmastoma multiforme (GBM) is a common primary brain tumor with poor prognosis and limited treatment options. In this study, the effect of propolis (EEP) combined with H. perforatum L. (HPE) on glioblastoma cell line U87MG was investigated for the first time. Anti-proliferative activity of EEP, HPE and their combination (EEP + HPE) was determined by a cytotoxicity test, DNA binding by [(3)H]-thymidine incorporation and cell migration assay. Anti-metastatic properties in U87MG treated with EEP, HPE and EEP + HPE were estimated on cells migration test (scratch assay) and metalloproteinases (MMP2 and MMP9) secretion (gelatin zymography). Combination of HPE and EEP extracts was found to have a time- and dose-dependent inhibitory effect on the viability of U87MG cells. This effect was significantly higher (p < 0.05) when compared to these two extracts applied separately, which was confirmed by the significant reduction of DNA synthesis and significantly higher mitochondrial membrane permeabilization. A significant decreasing in migration cells and in pro-MMP9 and pro-MMP2 secretion in U87MG cells were demonstrated after exposure to combination of EEP (30 μg/ml) with HPE (6.25 μg/ml). In this study, the combination of ethanolic extract from propolis and ethanolic extract of fresh-cut H. perforatum L. was proved the ability to reduce invasiveness of glioma cells through the inhibition of MMP2 and MMP9 secretion and suppression of cell migration. It has a more potent anti-proliferative effect on U87MG glioma cell line compared to using propolis and H. perforatum L. separately. Further studies are required to verify whether the examined extracts can activate apoptotic pathways.

Physiological oxygen concentration alters glioma cell malignancy and responsiveness to photodynamic therapy in vitro.

PubMed

Albert, Ina; Hefti, Martin; Luginbuehl, Vera

2014-11-01

The partial pressure of oxygen (pO2) in brain tumors ranges from 5 to 15%. Nevertheless, the majority of in vitro experiments with glioblastoma multiforme (GBM) cell lines are carried out under an atmospheric pO2 of 19 to 21%. Recently, 5-aminolevulinic acid (5-ALA), a precursor of protoporphyrin IX (PpIX), has been introduced to neurosurgery to allow for photodynamic diagnosis and photodynamic therapy (PDT) in high-grade gliomas. Here, we investigate whether low pO2 affects GBM cell physiology, PpIX accumulation, or PDT efficacy. GBM cell lines (U-87 MG and U-251 MG) were cultured under atmospheric (pO2  =  19%) and physiological (pO2  =  9%) oxygen concentrations. PpIX accumulation and localization were investigated, and cell survival and cell death were observed following in vitro PDT. A physiological pO2 of 9% stimulated GBM cell migration, increased hypoxia-inducible factor (HIF)-1 alpha levels, and elevated resistance to camptothecin in U-87 MG cells compared to cultivation at a pO2 of 19%. This oxygen reduction did not alter 5-ALA-induced intracellular PpIX accumulation. However, physiological pO2 changed the responsiveness of U-87 MG but not of U-251 MG cells to in vitro PDT. Around 20% more irradiation light was required to kill U-87 MG cells at physiological pO2, resulting in reduced lactate dehydrogenase (LDH) release (one- to two-fold) and inhibition of caspase 3 activation. Reduction of oxygen concentration from atmospheric to a more physiological level can influence the malignant behavior and survival of GBM cell lines after in vitro PDT. Therefore, precise oxygen concentration control should be considered when designing and performing experiments with GBM cells.

Protein alterations associated with temozolomide resistance in subclones of human glioblastoma cell lines.

PubMed

Sun, Stella; Wong, T S; Zhang, X Q; Pu, Jenny K S; Lee, Nikki P; Day, Philip J R; Ng, Gloria K B; Lui, W M; Leung, Gilberto K K

2012-03-01

Temozolomide (TMZ) is the standard chemotherapeutic agent for human malignant glioma, but intrinsic or acquired chemoresistance represents a major obstacle to successful treatment of this highly lethal group of tumours. Obtaining better understanding of the molecular mechanisms underlying TMZ resistance in malignant glioma is important for the development of better treatment strategies. We have successfully established a passage control line (D54-C10) and resistant variants (D54-P5 and D54-P10) from the parental TMZ-sensitive malignant glioma cell line D54-C0. The resistant sub-cell lines showed alterations in cell morphology, enhanced cell adhesion, increased migration capacities, and cell cycle arrests. Proteomic analysis identified a set of proteins that showed gradual changes in expression according to their 50% inhibitory concentration (IC(50)). Successful validation was provided by transcript profiling in another malignant glioma cell line U87-MG and its resistant counterparts. Moreover, three of the identified proteins (vimentin, cathepsin D and prolyl 4-hydroxylase, beta polypeptide) were confirmed to be upregulated in high-grade glioma. Our data suggest that acquired TMZ resistance in human malignant glioma is associated with promotion of malignant phenotypes, and our reported molecular candidates may serve not only as markers of chemoresistance but also as potential therapeutic targets in the treatment of TMZ-resistant human malignant glioma, providing a platform for future investigations.

Expression of R132H mutational IDH1 in human U87 glioblastoma cells affects the SREBP1a pathway and induces cellular proliferation.

PubMed

Zhu, Jian; Cui, Gang; Chen, Ming; Xu, Qinian; Wang, Xiuyun; Zhou, Dai; Lv, Shengxiang; Fu, Linshan; Wang, Zhong; Zuo, Jianling

2013-05-01

Sterol regulatory element-binding protein-1a (SREBP1a) is a member of the SREBP family of transcription factors, which mainly controls homeostasis of lipids. SREBP1a can also activate the transcription of isocitrate dehydrogenase 1 (IDH1) by binding to its promoter region. IDH1 mutations, especially R132H mutation of IDH1, are a common feature of a major subset of human gliomas. There are few data available on the relationship between mutational IDH1 expression and SREBP1a pathway. In this study, we investigated cellular effects and SREBP1a pathway alterations caused by R132H mutational IDH1 expression in U87 cells. Two glioma cell lines, stably expressing mutational (U87/R132H) or wild type (U87/wt) IDH1, were established. A cell line, stably transfected with pcDNA3.1(+) (U87/vector), was generated as a control. Click-iT EdU assay, sulforhodamine B assay, and wound healing assay respectively showed that the expression of R132H induced cellular proliferation, cell growth, and cell migration. Western blot revealed that SREBP1 was increased in U87/R132H compared with that in U87/wt. Elevated SREBP1a and several its target genes, but not SREBP1c, were detected by real-time polymerase chain reaction in U87/R132H. All these findings indicated that R132H mutational IDH1 is involved in the regulation of proliferation, growth, and migration of glioma cells. These effects may partially be mediated by SREBP1a pathway.

Antitumoral Cascade-Targeting Ligand for IL-6 Receptor-Mediated Gene Delivery to Glioma.

PubMed

Wang, Shanshan; Reinhard, Sören; Li, Chengyi; Qian, Min; Jiang, Huiling; Du, Yilin; Lächelt, Ulrich; Lu, Weiyue; Wagner, Ernst; Huang, Rongqin

2017-07-05

The effective treatment of glioma is largely hindered by the poor transfer of drug delivery systems across the blood-brain barrier (BBB) and the difficulty in distinguishing healthy and tumorous cells. In this work, for the first time, an interleukin-6 receptor binding I 6 P 7 peptide was exploited as a cascade-targeting ligand in combination with a succinoyl tetraethylene pentamine (Stp)-histidine oligomer-based nonviral gene delivery system (I 6 P 7 -Stp-His/DNA). The I 6 P 7 peptide provides multiple functions, including the cascade-targeting potential represented by a combined BBB-crossing and subsequent glioma-targeting ability, as well as a direct tumor-inhibiting effect. I 6 P 7 -Stp-His/DNA nanoparticles (NPs) mediated higher gene expression in human glioma U87 cells than in healthy human astrocytes and a deeper penetration into glioma spheroids than scrambled peptide-modified NPs. Transport of I 6 P 7 -modified, but not the control, NPs across the BBB was demonstrated in vitro in a transwell bEnd.3 cell model resulting in transfection of underlying U87 cells and also in vivo in glioma-bearing mice. Intravenous administration of I 6 P 7 -Stp-His/plasmid DNA (pDNA)-encoding inhibitor of growth 4 (pING4) significantly prolonged the survival time of orthotopic U87 glioma-bearing mice. The results denote that I 6 P 7 peptide is a roborant cascade-targeting ligand, and I 6 P 7 -modified NPs might be exploited for efficient glioma therapy. Copyright © 2017. Published by Elsevier Inc.

Magnolol Inhibits Human Glioblastoma Cell Migration by Regulating N-Cadherin.

PubMed

Cheng, Yu-Chen; Tsao, Min-Jen; Chiu, Chen-Yang; Kan, Po-Chieh; Chen, Ying

2018-06-01

Glioblastoma is a primary malignant brain tumor with a poor prognosis. An effective treatment for glioblastoma is needed. Magnolol is a natural compound from Magnolia officinalis suggested to have antiproliferative activity. The aim of this research was to investigate the anticancer effects of magnolol in glioma, with an emphasis on migration and the underlying mechanism. Magnolol decreased the expression of focal adhesion-related proteins and inhibited LN229 and U87MG glioma cell migration. The levels of phosphorylated myosin light chain (p-MLC), phosphorylated myosin light chain kinase and myosin phosphatase target subunit 1 were reduced in response to magnolol treatment. In addition, immunostaining and membrane fractionation showed that the distribution of N-cadherin at the glioma cell membrane was decreased by magnolol. In an orthotropic xenograft animal model, magnolol treatment not only inhibited tumor progression but also reduced p-MLC and N-cadherin protein expression. In conclusion, magnolol reduces cell migration, potentially through regulating focal adhesions and N-cadherin in glioma cells. Magnolol is a potential candidate for glioma treatment.

Single-wall carbon nanohorns (SWNHs) inhibited proliferation of human glioma cells and promoted its apoptosis

NASA Astrophysics Data System (ADS)

Li, Yunjun; Zhang, Jinqian; Zhao, Ming; Shi, Zujin; Chen, Xin; He, Xihui; Han, Nanyin; Xu, Ruxiang

2013-08-01

Although single-wall carbon nanohorns (SWNHs) have been demonstrated to accumulate to cytotoxic levels within organs of various animal models and cell types, they have been exploited for cancer therapies. The role of SWNHs in human glioma cell lines was unclear. To address this question, the research about direct role of SWNHs on the growth, proliferation, and apoptosis of human glioma cell lines (U87, U251, and U373) had been performed. Our results indicate that particle size of SWNHs in water is between 342 and 712 nm, the films of SEM show that SWNHs on PS surface are individual particles. SWNHs significantly delayed mitotic entry of human glioma cell lines cells, and inhibited its proliferation in a time- and dose-dependent manner. SWNHs induced a significant increase in G1 phase and inhibition of S phase followed the gradually increasing concentrations. SWNHs in human glioma cell lines cells significantly induced apoptosis followed by their gradually increasing concentrations. The TEM images showed that individual spherical SWNHs particles smaller than 100 nm in diameters were localized inside lysosomes of human glioma cell lines. SWNHs inhibited mitotic entry, growth, and proliferation of human glioma cell lines, and promoted its apoptosis. SWNHs may be a novel opportunity or method for the research on treatment of human glioma.

Convection-enhanced delivery of a synthetic retinoid Am80, loaded into polymeric micelles, prolongs the survival of rats bearing intracranial glioblastoma xenografts.

PubMed

Yokosawa, Michiko; Sonoda, Yukihiko; Sugiyama, Shin-ichiro; Saito, Ryuta; Yamashita, Yoji; Nishihara, Masamichi; Satoh, Taku; Kumabe, Toshihiro; Yokoyama, Masayuki; Tominaga, Teiji

2010-08-01

Prognosis for the patients with glioblastoma, the most common malignant brain tumor, remains dismal. A major barrier to progress in treatment of glioblastoma is the relative inaccessibility of tumors to chemotherapeutic agents. Convection-enhanced delivery (CED) is a direct intracranial drug infusion technique to deliver chemotherapeutic agents to the central nervous system, circumventing the blood-brain barrier and reducing systemic side effects. CED can provide wider distribution of infused agents compared to simple diffusion. We have reported that CED of a polymeric micelle carrier system could yield a clinically relevant distribution of encapsulated agents in the rat brain. Our aim was to evaluate the efficacy of CED of polymeric micellar Am80, a synthetic agonist with high affinity to nuclear retinoic acid receptor, in a rat model of glioblastoma xenografts. We also used systemic administration of temozolomide, a DNA-alkylating agent, which has been established as the standard of care for newly diagnosed malignant glioma. U87MG human glioma cells were injected into the cerebral hemisphere of nude rats. Rats bearing U87MG xenografts were treated with CED of micellar Am80 (2.4 mg/m(2)) on day 7 after tumor implantation. Temozolomide (200 mg/m(2)/day) was intraperitoneally administered daily for 5 days, starting on day 7 after tumor implantation. CED of micellar Am80 provided significantly longer survival than the control. The combination of CED of micellar Am80 and systemic administration of temozolomide provided significantly longer survival than single treatment. In conclusion, temozolomide combined with CED of micellar Am80 may be a promising method for the treatment of malignant gliomas.

Apigenin inhibits glioma cell growth through promoting microRNA-16 and suppression of BCL-2 and nuclear factor-κB/MMP‑9.

PubMed

Chen, Xin-Jun; Wu, Mian-Yun; Li, Deng-Hui; You, Jin

2016-09-01

The present study aimed to investigate the effect of apigenin on glioma cells and to explore its potential mechanism. U87 human glioma cells treated with apigenin were used in the current study. Cell Counting Kit‑8 solution and Annexin V-fluorescein isothiocyanate/propidium iodide Apoptosis Detection kit were used to analyze the effect of apigenin on U87 cell viability and apoptotic cell death. Reverse transcription‑quantitative polymerase chain reaction analysis was also used to determine microRNA‑16 (miR‑16) and MMP‑9 gene expression levels. Nuclear factor‑κB (NF‑κB) and B‑cell CLL/lymphoma 2 (BCL2) protein expression levels were determined using western blot analysis. An anti‑miR‑16 plasmid was constructed and transfected into U87 cells. The current study demonstrated that apigenin significantly decreased cell viability and induced apoptotic cell death of U87 cells in a dose‑dependent manner. Additionally, it was demonstrated that apigenin significantly increased miR‑16 levels, suppressed BCL2 protein expression and suppressed the NF‑κB/MMP9 signaling pathway in U87 cells. Furthermore, downregulation of miR‑16 using the anti‑miR‑16 plasmid reversed the effect of apigenin on cell viability, BCL2 protein expression and the NF‑κB/MMP‑9 pathway in U87 cells. The results of the present study suggested that apigenin inhibits glioma cell growth through promoting miR‑16 and suppression of BCL2 and NF-κB/MMP-9. In conclusion, the present study demonstrated the potential anticancer effects of apigenin on glioma cells.

Involvement of estrogen receptor β5 in the progression of glioma.

PubMed

Li, Wenjun; Winters, Ali; Poteet, Ethan; Ryou, Myoung-Gwi; Lin, Song; Hao, Shuyu; Wu, Zhen; Yuan, Fang; Hatanpaa, Kimmo J; Simpkins, James W; Yang, Shao-Hua

2013-03-29

Emerging evidence suggests a decline of ERβ expression in various peripheral cancers. ERβ has been proposed as a cancer brake that inhibits tumor proliferation. In the current study, we have identified ERβ5 as the predominant isoform of ERβ in human glioma and its expression was significantly increased in human glioma as compared with non-neoplastic brain tissue. Hypoxia and activation of hypoxia inducible factor (HIF) increased ERβ transcription in U87 cells, suggesting elevated ERβ expression in glioma might be induced by the hypoxic stress in the tumor. Over-expression of either ERβ1 or ERβ5 increased PTEN expression and inhibited activation of the PI3K/AKT/mTOR pathway. In addition, ERβ5 inhibited the MAPK/ERK pathway. In U87 cells, ERβ1 and ERβ5 inhibit cell proliferation and reduced cells in the S+G2/M phase. Our findings suggest hypoxia induced ERβ5 expression in glioma as a self-protective mechanism against tumor proliferation and that ERβ5 might serve as a therapeutic target for the treatment of glioma. Copyright © 2013 Elsevier B.V. All rights reserved.

PEGylated Polyamidoamine dendrimer conjugated with tumor homing peptide as a potential targeted delivery system for glioma.

PubMed

Jiang, Yan; Lv, Lingyan; Shi, Huihui; Hua, Yabing; Lv, Wei; Wang, Xiuzhen; Xin, Hongliang; Xu, Qunwei

2016-11-01

Glioblastoma multiforme (GBM) is the most common and aggressive primary central nervous system (CNS) tumor with a short survival time. The failure of chemotherapy is ascribed to the low transport of chemotherapeutics across the Blood Brain Tumor Barrier (BBTB) and poor penetration into tumor tissue. In order to overcome the two barriers, small nanoparticles with active targeted capability are urgently needed for GBM drug delivery. In this study, we proposed PEGylated Polyamidoamine (PAMAM) dendrimer nanoparticles conjugated with glioma homing peptides (Pep-1) as potential glioma targeting delivery system (Pep-PEG-PAMAM), where PEGylated PAMAM dendrimer nanoparticle was utilized as carrier due to its small size and perfect penetration into tumor and Pep-1 was used to overcome BBTB via interleukin 13 receptor α2 (IL-13Rα2) mediated endocytosis. The preliminary availability and safety of Pep-PEG-PAMAM as a nanocarrier for glioma was evaluated. In vitro results indicated that a significantly higher amount of Pep-PEG-PAMAM was endocytosed by U87 MG cells. In vivo fluorescence imaging of U87MG tumor-bearing mice confirmed that the fluorescence intensity at glioma site of targeted group was 2.02 folds higher than that of untargeted group (**p<0.01), and glioma distribution experiment further revealed that Pep-PEG-PAMAM exhibited a significantly enhanced accumulation and improved penetration at tumor site. In conclusion, Pep-1 modified PAMAM was a promising nanocarrier for targeted delivery of brain glioma. Copyright © 2016 Elsevier B.V. All rights reserved.

Incubation and application of transgenic green fluorescent nude mice in visualization studies on glioma tissue remodeling.

PubMed

Dong, Jun; Dai, Xing-liang; Lu, Zhao-hui; Fei, Xi-feng; Chen, Hua; Zhang, Quan-bin; Zhao, Yao-dong; Wang, Zhi-min; Wang, Ai-dong; Lan, Qing; Huang, Qiang

2012-12-01

The primary reasons for local recurrence and therapeutic failure in the treatment of malignant gliomas are the invasion and interactions of tumor cells with surrounding normal brain cells. However, these tumor cells are hard to be visualized directly in histopathological preparations, or in experimental glioma models. Therefore, we developed an experimental human dual-color in vivo glioma model, which made tracking solitary invasive glioma cells possible, for the purpose of visualizing the interactions between red fluorescence labeled human glioma cells and host brain cells. This may offer references for further studying the roles of tumor microenvironment during glioma tissue remodeling. Transgenic female C57BL/6 mice expressing enhanced green fluorescent protein (EGFP) were crossed with male Balb/c nude mice. Then sib mating was allowed to occur continuously in order to establish an inbred nude mice strain with 50% of their offspring that are EGFP positive. Human glioma cell lines U87-MG and SU3 were transfected with red fluorescent protein (RFP) gene, and a rat C6 glioma cell line was stained directly with CM-DiI, to establish three glioma cell lines emitting red fluorescence (SU3-RFP, U87-RFP, and C6-CM-DiI). Red fluorescence tumor cells were inoculated via intra-cerebral injection into caudate nucleus of the EGFP nude mice. Tumor-bearing mice were sacrificed when their clinical symptoms appeared, and the whole brain was harvested and snap frozen for further analysis. Confocal laser scanning microscopy was performed to monitor the mutual interactions between tumor cells and host brain cells. Almost all the essential tissues of the established EGFP athymic Balb/c nude mice, except hair and erythrocytes, fluoresced green under excitation using a blue light-emitting flashlight with a central peak of 470 nm, approximately 50% of the offsprings were nu/nu EGFP+. SU3-RFP, U87-RFP, and C6-CM-DiI almost 100% expressed red fluorescence under the fluorescence microscope. Under fluorescence microscopic view, RFP+ cells were observed growing wherever they arrived at, locating in the brain parenchyma, ventricles, and para-vascular region. The interactions between the transplanted tumor cells and host adjacent cells could be classified into three types: (1) interweaving; (2) mergence; and (3) fusion. Interweaving was observed in the early stage of tumor remodeling, in which both transplantable tumor cells and host cells were observed scattered in the tumor invading and spreading area without organic connections. Mergence was defined as mutual interactions between tumor cells and host stroma during tumorigenesis. Direct cell fusion between transplantable tumor cells and host cells could be observed occasionally. This study showed that self-established EGFP athymic nude mice offered the possibility of visualizing tumorigenesis of human xenograft tumor, and the dual-color xenograft glioma model was of considerable utility in studying the process of tumor remodeling. Based on this platform, mutual interactions between glioma cells and host tissues could be observed directly to further elucidate the development of tumor microenvironment.

Autophagy protein p62/SQSTM1 is involved in HAMLET-induced cell death by modulating apotosis in U87MG cells

PubMed Central

Zhang, Y-B; Gong, J-L; Xing, T-Y; Zheng, S-P; Ding, W

2013-01-01

HAMLET is a complex of oleic acids and decalcified α-lactalbumin that was discovered to selectively kill tumor cells both in vitro and in vivo. Autophagy is an important cellular process involved in drug-induced cell death of glioma cells. We treated U87MG human glioma cells with HAMLET and found that the cell viability was significantly decreased and accompanied with the activation of autophagy. Interestingly, we observed an increase in p62/SQSTM1, an important substrate of autophagosome enzymes, at the protein level upon HAMLET treatment for short periods. To better understand the functionality of autophagy and p62/SQSTM1 in HAMLET-induced cell death, we modulated the level of autophagy or p62/SQSTM1 with biochemical or genetic methods. The results showed that inhibition of autophagy aggravated HAMLET-induced cell death, whereas activation of authophagy attenuated this process. Meanwhile, we found that overexpression of wild-type p62/SQSTM1 was able to activate caspase-8, and then promote HAMLET-induced apoptosis, whereas knockdown of p62/SQSTM1 manifested the opposite effect. We further demonstrated that the function of p62/SQSTM1 following HAMLET treatment required its C-terminus UBA domain. Our results indicated that in addition to being a marker of autophagy activation in HAMLET-treated glioma cells, p62/SQSTM1 could also function as an important mediator for the activation of caspase-8-dependent cell death. PMID:23519119

Autophagy protein p62/SQSTM1 is involved in HAMLET-induced cell death by modulating apotosis in U87MG cells.

PubMed

Zhang, Y-B; Gong, J-L; Xing, T-Y; Zheng, S-P; Ding, W

2013-03-21

HAMLET is a complex of oleic acids and decalcified α-lactalbumin that was discovered to selectively kill tumor cells both in vitro and in vivo. Autophagy is an important cellular process involved in drug-induced cell death of glioma cells. We treated U87MG human glioma cells with HAMLET and found that the cell viability was significantly decreased and accompanied with the activation of autophagy. Interestingly, we observed an increase in p62/SQSTM1, an important substrate of autophagosome enzymes, at the protein level upon HAMLET treatment for short periods. To better understand the functionality of autophagy and p62/SQSTM1 in HAMLET-induced cell death, we modulated the level of autophagy or p62/SQSTM1 with biochemical or genetic methods. The results showed that inhibition of autophagy aggravated HAMLET-induced cell death, whereas activation of authophagy attenuated this process. Meanwhile, we found that overexpression of wild-type p62/SQSTM1 was able to activate caspase-8, and then promote HAMLET-induced apoptosis, whereas knockdown of p62/SQSTM1 manifested the opposite effect. We further demonstrated that the function of p62/SQSTM1 following HAMLET treatment required its C-terminus UBA domain. Our results indicated that in addition to being a marker of autophagy activation in HAMLET-treated glioma cells, p62/SQSTM1 could also function as an important mediator for the activation of caspase-8-dependent cell death.

IDH1(R132H) mutation causes a less aggressive phenotype and radiosensitizes human malignant glioma cells independent of the oxygenation status.

PubMed

Kessler, Jacqueline; Güttler, Antje; Wichmann, Henri; Rot, Swetlana; Kappler, Matthias; Bache, Matthias; Vordermark, Dirk

2015-09-01

In malignant glioma the presence of the IDH1 mutation (IDH1(R132H)) is associated with better clinical outcome. However, it is unclear whether IDH1 mutation is associated with a less aggressive phenotype or directly linked to increased sensitivity to radiotherapy. We determined the influence of IDH1(R132H) mutant protein on proliferation and growth in 3D culture, migration, cell survival and radiosensitivity in vitro under normoxia (21% O2) and hypoxia (<1% O2) in a panel of human malignant glioma cell lines (U-251MG, U-343MG, LN-229) with stable overexpression of wild-type (IDH1(wt)) and mutated IDH1 (IDH1(R132H)). Overexpression of IDH1(R132H) in glioma cells resulted in slightly decreased cell proliferation, considerably reduced cell migration and caused differences in growth properties in 3D spheroid cultures. Furthermore, IDH1(R132H)-positive cells consistently demonstrated an increased radiosensitivity in human malignant glioma cells U-251MG (DMF10: 1.52, p<0.01 and 1.42, p<0.01), U-343MG (DMF10: 1.78, p<0.01 and 1.75, p<0.01) and LN-229 (DMF10: 1.41, p<0.05 and 1.68, p<0.01) under normoxia and hypoxia, respectively. Our data indicate that IDH1(R132H) mutation causes both a less aggressive biological behavior and direct radiosensitization of human malignant glioma cells. Targeting IDH1 appears to be an attractive approach in combination with radiotherapy. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Silibinin-induced glioma cell apoptosis by PI3K-mediated but Akt-independent downregulation of FoxM1 expression.

PubMed

Zhang, Mingjie; Liu, Yunhui; Gao, Yun; Li, Shaoyi

2015-10-15

The oncogenic transcription factor Forkhead box M1 (FoxM1) is overexpressed in many human tumors, including glioma. As a critical regulator of the cell cycle and apoptosis-related genes, FoxM1 is a potential therapeutic target against human malignant glioma. Silibinin, a flavonoid isolated from Silybum marianum, dose-dependently reduced glioma cell proliferation, promoted apoptosis, and downregulated FoxM1 expression. Knockdown of FoxM1 by small hairpin RNA (shRNA) transfection also promoted glioma cell apoptosis and augmented the antiproliferative and pro-apoptotic properties of silibinin. Moreover, silibinin increased caspase-3 activation, upregulated pro-apoptotic Bax, and suppressed anti-apoptotic Bcl-2 expression, effects enhanced by FoxM1 knockdown. Silibinin treatment suppressed U87 cell PI3K phospho-activation, and simultaneous silibinin exposure, FoxM1 knockdown, and PI3K inhibition additively increased U87 cell apoptosis. Furthermore, PI3K inhibition reduced FoxM1 expression. Akt activity was also suppressed by FoxM1 downregulation but Akt inhibition did not alter FoxM1 expression. Thus, silibinin likely inhibited glioma cell proliferation and induced apoptosis through inactivation of PI3K and FoxM1, leading to activation of the mitochondrial apoptotic pathway. FoxM1 may be a novel target for chemotherapy against human glioma. Copyright © 2015 Elsevier B.V. All rights reserved.

Mangiferin regulates proliferation and apoptosis in glioma cells by induction of microRNA-15b and inhibition of MMP-9 expression.

PubMed

Xiao, Jinsong; Liu, Li; Zhong, Zian; Xiao, Cheng; Zhang, Junjian

2015-06-01

Mangiferin, a flavonoid extracted from the leaves of the Anacardiaceae plant, the mango tree, has physiological activity and pharmacological effects in many aspects. The present study aimed to clarify the effect of mangiferin on proliferation and apoptosis of glioma cells and the mechanism of these curative effects of mangiferin. In this experiment, we detected the proliferation using 3-(4,5-dimethylthylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) assay. Then, cell apoptosis of U87 glioma cells was measured with the Annexin V-FITC/propidium iodide (PI) apoptosis detection kit, DAPI staining assay and the caspase-3 and caspase-9 activity assay kit. Next, quantitative real-time PCR and gelatin zymography were used to analyze the expression of microRNA-15b (miR-15b) and matrix metalloproteinase-9 (MMP-9), respectively. MMP-9 agonist, miR-15b mimics and anti-miR-15b mimics were added to the U87 glioma cells for elucidating the mechanisms involved in the curative effects of mangiferin. In the present study, mangiferin notably restrained the proliferation and increased the apoptosis of the U87 glioma cells. Meanwhile, mangiferin specifically promoted the expression of miR-15b and suppressed the level of MMP-9 in the U87 glioma cells. miR-15b regulated the expression of MMP-9 in the U87 glioma cells. MMP-9 agonist and anti-miR‑15b reduced the curative effects of mangiferin in the U87 glioma cells. In summary, mangiferin regulates proliferation and apoptosis in glioma cells by induction of miR-15b and inhibition of MMP-9 expression.

Sensitivity of human glioma U-373MG cells to radiation and the protein kinase C inhibitor, calphostin C.

PubMed

Acevedo-Duncan, M; Pearlman, J; Zachariah, B

2001-02-01

We assessed the radiosensitivity of the grade III human glioma cell line U-373MG by investigating the effects of radiation and the specific protein kinase C inhibitor, calphostin C on the cell cycle and cell proliferation. Irradiated glioma U-373MG cells progressed through G1-S and underwent an arrest in G2-M phase. The radiosensitivity of U-373MG cells to graded doses of either photons or electrons was determine by microculture tetrazolium assay. The data was fitted to the linear-quadratic model. The proliferation curves demonstrated that U-373MG cells appear to be highly radiation resistant since 8 Gy was required to achieve 50% cell mortality. Compared to radiation alone, exposure to calphostin C (250 nM) 1 h prior to radiation decreased the proliferation of U-373MG by 76% and calphostin C provoked a weakly synergistic effect in concert with radiation. Depending on the time of application following radiation, calphostin C produced an additive or less than additive effect on cell proliferation. We postulate that the enhanced radiosensitivity observed when cells are exposed to calphostin C prior to radiation may be due to direct or indirect inhibition of protein kinase C isozymes required for cell cycle progression.

Metabolic remodeling of malignant gliomas for enhanced sensitization during radiotherapy: an in vitro study.

PubMed

Colen, Chaim B; Seraji-Bozorgzad, Navid; Marples, Brian; Galloway, Matthew P; Sloan, Andrew E; Mathupala, Saroj P

2006-12-01

To investigate a novel method to enhance radiosensitivity of gliomas via modification of metabolite flux immediately before radiotherapy. Malignant gliomas are highly glycolytic and produce copious amounts of lactic acid, which is effluxed to the tumor microenvironment via lactate transporters. We hypothesized that inhibition of lactic acid efflux would alter glioma metabolite profiles, including those that are radioprotective. H magnetic resonance spectroscopy (MRS) was used to quantify key metabolites, including those most effective for induction of low-dose radiation-induced cell death. We inhibited lactate transport in U87-MG gliomas with alpha-cyano-4-hydroxycinnamic acid (ACCA). Flow cytometry was used to assess induction of cell death in treated cells. Cells were analyzed by MRS after ACCA treatment. Control and treated cells were subjected to low-dose irradiation, and the surviving fractions of cells were determined by clonogenic assays. MRS revealed changes to intracellular lactate on treatment with ACCA. Significant decreases in the metabolites taurine, glutamate, glutathione, alanine, and glycine were observed, along with inversion of the choline/phosphocholine profile. On exposure to low-dose radiation, ACCA-pretreated U-87MG cells underwent rapid morphological changes, which were followed by apoptotic cell death. Inhibition of lactate efflux in malignant gliomas results in alterations of glycolytic metabolism, including decreased levels of the antioxidants taurine and glutathione and enhanced radiosensitivity of ACCA-treated cells. Thus, in situ application of lactate transport inhibitors such as ACCA as a novel adjunctive therapeutic strategy against glial tumors may greatly enhance the level of radiation-induced cell killing during a combined radio- and chemotherapeutic regimen.

Effect of saw palmetto extract on PI3K cell signaling transduction in human glioma.

PubMed

Yang, Yang; Hui, Lv; Yuqin, Che; Jie, Li; Shuai, Hou; Tiezhu, Zhou; Wei, Wang

2014-08-01

Saw palmetto extract can induce the apoptosis of prostate cancer cells. The aim of the present study was to investigate the effect of saw palmetto extract on the phosphatidylinositol 3-kinase (PI3K)/Akt signaling transduction pathway in human glioma U87 and U251 cell lines. Suspensions of U87 and U251 cells in a logarithmic growth phase were seeded into six-well plates at a density of 10 4 cells/well. In the experimental group, 1 μl/ml saw palmetto extract was added, while the control group was cultured without a drug for 24 h. The expression levels of PI3K, B-cell lymphoma-extra large (Bcl-xL) and p53 were evaluated through western blot analysis. In the experimental group, the U87 and U251 cells exhibited a lower expression level of PI3K protein as compared with the control group (t=6.849; P<0.001). In addition, the two cell lines had a higher expression level of p53 protein in the experimental group as compared with the control group (t=40.810; P<0.001). Protein expression levels of Bcl-xL decreased significantly in the experimental group as compared with the control group (t=19.640; P=0.000). Therefore, saw palmetto extract induces glioma cell growth arrest and apoptosis via decreasing PI3K/Akt signal transduction.

Targeted Imaging and Chemo-Phototherapy of Brain Cancer by a Multifunctional Drug Delivery System.

PubMed

Hao, Yongwei; Wang, Lei; Zhao, Yalin; Meng, Dehui; Li, Dong; Li, Haixia; Zhang, Bingxiang; Shi, Jinjin; Zhang, Hongling; Zhang, Zhenzhong; Zhang, Yun

2015-11-01

The aim of this study was to develop multifunctional poly lactide-co-glycolide (PLGA) nanoparticles with the ability to simultaneously deliver indocyanine green (ICG) and docetaxel (DTX) to the brain by surface decoration with the brain-targeting peptide angiopep-2 to achieve combined chemo-phototherapy for glioma under near-infrared (NIR) imaging. ICG was selected as a near-infrared imaging and phototherapy agent and DTX was employed as a chemotherapeutic agent. ICG and DTX were simultaneously incorporated into PLGA nanoparticles with higher stability. These nanoparticles were further decorated with angiopep-2 via the outer maleimide group of 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethyleneglycol)-2000]-maleinimide incorporated in the nanoparticles. The NIR image-guided chemo-phototherapy of the angiopep-2 modified PLGA/DTX/ICG nanoparticles (ANG/PLGA/DTX/ICG NPs) not only highly induced U87MG cell death in vitro, but also efficiently prolonged the life span of the brain orthotopic U87MG glioma xenograft-bearing mice in vivo. Thus, this study suggests that ANG/PLGA/DTX/ICG NPs have the potential for combinatorial chemotherapy and phototherapy for glioma. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Hydrogen Peroxide-Induced Secreted Frizzled-Related Protein 1 Gene Demethylation Contributes to Hydrogen Peroxide-Induced Apoptosis in Human U251 Glioma Cells.

PubMed

Xing, Zhiguo; Ni, Yaping; Zhao, Junjie; Ma, Xudong

2017-05-01

Glioblastoma multiforme is a type of central nervous system tumor with extremely poor prognosis. Previously, hydrogen peroxide (H 2 O 2 ), which promotes the oxidative stress response, has been reported to induce the apoptosis of glioma cells. Recently, secreted frizzled-related protein 1 (SFRP1) has been shown to be associated with various types of malignant tumors and with H 2 O 2 -induced oxidative stress in cardiomyocytes by negatively regulating the Wnt signaling pathway. This study aimed to explore SFRP1 expression and its roles in H 2 O 2 -induced apoptosis in human glioma cells. We found that the SFRP1 level was decreased in several human glioma cell lines, including U87, U251, and SW1783 cells. In U251 cells, SFRP1 could function as a cancer suppressor gene, and the growth of U251 cells could be inhibited not only by H 2 O 2 but also by the overexpression of SFRP1. Furthermore, we demonstrated that H 2 O 2 -induced SFRP1 gene demethylation partially contributed to H 2 O 2 -induced U251 cell apoptosis, which was verified by studies using an SFRP inhibitor (WAY-316606). Our research identified that H 2 O 2 -induced SFRP1 gene demethylation contributes to H 2 O 2 -induced apoptosis in human U251 glioma cells.

In vitro and in vivo anti-glioma activity of a chalcone-quinoxaline hybrid.

PubMed

Loch-Neckel, Gecioni; Bicca, Maíra Assunção; Leal, Paulo César; Mascarello, Alessandra; Siqueira, Jarbas Mota; Calixto, João B

2015-01-27

Chalcones are important compounds that exhibit multiple biological activities, including anti-inflammatory, antimitotic and antibacterial properties. In the present study, we have analyzed the potential anti-cancer activity of a chalcone named N9 (a hybrid chalcone-quinoxaline compound) using in vitro and in vivo experimental glioma models. Here, we report N9-induced inhibition of cell proliferation and also N9-induced cell death in a concentration-dependent manner in U87-MG glioma cells. These effects of N9 appear to be associated with its ability to inhibit the expression of cell cycle-associated proteins, and also the augmentation in the expression of the p21 (p21/Cip1) protein, a cyclin-dependent kinase inhibitor. Additionally, N9 also potentiates the production of the pro-apoptotic markers Bax and p53 via inhibition of MDM2. Moreover, our results show that N9 also significantly enhanced apoptosis of U87-MG cells with disruption of mitochondrial membrane potential, generation of ROS and caspase-9 activation. In vivo experiments carried out in a murine xenograft model of U87-MG revealed that N9 produced a significant reduction of tumors volume when compared to vehicle treated mice. Collectively, data demonstrate that N9 possess in vitro and in vivo anti-cancer activity, an effect that seems to involve the induction of p53 and p21 proteins, as well as, the activation of mitochondrial apoptosis pathway associated with the inhibition of protein MDM2. Overall, this study suggests N9 is affecting a variety of intracellular pathways related to tumor apoptosis. Perhaps N9 or derivate molecules could represent new potential drugs for cancer therapeutics. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

R132H mutation in IDH1 gene reduces proliferation, cell survival and invasion of human glioma by downregulating Wnt/β-catenin signaling.

PubMed

Cui, Daming; Ren, Jie; Shi, Jinlong; Feng, Lijing; Wang, Ke; Zeng, Tao; Jin, Yi; Gao, Liang

2016-04-01

Mutations in the isocitrate dehydrogenase 1 (IDH1) gene commonly occur in gliomas. Remarkably, the R132H mutation in IDH1 (IDH1-R132H) is associated with better prognosis and increased survival than patients lacking this mutation. The molecular mechanism underlying this phenomenon is largely unknown. In this study, we investigated potential cross-talk between IDH1-R132H and Wnt/β-catenin signaling in regulating the cellular properties of human glioma. Although aberrant nuclear accumulation of β-catenin is linked to the malignant progression of gliomas, its association with IDH1 remains unknown. We identified an inverse correlation between IDH1-R132H and the expression and activity of β-catenin in human gliomas. In addition, overexpression of IDH1-R132H in glioblastoma cell lines U87 and U251 led to reduced cell proliferation, migration and invasion, accompanied by increased apoptosis. At the molecular level, we detected a significant reduction in the expression, nuclear accumulation and activity of β-catenin following overexpression of IDH1-R132H. A microarray-based comparison of gene expression indicated that several mediators, effectors and targets of Wnt/β-catenin signaling are downregulated, while negative regulators are upregulated in IDH1-R132H gliomas. Further, overexpression of β-catenin in IDH1-R132H glioma cells restored the cellular phenotype induced by this mutation. Specifically, β-catenin abrogated the decrease in proliferation, invasion and migration, and the increase in apoptosis, triggered by overexpression of IDH1-R132H. Finally, we demonstrate that xenografts of IDH1-R132H overexpressing U87 cells can significantly decrease the growth of tumors in vivo. Altogether, our results strongly suggest that the R132H mutation in IDH1 serves a tumor suppressor function in human glioma by negatively regulating Wnt/β-catenin signaling. Copyright © 2016 Elsevier Ltd. All rights reserved.

Repression of Septin9 and Septin2 suppresses tumor growth of human glioblastoma cells.

PubMed

Xu, Dongchao; Liu, Ajuan; Wang, Xuan; Chen, Yidan; Shen, Yunyun; Tan, Zhou; Qiu, Mengsheng

2018-05-01

Glioblastoma (GBM) is the most common primary malignancy of the central nervous system (CNS) with <10% 5-year survival rate. The growth and invasion of GBM cells into normal brain make the resection and treatment difficult. A better understanding of the biology of GBM cells is crucial to the targeted therapies for the disease. In this study, we identified Septin9 (SEPT9) and Septin2 (SEPT2) as GBM-related genes through integrated multi-omics analysis across independent transcriptomic and proteomic studies. Further studies revealed that expression of SEPT9 and SEPT2 was elevated in glioma tissues and cell lines (A172, U87-MG). Knockdown of SEPT9 and SEPT2 in A172/U87-MG was able to inhibit GBM cell proliferation and arrest cell cycle progression in the S phase in a synergistic mechanism. Moreover, suppression of SEPT9 and SEPT2 decreased the GBM cell invasive capability and significantly impaired the growth of glioma xenografts in nude mice. Furthermore, the decrease in GBM cell growth caused by SEPT9 and SEPT2 RNAi appears to involve two parallel signaling pathway including the p53/p21 axis and MEK/ERK activation. Together, our integration of multi-omics analysis has revealed previously unrecognized synergistic role of SEPT9 and SEPT2 in GBM, and provided novel insights into the targeted therapy of GBM.

14-3-3β exerts glioma-promoting effects and is associated with malignant progression and poor prognosis in patients with glioma.

PubMed

Liu, Liang; Liu, Zhixiong; Wang, Hao; Chen, Long; Ruan, Fuqiang; Zhang, Jihui; Hu, Yi; Luo, Hengshan; Wen, Shuai

2018-03-01

Glioma is a type of tumor that affects the central nervous system. It has been demonstrated that 14-3-3β, a protein that is mainly concentrated in the brain, serves an important role in tumor regulation. However, the mechanism of action of 14-3-3β that underlies the pathogenesis of glioma remains to be elucidated. In the present study, 14-3-3β was silenced by RNA interference in the human glioma cell line U373-MG. Following knockdown of 14-3-3β, the proliferation, colony formation, cell cycle progression, migration and invasion of U373-MG cells were significantly decreased (P<0.01), whereas cell apoptosis was increased (P<0.01). Furthermore, in a tumor xenograft experiment, silencing 14-3-3β significantly inhibited the in vivo tumor growth of U373-MG cells (P<0.01). The results demonstrated that 14-3-3β levels were significantly higher in human glioma tissues compared with normal brain tissues (P<0.01) and high 14-3-3β expression was significantly associated with advanced pathological grade (P<0.03) and low Karnofsky performance scale (P<0.003). Patients with glioma who had high 14-3-3β levels had a significantly shorter survival time compared with those with low expression of 14-3-3β (P=0.031), suggesting that 14-3-3β may be an effective predictor of the prognosis of patients with glioma. The results of the present study indicate that 14-3-3β serves an oncogenic role in glioma, suggesting that 14-3-3β may have potential as a promising therapeutic target for glioma.

Epigenetic regulation of NOTCH1 and NOTCH3 by KMT2A inhibits glioma proliferation.

PubMed

Huang, Yin-Cheng; Lin, Sheng-Jia; Shih, Hung-Yu; Chou, Chung-Han; Chu, Hsiao-Han; Chiu, Ching-Chi; Yuh, Chiou-Hwa; Yeh, Tu-Hsueh; Cheng, Yi-Chuan

2017-09-08

Glioblastomas are among the most fatal brain tumors; however, the molecular determinants of their tumorigenic behavior are not adequately defined. In this study, we analyzed the role of KMT2A in the glioblastoma cell line U-87 MG. KMT2A knockdown promoted cell proliferation. Moreover, it increased the DNA methylation of NOTCH1 and NOTCH3 and reduced the expression of NOTCH1 and NOTCH3 . NOTCH1 or NOTCH3 activation inhibited U-87 MG cell proliferation, whereas NOTCH1 and NOTCH3 inhibition by shRNAs induced cell proliferation, thus demonstrating the tumor-suppressive ability of NOTCH1 and NOTCH3 in U-87 MG cells. The induced cell proliferation caused by KMT2A knockdown could be nullified by using either constitutively active NOTCH1 or constitutively active NOTCH3. This result demonstrates that KMT2A positively regulates NOTCH1 and NOTCH3 and that this mechanism is essential for inhibiting the U-87 MG cell proliferation. The role of KMT2A knockdown in promoting tumor growth was further confirmed in vivo by transplanting U-87 MG cells into the brains of zebrafish larvae. In conclusion, we identified KMT2A-NOTCH as a negative regulatory cascade for glioblastoma cell proliferation, and this result provides important information for KMT2A- or NOTCH-targeted therapeutic strategies for brain tumors.

Honokiol induces autophagic cell death in malignant glioma through reactive oxygen species-mediated regulation of the p53/PI3K/Akt/mTOR signaling pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lin, Chien-Ju

Honokiol, an active constituent extracted from the bark of Magnolia officinalis, possesses anticancer effects. Apoptosis is classified as type I programmed cell death, while autophagy is type II programmed cell death. We previously proved that honokiol induces cell cycle arrest and apoptosis of U87 MG glioma cells. Subsequently in this study, we evaluated the effect of honokiol on autophagy of glioma cells and examined the molecular mechanisms. Administration of honokiol to mice with an intracranial glioma increased expressions of cleaved caspase 3 and light chain 3 (LC3)-II. Exposure of U87 MG cells to honokiol also induced autophagy in concentration- andmore » time-dependent manners. Results from the addition of 3-methyladenine, an autophagy inhibitor, and rapamycin, an autophagy inducer confirmed that honokiol-induced autophagy contributed to cell death. Honokiol decreased protein levels of PI3K, phosphorylated (p)-Akt, and p-mammalian target of rapamycin (mTOR) in vitro and in vivo. Pretreatment with a p53 inhibitor or transfection with p53 small interfering (si)RNA suppressed honokiol-induced autophagy by reversing downregulation of p-Akt and p-mTOR expressions. In addition, honokiol caused generation of reactive oxygen species (ROS), which was suppressed by the antioxidant, vitamin C. Vitamin C also inhibited honokiol-induced autophagic and apoptotic cell death. Concurrently, honokiol-induced alterations in levels of p-p53, p53, p-Akt, and p-mTOR were attenuated following vitamin C administration. Taken together, our data indicated that honokiol induced ROS-mediated autophagic cell death through regulating the p53/PI3K/Akt/mTOR signaling pathway. - Highlights: • Exposure of mice with intracranial gliomas to honokiol induces cell apoptosis and autophagy. • Honokiol triggers autophagy of human glioma cells via the PISK/AKT/mTOR signaling pathway. • P53 induces autophagy via regulating the AKT/mTOR pathway in honokiol-treated glioma cells. • ROS participates in honokiol-induced cell death through the p53-mediated signaling pathway. • Honokiol induces ROS-mediated autophagic cell death via the p53/PI3K/Akt/mTOR mechanism.« less

Destruction of vasculogenic mimicry channels by targeting epirubicin plus celecoxib liposomes in treatment of brain glioma

PubMed Central

Ju, Rui-Jun; Zeng, Fan; Liu, Lei; Mu, Li-Min; Xie, Hong-Jun; Zhao, Yao; Yan, Yan; Wu, Jia-Shuan; Hu, Ying-Jie; Lu, Wan-Liang

2016-01-01

The efficacy of chemotherapy for brain glioma is restricted by the blood–brain barrier (BBB), and surgery or radiotherapy cannot eliminate the glioma cells because of their unique location. Residual brain glioma cells can form vasculogenic mimicry (VM) channels that can cause a recurrence of brain glioma. In the present study, targeting liposomes incorporating epirubicin and celecoxib were prepared and used for the treatment of brain glioma, along with the destruction of their VM channels. Evaluations were performed on the human brain glioma U87MG cells in vitro and on intracranial brain glioma-bearing nude mice. Targeting epirubicin plus celecoxib liposomes in the circulatory blood system were able to be transported across the BBB, and accumulated in the brain glioma region. Then, the liposomes were internalized by brain glioma cells and killed glioma cells by direct cytotoxic injury and the induction of apoptosis. The induction of apoptosis was related to the activation of caspase-8- and -3-signaling pathways, the activation of the proapoptotic protein Bax, and the suppression of the antiapoptotic protein Mcl-1. The destruction of brain glioma VM channels was related to the downregulation of VM channel-forming indictors, which consisted of MMP-2, MMP-9, FAK, VE-Cad, and VEGF. The results demonstrated that the targeting epirubicin plus celecoxib liposomes were able to effectively destroy the glioma VM channels and exhibited significant efficacy in the treatment of intracranial glioma-bearing nude mice. Therefore, targeting epirubicin plus celecoxib liposomes could be a potential nanostructured formulation to treat gliomas and destroy their VM channels. PMID:27042063

Anti-glioma activity and the mechanism of cellular uptake of asiatic acid-loaded solid lipid nanoparticles.

PubMed

Garanti, Tanem; Stasik, Aneta; Burrow, Andrea Julie; Alhnan, Mohamed A; Wan, Ka-Wai

2016-03-16

Asiatic acid (AA), a pentacyclic triterpene found in Centella Asiatica, has shown neuroprotective and anti-cancer activity against glioma. However, owing to its poor aqueous solubility, effective delivery and absorption across biological barriers, in particular the blood brain barrier (BBB), are challenging. Solid lipid nanoparticles (SLNs) have shown a promising potential as a drug delivery system to carry lipophilic drugs across the BBB, a major obstacle in brain cancer therapy. Nevertheless, limited information is available about the cytotoxic mechanisms of nano-lipidic carriers with AA on normal and glioma cells. This study assessed the anti-cancer efficacy of AA-loaded SLNs against glioblastoma and their cellular uptake mechanism in comparison with SVG P12 (human foetal glial) cells. SLNs were systematically investigated for three different solid lipids; glyceryl monostearate (MS), glyceryl distearate (DS) and glyceryl tristearate (TS). The non-drug containing MS-SLNs (E-MS-SLNs) did not show any apparent toxicity towards normal SVG P12 cells, whilst the AA-loaded MS-SLNs (AA-MS-SLNs) displayed a more favourable drug release profile and higher cytotoxicity towards U87 MG cells. Therefore, MS-SLNs were chosen for further in vitro studies. Cytotoxicity studies of SLNs (± AA) were performed using MTT assay where AA-SLNs showed significantly higher cytotoxicity towards U87 MG cells than SVG P12 normal cells, as confirmed by flow cell cytometry. Cellular uptake of SLNs also appeared to be preferentially facilitated by energy-dependent endocytosis as evidenced by fluorescence imaging and flow cell cytometry. Using the Annexin V-PI double staining technique, it was found that these AA-MS-SLNs displayed concentration-dependent apoptotic activity on glioma cells, which further confirms the potential of exploiting these AA-loaded MS-SLNs for brain cancer therapy. Copyright © 2016 Elsevier B.V. All rights reserved.

Effect of saw palmetto extract on PI3K cell signaling transduction in human glioma

PubMed Central

YANG, YANG; HUI, LV; YUQIN, CHE; JIE, LI; SHUAI, HOU; TIEZHU, ZHOU; WEI, WANG

2014-01-01

Saw palmetto extract can induce the apoptosis of prostate cancer cells. The aim of the present study was to investigate the effect of saw palmetto extract on the phosphatidylinositol 3-kinase (PI3K)/Akt signaling transduction pathway in human glioma U87 and U251 cell lines. Suspensions of U87 and U251 cells in a logarithmic growth phase were seeded into six-well plates at a density of 104 cells/well. In the experimental group, 1 μl/ml saw palmetto extract was added, while the control group was cultured without a drug for 24 h. The expression levels of PI3K, B-cell lymphoma-extra large (Bcl-xL) and p53 were evaluated through western blot analysis. In the experimental group, the U87 and U251 cells exhibited a lower expression level of PI3K protein as compared with the control group (t=6.849; P<0.001). In addition, the two cell lines had a higher expression level of p53 protein in the experimental group as compared with the control group (t=40.810; P<0.001). Protein expression levels of Bcl-xL decreased significantly in the experimental group as compared with the control group (t=19.640; P=0.000). Therefore, saw palmetto extract induces glioma cell growth arrest and apoptosis via decreasing PI3K/Akt signal transduction. PMID:25009620

Aptamer-conjugated PEGylated quantum dots targeting epidermal growth factor receptor variant III for fluorescence imaging of glioma.

PubMed

Tang, Jiaze; Huang, Ning; Zhang, Xiang; Zhou, Tao; Tan, Ying; Pi, Jiangli; Pi, Li; Cheng, Si; Zheng, Huzhi; Cheng, Yuan

2017-01-01

The extent of resection is a significant prognostic factor in glioma patients. However, the maximum safe resection level is difficult to determine due to the inherent infiltrative character of tumors. Recently, fluorescence-guided surgery has emerged as a new technique that allows safe resection of glioma. In this study, we constructed a new kind of quantum dot (QD)-labeled aptamer (QD-Apt) nanoprobe by conjugating aptamer 32 (A32) to the QDs surface, which can specially bind to the tumors. A32 is a single-stranded DNA capable of binding to the epidermal growth factor receptor variant III (EGFRvIII) specially distributed on the surface of glioma cells. To detect the expression of EGFRvIII in human brain tissues, 120 specimens, including 110 glioma tissues and 10 normal brain tissues, were examined by immunohistochemistry, and the results showed that the rate of positive expression of EGFRvIII in the glioma tissues was 41.82%, and 0.00% in normal brain tissues. Besides, the physiochemical properties of QD-Apt nanoparticles (NPs) were thoroughly characterized. Biocompatibility of the NPs was evaluated, and the results suggested that the QD-Apt was nontoxic in vivo and vitro. Furthermore, the use of the QD-Apt in labeling glioma cell lines and human brain glioma tissues, and target gliomas in situ was also investigated. We found that not only could QD-Apt specially bind to the U87-EGFRvIII glioma cells but also bind to human glioma tissues in vitro. Fluorescence imaging in vivo with orthotopic glioma model mice bearing U87-EGFRvIII showed that QD-Apt could penetrate the blood-brain barrier and then selectively accumulate in the tumors through binding to EGFRvIII, and consequently, generate a strong fluorescence, which contributed to the margins of gliomas that were visualized clearly, and thus, help the surgeons realize the maximum safe resection of glioma. In addition, QD-Apt can also be applied in preoperative diagnosis and postoperative examination of glioma. Therefore, these achievements facilitate the use of tumor-targeted fluorescence imaging in the diagnosis, surgical resection, and postoperative examination of glioma.

Aptamer-conjugated PEGylated quantum dots targeting epidermal growth factor receptor variant III for fluorescence imaging of glioma

PubMed Central

Tang, Jiaze; Huang, Ning; Zhang, Xiang; Zhou, Tao; Tan, Ying; Pi, Jiangli; Pi, Li; Cheng, Si; Zheng, Huzhi; Cheng, Yuan

2017-01-01

The extent of resection is a significant prognostic factor in glioma patients. However, the maximum safe resection level is difficult to determine due to the inherent infiltrative character of tumors. Recently, fluorescence-guided surgery has emerged as a new technique that allows safe resection of glioma. In this study, we constructed a new kind of quantum dot (QD)-labeled aptamer (QD-Apt) nanoprobe by conjugating aptamer 32 (A32) to the QDs surface, which can specially bind to the tumors. A32 is a single-stranded DNA capable of binding to the epidermal growth factor receptor variant III (EGFRvIII) specially distributed on the surface of glioma cells. To detect the expression of EGFRvIII in human brain tissues, 120 specimens, including 110 glioma tissues and 10 normal brain tissues, were examined by immunohistochemistry, and the results showed that the rate of positive expression of EGFRvIII in the glioma tissues was 41.82%, and 0.00% in normal brain tissues. Besides, the physiochemical properties of QD-Apt nanoparticles (NPs) were thoroughly characterized. Biocompatibility of the NPs was evaluated, and the results suggested that the QD-Apt was nontoxic in vivo and vitro. Furthermore, the use of the QD-Apt in labeling glioma cell lines and human brain glioma tissues, and target gliomas in situ was also investigated. We found that not only could QD-Apt specially bind to the U87-EGFRvIII glioma cells but also bind to human glioma tissues in vitro. Fluorescence imaging in vivo with orthotopic glioma model mice bearing U87-EGFRvIII showed that QD-Apt could penetrate the blood–brain barrier and then selectively accumulate in the tumors through binding to EGFRvIII, and consequently, generate a strong fluorescence, which contributed to the margins of gliomas that were visualized clearly, and thus, help the surgeons realize the maximum safe resection of glioma. In addition, QD-Apt can also be applied in preoperative diagnosis and postoperative examination of glioma. Therefore, these achievements facilitate the use of tumor-targeted fluorescence imaging in the diagnosis, surgical resection, and postoperative examination of glioma. PMID:28579776

Fluorescence of Pc 4 in U87 cells following photodynamic therapy

NASA Astrophysics Data System (ADS)

Varghai, Davood; Azizuddin, Kashif; Ahmad, Yusra; Oleinick, Nancy L.; Dean, David

2007-02-01

Introduction: Given the length of procedures and the brightness of operating room lights, there is concern that photosensitizers used to locate brain tumors and treat them with photodynamic therapy (PDT) may photobleach before they can be fully utilized. The phthalocyanine photosensitizer Pc 4 is resistant to photobleaching. In this study, we tested the hypothesis that exposure of Pc 4-loaded glioma cells to photoactivating light will result in continuing fluorescence of Pc 4. Methods: U87 human glioma cells were cultured in MEM with 5% penicillin/streptomycin, 5% sodium pyruvate, 10% fetal bovine serum, and 25 mM HEPES. These cultures were given 0 or 125 nM Pc 4, followed 2 hours later by three separate exposures of 200 J/cm2 of red light (λ max = 675 nm). Confocal fluorescence images were collected before and after each exposure. Results: Pc 4 fluorescence was localized to cytoplasmic membranes of the U87 glioma cells, as previously seen in other types of cells. After exposure to PDT, Pc 4 fluorescence was not reduced and even increased. Discussion: Pc 4 may be useful for the intra-operative detection of glioma by fluorescence and for PDT, since neither Pc 4 level nor its fluorescence is likely to decrease during exposure to operating room lights.

Ferrociphenol lipid nanocapsule delivery by mesenchymal stromal cells in brain tumor therapy.

PubMed

Roger, Mathilde; Clavreul, Anne; Huynh, Ngoc Trinh; Passirani, Catherine; Schiller, Paul; Vessières, Anne; Montero-Menei, Claudia; Menei, Philippe

2012-02-14

The prognosis of patients with malignant glioma remains extremely poor despite surgery and improvements in radio- and chemo-therapies. Thus, treatment strategies that specifically target these tumors have the potential to greatly improve therapeutic outcomes. "Marrow-isolated adult multilineage inducible" cells (MIAMI cells) are a subpopulation of mesenchymal stromal cells (MSCs) which possess the ability to migrate to brain tumors. We have previously shown that MIAMI cells were able to efficiently incorporate lipid nanocapsules (LNCs) without altering either their stem cell properties or their migration capacity. In this study, we assessed whether the cytotoxic effects of MIAMI cells loaded with LNCs containing an organometallic complex (ferrociphenol or Fc-diOH) could be used to treat brain tumors. The results showed that MIAMI cells internalized Fc-diOH-LNCs and that this internalization did not induce MIAMI cell death. Furthermore, Fc-diOH-LNC-loaded MIAMI cells produced a cytotoxic effect on U87MG glioma cells in vitro. This cytotoxic effect was validated in vivo after intratumoral injection of Fc-diOH-LNC-loaded MIAMI cells in a heterotopic U87MG glioma model in nude mice. These promising results open up a new field of treatment in which cellular vehicles and nanoparticles can be combined to treat brain tumors. Copyright © 2011 Elsevier B.V. All rights reserved.

miR-221/222 overexpession in human glioblastoma increases invasiveness by targeting the protein phosphate PTPμ

PubMed Central

Quintavalle, C; Garofalo, M; Zanca, C; Romano, G; Iaboni, M; De Caro, M del Basso; Martinez-Montero, JC; Incoronato, M; Nuovo, G; Croce, CM; Condorelli, G

2015-01-01

Glioblastoma is the most frequent brain tumor in adults and is the most lethal form of human cancer. Despite the improvements in treatments, survival of patients remains poor. In order to identify microRNAs (miRs) involved in glioma tumorigenesis, we evaluated, by a miRarray, differential expression of miRs in the tumorigenic glioma LN-18, LN-229 and U87MG cells compared with the non-tumorigenic T98G cells. Among different miRs we focused our attention on miR-221 and -222. We demonstrated the presence of a binding site for these two miRs in the 3′ untranslated region of the protein tyrosine phosphatase μ (PTPμ). Previous studies indicated that PTPμ suppresses cell migration and is downregulated in glioblastoma. Significantly, we found that miR-221 and -222 over-expression induced a downregulation of PTPμ as analyzed by both western blot and real-time PCR. Furthermore, miR-222 and -221 induced an increase in cell migration and growth in soft agar in glioma cells. Interestingly, the re-expression of PTPμ gene was able to revert the miR-222 and -221 effects on cell migration. Furthermore, we found an inverse correlation between miR-221 and -222 and PTPμ in human glioma cancer samples. In conclusion, our results suggest that miR-221 and -222 regulate glioma tumorigenesis at least in part through the control of PTPμ protein expression. PMID:21743492

Down-Regulation of AQP4 Expression via p38 MAPK Signaling in Temozolomide-Induced Glioma Cells Growth Inhibition and Invasion Impairment.

PubMed

Chen, Yuqin; Gao, Fei; Jiang, Rong; Liu, Hui; Hou, Jiaojiao; Yi, Yaoxing; Kang, Lili; Liu, Xueyuan; Li, Yuan; Yang, Mei

2017-12-01

Glioma is the most common and lethal central nervous system tumors. Temozolomide (TMZ) is an effective drug for malignant glioma, however, the intracellular and molecular mechanisms behind this anti-cancer effect have yet to be fully understood. The aim of the present study was to determine whether TMZ inhibits proliferation, invasion of glioma cells in vitro and whether these effects can be mediated through modulation of aquaporin 4 (AQP4) and phosphorylation of the MAPK pathway. The viability of U87 and U251 human glioma cells was evaluated using MTT assay. The cell cycle distribution was detected with flow cytometry. Migration ability and invasion ability were tested by scratch assays and transwell assays, respectively. The levels of AQP4 and MAPK were measured using immunoblot analyses. Our results showed that TMZ inhibited proliferation, migration and invasion, and induced G2/M arrest in U87 and U251 glioma cell lines. These changes were associated with a decrease in the levels of AQP4 expression as well as activation phosphorylated level of p38. Treatment with a p38 chemical activator (anisomycin) resulted in similar effects as TMZ treatment on glioma cells. And p38 chemical inhibitor (SB203580) could block these effects in glioma treated with TMZ, suggesting a direct up-regulation of the p38 signaling pathway. Therefore, we identified that TMZ might have therapeutic potential for controlling proliferation, invasion of malignant glioma by inhibiting AQP4 expression through activation of p38 signal transduction pathway. J. Cell. Biochem. 118: 4905-4913, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Replication-Dependent Radiosensitization of Human Glioma Cells by Inhibition of Poly(ADP-Ribose) Polymerase: Mechanisms and Therapeutic Potential

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dungey, Fiona A.; Loeser, Dana A.; Chalmers, Anthony J.

2008-11-15

Purpose: Current treatments for glioblastoma multiforme are inadequate and limited by the radiation sensitivity of normal brain. Because glioblastoma multiforme are rapidly proliferating tumors within nondividing normal tissue, the therapeutic ratio might be enhanced by combining radiotherapy with a replication-specific radiosensitizer. KU-0059436 (AZD2281) is a potent and nontoxic inhibitor of poly(ADP-ribose) polymerase-1 (PARP-1) undergoing a Phase II clinical trial as a single agent. Methods and Materials: Based on previous observations that the radiosensitizing effects of PARP inhibition are more pronounced in dividing cells, we investigated the mechanisms underlying radiosensitization of human glioma cells by KU-0059436, evaluating the replication dependence ofmore » this effect and its therapeutic potential. Results: KU-0059436 increased the radiosensitivity of four human glioma cell lines (T98G, U373-MG, UVW, and U87-MG). Radiosensitization was enhanced in populations synchronized in S phase and abrogated by concomitant exposure to aphidicolin. Sensitization was further enhanced when the inhibitor was combined with a fractionated radiation schedule. KU-0059436 delayed repair of radiation-induced DNA breaks and was associated with a replication-dependent increase in {gamma}H2AX and Rad51 foci. Conclusion: The results of our study have shown that KU-0059436 increases radiosensitivity in a replication-dependent manner that is enhanced by fractionation. A mechanism is proposed whereby PARP inhibition increases the incidence of collapsed replication forks after ionizing radiation, generating persistent DNA double-strand breaks. These observations indicate that KU-0059436 is likely to enhance the therapeutic ratio achieved by radiotherapy in the treatment of glioblastoma multiforme. A Phase I clinical trial is in development.« less

3-Bromopyruvate inhibits cell proliferation and induces apoptosis in CD133+ population in human glioma.

PubMed

Xu, Dong-Qiang; Tan, Xiao-Yu; Zhang, Bao-Wei; Wu, Tao; Liu, Ping; Sun, Shao-Jun; Cao, Yin-Guang

2016-03-01

The study was aimed to investigate the role of 3-bromopyruvate in inhibition of CD133+ U87 human glioma cell population growth. The results demonstrated that 3-bromopyruvate inhibited the viability of both CD133+ and parental cells derived from U87 human glioma cell line. However, the 3-bromopyruvate-induced inhibition in viability was more prominent in CD133+ cells at 10 μM concentration after 48 h. Treatment of CD133+ cells with 3-bromopyruvate caused reduction in cell population and cell size, membrane bubbling, and degradation of cell membranes. Hoechst 33258 staining showed condensation of chromatin material and fragmentation of DNA in treated CD133+ cells after 48 h. 3-Bromopyruvate inhibited the migration rate of CD133+ cells significantly compared to the parental cells. Flow cytometry revealed that exposure of CD133+ cells to 3-bromopyruvate increased the cell population in S phase from 24.5 to 37.9 % with increase in time from 12 to 48 h. In addition, 3-bromopyruvate significantly enhanced the expression of Bax and cleaved caspase 3 in CD133+ cells compared to the parental cells. Therefore, 3-bromopyruvate is a potent chemotherapeutic agent for the treatment of glioma by targeting stem cells selectively.

Phosphatidylcholine-specific phospholipase C inhibition down- regulates CXCR4 expression and interferes with proliferation, invasion and glycolysis in glioma cells

PubMed Central

Ricci, Alessandro; Pacella, Aurora; Cigliana, Giovanni; Bozzuto, Giuseppina; Podo, Franca; Carpinelli, Giulia

2017-01-01

Background The chemokine receptor CXCR4 plays a crucial role in tumors, including glioblastoma multiforme (GBM), the most aggressive glioma. Phosphatidylcholine-specific phospholipase C (PC-PLC), a catabolic enzyme of PC metabolism, is involved in several aspects of cancer biology and its inhibition down-modulates the expression of growth factor membrane receptors interfering with their signaling pathways. In the present work we investigated the possible interplay between CXCR4 and PC-PLC in GBM cells. Methods Confocal microscopy, immunoprecipitation, western blot analyses, and the evaluation of migration and invasion potential were performed on U87MG cells after PC-PLC inhibition with the xanthate D609. The intracellular metabolome was investigated by magnetic resonance spectroscopy; lactate levels and lactate dehydrogenase (LDH) activity were analyzed by colorimetric assay. Results Our studies demonstrated that CXCR4 and PC-PLC co-localize and are associated on U87MG cell membrane. D609 reduced CXCR4 expression, cell proliferation and invasion, interfering with AKT and EGFR activation and expression. Metabolic analyses showed a decrease in intracellular lactate concentration together with a decrement in LDH activity. Conclusions Our data suggest that inhibition of PC-PLC could represent a new molecular approach in glioma biology not only for its ability in modulating cell metabolism, glioma growth and motility, but also for its inhibitory effect on crucial molecules involved in cancer progression. PMID:28423060

Homozygously deleted gene DACH1 regulates tumor-initiating activity of glioma cells

PubMed Central

Watanabe, Akira; Ogiwara, Hideki; Ehata, Shogo; Mukasa, Akitake; Ishikawa, Shumpei; Maeda, Daichi; Ueki, Keisuke; Ino, Yasushi; Todo, Tomoki; Yamada, Yasuhiro; Fukayama, Masashi; Saito, Nobuhito; Miyazono, Kohei; Aburatani, Hiroyuki

2011-01-01

Loss or reduction in function of tumor suppressor genes contributes to tumorigenesis. Here, by allelic DNA copy number analysis using single-nucleotide polymorphism genotyping array and mass spectrometry, we report homozygous deletion in glioblastoma multiformes at chromosome 13q21, where DACH1 gene is located. We found decreased cell proliferation of a series of glioma cell lines by forced expression of DACH1. We then generated U87TR-Da glioma cells, where DACH1 expression could be activated by exposure of the cells to doxycycline. Both ex vivo cellular proliferation and in vivo growth of s.c. transplanted tumors in mice are reduced in U87TR-Da cells with DACH1 expression (U87-DACH1-high), compared with DACH1-nonexpressing U87TR-Da cells (U87-DACH1-low). U87-DACH1-low cells form spheroids with CD133 and Nestin expression in serum-free medium but U87-DACH1-high cells do not. Compared with spheroid-forming U87-DACH1-low cells, adherent U87-DACH1-high cells display lower tumorigenicity, indicating DACH1 decreases the number of tumor-initiating cells. Gene expression analysis and chromatin immunoprecipitation assay reveal that fibroblast growth factor 2 (FGF2/bFGF) is transcriptionally repressed by DACH1, especially in cells cultured in serum-free medium. Exogenous bFGF rescues spheroid-forming activity and tumorigenicity of the U87-DACH1-high cells, suggesting that loss of DACH1 increases the number of tumor-initiating cells through transcriptional activation of bFGF. These results illustrate that DACH1 is a distinctive tumor suppressor, which does not only suppress growth of tumor cells but also regulates bFGF-mediated tumor-initiating activity of glioma cells. PMID:21750150

Perfluorocarbon-Loaded Lipid Nanocapsules to Assess the Dependence of U87-Human Glioblastoma Tumor pO2 on In Vitro Expansion Conditions

PubMed Central

Lemaire, Laurent; Nel, Janske; Franconi, Florence; Bastiat, Guillaume; Saulnier, Patrick

2016-01-01

Growing tumor cell lines, such as U87-MG glioma cells, under mild hypoxia (3% O2) leads to a ca. 40% reduction in growth rate once implanted in the brain of nude mice, as compared to normoxia (21% O2) grown cells, wherein the former over-express HIF-1 and VEGF-A. Despite developing differently, the tumors have similar: blood perfusion, oxygen consumption, and vascular surface area parameters, whereas the number of blood vessels is nearly doubled in the tumor arising from normoxia cultured cells. Interestingly, tumor oxygen tension, measured using 19F-oximetry, showed that the normoxia grown cells led to tumors characterized by mild hypoxic environment (approximately 4%) conditions, whilst the hypoxia grown cells led to tumors characterized by physioxic environment (approximately 6%) conditions. This reversal in oxygen concentration may be responsible for the apparent paradoxical growth profiles. PMID:27788227

Dominant Negative Pleiotrophin Induces Tetraploidy and Aneuploidy in U87MG Human Glioblastoma Cells

PubMed Central

Chang, Yunchao; Berenson, James R.; Wang, Zhaoyi; Deuel, Thomas F.

2007-01-01

Summary Pleiotrophin (PTN, Ptn) is an 18 kD secretory cytokine that is expressed in many human cancers, including glioblastoma. In previous experiments, interruption of the constitutive PTN signaling in human U87MG glioblastoma cells that inappropriately express endogenous Ptn reversed their rapid growth in vitro and their malignant phenotype in vivo. To seek a mechanism for the effect of the dominant negative PTN, flow cytometry was used to compare the profiles of U87MG cells and four clones of U87MG cells that express the dominant negative PTN (U87MG/PTN 1–40 cells); here, we report that the dominant negative PTN in U87MG cells induces tetraploidy and aneuploidy and arrests the tetraploid and aneuploid cells in the G1 phase of the cell cycle. The data suggest that PTN signaling may have a critical role in chromosomal segregation and cell cycle progression; the data suggest induction of tetraploidy and aneuploidy in U87MG glioblastoma cells may be an important mechanism that contributes to the loss of the malignant phenotype of U87MG cells. PMID:17067552

MiR-320 inhibits the growth of glioma cells through downregulating PBX3.

PubMed

Pan, Cuicui; Gao, Hua; Zheng, Ni; Gao, Qi; Si, Yuanquan; Zhao, Yueran

2017-09-21

MiR-320 is downregulated in multiple cancers, including glioma and acts as tumor suppressor through inhibiting tumor cells proliferation and inducing apoptosis. PBX3 (Pre-B cell leukemia homeobox 3), a putative target gene of miR-320, has been reported to be upregulated in various tumors and promote tumor cell growth through regulating MAKP/ERK pathway. This study aimed to verify whether miR-320 influences glioma cells growth through regulating PBX3. Twenty-four human glioma and paired adjacent nontumorous tissues were collected for determination of miR-320 and PBX3 expression using RT-qPCR and western blot assays. Luciferase reporter assay was performed to verify the interaction between miR-320 and its targeting sequence in the 3' UTR of PBX3 in glioma cells U87 and U251. Increased miR-320 level in U87 and U251 cells was achieved through miR-320 mimic transfection and the effect of which on glioma cells growth, proliferation, cell cycle, apoptosis and activation of Raf-1/MAPK pathway was determined using MTT, colony formation, flow cytometry and western blot assays. PBX3 knockdown was performed using shPBX3 and the influence on MAPK pathway activation was evaluated. MiR-320 downregulation and PBX3 upregulation was found in glioma tissues. Luciferase reporter assays identified miR-320 directly blinds to the 3' UTR of PBX3 in glioma cells. MiR-320 mimic transfection suppressed glioma cells proliferation, and induced cell cycle arrest and apoptosis. Both miR-320 overexpression and PBX3 knockdown inhibited Raf-1/MAPK activation. MiR-320 may suppress glioma cells growth and induced apoptosis through the PBX3/Raf-1/MAPK axis, and miR-320 oligonucleotides may be a potential cancer therapeutic for glioma.

Microglia Activate Migration of Glioma Cells through a Pyk2 Intracellular Pathway

PubMed Central

Rolón-Reyes, Kimberleve; Kucheryavykh, Yuriy V.; Cubano, Luis A.; Inyushin, Mikhail; Skatchkov, Serguei N.; Eaton, Misty J.; Harrison, Jeffrey K.; Kucheryavykh, Lilia Y.

2015-01-01

Glioblastoma is one of the most aggressive and fatal brain cancers due to the highly invasive nature of glioma cells. Microglia infiltrate most glioma tumors and, therefore, make up an important component of the glioma microenvironment. In the tumor environment, microglia release factors that lead to the degradation of the extracellular matrix and stimulate signaling pathways to promote glioma cell invasion. In the present study, we demonstrated that microglia can promote glioma migration through a mechanism independent of extracellular matrix degradation. Using western blot analysis, we found upregulation of proline rich tyrosine kinase 2 (Pyk2) protein phosphorylated at Tyr579/580 in glioma cells treated with microglia conditioned medium. This upregulation occurred in rodent C6 and GL261 as well as in human glioma cell lines with varying levels of invasiveness (U-87MG, A172, and HS683). siRNA knock-down of Pyk2 protein and pharmacological blockade by the Pyk2/focal-adhesion kinase (FAK) inhibitor PF-562,271 reversed the stimulatory effect of microglia on glioma migration in all cell lines. A lower concentration of PF-562,271 that selectively inhibits FAK, but not Pyk2, did not have any effect on glioma cell migration. Moreover, with the use of the CD11b-HSVTK microglia ablation mouse model we demonstrated that elimination of microglia in the implanted tumors (GL261 glioma cells were used for brain implantation) by the local in-tumor administration of Ganciclovir, significantly reduced the phosphorylation of Pyk2 at Tyr579/580 in implanted tumor cells. Taken together, these data indicate that microglial cells activate glioma cell migration/dispersal through the pro-migratory Pyk2 signaling pathway in glioma cells. PMID:26098895

Radiosensitivity of Patient-Derived Glioma Stem Cell 3-Dimensional Cultures to Photon, Proton, and Carbon Irradiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chiblak, Sara; Tang, Zili; Molecular and Translational Radiation Oncology, Heidelberg Ion Therapy Center, Heidelberg Institute of Radiation Oncology, University of Heidelberg Medical School and National Center for Tumor Diseases, German Cancer Research Center, Heidelberg

Purpose: To investigate the radiosensitivity of primary glioma stem cell (GSC) cultures with different CD133 status in a 3-dimensional (3D) model after photon versus proton versus carbon irradiation. Methods and Materials: Human primary GSC spheroid cultures were established from tumor specimens of six consented glioblastoma patients. Human U87MG was used as a classical glioblastoma radioresistant cell line. Cell suspensions were generated by mechanical dissociation of GSC spheroids and embedded in a semi-solid 3D matrix before irradiation. Spheroid-like colonies were manually counted by microscopy. Cells were also recovered and quantified by fluorescence. CD133 expression and DNA damage were evaluated by flow cytometry.more » Results: The fraction of CD133{sup +} cells varied between 0.014% and 96% in the six GSC cultures and showed a nonsignificant correlation with plating efficiency and survival fractions. The 4 most photon-radioresistant GSC cultures were NCH644, NCH421k, NCH441, and NCH636. Clonogenic survival for proton irradiation revealed relative biologic effectiveness (RBE) in the range of 0.7-1.20. However, carbon irradiation rendered the photon-resistant GSC cultures sensitive, with average RBE of 1.87-3.44. This effect was partly attributed to impaired capability of GSC to repair carbon ion–induced DNA double-strand breaks as determined by residual DNA repair foci. Interestingly, radiosensitivity of U87 cells was comparable to GSC cultures using clonogenic survival as the standard readout. Conclusions: Carbon irradiation is effective in GSC eradication with similar RBE ranges approximately 2-3 as compared with non-stem GSC cultures (U87). Our data strongly suggest further exploration of GSC using classic radiobiology endpoints such as the here-used 3D clonogenic survival assay and integration of additional GSC-specific markers.« less

Procollagen-lysine 2-oxoglutarate 5-dioxygenase 2 promotes hypoxia-induced glioma migration and invasion

PubMed Central

Xu, Yangyang; Zhang, Lin; Wei, Yuzhen; Zhang, Xin; Xu, Ran; Han, Mingzhi; Huang, Bing; Chen, Anjing; Li, Wenjie; Zhang, Qing; Li, Gang; Wang, Jian; Zhao, Peng; Li, Xingang

2017-01-01

Poor prognosis of glioblastoma multiforme is strongly associated with the ability of tumor cells to invade the brain parenchyma, which is believed to be the major factor responsible for glioblastoma recurrence. Therefore, identifying the molecular mechanisms driving invasion may lead to the development of improved therapies for glioblastoma patients. Here, we investigated the role of procollagen-lysine 2-oxoglutarate 5-dioxygenase 2 (PLOD2), an enzyme catalyzing collagen cross-linking, in the biology of glioblastoma invasion. PLOD2 mRNA was significantly overexpressed in glioblastoma compared to low-grade tumors based on the Oncomine datasets and REMBRANDT database for human gliomas. Kaplan-Meier estimates based on the TCGA dataset demonstrated that high PLOD2 expression was associated with poor prognosis. In vitro, hypoxia upregulated PLOD2 protein in U87 and U251 human glioma cell lines. siRNA knockdown of endogenous HIF-1α or treatment of cells with the HIF-1α inhibitor PX-478 largely abolished the hypoxia-mediated PLOD2 upregulation. Knockdown of PLOD2 in glioma cell lines led to decreases in migration and invasion under normoxia and hypoxia. In addition, levels of phosphorylated FAK (Tyr 397), an important kinase mediating cell adhesion, were reduced in U87-shPLOD2 and U251-shPLOD2 cells, particularly under hypoxic conditions. Finally, orthotopic U251-shPLOD2 xenografts were circumscribed rather than locally invasive. In conclusion, the results indicated that PLOD2 was a gene of clinical relevance with implications in glioblastoma invasion and treatment strategies. PMID:28423580

Targeting TWIST1 through loss of function inhibits tumorigenicity of human glioblastoma.

PubMed

Mikheev, Andrei M; Mikheeva, Svetlana A; Severs, Liza J; Funk, Cory C; Huang, Lei; McFaline-Figueroa, José L; Schwensen, Jeanette; Trapnell, Cole; Price, Nathan D; Wong, Stephen; Rostomily, Robert C

2018-05-13

Twist1 (TW) is a bHLH transcription factor (TF) and master regulator of the epithelial to mesenchymal transition (EMT). In vitro, TW promotes mesenchymal change, invasion and self-renewal in glioblastoma (GBM) cells. However the potential therapeutic relevance of TW has not been established through loss of function studies in human GBM cell xenograft models. The effects of TW loss of function (gene editing and knock down) on inhibition of tumorigenicity of U87MG and GBM4 glioma stem cells were tested in orthotopic xenograft models and conditional knockdown in established flank xenograft tumors. RNAseq and the analysis of tumors investigated putative TW associated mechanisms. Multiple bioinformatics tools revealed significant alteration of ECM, membrane receptors, signaling transduction kinases and cytoskeleton dynamics leading to identification of PI3K/AKT signaling. We experimentally show alteration of AKT activity and periostin (POSTN) expression in vivo and/or in vitro. For the first time we show that effect of TW knockout inhibits AKT activity in U87MG cells in vivo independent of PTEN mutation. The clinical relevance of TW and candidate mechanisms was established by analysis of the TCGA and ENCODE databases. TW expression was associated with decreased patient survival and LASSO regression analysis identified POSTN as one of top targets of TW in human GBM. While we previously demonstrated the role of TW in promoting EMT and invasion of glioma cells, these studies provide direct experimental evidence supporting pro-tumorigenic role of TW independent of invasion in vivo and the therapeutic relevance of targeting TW in human GBM. Further, the role of TW driving POSTN expression and AKT signaling suggests actionable targets, which could be leveraged to mitigate the oncogenic effects of TW in GBM. Molecular Oncology (2018) © 2018 The Authors. Published by FEBS Press and John Wiley & Sons Ltd.

A choline derivate-modified nanoprobe for glioma diagnosis using MRI

NASA Astrophysics Data System (ADS)

Li, Jianfeng; Huang, Shixian; Shao, Kun; Liu, Yang; An, Sai; Kuang, Yuyang; Guo, Yubo; Ma, Haojun; Wang, Xuxia; Jiang, Chen

2013-04-01

Gadolinium (Gd) chelate contrast-enhanced magnetic resonance imaging (MRI) is a preferred method of glioma detection and preoperative localisation because it offers high spatial resolution and non-invasive deep tissue penetration. Gd-based contrast agents, such as Gd-diethyltriaminepentaacetic acid (DTPA-Gd, Magnevist), are widely used clinically for tumor diagnosis. However, the Gd-based MRI approach is limited for patients with glioma who have an uncompromised blood-brain barrier (BBB). Moreover, the rapid renal clearance and non-specificity of such contrast agents further hinders their prevalence. We present a choline derivate (CD)-modified nanoprobe with BBB permeability, glioma specificity and a long blood half-life. Specific accumulation of the nanoprobe in gliomas and subsequent MRI contrast enhancement are demonstrated in vitro in U87 MG cells and in vivo in a xenograft nude model. BBB and glioma dual targeting by this nanoprobe may facilitate precise detection of gliomas with an uncompromised BBB and may offer better preoperative and intraoperative tumor localization.

Hypoxic regulation of the expression of genes encoded estrogen related proteins in U87 glioma cells: eff ect of IRE1 inhibition.

PubMed

Minchenko, D O; Riabovol, O O; Ratushna, O O; Minchenko, O H

2017-01-01

The aim of the present study was to examine the effect of inhibition of endoplasmic reticulum stress signaling, mediated by IRE1 (inositol requiring enzyme 1), which is a central mediator of the unfolded protein response on the expression of genes encoded estrogen related proteins (NRIP1/RIP140, TRIM16/EBBP, ESRRA/NR3B1, FAM162A/E2IG5, PGRMC2/PMBP, and SLC39A6/LIV-1) and their hypoxic regulation in U87 glioma cells for evaluation of their possible significance in the control of glioma cells proliferation. The expression of NRIP1, EBBP, ESRRA, E2IG5, PGRMC2, and SLC39A6 genes in U87 glioma cells, transfected by empty vector pcDNA3.1 (control) and cells without IRE1 signaling enzyme function (transfected by dnIRE1) upon hypoxia, was studied by a quantitative polymerase chain reaction. Inhibition of both enzymatic activities (kinase and endoribonuclease) of IRE1 signaling enzyme function up-regulates the expression of EBBP, E2IG5, PGRMC2, and SLC39A6 genes is in U87 glioma cells in comparison with the control glioma cells, with more significant changes for E2IG5 and PGRMC2 genes. At the same time, the expression of NRIP1 and ESRRA genes is strongly down-regulated in glioma cells upon inhibition of IRE1. We also showed that hypoxia increases the expression of E2IG5, PGRMC2, and EBBP genes and decreases NRIP1 and ESRRA genes expression in control glioma cells. Furthermore, the inhibition of IRE1 in U87 glioma cells decreases the eff ect of hypoxia on the expression of E2IG5 and PGRMC2 genes, eliminates hypoxic regulation of NRIP1 gene, and enhances the sensitivity of ESRRA gene to hypoxic condition. Furthermore, the expression of SLC39A6 gene is resistant to hypoxia in both the glioma cells with and without IRE1 signaling enzyme function. Results of this investigation demonstrate that inhibition of IRE1 signaling enzyme function affects the expression of NRIP1, EBBP, ESRRA, E2IG5, PGRMC2, and SLC39A6 genes in U87 glioma cells in gene specific manner and these changes possibly contribute to the suppression of the cell proliferation. Most of these genes are regulated by hypoxia and preferentially through IRE1 signaling pathway of endoplasmic reticulum stress.

miR-141-3p functions as a tumor suppressor modulating activating transcription factor 5 in glioma.

PubMed

Wang, Mengyuan; Hu, Ming; Li, Zhaohua; Qian, Dongmeng; Wang, Bin; Liu, David X

2017-09-02

Glioma is the most common malignant primary brain tumor which arises from the central nervous system. Our studies reported that an anti-apoptotic factor, activating transcription factor 5 (ATF5), is highly expressed in malignant glioma specimens and cell lines. Downregulation by dominant-negetive ATF5 could repress glioma cell proliferation and accelerate apoptosis. Here, we further investigate the upstream factor which regulates ATF5 expression. Bioinformatic analysis showed that ATF5 was a potential target of miR-141-3p. Luciferase reporter assay verified that miR-141-3p specifically targeted the ATF5 3'-UTR in glioma cells. Functional studied suggested that miR-141-3p overexpression inhibited proliferation and promoted apoptosis of glioma cells (U87MG and U251). Xenograft experiments proved the inhibition of miR-141-3p on glioma growth in vivo. Moreover, exogenous ATF5 without 3'-UTR restored the cell proliferation inhibition triggered by miR-141-3p. Taken together, we put forward that miR-141-3p is a new upstream target towards ATF5. It can serve as a crucial tumor suppressor in regulating the ATF5-regulated growth of malignant glioma. Copyright © 2017 Elsevier Inc. All rights reserved.

Asiatic Acid Inhibits Pro-Angiogenic Effects of VEGF and Human Gliomas in Endothelial Cell Culture Models

PubMed Central

Kavitha, Chandagirikoppal V.; Agarwal, Chapla; Agarwal, Rajesh; Deep, Gagan

2011-01-01

Malignant gliomas are one of the most devastating and incurable tumors. Sustained excessive angiogenesis by glioma cells is the major reason for their uncontrolled growth and resistance toward conventional therapies resulting in high mortality. Therefore, targeting angiogenesis should be a logical strategy to prevent or control glioma cell growth. Earlier studies have shown that Asiatic Acid (AsA), a pentacyclic triterpenoid, is effective against glioma and other cancer cells; however, its efficacy against angiogenesis remains unknown. In the present study, we examined the anti-angiogenic efficacy of AsA using human umbilical vein endothelial cells (HUVEC) and human brain microvascular endothelial cells (HBMEC). Our results showed that AsA (5–20 µM) inhibits HUVEC growth and induces apoptotic cell death by activating caspases (3 and 9) and modulating the expression of apoptosis regulators Bad, survivin and pAkt-ser473. Further, AsA showed a dose-dependent inhibition of HUVEC migration, invasion and capillary tube formation, and disintegrated preformed capillary network. AsA also inhibited the VEGF-stimulated growth and capillary tube formation by HUVEC and HBMEC. Next, we analyzed the angiogenic potential of conditioned media collected from human glioma LN18 and U87-MG cells treated with either DMSO (control conditioned media, CCM) or AsA 20 µM (AsA20 conditioned media, AsA20CM). CCM from glioma cells significantly enhanced the capillary tube formation in both HUVEC and HBMEC, while capillary tube formation in both endothelial cell lines was greatly compromised in the presence of AsA20CM. Consistent with these results, VEGF expression was lesser in AsA20CM compared to CCM, and indeed AsA strongly inhibited VEGF level (both cellular and secreted) in glioma cells. AsA also showed dose-dependent anti-angiogenic efficacy in Matrigel plug assay, and inhibited the glioma cells potential to attract HUVEC/HBMEC. Overall, the present study clearly showed the strong anti-angiogenic potential of AsA and suggests its usefulness against malignant gliomas. PMID:21826202

Influence of in vivo growth on human glioma cell line gene expression: Convergent profiles under orthotopic conditions

PubMed Central

Camphausen, Kevin; Purow, Benjamin; Sproull, Mary; Scott, Tamalee; Ozawa, Tomoko; Deen, Dennis F.; Tofilon, Philip J.

2005-01-01

Defining the molecules that regulate tumor cell survival is an essential prerequisite for the development of targeted approaches to cancer treatment. Whereas many studies aimed at identifying such targets use human tumor cells grown in vitro or as s.c. xenografts, it is unclear whether such experimental models replicate the phenotype of the in situ tumor cell. To begin addressing this issue, we have used microarray analysis to define the gene expression profile of two human glioma cell lines (U251 and U87) when grown in vitro and in vivo as s.c. or as intracerebral (i.c.) xenografts. For each cell line, the gene expression profile generated from tissue culture was significantly different from that generated from the s.c. tumor, which was significantly different from those grown i.c. The disparity between the i.c gene expression profiles and those generated from s.c. xenografts suggests that whereas an in vivo growth environment modulates gene expression, orthotopic growth conditions induce a different set of modifications. In this study the U251 and U87 gene expression profiles generated under the three growth conditions were also compared. As expected, the profiles of the two glioma cell lines were significantly different when grown as monolayer cultures. However, the glioma cell lines had similar gene expression profiles when grown i.c. These results suggest that tumor cell gene expression, and thus phenotype, as defined in vitro is affected not only by in vivo growth but also by orthotopic growth, which may have implications regarding the identification of relevant targets for cancer therapy. PMID:15928080

Comparison of Biological Properties of 99mTc-Labeled Cyclic RGD Peptide Trimer and Dimer Useful as SPECT Radiotracers for Tumor Imaging

PubMed Central

Zhao, Zuo-Quan; Yang, Yong; Fang, Wei; Liu, Shuang

2016-01-01

Introduction This study sought to evaluate a 99mTc-labeled trimeric cyclic RGD peptide (99mTc-4P-RGD3) as the new radiotracer for tumor imaging. The objective was to compare its biological properties with those of 99mTc-3P-RGD2 in the same animal model. Methods HYNIC-4P-RGD3 was prepared by reacting 4P-RGD3 with excess HYNIC-OSu in the presence of diisopropylethylamine. 99mTc-4P-RGD3 was prepared using a kit formulation, and evaluated for its tumor-targeting capability and biodistribution properties in the BALB/c nude mice with U87MG human glioma xenografts. Planar and SPECT imaging studies were performed in athymic nude mice with U87MG glioma xenografts. For comparison purpose, 99mTc-3P-RGD2 (a αvβ3-targeted radiotracer currently under clinical evaluation for tumor imaging in cancer patients) was also evaluated in the same animal models. Blocking experiments were used to demonstrate the αvβ3 specificity of 99mTc-4P-RGD3. Results 99mTc-4P-RGD3 was prepared with >95% RCP and high specific activity (~200 GBq/µmol). 99mTc-4P-RGD3 and 99mTc-3P-RGD2 shared almost identical tumor uptake and similar biodistribution properties. 99mTc-4P-RGD3 had higher uptake than 99mTc-3P-RGD2 in the intestines and kidneys; but it showed better metabolic stability. The U87MG tumors were clearly visualized by SPECT with excellent contrast with 99mTc-4P-RGD3 and 99mTc-3P-RGD2. Conclusion Increasing peptide multiplicity from 3P-RGD2 to 4P-RGD3 offers no advantages with respect to the tumor-targeting capability. 99mTc-4P-RGD3 is as good a SPECT radiotracer as 99mTc-3P-RGD2 for imaging αvβ3-positive tumors. PMID:27556955

Sensitization of cerebral tissue in nude mice with photodynamic therapy induces ADAM17/TACE and promotes glioma cell invasion

PubMed Central

Zheng, Xuguang; Jiang, Feng; Katakowski, Mark; Zhang, Xuepeng; Jiang, Hao; Zhang, Zheng Gang; Chopp, Michael

2008-01-01

In the present study, we tested the hypothesis that a mild cerebral tissue injury promotes subsequent glioma invasion via activation of the ADAM17-EGFR-PI3K-Akt pathway. Mild injury was induced by Photodynamic therapy (PDT), which employs tissue-penetrating laser light exposure following systemic administration of a tumor-localizing photosensitizer. Athymic nude mice were treated with sublethal PDT (80J/cm2 with 2mg/kg Photofrin). Hypoxic stress and ADAM17-EGFR-PI3K-Akt were measured using Western blot and immunostaining. Additional groups with/without pro-sublethal PDT were subsequently implanted with U87 glioma tumor cell. Tumor invasion and ADAM17-EGFR-PI3K-Akt pathway in tumor area were measured. After a sublethal dose of PDT, HIF-1α expression was increased by a factor of three in PDT-treated normal brain tissue compared to contralateral control brain tissue. PDT-treated brain tissue exhibited a significant increase in ADAM17, p-EGFR, p-Akt expression compared to non-treated tissue. ADAM17 positive area significantly increased from 1.78% to 10.89%. The percentage of p-EGFR and p-Akt positive cells significantly increased from 9.50% and 14.50% to 21.31% and 32.29%,respectively, PDT treatment significantly increased subsequent implanted U87 glioma cell invasion by 3.68-fold and increased ADAM17, EGFR, p-EGFR, Akt, p-Akt expression by 178%, 43.9%,152.7%, 89.6%,and 164.2%, respectively, compared to control group. Our data showed that a sublethal sensitization of cerebral tissue with PDT significantly increased U87 cell invasion in nude mice, and that glioma cell invasion is highly correlated with activation of the ADAM17-EGFR-PI3K-Akt pathway (r=0.928, 0.775, 0.870, 0.872, and 0.883, respectively), most likely via HIF-1α. PMID:18358600

SIRT6 suppresses glioma cell growth via induction of apoptosis, inhibition of oxidative stress and suppression of JAK2/STAT3 signaling pathway activation.

PubMed

Feng, Jun; Yan, Peng-Fei; Zhao, Hong-Yang; Zhang, Fang-Cheng; Zhao, Wo-Hua; Feng, Min

2016-03-01

Sirtuin 6 (SIRT6) is a member of the mammalian NAD+‑dependent deacetylase sirtuin family that acts to maintain genomic stability and to repress genes. SIRT6 has recently been reported to be a tumor suppressor that controls cancer metabolism, although this effect of SIRT6 is still in dispute. Moreover, the role of SIRT6 in glioma is largely unknown. In the present study, we found that overexpression of SIRT6 using an adenovirus inhibited glioma cell growth and induced marked cell injury in two glioma cell lines (U87‑MG and T98G). Fluorescent terminal deoxyribonucleotidyl transferase (TdT)‑mediated biotin‑16‑dUTP nick‑end labelling (TUNEL) assay showed that SIRT6 overexpression induced obvious apoptosis in the T98G glioma cells. Immunoblotting and immunofluorescent staining demonstrated that SIRT6 overexpression promoted the mitochondrial-to‑nuclear translocation of apoptosis‑inducing factor (AIF), a potent apoptosis inducer. Moreover, we found that SIRT6 overexpression largely reduced oxidative stress and suppressed the activation of the JAK2/STAT3 signaling pathway in glioma cells. Finally, we showed that SIRT6 mRNA and protein levels in human glioblastoma multiforme tissues were significantly lower than the levels in peritumor tissues. In summary, our data suggest that SIRT6 suppresses glioma cell growth via induction of apoptosis, inhibition of oxidative stress and inhibition of the activation of the JAK2/STAT3 signaling pathway. These results indicate that SIRT6 may be a promising therapeutic target for glioma treatment.

Increased betulinic acid induced cytotoxicity and radiosensitivity in glioma cells under hypoxic conditions.

PubMed

Bache, Matthias; Zschornak, Martin P; Passin, Sarina; Kessler, Jacqueline; Wichmann, Henri; Kappler, Matthias; Paschke, Reinhard; Kaluđerović, Goran N; Kommera, Harish; Taubert, Helge; Vordermark, Dirk

2011-09-09

Betulinic acid (BA) is a novel antineoplastic agent under evaluation for tumor therapy. Because of the selective cytotoxic effects of BA in tumor cells (including gliomas), the combination of this agent with conservative therapies (such as radiotherapy and chemotherapy) may be useful. Previously, the combination of BA with irradiation under hypoxic conditions had never been studied. In this study, the effects of 3 to 30 μM BA on cytotoxicity, migration, the protein expression of PARP, survivin and HIF-1α, as well as radiosensitivity under normoxic and hypoxic conditions were analyzed in the human malignant glioma cell lines U251MG and U343MG. Cytotoxicity and radiosensitivity were analyzed with clonogenic survival assays, migration was analyzed with Boyden chamber assays (or scratch assays) and protein expression was examined with Western blot analyses. Under normoxic conditions, a half maximal inhibitory concentration (IC50) of 23 μM was observed in U251MG cells and 24 μM was observed in U343MG cells. Under hypoxic conditions, 10 μM or 15 μM of BA showed a significantly increased cytotoxicity in U251MG cells (p = 0.004 and p = 0.01, respectively) and U343MG cells (p < 0.05 and p = 0.01, respectively). The combination of BA with radiotherapy resulted in an additive effect in the U343MG cell line under normoxic and hypoxic conditions. Weak radiation enhancement was observed in U251MG cell line after treatment with BA under normoxic conditions. Furthermore, under hypoxic conditions, the incubation with BA resulted in increased radiation enhancement. The enhancement factor, at an irradiation dose of 15 Gy after treatment with 10 or 15 μM BA, was 2.20 (p = 0.02) and 4.50 (p = 0.03), respectively. Incubation with BA led to decreased cell migration, cleavage of PARP and decreased expression levels of survivin in both cell lines. Additionally, BA treatment resulted in a reduction of HIF-1α protein under hypoxic conditions. Our results suggest that BA is capable of improving the effects of tumor therapy in human malignant glioma cells, particularly under hypoxic conditions. Further investigations are necessary to characterize its potential as a radiosensitizer.

Erythropoietin Augments Survival of Glioma Cells After Radiation and Temozolomide

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hassouna, Imam; Sperling, Swetlana; Kim, Ella

2008-11-01

Purpose: Despite beneficial effects of irradiation/chemotherapy on survival of glioblastoma (GBM) patients, collateral damage to intact neural tissue leads to 'radiochemobrain' and reduced quality of life in survivors. For prophylactic neuroprotection, erythropoietin (EPO) is a promising candidate, provided that concerns regarding potential tumor promoting effects are alleviated. Methods and Materials: Human GBM-derived cell lines U87, G44, G112, and the gliosarcoma-derived line G28 were treated with EPO, with and without combinations of irradiation or temozolomide (TMZ). Responsiveness of glioma cells to EPO was measured by cell migration from spheroids, cell proliferation, and clonogenic survival. Implantation of U87 cells into brains ofmore » nude mice, followed 5 days later by EPO treatment (5,000 U/kg intraperitoneal every other day for 2 weeks) should reveal effects of EPO on tumor growth in vivo. Reverse transcriptase-polymerase chain reaction was performed for EPOR, HIF-1{alpha}, and epidermal growth factor receptor (EGFR)vIII in cell lines and 22 human GBM specimens. Results: EPO did not modulate basal glioma cell migration and stimulated proliferation in only one of four cell lines. Importantly, EPO did not enhance tumor growth in mouse brains. Preincubation of glioma cells with EPO for 3 h, followed by irradiation and TMZ for another 24 h, resulted in protection against chemoradiation-induced cytotoxicity in three cell lines. Conversely, EPO induced a dose-dependent decrease in survival of G28 gliosarcoma cells. In GBM specimens, expression of HIF-1{alpha} correlated positively with expression of EPOR and EGFRvIII. EPOR and EGFRvIII expression did not correlate. Conclusions: EPO is unlikely to appreciably influence basal glioma growth. However, concomitant use of EPO with irradiation/chemotherapy in GBM patients is not advisable.« less

Anti-miR21 oligonucleotide enhances chemosensitivity of T98G cell line to doxorubicin by inducing apoptosis

PubMed Central

Giunti, Laura; da Ros, Martina; Vinci, Serena; Gelmini, Stefania; Iorio, Anna Lisa; Buccoliero, Anna Maria; Cardellicchio, Stefania; Castiglione, Francesca; Genitori, Lorenzo; de Martino, Maurizio; Giglio, Sabrina; Genuardi, Maurizio; Sardi, Iacopo

2015-01-01

Various signal transduction pathways seem to be involved in chemoresistance mechanism of glioblastomas (GBMs). miR-21 is an important oncogenic miRNA which modulates drug resistance of tumor cells. We analyzed the expression of 5 miRNAs, previously found to be dysregulated in high grade gliomas, in 9 pediatric (pGBM) and in 5 adult (aGBM) GBMs. miR-21 was over-expressed, with a significant difference between pGBMs and aGBMs represented by a 4 times lower degree of expression in the pediatric compared to the adult series (p = 0.001). Doxorubicin (Dox) seems to be an effective anti-glioma agent with high antitumor activity also against glioblastoma stem cells. We therefore evaluated the chemosensitivity to Dox in 3 GBM cell lines (A172, U87MG and T98G). Dox had a cytotoxic effect after 48 h of treatment in A172 and U87MG, while T98G cells were resistant. TUNEL assay verified that Dox induced apoptosis in A172 and U87MG but not in T98G. miR-21 showed a low basal expression in treated cells and was over-expressed in untreated cells. To validate the possible association of miR-21 with drug resistance of T98G cells, we transfected anti-miR-21 inhibitor into the cells. The expression level of miR-21 was significantly lower in T98G transfected cells (than in the parental control cells). Transfected cells showed a high apoptotic rate compared to control after Dox treatment by TUNEL assay, suggesting that combined Dox and miR-21 inhibitor therapy can sensitize GBM resistant cells to anthracyclines by enhancing apoptosis. PMID:25628933

Soy Metabolites, Isoflavones in Cell Growth and Apoptosis

DTIC Science & Technology

2000-07-01

previously PROTEINS BY APIGENIN IS P21/WAF1 INDEPENDENT. M McVean, W C shown that genistein, at 5MM, can block invasion of glioblastoma multiforme into...may Kansas City, KS be involved in the invasion of glioblastoma multiforme into FBRA. These studies Apigenin , a nonmutagenic flavonoid, has been shown...p21/wafl in modulating cell cycle regulatory lion mechanisms. In C6 rat glioma cells and U87 human glioma cells treated with proteins during apigenin

Convection enhanced delivery of panobinostat (LBH589)-loaded pluronic nano-micelles prolongs survival in the F98 rat glioma model.

PubMed

Singleton, W G; Collins, A M; Bienemann, A S; Killick-Cole, C L; Haynes, H R; Asby, D J; Butts, C P; Wyatt, M J; Barua, N U; Gill, S S

2017-01-01

The pan-histone deacetylase inhibitor panobinostat is a potential therapy for malignant glioma, but it is water insoluble and does not cross the blood-brain barrier when administered systemically. In this article, we describe the in vitro and in vivo efficacy of a novel water-soluble nano-micellar formulation of panobinostat designed for administration by convection enhanced delivery (CED). The in vitro efficacy of panobinostat-loaded nano-micelles against rat F98, human U87-MG and M059K glioma cells and against patient-derived glioma stem cells was measured using a cell viability assay. Nano-micelle distribution in rat brain was analyzed following acute CED using rhodamine-labeled nano-micelles, and toxicity was assayed using immunofluorescent microscopy and synaptophysin enzyme-linked immunosorbent assay. We compared the survival of the bioluminescent syngenic F98/Fischer344 rat glioblastoma model treated by acute CED of panobinostat-loaded nano-micelles with that of untreated and vehicle-only-treated controls. Nano-micellar panobinostat is cytotoxic to rat and human glioma cells in vitro in a dose-dependent manner following short-time exposure to drug. Fluorescent rhodamine-labelled nano-micelles distribute with a volume of infusion/volume of distribution (Vi/Vd) ratio of four and five respectively after administration by CED. Administration was not associated with any toxicity when compared to controls. CED of panobinostat-loaded nano-micelles was associated with significantly improved survival when compared to controls (n=8 per group; log-rank test, P <0.001). One hundred percent of treated animals survived the 60-day experimental period and had tumour response on post-mortem histological examination. CED of nano-micellar panobinostat represents a potential novel therapeutic option for malignant glioma and warrants translation into the clinic.

Anti-invasive and antiangiogenic effects of MMI-166 on malignant glioma cells

PubMed Central

2010-01-01

Background The constitutive overexpression of matrix metalloproteinases (MMPs) is frequently observed in malignant tumours. In particular, MMP-2 and MMP-9 have been reported to be closely associated with invasion and angiogenesis in malignant gliomas. Our study aimed to evaluate the antitumour effects of MMI-166 (Nalpha-[4-(2-Phenyl-2H- tetrazole-5-yl) phenyl sulfonyl]-D-tryptophan), a third generation MMP inhibitor, on three human glioma cell lines (T98G, U87MG, and ONS12) in vitro and in vivo. Methods The effects of MMI-166 on the gelatinolytic activity was analysed by gelatine zymography. The anti-invasive effect of MMI-166 was analysed by an in vitro invasion assay. An in vitro angiogenesis assay was also performed. In vitro growth inhibition of glioma cells by MMI-166 was determined by the MTT assay. The effect of MMI-166 on an orthotropic implantation model using athymic mice was also evaluated. Results Gelatine zymography revealed that MMP-2 and MMP-9 activities were suppressed by MMI-166. The invasion of glioma cells was suppressed by MMI-166. The angiogenesis assay showed that MMI-166 had a suppressive effect on glioma cell-induced angiogenesis. However, MMI-166 did not suppress glioma cell proliferation in the MTT assay. In vivo, MMI-166 suppressed tumour growth in athymic mice implanted orthotropically with T98G cells and showed an inhibitory effect on tumour-induced angiogenesis and tumour growth. This is the first report of the effect of a third generation MMP inhibitor on malignant glioma cells. Conclusions These results suggest that MMI-166 may have potentially suppressive effects on the invasion and angiogenesis of malignant gliomas. PMID:20587068

Antiglioma effects of cytarabine on leptomeningeal metastasis of high-grade glioma by targeting the PI3K/Akt/mTOR pathway

PubMed Central

Zhao, Kai-Hong; Zhang, Can; Bai, Yue; Li, Yan; Kang, Xun; Chen, Jian-Xin; Yao, Kun; Jiang, Tao; Zhong, Xiao-Song; Li, Wen-Bin

2017-01-01

Leptomeningeal metastasis (LM) of high-grade glioma is a highly lethal disease requiring new effective therapeutic measures. For both de novo or relapsed glioma with LM, intrathecal cytarabine chemotherapy is not frequently used for first-line and relapse protocols. We encountered a clinical case demonstrating effective application of cytarabine in high-grade glioma with LM, prompting us to explore the effects of cytarabine on malignant glioma and molecular mechanisms of such effects through in vivo and in vitro experiments. The U87 cell line was selected to represent human glioma for studies. Cell viability was measured by MTT assay, plate colony formation assay, and trypan-blue dye exclusion test. Apoptosis was assessed by flow cytometry. Protein expression levels were detected by Western blot assay and immunohistochemistry. mRNA expression was examined by quantitative real-time reverse transcription polymerase chain reaction. Cytarabine inhibited tumor growth during the in vivo experiment. The present study confirmed that cytarabine inhibits proliferation and promotes apoptosis of U87 cells, and molecular analysis of this effect showed that cytarabine significantly reduces expression of phosphatidylinositol 3-kinase/serine/threonine kinase also known as the protein kinase B/mechanistic target of rapamycin (PI3K/Akt/mTOR) pathway, Ki-67, BCL2, and 4-1BB, and upregulates Bax and cleaved caspase-3. Our findings indicated that intrathecal administration of cytarabine manifests potential in prophylaxis and treatment of malignant glioma with LM. Effective medications for high-grade glioma with LM should contain cytarabine. PMID:28721010

Antiglioma effects of cytarabine on leptomeningeal metastasis of high-grade glioma by targeting the PI3K/Akt/mTOR pathway.

PubMed

Zhao, Kai-Hong; Zhang, Can; Bai, Yue; Li, Yan; Kang, Xun; Chen, Jian-Xin; Yao, Kun; Jiang, Tao; Zhong, Xiao-Song; Li, Wen-Bin

2017-01-01

Leptomeningeal metastasis (LM) of high-grade glioma is a highly lethal disease requiring new effective therapeutic measures. For both de novo or relapsed glioma with LM, intrathecal cytarabine chemotherapy is not frequently used for first-line and relapse protocols. We encountered a clinical case demonstrating effective application of cytarabine in high-grade glioma with LM, prompting us to explore the effects of cytarabine on malignant glioma and molecular mechanisms of such effects through in vivo and in vitro experiments. The U87 cell line was selected to represent human glioma for studies. Cell viability was measured by MTT assay, plate colony formation assay, and trypan-blue dye exclusion test. Apoptosis was assessed by flow cytometry. Protein expression levels were detected by Western blot assay and immunohistochemistry. mRNA expression was examined by quantitative real-time reverse transcription polymerase chain reaction. Cytarabine inhibited tumor growth during the in vivo experiment. The present study confirmed that cytarabine inhibits proliferation and promotes apoptosis of U87 cells, and molecular analysis of this effect showed that cytarabine significantly reduces expression of phosphatidylinositol 3-kinase/serine/threonine kinase also known as the protein kinase B/mechanistic target of rapamycin (PI3K/Akt/mTOR) pathway, Ki-67, BCL2, and 4-1BB, and upregulates Bax and cleaved caspase-3. Our findings indicated that intrathecal administration of cytarabine manifests potential in prophylaxis and treatment of malignant glioma with LM. Effective medications for high-grade glioma with LM should contain cytarabine.

Metabolic targeting of lactate efflux by malignant glioma inhibits invasiveness and induces necrosis: an in vivo study.

PubMed

Colen, Chaim B; Shen, Yimin; Ghoddoussi, Farhad; Yu, Pingyang; Francis, Todd B; Koch, Brandon J; Monterey, Michael D; Galloway, Matthew P; Sloan, Andrew E; Mathupala, Saroj P

2011-07-01

Glioblastoma multiforme (GBM) are the most malignant among brain tumors. They are frequently refractory to chemotherapy and radiotherapy with mean patient survival of approximately 6 months, despite surgical intervention. The highly glycolytic nature of glioblastomas describes their propensity to metabolize glucose to lactic acid at an elevated rate. To survive, GBMs efflux lactic acid to the tumor microenvironment through transmembrane transporters denoted monocarboxylate transporters (MCTs). We hypothesized that inhibition of MCT function would impair the glycolytic metabolism and affect both glioma invasiveness and survival. We examined the effect on invasiveness with α-cyano-4-hydroxy-cinnamic acid (ACCA, 4CIN, CHCA), a small-molecule inhibitor of lactate transport, through Matrigel-based and organotypic (brain) slice culture invasive assays using U87-MG and U251-MG glioma cells. We then conducted studies in immunodeficient rats by stereotaxic intracranial implantation of the glioma cells followed by programmed orthotopic application of ACCA through osmotic pumps. Effect on the implanted tumor was monitored by small-animal magnetic resonance imaging. Our assays indicated that glioma invasion was markedly impaired when lactate efflux was inhibited. Convection-enhanced delivery of inhibitor to the tumor bed caused tumor necrosis, with 50% of the animals surviving beyond the experimental end points (3 months after inhibitor exhaustion). Most importantly, control animals did not display any adverse neurologic effects during orthotopic administration of ACCA to brain through programmed delivery. These results indicate the clinical potential of targeting lactate efflux in glioma through delivery of small-molecule inhibitors of MCTs either to the tumor bed or to the postsurgical resection cavity.

Hydroxyapatite nanoparticles inhibit the growth of human glioma cells in vitro and in vivo.

PubMed

Chu, Sheng-Hua; Feng, Dong-Fu; Ma, Yan-Bin; Li, Zhi-Qiang

2012-01-01

Hydroxyapatite nanoparticles (nano-HAPs) have been reported to exhibit antitumor effects on various human cancers, but the effects of nano-HAPs on human glioma cells remain unclear. The aim of this study was to explore the inhibitory effect of nano-HAPs on the growth of human glioma U251 and SHG44 cells in vitro and in vivo. Nano-HAPs could inhibit the growth of U251 and SHG44 cells in a dose- and time-dependent manner, according to methyl thiazoletetrazolium assay and flow cytometry. Treated with 120 mg/L and 240 mg/L nano-HAPs for 48 hours, typical apoptotic morphological changes were noted under Hoechst staining and transmission electron microscopy. The tumor growth of cells was inhibited after the injection in vivo, and the related side effects significantly decreased in the nano-HAP-and-drug combination group. Because of the function of nano-HAPs, the expression of c-Met, SATB1, Ki-67, and bcl-2 protein decreased, and the expression of SLC22A18 and caspase-3 protein decreased noticeably. The findings indicate that nano-HAPs have an evident inhibitory action and induce apoptosis of human glioma cells in vitro and in vivo. In a drug combination, they can significantly reduce the adverse reaction related to the chemotherapeutic drug 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU).

Hydroxyapatite nanoparticles inhibit the growth of human glioma cells in vitro and in vivo

PubMed Central

Chu, Sheng-Hua; Feng, Dong-Fu; Ma, Yan-Bin; Li, Zhi-Qiang

2012-01-01

Hydroxyapatite nanoparticles (nano-HAPs) have been reported to exhibit antitumor effects on various human cancers, but the effects of nano-HAPs on human glioma cells remain unclear. The aim of this study was to explore the inhibitory effect of nano-HAPs on the growth of human glioma U251 and SHG44 cells in vitro and in vivo. Nano-HAPs could inhibit the growth of U251 and SHG44 cells in a dose- and time-dependent manner, according to methyl thiazoletetrazolium assay and flow cytometry. Treated with 120 mg/L and 240 mg/L nano-HAPs for 48 hours, typical apoptotic morphological changes were noted under Hoechst staining and transmission electron microscopy. The tumor growth of cells was inhibited after the injection in vivo, and the related side effects significantly decreased in the nano-HAP-and-drug combination group. Because of the function of nano-HAPs, the expression of c-Met, SATB1, Ki-67, and bcl-2 protein decreased, and the expression of SLC22A18 and caspase-3 protein decreased noticeably. The findings indicate that nano-HAPs have an evident inhibitory action and induce apoptosis of human glioma cells in vitro and in vivo. In a drug combination, they can significantly reduce the adverse reaction related to the chemotherapeutic drug 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). PMID:22888225

Effects of the nitric oxide donor JS-K on the blood-tumor barrier and on orthotopic U87 rat gliomas assessed by MRI

PubMed Central

Weidensteiner, Claudia; Reichardt, Wilfried; Shami, Paul J.; Saavedra, Joseph E.; Keefer, Larry K.; Baumer, Brunhilde; Werres, Anna; Jasinski, Robert; Osterberg, Nadja; Weyerbrock, Astrid

2013-01-01

Nitric oxide (NO) released from NO donors can be cytotoxic in tumor cells and can enhance the transport of drugs into brain tumors by altering blood-tumor barrier permeability. The NO donor JS-K [O2-(2,4-dinitrophenyl) 1-[(4-ethoxycarbonyl)piperazin-1-yl]diazen-1-ium-1,2-diolate] releases NO upon enzymatic activation selectively in cells overexpressing glutathione-S-transferases (GSTs) such as gliomas. Thus, JS-K-dependent NO effects - especially on cell viability and vascular permeability - were investigated in U87 glioma cells in vitro and in an orthotopic U87 xenograft model in vivo by magnetic resonance imaging (MRI). In vitro experiments showed dose-dependent antiproliferative and cytotoxic effects in U87 cells. In addition, treatment of U87 cells with JS-K resulted in a dose-dependent activation of soluble guanylate cyclase and intracellular accumulation of cyclic guanosine monophosphate (cGMP) which was irreversibly inhibited by the selective inhibitor of soluble guanylate cyclase ODQ (1H-[1,2,4]oxadiazolo(4,3a)quinoxaline-1-one). Using dynamic contrast enhanced MRI (DCE-MRI) as a minimally invasive technique, we demonstrated for the first time a significant increase in the DCE-MRI read-out initial area under the concentration curve (iAUC60) indicating an acute increase in blood-tumor barrier permeability after i.v. treatment with JS-K. Repeated MR imaging of animals with intracranial U87 gliomas under treatment with JS-K (3.5 μmol/kg JS-K 3×/week) and of untreated controls on day 12 and 19 after tumor inoculation revealed no significant changes in tumor growth, edema formation or tumor perfusion. Immunohistochemical workup of the brains showed a significant antiproliferative effect of JS-K in the gliomas. Taken together, in vitro and in vivo data suggest that JS-K has antiproliferative effects in U87 gliomas and opens the blood-tumor barrier by activation of the NO/cGMP signaling pathway. This might be a novel approach to facilitate entry of therapeutic drugs into brain tumors. DCE-MRI is a non-invasive, repeatable imaging modality to monitor biological effects of NO donors and other experimental therapeutics in intracranial tumor models. PMID:23370169

Effects of the nitric oxide donor JS-K on the blood-tumor barrier and on orthotopic U87 rat gliomas assessed by MRI.

PubMed

Weidensteiner, Claudia; Reichardt, Wilfried; Shami, Paul J; Saavedra, Joseph E; Keefer, Larry K; Baumer, Brunhilde; Werres, Anna; Jasinski, Robert; Osterberg, Nadja; Weyerbrock, Astrid

2013-04-01

Nitric oxide (NO) released from NO donors can be cytotoxic in tumor cells and can enhance the transport of drugs into brain tumors by altering blood-tumor barrier permeability. The NO donor JS-K [O(2)-(2,4-dinitrophenyl) 1-[(4-ethoxycarbonyl)piperazin-1-yl]diazen-1-ium-1,2-diolate] releases NO upon enzymatic activation selectively in cells overexpressing glutathione-S-transferases (GSTs) such as gliomas. Thus, JS-K-dependent NO effects - especially on cell viability and vascular permeability - were investigated in U87 glioma cells in vitro and in an orthotopic U87 xenograft model in vivo by magnetic resonance imaging (MRI). In vitro experiments showed dose-dependent antiproliferative and cytotoxic effects in U87 cells. In addition, treatment of U87 cells with JS-K resulted in a dose-dependent activation of soluble guanylate cyclase and intracellular accumulation of cyclic guanosine monophosphate (cGMP) which was irreversibly inhibited by the selective inhibitor of soluble guanylate cyclase ODQ (1H-[1,2,4]oxadiazolo(4,3a)quinoxaline-1-one). Using dynamic contrast enhanced MRI (DCE-MRI) as a minimally invasive technique, we demonstrated for the first time a significant increase in the DCE-MRI read-out initial area under the concentration curve (iAUC60) indicating an acute increase in blood-tumor barrier permeability after i.v. treatment with JS-K. Repeated MR imaging of animals with intracranial U87 gliomas under treatment with JS-K (3.5 μmol/kg JS-K 3×/week) and of untreated controls on day 12 and 19 after tumor inoculation revealed no significant changes in tumor growth, edema formation or tumor perfusion. Immunohistochemical workup of the brains showed a significant antiproliferative effect of JS-K in the gliomas. Taken together, in vitro and in vivo data suggest that JS-K has antiproliferative effects in U87 gliomas and opens the blood-tumor barrier by activation of the NO/cGMP signaling pathway. This might be a novel approach to facilitate entry of therapeutic drugs into brain tumors. DCE-MRI is a non-invasive, repeatable imaging modality to monitor biological effects of NO donors and other experimental therapeutics in intracranial tumor models. Copyright © 2013 Elsevier Inc. All rights reserved.

C-type natriuretic peptide-modified lipid vesicles: fabrication and use for the treatment of brain glioma.

PubMed

Wu, Jia-Shuan; Mu, Li-Min; Bu, Ying-Zi; Liu, Lei; Yan, Yan; Hu, Ying-Jie; Bai, Jing; Zhang, Jing-Ying; Lu, Weiyue; Lu, Wan-Liang

2017-06-20

Chemotherapy of brain glioma faces a major obstacle owing to the inability of drug transport across the blood-brain barrier (BBB). Besides, neovasculatures in brain glioma site result in a rapid infiltration, making complete surgical removal virtually impossible. Herein, we reported a novel kind of C-type natriuretic peptide (CNP) modified vinorelbine lipid vesicles for transferring drug across the BBB, and for treating brain glioma along with disrupting neovasculatures. The studies were performed on brain glioma U87-MG cells in vitro and on glioma-bearing nude mice in vivo. The results showed that the CNP-modified vinorelbine lipid vesicles could transport vinorelbine across the BBB, kill the brain glioma, and destroy neovasculatures effectively. The above mechanisms could be associated with the following aspects, namely, long circulation in the blood; drug transport across the BBB via natriuretic peptide receptor B (NPRB)-mediated transcytosis; elimination of brain glioma cells and disruption of neovasculatures by targeting uptake and cytotoxic injury. Besides, CNP-modified vinorelbine lipid vesicles could induce apoptosis of the glioma cells. The mechanisms could be related to the activations of caspase 8, caspase 3, p53, and reactive oxygen species (ROS), and inhibition of survivin. Hence, CNP-modified lipid vesicles could be used as a carrier material for treating brain glioma and disabling glioma neovasculatures.

Local convection-enhanced delivery of an anti-CD40 agonistic monoclonal antibody induces antitumor effects in mouse glioma models

PubMed Central

Shoji, Takuhiro; Saito, Ryuta; Chonan, Masashi; Shibahara, Ichiyo; Sato, Aya; Kanamori, Masayuki; Sonoda, Yukihiko; Kondo, Toru; Ishii, Naoto; Tominaga, Teiji

2016-01-01

Background Glioblastoma is one of the most malignant brain tumors in adults and has a dismal prognosis. In a previous report, we reported that CD40, a TNF-R-related cell surface receptor, and its ligand CD40L were associated with glioma outcomes. Here we attempted to activate CD40 signaling in the tumor and determine if it exerted therapeutic efficacy. Methods CD40 expression was examined in 3 mouse glioma cell lines (GL261, NSCL61, and bRiTs-G3) and 5 human glioma cell lines (U87, U251, U373, T98, and A172). NSCL61 and bRiTs-G3, as glioma stem cells, also expressed the glioma stem cell markers MELK and CD44. In vitro, we demonstrated direct antitumor effects of an anti-CD40 agonistic monoclonal antibody (FGK45) against the cell lines. The efficacy of FGK45 was examined by local convection-enhanced delivery of the monoclonal antibody against each glioma model. Results CD40 was expressed in all mouse and human cell lines tested and was found at the cell membrane of each of the 3 mouse cell lines. FGK45 administration induced significant, direct antitumor effects in vitro. The local delivery of FGK45 significantly prolonged survival compared with controls in the NSCL61 and bRiTs-G3 models, but the effect was not significant in the GL261 model. Increases in apoptosis and CD4+ and CD8+ T cell infiltration were observed in the bRiTs-G3 model after FGK45 treatment. Conclusions Local delivery of FGK45 significantly prolonged survival in glioma stem cell models. Thus, local delivery of this monoclonal antibody is promising for immunotherapy against gliomas. PMID:26917236

miR-489 inhibits proliferation, cell cycle progression and induces apoptosis of glioma cells via targeting SPIN1-mediated PI3K/AKT pathway.

PubMed

Li, Yan; Ma, Xiaolin; Wang, Yanpeng; Li, Guohua

2017-09-01

microRNA-489 (miR-489), a newly identified tumor-related miRNA, functions as an oncogene or tumor suppressor via regulating growth and metastasis of human cancers. But, the clinical significance, biological function and underlying mechanisms of miR-489 in glioma remain rarely known. Here, we showed that the levels of miR-489 in glioma tissues were notably underexpressed compared to corresponding non-tumor tissues. In accordance, the relative levels of miR-489 were decreased in glioma cell lines compared with NHA cells. Kaplan-Meier plots indicated that miR-489 low expressing glioma patients showed a prominent shorter overall survival. In addition, miR-489 overexpression prohibited proliferation and cell cycle progression, and promoted apoptosis in U251 cells. While, miR-489 knockdown showed opposite effects on these cellular processes of U87 cells. In vivo experiments demonstrated that miR-489 restoration reduced the tumor volume and weight of subcutaneous glioma xenografts in nude mice. Notably, Spindlin 1 (SPIN1) was inversely and directly regulated by miR-489 in glioma cells. A negative correlation between the expression of miR-489 and SPIN1 mRNA was confirmed in glioma tissues. Interestingly, miR-489 inversely modulated activation of PI3K/AKT pathway and expression of downstream targets including p-mTOR, Cyclin D1 and BCL-XL. SPIN1 re-expression abolished the effects of miR-489 on U251 cells with enhanced activation of PI3K/AKT pathway and malignant phenotype. Meanwhile, AKT inhibitor MK-2206 blocked activation of PI3K/AKT pathway and resulted in reduced proliferation, cell cycle arrest and increased apoptosis in miR-489 down-regulating U87 cells. Altogether, our data support that miR-489 loss facilitates malignant phenotype of glioma cells probably via SPIN1-mediated PI3K/AKT pathway. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Expression and prognostic role of orphan receptor GPR110 in glioma.

PubMed

Shi, Haiping; Zhang, Shiyuan

2017-09-16

Glioma is the most common type of malignancy in the central nervous system, which has a poor prognosis due to its rapid progression and diffuse invasion. Identification of novel biomarkers for glioma would be invaluable for studying disease mechanism and improving prognosis. Orphan G protein-coupled receptor 110 (GPR110) belongs to the subfamily VI of adhesion GPCR. The knowledge of the ligand, signaling pathway or physiology function of GPR110 is poorly elucidated. The potential role of GPR110 as an oncogene in mouse has been recently reported by mutagenesis screen. However, its expression and role in human glioma hasn't been identified. Here in the current study, we initially explored the RNA and protein expression of GPR110 in patients with glioma. Statistical analysis proved that GPR110 was highly expressed in some patients, which was correlated with advanced disease stages. Furthermore, univariate and multivariate analyses revealed its role as an independent prognostic biomarker for the overall survival of glioma patients. Interestingly, cellular studies showed that overexpression or knockdown of GPR110 in U87 cells didn't affect cell proliferation and migration. However, the invasion of U87 cells was significantly enhanced by GPR110-overepxression, while inhibited by GPR110-knockdown. The detailed mechanisms remain further investigation although our results suggested the possible participation of STAT3 instead of ERK in the GPR110 signaling pathways. Copyright © 2017 Elsevier Inc. All rights reserved.

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

PubMed

Sales, Thais Torquato; Resende, Fernando Francisco Borges; Chaves, Natália Lemos; Titze-De-Almeida, Simoneide Souza; Báo, Sônia Nair; Brettas, Marcella Lemos; Titze-De-Almeida, Ricardo

2016-10-01

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.

Quercetin-induced downregulation of phospholipase D1 inhibits proliferation and invasion in U87 glioma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, Mi Hee; Min, Do Sik, E-mail: minds@pusan.ac.kr

Highlights: {yields} Quercetin, a bioactive flavonoid, suppresses expression and enzymatic activity of phospholipase D1. {yields} Quercetin abolishes NFkB-induced phospholipase D1 expression via inhibition of NFkB transactivation. {yields} Quercetin-induced suppression of phospholipase D1 inhibits invasion and proliferation of human glioma cells. -- Abstract: Phospholipase D (PLD) has been recognized as a regulator of cell proliferation and tumorigenesis, but little is known about the molecules regulating PLD expression. Thus, the identification of small molecules inhibiting PLD expression would be an important advance in PLD-mediated physiology. Quercetin, a ubiquitous bioactive flavonoid, is known to inhibit proliferation and induce apoptosis in a variety ofmore » cancer cells. In the present study, we examined the effect of quercetin on the expression of PLD in U87 glioma cells. Quercetin significantly suppressed the expression of PLD1 at the transcriptional level. Moreover, quercetin abolished the protein expression of PLD1 in a time and dose-dependent manner, as well as inhibited PLD activity. Quercetin suppressed NF{kappa}B-induced PLD1 expression via inhibition of NFkB transactivation. Furthermore, quercetin inhibited activation and invasion of metalloproteinase-2 (MMP-2), a key modulator of glioma cell invasion, induced by phosphatidic acid (PA), a product of PLD activity. Taken together these data demonstrate that quercetin abolishes PLD1 expression and subsequently inhibits invasion and proliferation of glioma cells.« less

Antitumor activity of 7-O-succinyl macrolactin A tromethamine salt in the mouse glioma model.

PubMed

Jin, Jun; Choi, Suh Hee; Lee, Jung Eun; Joo, Jin-Deok; Han, Jung Ho; Park, Su-Young; Kim, Chae-Yong

2017-05-01

Chemoradiotherapy with temozolomide is the current standard treatment option for patients with glioblastoma. However, the majority of patients with glioblastoma survive for <2 years. Therefore, it is necessary to develop more effective therapeutic strategies for the treatment of glioblastoma. 7-O-succinyl macrolactin A tromethamine salt (SMA salt), a macrolactin compound, is known to possess an antiangiogenic activity. The present study investigated the antitumor effects of SMA salt in the treatment of glioblastoma by evaluating in vitro and in vivo antitumor effects of SMA salt in an experimental glioblastoma model. The antitumor effects of the drug on human glioblastoma U87MG, U251MG and LN229 cell lines were assessed using in vitro cell viability, migration and invasion assays. Nude mice with established U87MG glioblastoma were assigned to either the control or SMA salt treatment group. The volume of tumors and the duration of survival were also measured. SMA salt affected cell viability and caused a concentration-dependent inhibition effect on the migration and invasion of glioblastoma cell lines. Animals in the SMA salt treatment group demonstrated a significant reduction in tumor volume and an increase in survival (P<0.05). Treatment with SMA salt presented more cytotoxic effects as well as anti-migration and anti-invasion activity compared with the control group in vitro and in vivo . These results suggest that SMA salt has significant antitumor effects on glioblastoma.

LncRNA-TP53TG1 Participated in the Stress Response Under Glucose Deprivation in Glioma.

PubMed

Chen, Xin; Gao, Yang; Li, Deheng; Cao, Yiqun; Hao, Bin

2017-12-01

Gliomas are the most common brain tumors of the center nervous system. And long non-coding RNAs (lncRNAs) are non-protein coding transcripts, which have been considered as one type of gene expression regulator for cancer development. In this study, we investigated the role of lncRNA-TP53TG1 in response to glucose deprivation in human gliomas. The expression levels of TP53TG1 in glioma tissues and cells were analyzed by qRT-PCR. In addition, the influence of TP53TG1 on glucose metabolism related genes at the mRNA level during both high and low glucose treatment was detected by qRT-PCR. MTT, clonogenicity assays, and flow cytometry were performed to detect the cell proliferation and cell apoptosis. Furthermore, the migration of glioma cells was examined by Transwell assays. The expression of TP53TG1 was significantly higher in human glioma tissues or cell lines compared with normal brain tissue or NHA. Moreover, TP53TG1 and some tumor glucose metabolism related genes, such as GRP78, LDHA, and IDH1 were up-regulated significantly in U87 and LN18 cells under glucose deprivation. In addition, knockdown of TP53TG1 decreased cell proliferation and migration and down-regulated GRP78 and IDH1 expression levels and up-regulated PKM2 levels in U87 cells under glucose deprivation. However, over-expression of TP53TG1 showed the opposite tendency. Moreover, the effects of TP53TG1 were more remarkable in low glucose than that in high glucose. Our data showed that TP53TG1 under glucose deprivation may promote cell proliferation and migration by influencing the expression of glucose metabolism related genes in glioma. J. Cell. Biochem. 118: 4897-4904, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Metabolic Targeting of Lactate Efflux by Malignant Glioma Inhibits Invasiveness and Induces Necrosis: An In Vivo Study1

PubMed Central

Colen, Chaim B; Shen, Yimin; Ghoddoussi, Farhad; Yu, Pingyang; Francis, Todd B; Koch, Brandon J; Monterey, Michael D; Galloway, Matthew P; Sloan, Andrew E; Mathupala, Saroj P

2011-01-01

Glioblastoma multiforme (GBM) are the most malignant among brain tumors. They are frequently refractory to chemotherapy and radiotherapy with mean patient survival of approximately 6 months, despite surgical intervention. The highly glycolytic nature of glioblastomas describes their propensity to metabolize glucose to lactic acid at an elevated rate. To survive, GBMs efflux lactic acid to the tumor microenvironment through transmembrane transporters denoted monocarboxylate transporters (MCTs). We hypothesized that inhibition of MCT function would impair the glycolytic metabolism and affect both glioma invasiveness and survival. We examined the effect on invasiveness with α-cyano-4-hydroxy-cinnamic acid (ACCA, 4CIN, CHCA), a small-molecule inhibitor of lactate transport, through Matrigel-based and organotypic (brain) slice culture invasive assays using U87-MG and U251-MG glioma cells. We then conducted studies in immunodeficient rats by stereotaxic intracranial implantation of the glioma cells followed by programmed orthotopic application of ACCA through osmotic pumps. Effect on the implanted tumor was monitored by small-animal magnetic resonance imaging. Our assays indicated that glioma invasion was markedly impaired when lactate efflux was inhibited. Convection-enhanced delivery of inhibitor to the tumor bed caused tumor necrosis, with 50% of the animals surviving beyond the experimental end points (3 months after inhibitor exhaustion). Most importantly, control animals did not display any adverse neurologic effects during orthotopic administration of ACCA to brain through programmed delivery. These results indicate the clinical potential of targeting lactate efflux in glioma through delivery of small-molecule inhibitors of MCTs either to the tumor bed or to the postsurgical resection cavity. PMID:21750656

Efficacy of ribavirin against malignant glioma cell lines: Follow-up study

PubMed Central

Ochiai, Yushi; Sano, Emiko; Okamoto, Yutaka; Yoshimura, Sodai; Makita, Kotaro; Yamamuro, Shun; Ohta, Takashi; Ogino, Akiyoshi; Tadakuma, Hisashi; Ueda, Takuya; Nakayama, Tomohiro; Hara, Hiroyuki; Yoshino, Atsuo; Katayama, Yoichi

2018-01-01

Ribavirin, a nucleic acid analog, has been employed as an antiviral agent against RNA and DNA viruses and has become the standard agent used for chronic hepatitis C in combination with interferon-α2a. Furthermore, the potential antitumor efficacy of ribavirin has attracted increasing interest. Recently, we demonstrated a dose-dependent antitumor effect of ribavirin for seven types of malignant glioma cell lines. However, the mechanism underlying the antitumor effect of ribavirin has not yet been fully elucidated. Therefore, the main aim of the present study was to provide further relevant data using two types of malignant glioma cell lines (U-87MG and U-138MG) with different expression of MGMT. Dotted accumulations of γH2AX were found in the nuclei and increased levels of ATM and phosphorylated ATM protein expression were also observed following ribavirin treatment (10 µM of ribavirin, clinical relevant concentration) in both the malignant glioma cells, indicating double-strand breaks as one possible mechanism underlying the antitumor effect of ribavirin. In addition, based on assessements using FACS, ribavirin treatment tended to increase the G0/G1 phase, with a time-lapse, indicating the induction of G0/G1-phase arrest. Furthermore, an increased phosphorylated p53 and p21 protein expression was confirmed in both glioma cells. Additionally, analysis by FACS indicated that apoptosis was induced following ribavirin treatment and caspase cascade, downstream of the p53 pathway, which indicated the activation of both exogenous and endogenous apoptosis in both malignant glioma cell lines. These findings may provide an experimental basis for the clinical treatment of glioblastomas with ribavirin. PMID:29251333

The neural stem cell fate determinant TLX promotes tumorigenesis and genesis of cells resembling glioma stem cells.

PubMed

Park, Hyo-Jung; Kim, Jun-Kyum; Jeon, Hye-Min; Oh, Se-Yeong; Kim, Sung-Hak; Nam, Do-Hyun; Kim, Hyunggee

2010-11-01

A growing body of evidence indicates that deregulation of stem cell fate determinants is a hallmark of many types of malignancies. The neural stem cell fate determinant TLX plays a pivotal role in neurogenesis in the adult brain by maintaining neural stem cells. Here, we report a tumorigenic role of TLX in brain tumor initiation and progression. Increased TLX expression was observed in a number of glioma cells and glioma stem cells, and correlated with poor survival of patients with gliomas. Ectopic expression of TLX in the U87MG glioma cell line and Ink4a/Arf-deficient mouse astrocytes (Ink4a/Arf(-/-) astrocytes) induced cell proliferation with a concomitant increase in cyclin D expression, and accelerated foci formation in soft agar and tumor formation in in vivo transplantation assays. Furthermore, overexpression of TLX in Ink4a/Arf(-/-) astrocytes inhibited cell migration and invasion and promoted neurosphere formation and Nestin expression, which are hallmark characteristics of glioma stem cells, under stem cell culture conditions. Our results indicate that TLX is involved in glioma stem cell genesis and represents a potential therapeutic target for this type of malignancy.

The novel phloroglucinol derivative BFP induces apoptosis of glioma cancer through reactive oxygen species and endoplasmic reticulum stress pathways.

PubMed

Lu, Dah-Yuu; Chang, Chih-Shiang; Yeh, Wei-Lan; Tang, Chih-Hsin; Cheung, Chi-Wai; Leung, Yuk-Man; Liu, Ju-Fang; Wong, Kar-Lok

2012-09-15

Prenyl-phloroglucinol derivatives from hop plants have been shown to have anticancer activities. This study is the first to investigate the anticancer effects of the new phloroglucinol derivative (2,4-bis(4-fluorophenylacetyl)phloroglucinol; BFP). BFP induced cell death and anti-proliferation in three glioma, U251, U87 and C6 cells, but not in primary human astrocytes. BFP-induced concentration-dependently cell death in glioma cells was determined by MTT and SRB assay. Moreover, BFP-induced apoptotic cell death in glioma cells was measured by Hochest 33258 staining and fluorescence-activated cell sorter (FACS) of propidine iodine (PI) analysis. Treatment of U251 human glioma cells with BFP was also found to induce reactive oxygen species (ROS) generation, which was detected by a fluorescence dye used FACS analysis. Treatment of BFP also increased a number of signature endoplasmic reticulum (ER) stress markers glucose-regulated protein (GRP)-78, GRP-94, IRE1, phosphorylation of eukaryotic initiation factor-2α (eIF-2α) and up-regulation of CAAT/enhancer-binding protein homologous protein (CHOP). Moreover, treatment of BFP also increased the down-stream caspase activation, such as pro-caspase-7 and pro-caspase-12 degradation, suggesting the induction of ER stress. Furthermore, BFP also induced caspase-9 and caspase-3 activation as well as up-regulation of cleaved PARP expression. Treatment of antioxidants, or pre-transfection of cells with GRP78 or CHOP siRNA reduced BFP-mediated apoptotic-related protein expression. Taken together, the present study provides evidences to support that ROS generation, GRP78 and CHOP activation are mediating the BFP-induced human glioma cell apoptosis. Copyright © 2012 Elsevier GmbH. All rights reserved.

Osteopontin and splice variant expression level in human malignant glioma: radiobiologic effects and prognosis after radiotherapy.

PubMed

Güttler, Antje; Giebler, Maria; Cuno, Peter; Wichmann, Henri; Keßler, Jacqueline; Ostheimer, Christian; Söling, Ariane; Strauss, Christian; Illert, Jörg; Kappler, Matthias; Vordermark, Dirk; Bache, Matthias

2013-09-01

We investigated the role of the hypoxia-associated secreted glycoprotein osteopontin (OPN) in the response of malignant glioma to radiotherapy by characterizing OPN and its splice variants in vitro and in patient material. The effect of siRNA knockdown of OPN splice variants on cellular and radiobiologic behavior was analyzed in U251MG cells using OpnS siRNA (inhibition of all OPN splice variants) and OpnAC siRNA (knockdown only of OPNa and OPNc). OPN and splice variant mRNA levels were quantified in archival material of 41 glioblastoma tumor samples. Plasma OPN was prospectively measured in 33 malignant glioma patients. Inhibition of OPNa and OPNc (OpnAC) reduced clonogenic survival in U251MG cells but did not affect proliferation, migration or apoptosis. Knockdown of all OPN splice variants (OpnS) resulted in an even stronger inhibition of clonogenic survival, while cell proliferation and migration were reduced and rate of apoptosis was increased. Additional irradiation had additive effects with both siRNAs. Plasma OPN increased continuously in malignant glioma patients and was associated with poor survival. OPNb is partially able to compensate the effects of OPNa and OPNc knockdown in U251MG cells. High OPN plasma levels at the end of radiotherapy are associated with poor survival. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

L1 stimulation of human glioma cell motility correlates with FAK activation

PubMed Central

Yang, Muhua; Li, Yupei; Chilukuri, Kalyani; Brady, Owen A.; Boulos, Magdy I.; Kappes, John C.

2011-01-01

The neural adhesion/recognition protein L1 (L1CAM; CD171) has been shown or implicated to function in stimulation of cell motility in several cancer types, including high-grade gliomas. Our previous work demonstrated the expression and function of L1 protein in stimulation of cell motility in rat glioma cells. However, the mechanism of this stimulation is still unclear. This study further investigated the function of L1 and L1 proteolysis in human glioblastoma multiforme (GBM) cell migration and invasion, as well as the mechanism of this stimulation. L1 mRNA was found to be present in human T98G GBM cell line but not in U-118 MG grade III human glioma cell line. L1 protein expression, proteolysis, and release were found in T98G cells and human surgical GBM cells by Western blotting. Exosome-like vesicles released by T98G cells were purified and contained full-length L1. In a scratch assay, T98G cells that migrated into the denuded scratch area exhibited upregulation of ADAM10 protease expression coincident with loss of surface L1. GBM surgical specimen cells exhibited a similar loss of cell surface L1 when xenografted into the chick embryo brain. When lentivirally introduced shRNA was used to attenuate L1 expression, such T98G/shL1 cells exhibited significantly decreased cell motility by time lapse microscopy in our quantitative Super Scratch assay. These cells also showed a decrease in FAK activity and exhibited increased focal complexes. L1 binding integrins which activate FAK were found in T98G and U-118 MG cells. Addition of L1 ectodomain-containing media (1) rescued the decreased cell motility of T98G/shL1 cells and (2) increased cell motility of U-118 MG cells but (3) did not further increase T98G cell motility. Injection of L1-attenuated T98G/shL1 cells into embryonic chick brains resulted in the absence of detectable invasion compared to control cells which invaded brain tissue. These studies support a mechanism where glioma cells at the edge of a cell mass upregulate ADAM10 to proteolyze surface L1 and the resultant ectodomain increases human glioma cell migration and invasion by binding to integrin receptors, activating FAK, and increasing turnover of focal complexes. PMID:21373966

L1 stimulation of human glioma cell motility correlates with FAK activation.

PubMed

Yang, Muhua; Li, Yupei; Chilukuri, Kalyani; Brady, Owen A; Boulos, Magdy I; Kappes, John C; Galileo, Deni S

2011-10-01

The neural adhesion/recognition protein L1 (L1CAM; CD171) has been shown or implicated to function in stimulation of cell motility in several cancer types, including high-grade gliomas. Our previous work demonstrated the expression and function of L1 protein in stimulation of cell motility in rat glioma cells. However, the mechanism of this stimulation is still unclear. This study further investigated the function of L1 and L1 proteolysis in human glioblastoma multiforme (GBM) cell migration and invasion, as well as the mechanism of this stimulation. L1 mRNA was found to be present in human T98G GBM cell line but not in U-118 MG grade III human glioma cell line. L1 protein expression, proteolysis, and release were found in T98G cells and human surgical GBM cells by Western blotting. Exosome-like vesicles released by T98G cells were purified and contained full-length L1. In a scratch assay, T98G cells that migrated into the denuded scratch area exhibited upregulation of ADAM10 protease expression coincident with loss of surface L1. GBM surgical specimen cells exhibited a similar loss of cell surface L1 when xenografted into the chick embryo brain. When lentivirally introduced shRNA was used to attenuate L1 expression, such T98G/shL1 cells exhibited significantly decreased cell motility by time lapse microscopy in our quantitative Super Scratch assay. These cells also showed a decrease in FAK activity and exhibited increased focal complexes. L1 binding integrins which activate FAK were found in T98G and U-118 MG cells. Addition of L1 ectodomain-containing media (1) rescued the decreased cell motility of T98G/shL1 cells and (2) increased cell motility of U-118 MG cells but (3) did not further increase T98G cell motility. Injection of L1-attenuated T98G/shL1 cells into embryonic chick brains resulted in the absence of detectable invasion compared to control cells which invaded brain tissue. These studies support a mechanism where glioma cells at the edge of a cell mass upregulate ADAM10 to proteolyze surface L1 and the resultant ectodomain increases human glioma cell migration and invasion by binding to integrin receptors, activating FAK, and increasing turnover of focal complexes.

Preclinical Biosafety Evaluation of Genetically Modified Human Adipose Tissue-Derived Mesenchymal Stem Cells for Clinical Applications to Brainstem Glioma.

PubMed

Choi, Seung Ah; Yun, Jun-Won; Joo, Kyeung Min; Lee, Ji Yeoun; Kwak, Pil Ae; Lee, Young Eun; You, Ji-Ran; Kwon, Euna; Kim, Woo Ho; Wang, Kyu-Chang; Phi, Ji Hoon; Kang, Byeong-Cheol; Kim, Seung-Ki

2016-06-15

Stem-cell based gene therapy is a promising novel therapeutic approach for inoperable invasive tumors, including brainstem glioma. Previously, we demonstrated the therapeutic potential of human adipose tissue-derived mesenchymal stem cells (hAT-MSC) genetically engineered to express a secreted form of tumor necrosis factor-related apoptosis-inducing ligand (sTRAIL) against brainstem glioma. However, safety concerns should be comprehensively investigated before clinical applications of hAT-MSC.sTRAIL. At first, we injected stereotactically low (1.2 × 10(5) cells/18 μL), medium (2.4 × 10(5)/18 μL), or high dose (3.6 × 10(5)/18 μL) of hAT-MSC.sTRAIL into the brainstems of immunodeficient mice reflecting the plan of the future clinical trial. Local toxicity, systemic toxicity, secondary tumor formation, and biodistribution of hAT-MSC.sTRAIL were investigated. Next, presence of hAT-MSC.sTRAIL was confirmed in the brain and major organs at 4, 9, and 14 weeks in brainstem glioma-bearing mice. In the 15-week subchronic toxicity test, no serious adverse events in terms of body weight, food consumption, clinical symptom, urinalysis, hematology, clinical chemistry, organ weight, and histopathology were observed. In the 26-week tumorigenicity test, hAT-MSC.sTRAIL made no detectable tumors, whereas positive control U-87 MG cells made huge tumors in the brainstem. No remaining hAT-MSC.sTRAIL was observed in any organs examined, including the brainstem at 15 or 26 weeks. In brainstem glioma-bearing mice, injected hAT-MSC.sTRAIL was observed, but gradually decreased over time in the brain. The mRNA of human specific GAPDH and TRAIL was not detected in all major organs. These results indicate that the hAT-MSC.sTRAIL could be applicable to the future clinical trials in terms of biosafety.

Zinc substituted ferrite nanoparticles with Zn0.9Fe2.1O4 formula used as heating agents for in vitro hyperthermia assay on glioma cells

NASA Astrophysics Data System (ADS)

Hanini, Amel; Lartigue, Lenaic; Gavard, Julie; Kacem, Kamel; Wilhelm, Claire; Gazeau, Florence; Chau, François; Ammar, Souad

2016-10-01

In this paper we investigate the ability of zinc rich ferrite nanoparticles to induce hyperthermia on cancer cells using an alternating magnetic field (AMF). First, we synthesized ferrites and then we analyzed their physico-chemical properties by transmission electron microscopy, X-ray diffraction and magnetic and magnetocalorimetric measurements. We found that the polyol-made magnetically diluted particles are of 11 nm in size. They are superparamagnetic at body temperature (310 K) with a low but non-negligible magnetization. Interestingly, as nano-ferrimagnets they exhibit a Curie temperature of 366 K, close to the therapeutic temperature range. Their effect on human healthy endothelial (HUVEC) and malignant glioma (U87-MG) cells was also evaluated using MTT viability assays. Incubated with the two cell lines, at doses ≤100 μg mL-1 and contact times ≤4 h, they exhibit a mild in vitro toxicity. In these same operating biological conditions and coupled to AMF (700 kHz and 34.4 Oe) for 1 h, they rapidly induce a net temperature increase. In the case of tumor cells it reaches 4 K, making the produced particles particularly promising for self-regulated magnetically-induced heating in local glioma therapy.

Local convection-enhanced delivery of an anti-CD40 agonistic monoclonal antibody induces antitumor effects in mouse glioma models.

PubMed

Shoji, Takuhiro; Saito, Ryuta; Chonan, Masashi; Shibahara, Ichiyo; Sato, Aya; Kanamori, Masayuki; Sonoda, Yukihiko; Kondo, Toru; Ishii, Naoto; Tominaga, Teiji

2016-08-01

Glioblastoma is one of the most malignant brain tumors in adults and has a dismal prognosis. In a previous report, we reported that CD40, a TNF-R-related cell surface receptor, and its ligand CD40L were associated with glioma outcomes. Here we attempted to activate CD40 signaling in the tumor and determine if it exerted therapeutic efficacy. CD40 expression was examined in 3 mouse glioma cell lines (GL261, NSCL61, and bRiTs-G3) and 5 human glioma cell lines (U87, U251, U373, T98, and A172). NSCL61 and bRiTs-G3, as glioma stem cells, also expressed the glioma stem cell markers MELK and CD44. In vitro, we demonstrated direct antitumor effects of an anti-CD40 agonistic monoclonal antibody (FGK45) against the cell lines. The efficacy of FGK45 was examined by local convection-enhanced delivery of the monoclonal antibody against each glioma model. CD40 was expressed in all mouse and human cell lines tested and was found at the cell membrane of each of the 3 mouse cell lines. FGK45 administration induced significant, direct antitumor effects in vitro. The local delivery of FGK45 significantly prolonged survival compared with controls in the NSCL61 and bRiTs-G3 models, but the effect was not significant in the GL261 model. Increases in apoptosis and CD4(+) and CD8(+) T cell infiltration were observed in the bRiTs-G3 model after FGK45 treatment. Local delivery of FGK45 significantly prolonged survival in glioma stem cell models. Thus, local delivery of this monoclonal antibody is promising for immunotherapy against gliomas. © The Author(s) 2016. Published by Oxford University Press on behalf of the Society for Neuro-Oncology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Convection enhanced delivery of panobinostat (LBH589)-loaded pluronic nano-micelles prolongs survival in the F98 rat glioma model

PubMed Central

Singleton, WG; Collins, AM; Bienemann, AS; Killick-Cole, CL; Haynes, HR; Asby, DJ; Butts, CP; Wyatt, MJ; Barua, NU; Gill, SS

2017-01-01

Background The pan-histone deacetylase inhibitor panobinostat is a potential therapy for malignant glioma, but it is water insoluble and does not cross the blood–brain barrier when administered systemically. In this article, we describe the in vitro and in vivo efficacy of a novel water-soluble nano-micellar formulation of panobinostat designed for administration by convection enhanced delivery (CED). Materials and methods The in vitro efficacy of panobinostat-loaded nano-micelles against rat F98, human U87-MG and M059K glioma cells and against patient-derived glioma stem cells was measured using a cell viability assay. Nano-micelle distribution in rat brain was analyzed following acute CED using rhodamine-labeled nano-micelles, and toxicity was assayed using immunofluorescent microscopy and synaptophysin enzyme-linked immunosorbent assay. We compared the survival of the bioluminescent syngenic F98/Fischer344 rat glioblastoma model treated by acute CED of panobinostat-loaded nano-micelles with that of untreated and vehicle-only-treated controls. Results Nano-micellar panobinostat is cytotoxic to rat and human glioma cells in vitro in a dose-dependent manner following short-time exposure to drug. Fluorescent rhodamine-labelled nano-micelles distribute with a volume of infusion/volume of distribution (Vi/Vd) ratio of four and five respectively after administration by CED. Administration was not associated with any toxicity when compared to controls. CED of panobinostat-loaded nano-micelles was associated with significantly improved survival when compared to controls (n=8 per group; log-rank test, P<0.001). One hundred percent of treated animals survived the 60-day experimental period and had tumour response on post-mortem histological examination. Conclusion CED of nano-micellar panobinostat represents a potential novel therapeutic option for malignant glioma and warrants translation into the clinic. PMID:28260886

The nitric oxide donor JS-K sensitizes U87 glioma cells to repetitive irradiation.

PubMed

Heckler, Max; Osterberg, Nadja; Guenzle, Jessica; Thiede-Stan, Nina Kristin; Reichardt, Wilfried; Weidensteiner, Claudia; Saavedra, Joseph E; Weyerbrock, Astrid

2017-06-01

As a potent radiosensitizer nitric oxide (NO) may be a putative adjuvant in the treatment of malignant gliomas which are known for their radio- and chemoresistance. The NO donor prodrug JS-K (O2-(2.4-dinitrophenyl) 1-[(4-ethoxycarbonyl) piperazin-1-yl] diazen-1-ium-1,2-diolate) allows cell-type specific intracellular NO release via enzymatic activation by glutathione-S-transferases overexpressed in glioblastoma multiforme. The cytotoxic and radiosensitizing efficacy of JS-K was assessed in U87 glioma cells in vitro focusing on cell proliferation, induction of DNA damage, and cell death. In vivo efficacy of JS-K and repetitive irradiation were investigated in an orthotopic U87 xenograft model in mice. For the first time, we could show that JS-K acts as a potent cytotoxic and radiosensitizing agent in U87 cells in vitro. This dose- and time-dependent effect is due to an enhanced induction of DNA double-strand breaks leading to mitotic catastrophe as the dominant form of cell death. However, this potent cytotoxic and radiosensitizing effect could not be confirmed in an intracranial U87 xenograft model, possibly due to insufficient delivery into the brain. Although NO donor treatment was well tolerated, neither a retardation of tumor growth nor an extended survival could be observed after JS-K and/or radiotherapy.

Bioconjugated PLGA-4-arm-PEG branched polymeric nanoparticles as novel tumor targeting carriers

NASA Astrophysics Data System (ADS)

Ding, Hong; Yong, Ken-Tye; Roy, Indrajit; Hu, Rui; Wu, Fang; Zhao, Lingling; Law, Wing-Cheung; Zhao, Weiwei; Ji, Wei; Liu, Liwei; Bergey, Earl J.; Prasad, Paras N.

2011-04-01

In this study, we have developed a novel carrier, micelle-type bioconjugated PLGA-4-arm-PEG branched polymeric nanoparticles (NPs), for the detection and treatment of pancreatic cancer. These NPs contained 4-arm-PEG as corona, and PLGA as core, the particle surface was conjugated with cyclo(arginine-glycine-aspartate) (cRGD) as ligand for in vivo tumor targeting. The hydrodynamic size of the NPs was determined to be 150-180 nm and the critical micellar concentration (CMC) was estimated to be 10.5 mg l - 1. Our in vitro study shows that these NPs by themselves had negligible cytotoxicity to human pancreatic cancer (Panc-1) and human glioblastoma (U87) cell lines. Near infrared (NIR) microscopy and flow cytometry demonstrated that the cRGD conjugated PLGA-4-arm-PEG polymeric NPs were taken up more efficiently by U87MG glioma cells, over-expressing the αvβ3 integrin, when compared with the non-targeted NPs. Whole body imaging showed that the cRGD conjugated PLGA-4-arm-PEG branched polymeric NPs had the highest accumulation in the pancreatic tumor site of mice at 48 h post-injection. Physical, hematological, and pathological assays indicated low in vivo toxicity of this NP formulation. These studies on the ability of these bioconjugated PLGA-4-arm-PEG polymeric NPs suggest that the prepared polymeric NPs may serve as a promising platform for detection and targeted drug delivery for pancreatic cancer.

Convection-enhanced delivery of SN-38-loaded polymeric micelles (NK012) enables consistent distribution of SN-38 and is effective against rodent intracranial brain tumor models.

PubMed

Zhang, Rong; Saito, Ryuta; Mano, Yui; Sumiyoshi, Akira; Kanamori, Masayuki; Sonoda, Yukihiko; Kawashima, Ryuta; Tominaga, Teiji

2016-10-01

Convection-enhanced delivery (CED) of therapeutic agents is a promising local delivery technique that has been extensively studied as a treatment for CNS diseases over the last two decades. One continuing challenge of CED is accurate and consistent delivery of the agents to the target. The present study focused on a new type of therapeutic agent, NK012, a novel SN-38-loaded polymeric micelle. Local delivery profiles of NK012 and SN-38 were studied using rodent brain and intracranial rodent brain tumor models. First, the cytotoxicity of NK012 against glioma cell lines was determined in vitro. Proliferations of glioma cells were significantly reduced after exposure to NK012. Then, the distribution and local toxicity after CED delivery of NK012 and SN-38 were evaluated in vivo. Volume of distribution of NK012 after CED was much larger than that of SN-38. Histological examination revealed minimum brain tissue damage in rat brains after delivery of 40 µg NK012 but severe damage with SN-38 at the same dose. Subsequently, the efficacy of NK012 delivered via CED was tested in 9L and U87MG rodent orthotopic brain tumor models. CED of NK012 displayed excellent efficacy in the 9L and U87MG orthotopic brain tumor models. Furthermore, NK012 and gadolinium diamide were co-delivered via CED to monitor the NK012 distribution using MRI. Volume of NK012 distribution evaluated by histology and MRI showed excellent agreement. CED of NK012 represents an effective treatment option for malignant gliomas. MRI-guided CED of NK012 has potential for clinical application.

Cross-link regulation of precursor N-cadherin and FGFR1 by GDNF increases U251MG cell viability.

PubMed

Tang, Chuan-Xi; Gu, Yan-Xia; Liu, Xin-Feng; Tong, Shu-Yan; Ayanlaja, Abiola A; Gao, Yue; Ji, Guang-Quan; Xiong, Ye; Huang, Lin-Yan; Gao, Dian-Shuai

2018-07-01

Glial cell line-derived neurotrophic factor (GDNF) is considered to be involved in the development of glioma. However, uncovering the underlying mechanism of the proliferation of glioma cells is a challenging work in progress. We have identified the binding of the precursor of N-cadherin (proN-cadherin) and GDNF on the cell membrane in previous studies. In the present study, we observed increased U251 Malignant glioma (U251MG) cell viability by exogenous GDNF (50 ng/ml). We also confirmed that the high expression of the proN-cadherin was stimulated by exogenous GDNF. Concurrently, we affirmed that lower expression of proN-cadherin correlated with reduced glioma cell viability. Additionally, we observed glioma cell U251MG viability as the phosphorylation level of FGFR1 at Y653 and Y654 was increased after exogenous GDNF treatment, which led to increased interaction between proN-cadherin and FGFR1 (pY653+Y654). Our experiments presented a new mechanism adopted by GDNF supporting glioma development and indicated a possible therapeutic potential via the inhibition of proN-cadherin/FGFR1 interaction.

PAMAM-RGD Conjugates Enhance siRNA Delivery Through a Multicellular Spheroid Model of Malignant Glioma

PubMed Central

Waite, Carolyn L.; Roth, Charles M.

2011-01-01

Generation 5 poly(amidoamine) (PAMAM) dendrimers were modified by the addition of cyclic RGD targeting peptides and were evaluated for their ability to associate with siRNA and mediate siRNA delivery to U87 malignant glioma cells. PAMAM-RGD conjugates were able to complex with siRNA to form complexes of approximately 200 nm in size. Modest siRNA delivery was observed in U87 cells using either PAMAM or PAMAM-RGD conjugates. PAMAM-RGD conjugates prevented the adhesion of U87 cells to fibrinogen coated plates, in a manner that depends on the number of RGD ligands per dendrimer. The delivery of siRNA through three-dimensional multicellular spheroids of U87 cells was enhanced using PAMAM-RGD conjugates compared to the native PAMAM dendrimers, presumably by interfering with integrin-ECM contacts present in a three-dimensional tumor model. PMID:19775120

IDH1(R132H) mutation increases U87 glioma cell sensitivity to radiation therapy in hypoxia.

PubMed

Wang, Xiao-Wei; Labussière, Marianne; Valable, Samuel; Pérès, Elodie A; Guillamo, Jean-Sébastien; Bernaudin, Myriam; Sanson, Marc

2014-01-01

IDH1 codon 132 mutation (mostly Arg132His) is frequently found in gliomas and is associated with longer survival. However, it is still unclear whether IDH1 mutation renders the cell more vulnerable to current treatment, radio- and chemotherapy. We transduced U87 with wild type IDH1 or IDH1 (R132H) expressing lentivirus and analyzed the radiosensitivity (dose ranging 0 to 10 Gy) under normoxia (20% O2) and moderate hypoxia (1% O2). We observed that IDH1 (R132H) U87 cells grow faster in hypoxia and were more sensitive to radiotherapy (in terms of cell mortality and colony formation assay) compared to nontransduced U87 and IDH1 (wt) cells. This effect was not observed in normoxia. These data suggest that IDH1 (R132H) mutation increases radiosensitivity in mild hypoxic conditions.

Essential role of TRPC6 channels in G2/M phase transition and development of human glioma.

PubMed

Ding, Xia; He, Zhuohao; Zhou, Kechun; Cheng, Ju; Yao, Hailan; Lu, Dongliang; Cai, Rong; Jin, Yening; Dong, Bin; Xu, Yinghui; Wang, Yizheng

2010-07-21

Patients with glioblastoma multiforme, the most aggressive form of glioma, have a median survival of approximately 12 months. Calcium (Ca(2+)) signaling plays an important role in cell proliferation, and some members of the Ca(2+)-permeable transient receptor potential canonical (TRPC) family of channel proteins have demonstrated a role in the proliferation of many types of cancer cells. In this study, we investigated the role of TRPC6 in cell cycle progression and in the development of human glioma. TRPC6 protein and mRNA expression were assessed in glioma (n = 33) and normal (n = 17) brain tissues from patients and in human glioma cell lines U251, U87, and T98G. Activation of TRPC6 channels was tested by platelet-derived growth factor-induced Ca(2+) imaging. The effect of inhibiting TRPC6 activity or expression using the dominant-negative mutant TRPC6 (DNC6) or RNA interference, respectively, was tested on cell growth, cell cycle progression, radiosensitization of glioma cells, and development of xenografted human gliomas in a mouse model. The green fluorescent protein (GFP) and wild-type TRPC6 (WTC6) were used as controls. Survival of mice bearing xenografted tumors in the GFP, DNC6, and WTC6 groups (n = 13, 15, and 13, respectively) was compared using Kaplan-Meier analysis. All statistical tests were two-sided. Functional TRPC6 was overexpressed in human glioma cells. Inhibition of TRPC6 activity or expression attenuated the increase in intracellular Ca(2+) by platelet-derived growth factor, suppressed cell growth and clonogenic ability, induced cell cycle arrest at the G2/M phase, and enhanced the antiproliferative effect of ionizing radiation. Cyclin-dependent kinase 1 activation and cell division cycle 25 homolog C expression regulated the cell cycle arrest. Inhibition of TRPC6 activity also reduced tumor volume in a subcutaneous mouse model of xenografted human tumors (P = .014 vs GFP; P < .001 vs WTC6) and increased mean survival in mice in an intracranial model (P < .001 vs GFP or WTC6). In this preclinical model, TRPC6 channels were essential for glioma development via regulation of G2/M phase transition. This study suggests that TRPC6 might be a new target for therapeutic intervention of human glioma.

T-lymphokine-activated killer cell-originated protein kinase (TOPK) as a prognostic factor and a potential therapeutic target in glioma

PubMed Central

Duan, Qiuhong; Yuan, Ping; Xue, Peipei; Lu, Hui; Yan, Meng; Guo, Dongsheng; Xu, Sanpeng; Zhang, Xiaohui; Lin, Xuan; Wang, Yong; Dogan, Soner; Zhang, Jianmin; Zhu, Feng; Ke, Changshu; Liu, Lin

2018-01-01

TOPK is overexpressed in various types of cancer and associated with poor outcomes in different types of cancer. In this study, we first found that the expression of T-lymphokine-activated killer cell-originated protein kinase (TOPK) was significantly higher in Grade III or Grade IV than that in Grade II in glioma (P = 0.007 and P < 0.001, respectively). Expression of TOPK was positively correlated with Ki67 (P < 0.001). Knockdown of TOPK significantly inhibited cell growth, colony formation and increased sensitivities to temozolomide (TMZ) in U-87 MG or U-251 cells, while TOPK overexpression promoted cell growth and colony formation in Hs 683 or A-172 cells. Glioma patients expressing high levels of TOPK have poor survival compared with those expressing low levels of TOPK in high-grade or low-grade gliomas (hazard ratio = 0.2995; 95% CI, 0.1262 to 0.7108; P = 0.0063 and hazard ratio = 0.1509; 95% CI, 0.05928 to 0.3842; P < 0.0001, respectively). The level of TOPK was low in TMZ-sensitive patients compared with TMZ-resistant patients (P = 0.0056). In TMZ-resistant population, patients expressing high TOPK have two months’ shorter survival time than those expressing low TOPK. Our findings demonstrated that TOPK might represent as a promising prognostic and predictive factor and potential therapeutic target for glioma. PMID:29487691

The combination of cannabidiol and Δ9-tetrahydrocannabinol enhances the anticancer effects of radiation in an orthotopic murine glioma model.

PubMed

Scott, Katherine A; Dalgleish, Angus G; Liu, Wai M

2014-12-01

High-grade glioma is one of the most aggressive cancers in adult humans and long-term survival rates are very low as standard treatments for glioma remain largely unsuccessful. Cannabinoids have been shown to specifically inhibit glioma growth as well as neutralize oncogenic processes such as angiogenesis. In an attempt to improve treatment outcome, we have investigated the effect of Δ(9)-tetrahydrocannabinol (THC) and cannabidiol (CBD) both alone and in combination with radiotherapy in a number of glioma cell lines (T98G, U87MG, and GL261). Cannabinoids were used in two forms, pure (P) and as a botanical drug substance (BDS). Results demonstrated a duration- and dose-dependent reduction in cell viability with each cannabinoid and suggested that THC-BDS was more efficacious than THC-P, whereas, conversely, CBD-P was more efficacious than CBD-BDS. Median effect analysis revealed all combinations to be hyperadditive [T98G 48-hour combination index (CI) at FU50, 0.77-1.09]. Similarly, pretreating cells with THC-P and CBD-P together for 4 hours before irradiation increased their radiosensitivity when compared with pretreating with either of the cannabinoids individually. The increase in radiosensitivity was associated with an increase in markers of autophagy and apoptosis. These in vitro results were recapitulated in an orthotopic murine model for glioma, which showed dramatic reductions in tumor volumes when both cannabinoids were used with irradiation (day 21: 5.5 ± 2.2 mm(3) vs. 48.7 ± 24.9 mm(3) in the control group; P < 0.01). Taken together, our data highlight the possibility that these cannabinoids can prime glioma cells to respond better to ionizing radiation, and suggest a potential clinical benefit for glioma patients by using these two treatment modalities. ©2014 American Association for Cancer Research.

Synergistic inhibition of glioma cell proliferation by Withaferin A and tumor treating fields.

PubMed

Chang, Edwin; Pohling, Christoph; Beygui, Nooshin; Patel, Chirag B; Rosenberg, Jarrett; Ha, Dong Ho; Gambhir, Sanjiv S

2017-09-01

Glioblastoma (GBM) is the most aggressive and lethal form of brain cancer. Standard therapies are non-specific and often of limited effectiveness; thus, efforts are underway to uncover novel, unorthodox therapies against GBM. In previous studies, we investigated Withaferin A, a steroidal lactone from Ayurvedic medicine that inhibits proliferation in cancers including GBM. Another novel approach, tumor treating fields (TTFields), is thought to disrupt mitotic spindle formation and stymie proliferation of actively dividing cells. We hypothesized that combining TTFields with Withaferin A would synergistically inhibit proliferation in glioblastoma. Human glioblastoma cells (GBM2, GBM39, U87-MG) and human breast adenocarcinoma cells (MDA-MB-231) were isolated from primary tumors. The glioma cell lines were genetically engineered to express firefly luciferase. Proliferative potential was assessed either by bioluminescence imaging or cell counting via hemocytometer. TTFields (4 V/cm) significantly inhibited growth of the four cancer cell lines tested (n = 3 experiments per time point, four measurements per sample, p < 0.02 at least; 2-way ANOVA, control vs. treatment). The combination of Withaferin A (10-100 nM) with TTFields significantly inhibited the growth of the glioma cells to a degree beyond that of Withaferin A or TTFields alone. The interaction of the Withaferin A and TTFields on glioma cells was found to be synergistic in nature (p < 0.01, n = 3 experiments). These findings were validated by both bioluminescence and hemocytometric measurements. The combination of Withaferin A with TTFields represents a novel approach to treat GBM in a manner that is likely better than either treatment alone and that is synergistic.

Stimulation of glioma cell motility by expression, proteolysis, and release of the L1 neural cell recognition molecule.

PubMed

Yang, Muhua; Adla, Shalini; Temburni, Murali K; Patel, Vivek P; Lagow, Errin L; Brady, Owen A; Tian, Jing; Boulos, Magdy I; Galileo, Deni S

2009-10-29

Malignant glioma cells are particularly motile and can travel diffusely through the brain parenchyma, apparently without following anatomical structures to guide their migration. The neural adhesion/recognition protein L1 (L1CAM; CD171) has been implicated in contributing to stimulation of motility and metastasis of several non-neural cancer types. We explored the expression and function of L1 protein as a stimulator of glioma cell motility using human high-grade glioma surgical specimens and established rat and human glioma cell lines. L1 protein expression was found in 17 out of 18 human high-grade glioma surgical specimens by western blotting. L1 mRNA was found to be present in human U-87/LacZ and rat C6 and 9L glioma cell lines. The glioma cell lines were negative for surface full length L1 by flow cytometry and high resolution immunocytochemistry of live cells. However, fixed and permeablized cells exhibited positive staining as numerous intracellular puncta. Western blots of cell line extracts revealed L1 proteolysis into a large soluble ectodomain (~180 kDa) and a smaller transmembrane proteolytic fragment (~32 kDa). Exosomal vesicles released by the glioma cell lines were purified and contained both full-length L1 and the proteolyzed transmembrane fragment. Glioma cell lines expressed L1-binding alphavbeta5 integrin cell surface receptors. Quantitative time-lapse analyses showed that motility was reduced significantly in glioma cell lines by 1) infection with an antisense-L1 retroviral vector and 2) L1 ectodomain-binding antibodies. Our novel results support a model of autocrine/paracrine stimulation of cell motility in glioma cells by a cleaved L1 ectodomain and/or released exosomal vesicles containing L1. This mechanism could explain the diffuse migratory behavior of high-grade glioma cancer cells within the brain.

Stimulation of glioma cell motility by expression, proteolysis, and release of the L1 neural cell recognition molecule

PubMed Central

Yang, Muhua; Adla, Shalini; Temburni, Murali K; Patel, Vivek P; Lagow, Errin L; Brady, Owen A; Tian, Jing; Boulos, Magdy I; Galileo, Deni S

2009-01-01

Background Malignant glioma cells are particularly motile and can travel diffusely through the brain parenchyma, apparently without following anatomical structures to guide their migration. The neural adhesion/recognition protein L1 (L1CAM; CD171) has been implicated in contributing to stimulation of motility and metastasis of several non-neural cancer types. We explored the expression and function of L1 protein as a stimulator of glioma cell motility using human high-grade glioma surgical specimens and established rat and human glioma cell lines. Results L1 protein expression was found in 17 out of 18 human high-grade glioma surgical specimens by western blotting. L1 mRNA was found to be present in human U-87/LacZ and rat C6 and 9L glioma cell lines. The glioma cell lines were negative for surface full length L1 by flow cytometry and high resolution immunocytochemistry of live cells. However, fixed and permeablized cells exhibited positive staining as numerous intracellular puncta. Western blots of cell line extracts revealed L1 proteolysis into a large soluble ectodomain (~180 kDa) and a smaller transmembrane proteolytic fragment (~32 kDa). Exosomal vesicles released by the glioma cell lines were purified and contained both full-length L1 and the proteolyzed transmembrane fragment. Glioma cell lines expressed L1-binding αvβ5 integrin cell surface receptors. Quantitative time-lapse analyses showed that motility was reduced significantly in glioma cell lines by 1) infection with an antisense-L1 retroviral vector and 2) L1 ectodomain-binding antibodies. Conclusion Our novel results support a model of autocrine/paracrine stimulation of cell motility in glioma cells by a cleaved L1 ectodomain and/or released exosomal vesicles containing L1. This mechanism could explain the diffuse migratory behavior of high-grade glioma cancer cells within the brain. PMID:19874583

Pharmacodynamic and Therapeutic Investigation of Focused Ultrasound-Induced Blood-Brain Barrier Opening for Enhanced Temozolomide Delivery in Glioma Treatment

PubMed Central

Liu, Hao-Li; Huang, Chiung-Yin; Chen, Ju-Yu; Wang, Hay-Yan Jack; Chen, Pin-Yuan; Wei, Kuo-Chen

2014-01-01

Focused ultrasound (FUS) exposure with the presence of microbubbles has been shown to transiently open the blood-brain barrier (BBB), and thus has potential to enhance the delivery of various kinds of therapeutic agents into brain tumors. The purpose of this study was to assess the preclinical therapeutic efficacy of FUS-BBB opening for enhanced temozolomide (TMZ) delivery in glioma treatment. FUS exposure with microbubbles was delivered to open the BBB of nude mice that were either normal or implanted with U87 human glioma cells. Different TMZ dose regimens were tested, ranging from 2.5 to 25 mg/kg. Plasma and brain samples were obtained at different time-points ranging from 0.5 to 4 hours, and the TMZ concentration within samples was quantitated via a developed LC-MS/MS procedure. Tumor progression was followed with T2-MRI, and animal survival and brain tissue histology were conducted. Results demonstrated that FUS-BBB opening caused the local TMZ accumulation in the brain to increase from 6.98 to 19 ng/mg. TMZ degradation time in the tumor core was found to increase from 1.02 to 1.56 hours. Improved tumor progression and animal survival were found at different TMZ doses (up to 15% and 30%, respectively). In conclusion, this study provides preclinical evidence that FUS-BBB opening increases the local concentration of TMZ to improve the control of tumor progression and animal survival, suggesting the potential for clinical application to improve current brain tumor treatment. PMID:25490097

The interaction of bee products with temozolomide in human diffuse astrocytoma, glioblastoma multiforme and astroglia cell lines.

PubMed

Borawska, Maria H; Markiewicz-Żukowska, Renata; Naliwajko, Sylwia K; Moskwa, Justyna; Bartosiuk, Emilia; Socha, Katarzyna; Surażyński, Arkadiusz; Kochanowicz, Jan; Mariak, Zenon

2014-01-01

In the present study, we investigated the influence of extracts from Salix spp. honey (ESH), beebread (EBB), and royal jelly (ERJ) with and without temozolomide (TMZ) on cell lines derived from a patient with diffuse astrocytoma (DASC), human glioblastoma multiforme (U87MG), and normal human astroglia (SVGp12). DASC was identified by immunocytochemistry. TMZ (20 μM) in combination with ESH (30 μg/mL), EBB (50 μg/mL), and ERJ (30 μg/mL) has stronger cytotoxic activity on U87MG cells after 72 h (20.0, 26.5, and 29.3% of control, respectively) than TMZ alone (about 6% of control). An increase of the cytotoxic effect and inhibition of DNA synthesis in SVGp12 were detected after administering TMZ with the studied extracts. NF-κB p50 subunit was reduced in U87MG cells after treatment with ESH (70.9%) and ESH + TMZ (74.7%). A significant decline of MMP-9 and MMP-2 secretion in cultured U87MG was detected after incubation with EBB (42.9% and 73.0%, respectively) and EBB + TMZ (38.4% and 68.5%, respectively). In conclusion, the use of bee products may increase the cytotoxic effect of TMZ in U87MG but also in SVGp12 cell line. It is important to note that the U87MG cells were sensitive to natural bee products, although there was no influence of natural bee products on the DASC cells.

Proteasome inhibition with bortezomib induces cell death in GBM stem-like cells and temozolomide-resistant glioma cell lines, but stimulates GBM stem-like cells' VEGF production and angiogenesis.

PubMed

Bota, Daniela A; Alexandru, Daniela; Keir, Stephen T; Bigner, Darell; Vredenburgh, James; Friedman, Henry S

2013-12-01

Recurrent malignant gliomas have inherent resistance to traditional chemotherapy. Novel therapies target specific molecular mechanisms involved in abnormal signaling and resistance to apoptosis. The proteasome is a key regulator of multiple cellular functions, and its inhibition in malignant astrocytic lines causes cell growth arrest and apoptotic cell death. The proteasome inhibitor bortezomib was reported to have very good in vitro activity against malignant glioma cell lines, with modest activity in animal models as well as in clinical trials as a single agent. In this paper, the authors describe the multiple effects of bortezomib in both in vitro and in vivo glioma models and offer a novel explanation for its seeming lack of activity. Glioma stem-like cells (GSCs) were obtained from resected glioblastomas (GBMs) at surgery and expanded in culture. Stable glioma cell lines (U21 and D54) as well as temozolomide (TMZ)-resistant glioma cells derived from U251 and D54-MG were also cultured. GSCs from 2 different tumors, as well as D54 and U251 cells, were treated with bortezomib, and the effect of the drug was measured using an XTT cell viability assay. The activity of bortezomib was then determined in D54-MG and/or U251 cells using apoptosis analysis as well as caspase-3 activity and proteasome activity measurements. Human glioma xenograft models were created in nude mice by subcutaneous injection. Bevacizumab was administered via intraperitoneal injection at a dose of 5 mg/kg daily. Bortezomib was administered by intraperitoneal injection 1 hour after bevacizumab administration in doses of at a dose of 0.35 mg/kg on days 1, 4, 8, and 11 every 21 days. Tumors were measured twice weekly. Bortezomib induced caspase-3 activation and apoptotic cell death in stable glioma cell lines and in glioma stem-like cells (GSCs) derived from malignant tumor specimens Furthermore, TMZ-resistant glioma cell lines retained susceptibility to the proteasome inhibition. The bortezomib activity was directly proportional with the cells' baseline proteasome activity. The proteasome inhibition stimulated both hypoxia-inducible factor (HIF)-1α and vascular endothelial growth factor (VEGF) production in malignant GSCs. As such, the VEGF produced by GSCs stimulated endothelial cell growth, an effect that could be prevented by the addition of bevacizumab (VEGF antibody) to the media. Similarly, administration of bortezomib and bevacizumab to athymic mice carrying subcutaneous malignant glioma xenografts resulted in greater tumor inhibition and greater improvement in survival than administration of either drug alone. These data indicate that simultaneous proteasome inhibition and VEGF blockade offer increased benefit as a strategy for malignant glioma therapy. The results of this study indicate that combination therapies based on bortezomib and bevacizumab might offer an increased benefit when the two agents are used in combination. These drugs have a complementary mechanism of action and therefore can be used together to treat TMZ-resistant malignant gliomas.

Dimethyl phenyl piperazine iodide (DMPP) induces glioma regression by inhibiting angiogenesis

DOE Office of Scientific and Technical Information (OSTI.GOV)

He, Yan-qing; Li, Yan; Wang, Xiao-yu

1,1-Dimethyl-4-phenyl piperazine iodide (DMPP) is a synthetic nicotinic acetylcholine receptor (nAChR) agonist that could reduce airway inflammation. In this study, we demonstrated that DMPP could dramatically inhibit glioma size maintained on the chick embryonic chorioallantoic membrane (CAM). We first performed MTT and BrdU incorporation experiments on U87 glioma cells in vitro to understand the mechanism involved. We established that DMPP did not significantly affect U87 cell proliferation and survival. We speculated that DMPP directly caused the tumor to regress by affecting the vasculature in and around the implanted tumor on our chick CAM model. Hence, we conducted detailed analysis ofmore » DMPP's inhibitory effects on angiogenesis. Three vasculogenesis and angiogenesis in vivo models were used in the study which included (1) early chick blood islands formation, (2) chick yolk-sac membrane (YSW) and (3) CAM models. The results revealed that DMPP directly suppressed all developmental stages involved in vasculogenesis and angiogenesis – possibly by acting through Ang-1 and HIF-2α signaling. In sum, our results show that DMPP could induce glioma regression grown on CAM by inhibiting vasculogenesis and angiogenesis. - Highlights: ●We demonstrated that DMPP inhibited the growth of glioma cells on chick CAM. ●DMPP did not significantly affect the proliferation and survival of U87 cells. ●We revealed that DMPP suppressed vasculogenesis and angiogenesis in chick embryo. ●Angiogenesis in chick CAM was inhibited by DMPP via most probably Ang-1 and HIF-2α. ●DMPP could be potentially developed as an anti-tumor drug in the future.« less

In vitro modulation of microglia motility by glioma cells is mediated by hepatocyte growth factor/scatter factor.

PubMed

Badie, B; Schartner, J; Klaver, J; Vorpahl, J

1999-05-01

Considered as immune effector cells of the central nervous system, microglia represent a major component of the inflammatory cells found in malignant gliomas. Although their role in brain tumor biology is unclear, accumulation of microglia in malignant brain tumors may be mediated through active secretion of cytokines by glioma cells. Because hepatocyte growth factor/scatter factor (HGF/SF) has been shown to modulate glioma motility through an autocrine mechanism, and because microglia have been reported to express the HGF/SF receptor Met, we hypothesized that microglia recruitment by gliomas may also occur through the secretion of HGF/SF. The effect of glioma cells in augmenting BV-2 murine microglia motility was studied by using an in vitro Boyden chamber migration assay. To determine the chemokines involved in microglia migration, neutralizing monoclonal antibodies against monocyte chemotactic protein-1 and HGF/SF were tested. Immunoblotting was used to check for the expression of HGF/SF by glioma cells, and the expression of Met by BV-2 cells was examined by flow cytometry. BV-2 migration was noted within 7 hours of incubation with both human (U251 MG and U373 MG) and murine (GL261) glioma cell lines. This migration corresponded to HGF/SF secretion by glioma cells and was completely inhibited by neutralizing monoclonal antibody against HGF/SF, but not monocyte chemotactic protein-1. Exposure of BV-2 cells to recombinant HGF/SF, but not monocyte chemotactic protein-1, resulted in their migration and down-regulation of Met in a dose-dependent fashion. HGF/SF, which plays a role in glioma motility and mitogenesis, may also act as a chemokine for microglia and may be responsible for the microglia infiltration in malignant gliomas. This active recruitment of microglia may play an important role in glioma biology.

System analysis identifies distinct and common functional networks governed by transcription factor ASCL1, in glioma and small cell lung cancer.

PubMed

Donakonda, Sainitin; Sinha, Swati; Dighe, Shrinivas Nivrutti; Rao, Manchanahalli R Satyanarayana

2017-07-25

ASCL1 is a basic Helix-Loop-Helix transcription factor (TF), which is involved in various cellular processes like neuronal development and signaling pathways. Transcriptome profiling has shown that ASCL1 overexpression plays an important role in the development of glioma and Small Cell Lung Carcinoma (SCLC), but distinct and common molecular mechanisms regulated by ASCL1 in these cancers are unknown. In order to understand how it drives the cellular functional network in these two tumors, we generated a gene expression profile in a glioma cell line (U87MG) to identify ASCL1 gene targets by an si RNA silencing approach and then compared this with a publicly available dataset of similarly silenced SCLC (NCI-H1618 cells). We constructed TF-TF and gene-gene interactions, as well as protein interaction networks of ASCL1 regulated genes in glioma and SCLC cells. Detailed network analysis uncovered various biological processes governed by ASCL1 target genes in these two tumor cell lines. We find that novel ASCL1 functions related to mitosis and signaling pathways influencing development and tumor growth are affected in both glioma and SCLC cells. In addition, we also observed ASCL1 governed functional networks that are distinct to glioma and SCLC.

Syndecan-1 knockdown inhibits glioma cell proliferation and invasion by deregulating a c-src/FAK-associated signaling pathway.

PubMed

Shi, Shuang; Zhong, Dong; Xiao, Yao; Wang, Bing; Wang, Wentao; Zhang, Fu'an; Huang, Haoyang

2017-06-20

Recent studies have shown that increased syndecan-1 (SDC1) expression in human glioma is associated with higher tumor grades and poor prognoses, but its oncogenic functions and the underlying molecular mechanisms remain unknown. Here, we examined SDC1 expression in datasets from The Cancer Genome Atlas and the National Center for Biotechnology Information Gene Expression Omnibus. Elevated SDC1 expression in glioma was closely associated with increases in tumor progression and shorter survival. We also examined SDC1 expression and evaluated the effects of stable SDC1 knockdown in glioma cell lines. SDC1 knockdown attenuated proliferation and invasion by glioma cells and markedly decreased PCNA and MMP-9 mRNA and protein expression. In a xenograft model, SDC1 knockdown suppressed the tumorigenic effects of U87 cells in vivo. SDC1 knockdown decreased phosphorylation of the c-src/FAK complex and its downstream signaling molecules, Erk, Akt and p38 MAPK. These results suggest that SDC1 may be a novel therapeutic target in the treatment of glioma.

Effect of hypoxia on the expression of genes encoding insulin-like growth factors and some related proteins in U87 glioma cells without IRE1 function.

PubMed

Minchenko, Dmytro O; Kharkova, A P; Halkin, O V; Karbovskyi, L L; Minchenko, O H

2016-04-01

The aim of the present study was to investigate the effect of hypoxia on the expression of genes encoding insulin-like growth factors (IGF1 and IGF2), their receptor (IGF1R), binding protein-4 (IGFBP4), and stanniocalcin 2 (STC2) in U87 glioma cells in relation to inhibition of endoplasmic reticulum stress signaling mediated by IRE1 (inositol requiring enzyme 1) for evaluation of their possible significance in the control of tumor growth. The expression of IGF1, IGF2, IGF1R, IGFBP4, and STC2 genes in U87 glioma cells transfected by empty vector pcDNA3.1 (control) and cells without IRE1 signaling enzyme function (transfected by dnIRE1) upon hypoxia was studied by qPCR. The expression of IGF1 and IGF2 genes is down-regulated in glioma cells without IRE1 signaling enzyme function in comparison with the control cells. At the same time, the expression of IGF1R, IGFBP4, and STC2 genes was up-regulated in glioma cells upon inhibition of IRE1, with more significant changes for IGFBP4 and STC2 genes. We also showed that hypoxia does not change significantly the expression of IGF1, IGF2, and IGF1R genes but up-regulated IGFBP4 and STC2 genes expression in control glioma cells. Moreover, the inhibition of both enzymatic activities (kinase and endoribonuclease) of IRE1 in glioma cells does not change significantly the effect of hypoxia on the expression of IGF1, IGF1R, and IGFBP4 genes but introduces sensitivity of IGF2 gene to hypoxic condition. Thus, the expression of IGF2 gene is resistant to hypoxia only in control glioma cells and significantly down-regulated in cells without functional activity of IRE1 signaling enzyme, which is central mediator of the unfolded protein response and an important component of the tumor growth as well as metabolic diseases. Results of this study demonstrate that the expression of IGF1 and IGF1R genes is resistant to hypoxic condition both in control U87 glioma cells and cells without IRE1 signaling enzyme function. However, hypoxia significantly up-regulates the expression of IGFBP4 gene independently on the inhibition of IRE1 enzyme. These data show that proteins encoded by these genes are resistant to hypoxia except IGFBP4 and participate in the regulation of metabolic and proliferative processes through IRE1 signaling.

2-[1-hexyloxyethyl]-2-devinyl pyropheophorbide-a (HPPH) in a nude rat glioma model: implications for photodynamic therapy.

PubMed

Lobel, J; MacDonald, I J; Ciesielski, M J; Barone, T; Potter, W R; Pollina, J; Plunkett, R J; Fenstermaker, R A; Dougherty, T J

2001-01-01

In this study, we evaluated 2-[1-hexyloxyethyl]-2-devinyl pyropheophorbide-alpha (HPPH or Photochlor) as a photosensitizer for the treatment of malignant gliomas by photodynamic therapy (PDT). We performed in vivo reflection spectroscopy in athymic rats to measure the attenuation of light in normal brain tissue. We also studied HPPH pharmacokinetics and PDT effects in nude rats with brain tumors derived from stereotactically implanted U87 human glioma cells. Rats implanted with tumors were sacrificed at designated time points to determine the pharmacokinetics of HPPH in serum, tumor, normal brain, and brain adjacent to tumor (BAT). HPPH concentrations in normal brain, BAT and tumor were determined using fluorescence spectroscopy. Twenty-four hours after intravenous injection of HPPH, we administered interstitial PDT treatment at a wavelength of 665 nm. Light was given in doses of 3.5, 7.5 or 15 J/cm at the tumor site and at a rate of 50 mW/cm. In vivo spectroscopy of normal brain tissue showed that the attenuation depth of 665 nm light is approximately 30% greater than that of 630 nm light used to activate Photofrin, which is currently being evaluated for PDT as an adjuvant to surgery for malignant gliomas. The t1/2 of disappearance of drug from serum and tumor was 25 and 30 hours, respectively. Twenty-four hours after injection of 0.5 mg/kg HPPH, tumor-to-brain drug ratios ranged from 5:1 to 15:1. Enhanced survival was observed in each of the HPPH/PDT-treated animal groups. These data suggest that HPPH may be a useful adjuvant for the treatment of malignant gliomas.

Effect of Hyp delivery system on PKCα activity: What will happen after pkcα gene silencing and Hyp photo-activation?

NASA Astrophysics Data System (ADS)

Misuth, Matus; Joniova, Jaroslava; Ferencakova, Michaela; Miskovsky, Pavol; Nadova, Zuzana

2015-08-01

Low density lipoproteins (LDL) are considered as suitable natural in vivo delivery system for hydrophobic photosensitizers (pts) such as hypericin (Hyp) and it was shown that over expression of LDL-receptors in tumor cells can be used for specific targeting. Activation of pts by irradiation results in a formation of reactive oxygen species (ROS) at the place of light application and starts destructive mechanism. PKCα plays a key role in the cell survival and its overexpression was observed in glioma cell lines. In the present study we aim to present the effectivity of the pts delivery in the glioma cells and consequences of silencing pkcα gene on cell death/survival after Hyp photo-activation. Pts can be delivered through two pathways: endocytosis - when cells are incubated with LDL/Hyp complex and Hyp transport through cellular membrane without any carrier. Preliminary results show that incubation of cells with or without LDL leads to PKCα activation. Photo-activated Hyp seems to be more effective in terms of apoptosis induction when compared to photo-activated LDL/Hyp complex. We have evaluated the influence of photo-activated Hyp on cell death in non-transfected and transfected (PKCα-) human glioma cells (U87-MG). Level of ROS production and type of cell death was notably affected by silencing pkca gene resulting in significant increase of necrosis after Hyp photo-activation.

Cytotoxic human peripheral blood-derived γδT cells kill glioblastoma cell lines: implications for cell-based immunotherapy for patients with glioblastoma.

PubMed

Nakazawa, Tsutomu; Nakamura, Mitsutoshi; Park, Young Soo; Motoyama, Yasushi; Hironaka, Yasuo; Nishimura, Fumihiko; Nakagawa, Ichiro; Yamada, Shuichi; Matsuda, Ryosuke; Tamura, Kentaro; Sugimoto, Tadashi; Takeshima, Yasuhiro; Marutani, Akiko; Tsujimura, Takahiro; Ouji, Noriko; Ouji, Yukiteru; Yoshikawa, Masahide; Nakase, Hiroyuki

2014-01-01

Glioblastoma (GBM) is a highly aggressive brain tumor for which novel therapeutic approaches, such as immunotherapy, are urgently needed. Zoledronate (ZOL), an inhibitor of osteoclastic activity, is known to stimulate peripheral blood-derived γδT cells and sensitize tumors to γδT cell-mediated killing. To investigate the feasibility of γδT cell-based immunotherapy for patients with GBM, we focused on the killing of GBM cell lines by γδT cells and the molecular mechanisms involved in these cell-cell interactions. Peripheral blood mononuclear cells were expanded in ZOL and interleukin (IL)-2 for 14 days, and γδT cells were enriched in the expanded cells by the immunomagnetic depletion of αβT cells. Gliomas are resistant to NK cells but susceptible to lymphokine-activated killer cells and some cytotoxic T lymphocytes. When the γδT cell-mediated killing of three GBM cell lines (U87MG, U138MG and A172 cells) and an NK-sensitive leukemia cell line (K562 cells) were tested, 32% U87MG, 15% U138MG, 1% A172, and 50% K562 cells were killed at an effector:target ratio of 5:1. The γδT cell-mediated killing of all three GBM cell lines was significantly enhanced by ZOL and this ZOL-enhanced killing was blocked by an anti-T cell receptor (TcR) antibody. These results indicated that TcR γδ is crucial for the recognition of ZOL-treated GBM cells by γδT cells. Since the low level killing of GBM cells by the γδT cells was enhanced by ZOL, γδT cell-targeting therapy in combination with ZOL treatment could be effective for patients with GBM.

Functional analysis of a novel glioma antigen, EFTUD1

PubMed Central

Saito, Katsuya; Iizuka, Yukihiko; Ohta, Shigeki; Takahashi, Satoshi; Nakamura, Kenta; Saya, Hideyuki; Yoshida, Kazunari; Kawakami, Yutaka; Toda, Masahiro

2014-01-01

Background A cDNA library made from 2 glioma cell lines, U87MG and T98G, was screened by serological identification of antigens by recombinant cDNA expression (SEREX) using serum from a glioblastoma patient. Elongation factor Tu GTP binding domain containing protein 1 (EFTUD1), which is required for ribosome biogenesis, was identified. A cancer microarray database showed overexpression of EFTUD1 in gliomas, suggesting that EFTUD1 is a candidate molecular target for gliomas. Methods EFTUD1 expression in glioma cell lines and glioma tissue was assessed by Western blot, quantitative PCR, and immunohistochemistry. The effect on ribosome biogenesis, cell growth, cell cycle, and induction of apoptosis and autophagy in glioma cells during the downregulation of EFTUD1 was investigated. To reveal the role of autophagy, the autophagy-blocker, chloroquine (CQ), was used in glioma cells downregulating EFTUD1. The effect of combining CQ with EFTUD1 inhibition in glioma cells was analyzed. Results EFTUD1 expression in glioma cell lines and tissue was higher than in normal brain tissue. Downregulating EFTUD1 induced G1 cell-cycle arrest and apoptosis, leading to reduced glioma cell proliferation. The mechanism underlying this antitumor effect was impaired ribosome biogenesis via EFTUD1 inhibition. Additionally, protective autophagy was induced by glioma cells as an adaptive response to EFTUD1 inhibition. The antitumor effect induced by the combined treatment was significantly higher than that of either EFTUD1 inhibition or CQ alone. Conclusion These results suggest that EFTUD1 represents a novel therapeutic target and that the combination of EFTUD1 inhibition with autophagy blockade may be effective in the treatment of gliomas. PMID:25015090

Intracranial AAV-sTRAIL combined with lanatoside C prolongs survival in an orthotopic xenograft mouse model of invasive glioblastoma.

PubMed

Crommentuijn, Matheus H W; Maguire, Casey A; Niers, Johanna M; Vandertop, W Peter; Badr, Christian E; Würdinger, Thomas; Tannous, Bakhos A

2016-04-01

Glioblastoma (GBM) is the most common malignant brain tumor in adults. We designed an adeno-associated virus (AAV) vector for intracranial delivery of secreted, soluble tumor necrosis factor-related apoptosis-inducing ligand (sTRAIL) to GBM tumors in mice and combined it with the TRAIL-sensitizing cardiac glycoside, lanatoside C (lan C). We applied this combined therapy to two different GBM models using human U87 glioma cells and primary patient-derived GBM neural spheres in culture and in orthotopic GBM xenograft models in mice. In U87 cells, conditioned medium from AAV2-sTRAIL expressing cells combined with lan C induced 80% cell death. Similarly, lan C sensitized primary GBM spheres to sTRAIL causing over 90% cell death. In mice bearing intracranial U87 tumors treated with AAVrh.8-sTRAIL, administration of lan C caused a decrease in tumor-associated Fluc signal, while tumor size increased within days of stopping the treatment. Another round of lan C treatment re-sensitized GBM tumor to sTRAIL-induced cell death. AAVrh.8-sTRAIL treatment alone and combined with lanatoside C resulted in a significant decrease in tumor growth and longer survival of mice bearing orthotopic invasive GBM brain tumors. In summary, AAV-sTRAIL combined with lanatoside C induced cell death in U87 glioma cells and patient-derived GBM neural spheres in culture and in vivo leading to an increased in overall mice survival. Copyright © 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Gracilaria lemaneiformis polysaccharide as integrin-targeting surface decorator of selenium nanoparticles to achieve enhanced anticancer efficacy.

PubMed

Jiang, Wenting; Fu, Yuanting; Yang, Fang; Yang, Yufeng; Liu, Ting; Zheng, Wenjie; Zeng, Lilan; Chen, Tianfeng

2014-08-27

The poor permeability of glioma parenchyma represents a major limit for antiglioblastoma drug delivery. Gracilaria lemaneiformis polysaccharide (GLP), which has a high binding affinity to αvβ3 integrin overexpressed in glioma cells, was employed in the present study to functionalize selenium nanoparticles (SeNPs) to achieve antiglioblastoma efficacy. GLP-SeNPs showed satisfactory size distribution, high stability, and selectivity between cancer and normal cells. In U87 glioma cell membrane, which has a high integrin expression level, GLP-SeNPs exhibited significantly higher cellular uptake than unmodified SeNPs. As expected, U87 cells exhibited a greater uptake of GLP-SeNPs than C6 cells with low integrin expression level. Furthermore, the internalization of GLP-SeNPs was inhibited by cyclo-(Arg-Gly-Asp-Phe-Lys) peptides, suggesting that cellular uptake into U87 cells and C6 cells occurred via αvβ3 integrin-mediated endocytosis. For U87 cells, the cytotoxicity of SeNPs decorated by GLP was enhanced significantly because of the induction of various apoptosis signaling pathways. Internalized GLP-SeNPs triggered intracellular reactive oxygen species downregulation. Therefore, p53, MAPKs, and AKT pathways were activated to advance cell apoptosis. These findings suggest that surface decoration of nanomaterials with GLP could be an efficient strategy for design and preparation of glioblastoma targeting nanodrugs.

Identification, characterization and potent antitumor activity of ECO-4601, a novel peripheral benzodiazepine receptor ligand.

PubMed

Gourdeau, Henriette; McAlpine, James B; Ranger, Maxime; Simard, Bryan; Berger, Francois; Beaudry, Francis; Farnet, Chris M; Falardeau, Pierre

2008-05-01

ECO-4601 is a structurally novel farnesylated dibenzodiazepinone discovered through DECIPHER technology, Thallion's proprietary drug discovery platform. The compound was shown to have a broad cytotoxic activity in the low micromolar range when tested in the NCI 60 cell line panel. In the work presented here, ECO-4601 was further evaluated against brain tumor cell lines. Preliminary mechanistic studies as well as in vivo antitumor evaluation were performed. Since ECO-4601 has a benzodiazepinone moiety, we first investigated if it binds the central and/or peripheral benzodiazepine receptors. ECO-4601 was tested in radioligand binding assays on benzodiazepine receptors obtained from rat hearts. The ability of ECO-4601 to inhibit the growth of CNS cancers was evaluated on a panel of mouse, rat and human glioma cell lines using a standard MTT assay. Antitumor efficacy studies were performed on gliomas (rat and human), human breast and human prostate mouse tumor xenografts. Antitumor activity and pharmacokinetic analysis of ECO-4601 was evaluated following intravenous (i.v.), subcutaneous (s.c.), and intraperitoneal (i.p.) bolus administrations. ECO-4601 was shown to bind the peripheral but not the central benzodiazepine receptor and inhibited the growth of CNS tumor cell lines. Bolus s.c. and i.p. administration gave rise to low but sustained drug exposure, and resulted in moderate to significant antitumor activity at doses that were well tolerated. In a rat glioma (C6) xenograft model, ECO-4601 produced up to 70% tumor growth inhibition (TGI) while in a human glioma (U-87MG) xenograft, TGI was 34%. Antitumor activity was highly significant in both human hormone-independent breast (MDA-MB-231) and prostate (PC-3) xenografts, resulting in TGI of 72 and 100%, respectively. On the other hand, i.v. dosing was followed by rapid elimination of the drug and was ineffective. Antitumor efficacy of ECO-4601 appears to be associated with the exposure parameter AUC and/or sustained drug levels rather than C (max). These in vivo data constitute a rationale for clinical studies testing prolonged continuous administration of ECO-4601.

A super gene expression system enhances the anti-glioma effects of adenovirus-mediated REIC/Dkk-3 gene therapy

NASA Astrophysics Data System (ADS)

Oka, Tetsuo; Kurozumi, Kazuhiko; Shimazu, Yosuke; Ichikawa, Tomotsugu; Ishida, Joji; Otani, Yoshihiro; Shimizu, Toshihiko; Tomita, Yusuke; Sakaguchi, Masakiyo; Watanabe, Masami; Nasu, Yasutomo; Kumon, Hiromi; Date, Isao

2016-09-01

Reduced expression in immortalized cells/Dickkopf-3 (REIC/Dkk-3) is a tumor suppressor and therapeutic gene in many human cancers. Recently, an adenovirus REIC vector with the super gene expression system (Ad-SGE-REIC) was developed to increase REIC/Dkk-3 expression and enhance therapeutic effects compared with the conventional adenoviral vector (Ad-CAG-REIC). In this study, we investigated the in vitro and in vivo effects of Ad-SGE-REIC on malignant glioma. In U87ΔEGFR and GL261 glioma cells, western blotting confirmed that robust upregulation of REIC/Dkk-3 expression occurred in Ad-SGE-REIC-transduced cells, most notably after transduction at a multiplicity of infection of 10. Cytotoxicity assays showed that Ad-SGE-REIC resulted in a time-dependent and significant reduction in the number of malignant glioma cells attaching to the bottom of culture wells. Xenograft and syngeneic mouse intracranial glioma models treated with Ad-SGE-REIC had significantly longer survival than those treated with the control vector Ad-LacZ or with Ad-CAG-REIC. This study demonstrated the anti-glioma effect of Ad-SGE-REIC, which may represent a promising strategy for the treatment of malignant glioma.

NUMB does not impair growth and differentiation status of experimental gliomas

DOE Office of Scientific and Technical Information (OSTI.GOV)

Euskirchen, Philipp, E-mail: philipp.euskirchen@charite.de; Laboratory for Gene Therapy and Molecular Imaging, Max-Planck-Institute for Neurological Research, Cologne; Skaftnesmo, Kai-Ove

2011-12-10

The cell fate determinant NUMB orchestrates asymmetric cell division in flies and mammals and has lately been suggested to have a tumor suppressor function in breast and lung cancer. Here, we studied NUMB in the context of malignant gliomas. We used ectopic expression of NUMB in order to inhibit proliferation and induce differentiation in glioma cells by alteration of Notch, Hedgehog and p53 signaling. We found that NUMB is consistently expressed in glioma biopsies with predominance of NUMB2/4 isoforms as determined by isoform-specific real-time PCR and Western blotting. Upon lentiviral overexpression, in vitro proliferation rate and the grade of differentiationmore » as assessed by morphology and expression of neural and glial markers remained unchanged. Orthotopic xenografts of NUMB-transduced human U87 glioma cells could be established in nude rats without impairing engraftment or causing significant changes in morphology based on magnetic resonance imaging (MRI). The previously reported alteration of Hedgehog and p53 signaling by NUMB could not be recapitulated in glioma cells. We thus show that in experimental gliomas, NUMB overexpression most likely does not exert a tumor suppressor function such as seen in epithelial cancers.« less

EMMPRIN expression positively correlates with WHO grades of astrocytomas and meningiomas.

PubMed

Tsai, Wen-Chiuan; Chen, Ying; Huang, Li-Chun; Lee, Herng-Sheng; Ma, Hsin-I; Huang, Shih-Ming; Sytwu, Huey-Kang; Hueng, Dueng-Yuan

2013-09-01

High-grade primary brain tumors possessed poor outcome due to invasiveness. Extracellular matrix metalloproteinase inducer (EMMPRIN) stimulates peri-tumoral fibroblasts to secrete matrix metalloproteinase and promote invasiveness. This study hypothesized that high-grade brain tumors overexpress EMMPRIN. Analyzing the public delinked database from the Gene Expression Omnibus profile, the results showed that the EMMPRIN mRNA level was higher in WHO grade IV (n = 81) than in grade III (n = 19, p < 0.0005) astrocytomas and non-tumor brain tissue controls (n = 23, p < 0.00001). The results of tissue microarray-based immunohistochemical (IHC) staining revealed that EMMPRIN levels positively correlated with WHO grades for astrocytomas (p = 0.008) and meningiomas (p = 0.048). EMMPRIN mRNA levels in conventional glioma cell lines (n = 36) was not less than those in glioma primary culture cells (n = 27) and glioblastoma stem-like cells (n = 12). The GBM8401, U87MG, and LN229 human glioma cell lines also overexpressed EMMPRIN. Hematoxylin and eosin, IHC, and immunofluorescence staining of xenografts confirmed that high-grade brain tumors overexpressed EMMPRIN. Lastly, Kaplan-Meier analysis revealed poorer survival in WHO grade IV (n = 56) than in grade III astrocytomas (n = 21, by log-rank test; p = 0.0001, 95 % CI: 1.842-3.053). However, in high-grade astrocytomas, there was no difference in survival between high and low EMMPRIN mRNA levels. Thus, this study identified that high-grade brain tumors overexpress EMMPRIN, which positively correlates with WHO grades in human astrocytomas and meningiomas, and suggests that EMMPRIN may be a therapeutic target of brain tumor.

Integrin-mediated human glioblastoma cells adhesion, migration and invasion by native and recombinant phospholipases of Scorpio maurus venom glands.

PubMed

Krayem, Najeh; Abdelkefi-Koubaa, Zaineb; Gargouri, Youssef; Luis, José

2018-05-01

Integrins are a large family of cell surface receptors mediating the interaction of cells with their microenvironment and they play an important role in glioma biology. In the present work, we reported the anti-tumor effect of Sm-PLGV a phospholipase A 2 from Tunisian scorpion venom glands-as well as its recombinant forms expressed in Escherichia coli-through interference with integrin receptor function in malignant glioma cells U87. These phospholipases inhibited in a dose dependent manner the adhesion, migration and invasion onto fibrinogen and fibronectin without any cytotoxicity. We showed that Sm-PLGV and its recombinant constructs blocked U87 migration by reducing their velocity and directional persistence. The inhibitory effect was related to a blockage of the integrins αvβ3 and α5β1 function. Inactivation of the enzymatic activity of Sm-PLGV by chemical modification with p-bromophenacyl bromide did not affect its anti-tumor properties, suggesting the presence of 'pharmacological sites' distinct from the catalytic site in scorpion venom phospholipases A 2 . Copyright © 2018 Elsevier Inc. All rights reserved.

IDH1 R132H Mutation Enhances Cell Migration by Activating AKT-mTOR Signaling Pathway, but Sensitizes Cells to 5-FU Treatment as NADPH and GSH Are Reduced.

PubMed

Zhu, Huixia; Zhang, Ye; Chen, Jianfeng; Qiu, Jiangdong; Huang, Keting; Wu, Mindan; Xia, Chunlin

2017-01-01

Mutations of isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2) gene were recently discovered in vast majority of World Health Organization (WHO) grade II/III gliomas. This study is to understand the effects of IDH1 R132H mutation in gliomagenesis and to develop new strategies to treat glioma with IDH1 R132H mutation. Over expression of IDH1 R132H in U87MG cells was done by transfecting cells with IDH1 R132H plasmid. MTT assay, scratch repair assay and western blot were performed to study effects of IDH1 R132H mutation on cell proliferation, migration, regulating AKT-mTOR signaling pathway and cell death respectively. NADP+/NADPH and GSH quantification assays were performed to evaluate effects of IDH1 R132H mutation on the production of antioxidant NADPH and GSH. We found that over expression of IDH1 R132H mutation decreased cell proliferation consistent with previous reports; however, it increased cell migration and enhanced AKT-mTOR signaling pathway activation. Mutations in isocitrate dehydrogenase (IDH) 1 also change the function of the enzymes and cause them to produce 2-hydroxyglutarate and not produce NADPH. We tested the level of NADPH and GSH and demonstrated that IDH1 R132H mutant stable cells had significantly low NADPH and GSH level compared to control or IDH1 wild type stable cells. The reduced antioxidants (NADPH and GSH) sensitized U87MG cells with IDH R132H mutant to 5-FU treatment. Our study highlights the important role of IHD1 R132H mutant in up- regulating AKT-mTOR signaling pathway and enhancing cell migration. Furthermore, we demonstrate that IDH1 R132H mutation affects cellular redox status and sensitizes gliomas cells with IDH1 R132H mutation to 5FU treatment.

Glioblastoma recurrence correlates with NLGN3 levels.

PubMed

Liu, Rui; Qin, Xing-Ping; Zhuang, Yang; Zhang, Ya; Liao, Hua-Bao; Tang, Jun-Chun; Pan, Meng-Xian; Zeng, Fei-Fei; Lei, Yang; Lei, Rui-Xue; Wang, Shu; Liu, An-Chun; Chen, Juan; Zhang, Zhi-Feng; Zhao, Dan; Wu, Song-Lin; Liu, Ren-Zhong; Wang, Ze-Fen; Wan, Qi

2018-05-18

Glioblastoma (GBM) is the most aggressive glioma in the brain. Recurrence of GBM is almost inevitable within a short term after tumor resection. In a retrospective study of 386 cases of GBM collected between 2013 and 2016, we found that recurrence of GBM mainly occurs in the deep brain regions, including the basal ganglia, thalamus, and corpus callosum. But the mechanism underlying this phenomenon is not clear. Previous studies suggest that neuroligin-3 (NLGN3) is necessary for GBM growth. Our results show that the levels of NLGN3 in the cortex are higher than those in the deep regions in a normal human brain, and similar patterns are also found in a normal mouse brain. In contrast, NLGN3 levels in the deep brain regions of GBM patients are high. We also show that an increase in NLGN3 concentration promotes the growth of U251 cells and U87-MG cells. Respective use of the cortex neuron culture medium (C-NCM) and basal ganglia neuron culture medium (BG-NCM) with DMEM to cultivate U251, U87-MG and GBM cells isolated from patients, we found that these cells grew faster after treatment with C-NCM and BG-NCM in which the cells treated with C-NCM grew faster than the ones treated with BG-NCM group. Inhibition of NLGN3 release by ADAM10i prevents NCM-induced cell growth. Together, this study suggests that increased levels of NLGN3 in the deep brain region under the GBM pathological circumstances may contribute to GBM recurrence in the basal ganglia, thalamus, and corpus callosum. © 2018 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.

Pc 4 photodynamic therapy of U87 (human glioma) orthotopic tumor in nude rat brain

NASA Astrophysics Data System (ADS)

Dean, David; George, John E., III; Ahmad, Yusra; Wolfe, Michael S.; Lilge, Lothar; Morris, Rachel L.; Peterson, Allyn; Lust, W. D.; Totonchi, Ali; Varghai, Davood; Li, Xiaolin; Hoppel, Charles L.; Sun, Jiayang; Oleinick, Nancy L.

2005-04-01

Introduction: Photodynamic therapy (PDT) for Barrett"s esophagus, advanced esophageal cancer, and both early and late inoperable lung carcinoma is now FDA-approved using the first generation photosensitizer PhotofrinTM (Axcan Pharma, Birmingham, AL). Photofrin-mediated PDT of glioma is now in Phase III clinical trials. A variety of second generation photosensitizers have been developed to provide improved: (1) specificity for the target tissue, (2) tumoricidal capability, and (3) rapid clearance the vascular compartment, skin, and eyes. The phthalocyanine Pc 4 is a second generation photosensitizer that is in early phase I clinical trials for skin cancer. We have undertaken a preclinical study that seeks to determine if Pc 4-mediated PDT can be of benefit for the intra-operative localization and treatment of glioma. Methods: Using a stereotactic frame, 250,000 U87 cells were injected via Hamilton syringe through a craniotomy, and the dura, 1-2 mm below the cortical surface of nude (athymic) rat brains (N=91). The craniotomy was filled with a piece of surgical PVC and the scalp closed. After two weeks of tumor growth, the animals received 0.5 mg/kg Pc 4 via tail vein injection. One day later the scalp was re-incised, and the PVC removed. The tumor was then illuminated with either 5 or 30 Joule/cm2 of 672-nm light from a diode laser at 50 mW/cm2. The animals were sacrificed one day later and the brain was cold-perfused with formaldehyde. Two thirds of the explanted brains are now being histologically surveyed for necrosis after staining with hematoxylin and eosin and for apoptosis via immunohistochemistry (i.e., TUNEL assay). The other third were analyzed by HPLC-mass spectrometry for the presence of drug in tumor, normal brain, and plasma at sacrifice. Initial histological results show PDT-induced apoptosis and necrosis confined to the growing (live) portion of the tumor. Preliminary analysis shows an average selectivity of Pc 4 uptake in the bulk tumor to be 3.8 times greater than in normal brain tissue. Discussion: The observed specificity and tumoricidal activity of Pc 4 warrants further preclinical studies to determine the preferred Pc 4 drug and light dose for future glioma patient clinical trials.

Methotrexate loaded polyether-copolyester dendrimers for the treatment of gliomas: enhanced efficacy and intratumoral transport capability.

PubMed

Dhanikula, Renu Singh; Argaw, Anteneh; Bouchard, Jean-Francois; Hildgen, Patrice

2008-01-01

Therapeutic benefit in glial tumors is often limited due to low permeability of delivery systems across the blood-brain barrier (BBB), drug resistance, and poor penetration into the tumor tissue. In an attempt to overcome these hurdles, polyether-copolyester (PEPE) dendrimers were evaluated as drug carriers for the treatment of gliomas. Dendrimers were conjugated to d-glucosamine as the ligand for enhancing BBB permeability and tumor targeting. The efficacy of methotrexate (MTX)-loaded dendrimers was established against U87 MG and U 343 MGa cells. Permeability of rhodamine-labeled dendrimers and MTX-loaded dendrimers across the in vitro BBB model and their distribution into avascular human glioma tumor spheroids was also studied. Glucosylated dendrimers were found to be endocytosed in significantly higher amounts than nonglucosylated dendrimers by both the cell lines. IC 50 of MTX after loading in dendrimers was lower than that of the free MTX, suggesting that loading MTX in PEPE dendrimers increased its potency. Similar higher activity of MTX-loaded glucosylated and nonglucosylated dendrimers was found in the reduction of tumor spheroid size. These MTX-loaded dendrimers were able to kill even MTX-resistant cells highlighting their ability to overcome MTX resistance. In addition, the amount of MTX-transported across BBB was three to five times more after loading in the dendrimers. Glucosylation further increased the cumulative permeation of dendrimers across BBB and hence increased the amount of MTX available across it. Glucosylated dendrimers distributed through out the avascular tumor spheroids within 6 h, while nonglucosylated dendrimers could do so in 12 h. The results show that glucosamine can be used as an effective ligand not only for targeting glial tumors but also for enhanced permeability across BBB. Thus, glucosylated PEPE dendrimers can serve as potential delivery system for the treatment of gliomas.

Cytopathic Effects of X-ray Irradiation and MnO Nanoparticles on Human Glioblastoma (U87)

NASA Astrophysics Data System (ADS)

Kuper, K. E.; Zavjalov, E. L.; Razumov, I. A.; Romaschenko, A. V.; Stupak, A. S.; Troicky, S. Yu; Goldenberg, B. G.; Legkodymov, A. G.; Lemzyakov, A. A.; Moshkin, M. P.

Glioblastoma is a leader among the most malignant brain tumors with the average lifespan of patients around 9-12 months. For prevention and treatment of neuropathology, a variety of therapeutic and surgical approaches are being developed and improved, including radiation and chemical therapy methods. In our work, we investigated cytopathic effect of X-ray irradiation with application of metal oxides nanoparticles such as manganese oxide (MnO) on U87 human glioblastoma cells. We used the X-ray irradiation dose of 0.5, 4, 40 and 100 Gy in combination with nanoparticles at the concentration of 0.5 ng/ml. The irradiation of glioma cell was carried out at the synchrotron radiation source VEPP-4. After cells treatments with nanoparticles for about 24 h and radiation the results were assessed by MTT assay test with 106/ml cells densities. We demonstrate that preincubation of the glioblastoma cell lines U87 with MnO nanoparticles allows reducing dose of irradiation. This combination of nanoparticles and X-ray irradiation provides new possibilities for the treatment of brain tumors.

Acrylamide-induced apoptosis in rat primary astrocytes and human astrocytoma cell lines.

PubMed

Lee, Jiann-Gwu; Wang, Yan-Shiu; Chou, Chin-Cheng

2014-06-01

This study aimed to evaluate the acrylamide (ACR)-induced apoptotic effects on rat primary astrocytes and three human astrocytoma-derived cell lines (U-1240 MG, U-87 MG, and U-251 MG). As determined through the MTT assay, treatment with 1 and 2 mM ACR for 24-72 h resulted in decreased cell viability in all cells. Decreases in cell viability could be blocked in all cells with the exception of U-251 MG cells by Z-DEVD FMK. ACR-induced dose-dependent apoptotic effects were also demonstrated by increases in the sub-G1 phase cell population in all cells. The decreased expressions of pro-caspase 3, 8, and 9 and the interruption of the mitochondrial membrane potential were observed in all cells. Exposure to 2 mM ACR for 48 h resulted in increased Bax/Bcl-2 ratios in primary astrocytes and U-87 MG cells, whereas the overexpression of Bcl-2 was observed in U-1240 MG and U-251 MG cells. The ACR-induced increases in the levels of p53 and pp53 in primary astrocytes could be attenuated by caffeine. These results suggest the existence of a common apoptotic pathway among all cell types and that U-87 MG cells may be a suitable substitute in vitro model for primary astrocytes in future studies on ACR-induced neurotoxicity. Copyright © 2014 Elsevier Ltd. All rights reserved.

Saponin 6 derived from Anemone taipaiensis induces U87 human malignant glioblastoma cell apoptosis via regulation of Fas and Bcl‑2 family proteins.

PubMed

Ji, Chen-Chen; Tang, Hai-Feng; Hu, Yi-Yang; Zhang, Yun; Zheng, Min-Hua; Qin, Hong-Yan; Li, San-Zhong; Wang, Xiao-Yang; Fei, Zhou; Cheng, Guang

2016-07-01

Glioblastoma multiforme (GBM) is the most common and aggressive type of brain tumor, and is associated with a poor prognosis. Saponin 6, derived from Anemone taipaiensis, exerts potent cytotoxic effects against the human hepatocellular carcinoma HepG2 cell line and the human promyelocytic leukemia HL‑60 cell line; however, the effects of saponin 6 on glioblastoma remain unknown. The present study aimed to evaluate the effects of saponin 6 on human U87 malignant glioblastoma (U87 MG) cells. The current study revealed that saponin 6 induced U87 MG cell death in a dose‑ and time‑dependent manner, with a half maximal inhibitory concentration (IC50) value of 2.83 µM after treatment for 48 h. However, saponin 6 was needed to be used at a lesser potency in HT‑22 cells, with an IC50 value of 6.24 µM. Cell apoptosis was assessed by flow cytometry using Annexin V‑fluorescein isothiocyanate/propidium iodide double staining. DNA fragmentation and alterations in nuclear morphology were examined by terminal deoxynucleotidyl transferase‑mediated dUTP nick end labeling and transmission electron microscopy, respectively. The present study demonstrated that treatment with saponin 6 induced cell apoptosis in U87 MG cells, and resulted in DNA fragmentation and nuclear morphological alterations typical of apoptosis. In addition, flow cytometric analysis revealed that saponin 6 was able to induce cell cycle arrest. The present study also demonstrated that saponin 6‑induced apoptosis of U87 MG cells was attributed to increases in the protein expression levels of Fas, Fas ligand, and cleaved caspase‑3, ‑8 and ‑9, and decreases in the levels of B‑cell lymphoma 2. The current study indicated that saponin 6 may exhibit selective cytotoxicity toward U87 MG cells by activating apoptosis via the extrinsic and intrinsic pathways. Therefore, saponin 6 derived from A. taipaiensis may possess therapeutic potential for the treatment of GBM.

4'-Acetoamido-4-hydroxychalcone, a chalcone derivative, inhibits glioma growth and invasion through regulation of the tropomyosin 1 gene

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ku, Bo Mi; Ryu, Hyung Won; Lee, Yeon Kyung

2010-11-19

Research highlights: {yields} 4'-Acetoamido-4-hydroxychalcone (AHC) has anti-cancer property for glioma. {yields} 4'-Acetoamido-4-hydroxychalcone (AHC) increased tropomyosin expreesion through activattion of PKA signaling. {yields} 4'-Acetoamido-4-hydroxychalcone (AHC) inhibits glioma cell migration and invasion. {yields} In vivo administration of 4'-acetoamido-4-hydroxychalcone (AHC) reduced tumor growth. -- Abstract: Chalcones are precursors of flavonoids and have been shown to have anti-cancer activity. Here, we identify the synthetic chalcone derivative 4'-acetoamido-4-hydroxychalcone (AHC) as a potential therapeutic agent for the treatment of glioma. Treatment with AHC reduced glioma cell invasion, migration, and colony formation in a concentration-dependent manner. In addition, AHC inhibited vascular endothelial growth factor-induced migration, invasion, andmore » tube formation in HUVECs. To determine the mechanism underlying the inhibitory effect of AHC on glioma cell invasion and migration, we investigated the effect of AHC on the gene expression change and found that AHC affects actin dynamics in U87MG glioma cells. In actin cytoskeleton regulating system, AHC increased tropomyosin expression and stress fiber formation, probably through activation of PKA. Suppression of tropomyosin expression by siRNA or treatment with the PKA inhibitor H89 reduced the inhibitory effects of AHC on glioma cell invasion and migration. In vivo experiments also showed that AHC inhibited tumor growth in a xenograft mouse tumor model. Together, these data suggest that the synthetic chalcone derivative AHC has potent anti-cancer activity through inhibition of glioma proliferation, invasion, and angiogenesis and is therefore a potential chemotherapeutic candidate for the treatment of glioma.« less

Identification of NVP-BKM120 as a Potent, Selective, Orally Bioavailable Class I PI3 Kinase Inhibitor for Treating Cancer.

PubMed

Burger, Matthew T; Pecchi, Sabina; Wagman, Allan; Ni, Zhi-Jie; Knapp, Mark; Hendrickson, Thomas; Atallah, Gordana; Pfister, Keith; Zhang, Yanchen; Bartulis, Sarah; Frazier, Kelly; Ng, Simon; Smith, Aaron; Verhagen, Joelle; Haznedar, Joshua; Huh, Kay; Iwanowicz, Ed; Xin, Xiaohua; Menezes, Daniel; Merritt, Hanne; Lee, Isabelle; Wiesmann, Marion; Kaufman, Susan; Crawford, Kenneth; Chin, Michael; Bussiere, Dirksen; Shoemaker, Kevin; Zaror, Isabel; Maira, Sauveur-Michel; Voliva, Charles F

2011-10-13

Phosphoinositide-3-kinases (PI3Ks) are important oncology targets due to the deregulation of this signaling pathway in a wide variety of human cancers. Herein we describe the structure guided optimization of a series of 2-morpholino, 4-substituted, 6-heterocyclic pyrimidines where the pharmacokinetic properties were improved by modulating the electronics of the 6-position heterocycle, and the overall druglike properties were fine-tuned further by modification of the 4-position substituent. The resulting 2,4-bismorpholino 6-heterocyclic pyrimidines are potent class I PI3K inhibitors showing mechanism modulation in PI3K dependent cell lines and in vivo efficacy in tumor xenograft models with PI3K pathway deregulation (A2780 ovarian and U87MG glioma). These efforts culminated in the discovery of 15 (NVP-BKM120), currently in Phase II clinical trials for the treatment of cancer.

Identification of NVP-BKM120 as a Potent, Selective, Orally Bioavailable Class I PI3 Kinase Inhibitor for Treating Cancer

PubMed Central

2011-01-01

Phosphoinositide-3-kinases (PI3Ks) are important oncology targets due to the deregulation of this signaling pathway in a wide variety of human cancers. Herein we describe the structure guided optimization of a series of 2-morpholino, 4-substituted, 6-heterocyclic pyrimidines where the pharmacokinetic properties were improved by modulating the electronics of the 6-position heterocycle, and the overall druglike properties were fine-tuned further by modification of the 4-position substituent. The resulting 2,4-bismorpholino 6-heterocyclic pyrimidines are potent class I PI3K inhibitors showing mechanism modulation in PI3K dependent cell lines and in vivo efficacy in tumor xenograft models with PI3K pathway deregulation (A2780 ovarian and U87MG glioma). These efforts culminated in the discovery of 15 (NVP-BKM120), currently in Phase II clinical trials for the treatment of cancer. PMID:24900266

Functionalized cell nucleus-penetrating peptide combined with doxorubicin for synergistic treatment of glioma.

PubMed

Zhang, Li; Zhang, Yanyu; Tai, Lingyu; Jiang, Kuan; Xie, Cao; Li, Zhuoquan; Lin, Yao-Zhong; Wei, Gang; Lu, Weiyue; Pan, Weisan

2016-09-15

Clinical application of cell-penetrating peptides (CPPs) in cancer therapy is greatly restricted due to lack of tissue selectivity and tumor-targeting ability. CB5005, a rationally designed CPP that targets and inhibits intracellular NF-κB activation, is constituted by a unique membrane-permeable sequence (CB5005M) cascading to a NF-κB nuclear localization sequence (CB5005N). In vitro cellular evaluation confirmed that CB5005 was effectively taken up by brain capillary endothelial cell bEnd.3 and glioma cells U87. The intracellular localization analysis further demonstrated that CB5005 could not only penetrate into the cells but also enter into their nuclei. More interestingly, CB5005 permeated deeply into the tumor spheroids of U87 cell. In vivo imaging illustrated that the fluorescence-labeled CB5005 distributed itself into the brain and accumulated at the tumor site after intravenous injection. Given the important role of over expressed NF-κB in tumor growth and development, we further investigated CB5005 for its potential in treatment of glioma. When combined administration in vitro with doxorubicin (DOX), CB5005 exhibited a synergistic effect in killing U87 cells. In a nude mice xenograft model, CB5005 inhibited the growth of tumor when applied alone, and displayed a synergistic anti-tumor effect with DOX. In conclusion, CB5005 functioned simultaneously as a cell penetrating peptide and a tumor growth inhibitor, therefore can work as a potential synergist for chemotherapy of human tumor. Clinical application of cell-penetrating peptides in cancer therapy is restricted due to lack of tissue selectivity and tumor-targeting ability. In this manuscript, we reported a rationally designed peptide, named CB5005, which had an attractive capability of translocation into the cell nucleus and blocking nuclear translocation of endogenous NF-κB protein. CB5005 had unique affinity with brain and glioma, and could rapidly accumulate in these tissues after intravenous injection. Furthermore, CB5005 showed a synergistic effect on inhibiting gliomas when administrated with doxorubicin. This is the first literature report on this multi-functionalized peptide, which can work as a potential synergist for chemotherapy of tumor. This work should be of general interest to scientists in the fields of biomaterials, biology, pharmacy, and oncology. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

The neuro-steroid, 3β androstene 17α diol exhibits potent cytotoxic effects on human malignant glioma and lymphoma cells through different programmed cell death pathways

PubMed Central

Graf, M R; Jia, W; Loria, R M

2007-01-01

The neuro-steroids 3β-androstene-17α-diol (17α-AED), 3β-androstene-17β-diol (17β-AED), 3β-androstene-7α,-17β-triol (7α-AET) and 3β-androstene-7β,-17β-triol (7β-AET) are metabolites of dehydroepiandrosterone and are produced in neuro-ectodermal tissue. Both epimers of androstenediols (17α-AED and 17β-AED) and androstenetriols (7α-AET and 7β-AET) have markedly different biological functions of their chemical analogue. We investigated the cytotoxic activity of these neuro-steroids on human T98G and U251MG glioblastoma and U937 lymphoma cells. Proliferation studies showed that 17α-AED is the most potent inhibitor, with an IC50 ∼15 μM. For T98G glioma, 90% inhibition was achieved with 25 μM of 17α-AED. Other neuro-steroids tested only marginally suppressed cell proliferation. Reduced cell adherence and viability could be detected after 18 h of 17α-AED exposure. Treatment with 17α-AED induced a significant level of apoptosis in U937 lymphoma cells, but not in the glioma cells. Cytopathology of 17α-AED-treated T98G cells revealed the presence of multiple cytoplasmic vacuoles. Acridine orange staining demonstrated the formation of acidic vesicular organelles in 17α-AED-treated T98G and U251MG, which was inhibited by bafilomycin A1. These findings indicate that 17α-AED bears the most potent cytotoxic activity of the neuro-steroids tested, and the effectiveness may depend on the number of hydroxyls and their position on the androstene molecule. These cytotoxic effects may utilize a non-apoptotic pathway in malignant glioma cells. PMID:17637679

Antitumor efficacy profile of PKI-402, a dual phosphatidylinositol 3-kinase/mammalian target of rapamycin inhibitor.

PubMed

Mallon, Robert; Hollander, Irwin; Feldberg, Larry; Lucas, Judy; Soloveva, Veronica; Venkatesan, Aranapakam; Dehnhardt, Christoph; Delos Santos, Efren; Chen, Zecheng; Dos Santos, Osvaldo; Ayral-Kaloustian, Semiramis; Gibbons, Jay

2010-04-01

PKI-402 is a selective, reversible, ATP-competitive, equipotent inhibitor of class I phosphatidylinositol 3-kinases (PI3K), including PI3K-alpha mutants, and mammalian target of rapamycin (mTOR; IC(50) versus PI3K-alpha = 2 nmol/L). PKI-402 inhibited growth of human tumor cell lines derived from breast, brain (glioma), pancreas, and non-small cell lung cancer tissue and suppressed phosphorylation of PI3K and mTOR effector proteins (e.g., Akt at T308) at concentrations that matched those that inhibited cell growth. In MDA-MB-361 [breast: Her2(+) and PIK3CA mutant (E545K)], 30 nmol/L PKI-402 induced cleaved poly(ADP-ribose) polymerase (PARP), a marker for apoptosis. In vivo, PKI-402 inhibited tumor growth in MDA-MB-361, glioma (U87MG), and lung (A549) xenograft models. In MDA-MB-361, PKI-402 at 100 mg/kg (daily for 5 days, one round) reduced initial tumor volume of 260 mm(3) to 129 mm(3) and prevented tumor regrowth for 70 days. In MDA-MB-361 tumors, PKI-402 (100 mg/kg, single dose) suppressed Akt phosphorylation (at T308) and induced cleaved PARP. Suppression of phosphorylated Akt (p-Akt) was complete at 8 hours and still evident at 24 hours. Cleaved PARP was evident at 8 and 24 hours. In normal tissue (heart and lung), PKI-402 (100 mg/kg) had minimal effect on p-Akt, with no detectable cleaved PARP. Preferential accumulation of PKI-402 in tumor tissue was observed. Complete, sustained suppression of Akt phosphorylation may cause tumor regression in MDA-MB-361 and other xenograft models. We are testing whether dual PI3K/mTOR inhibitors can durably suppress p-Akt, induce cleaved PARP, and cause tumor regression in a diverse set of human tumor xenograft models. Mol Cancer Ther; 9(4); 976-84. (c)2010 AACR.

A long noncoding RNA UCA1 promotes proliferation and predicts poor prognosis in glioma.

PubMed

Zhao, W; Sun, C; Cui, Z

2017-06-01

Acting as a proto-oncogene, long noncoding RNAs (lncRNAs) urothelial carcinoembryonic antigen 1 (UCA1) plays a key role in the occurrence and development of several human tumors. However, the expression and biological functions of UCA1 in glioma are less known. This study discussed the expression of UCA1 in glioma and its effect on the proliferation and cell cycle of glioma cells. LncRNA UCA1 expressions in 64 glioma samples (Grade I-II in 22 cases and Grade III-IV in 42 cases, according to WHO criteria) and 10 normal brain samples were detected using real-time fluorescence quantitative PCR. On this basis, the correlations of UCA1 to clinicopathological characteristics and prognosis of glioma were assessed. Then, using qPCR, the lncRNA UCA1 expressions in glioma cell lines and astrocytes were detected. UCA1-overexpressing glioma cell lines U87 and U251 were further detected after siRNA transfection of these two cell lines, and the impact on cell proliferation and cell cycle was assessed with CCK-8 (cell counting kit-8) assay and flow cytometry method (FCM), respectively. The expression of cyclin D1, a cell cycle-related protein, was detected using Western Blot. LncRNA UCA1 expression in the glioma samples was obviously higher as compared with the normal brain samples (P < 0.001), and the expression was correlated significantly with grading of the tumors (P < 0.05). However, lncRNA UCA1 expression was not correlated with age, gender, tumor size and KPS score (P > 0.05). After interference of UCA1 expression by siRNA transfection, the proliferation of both U251 and SHG-44 cells was inhibited (P < 0.05), with more cells arrested in G0/G1 (P < 0.05). Moreover, cyclin D1 expression was also downregulated considerably. LncRNA UCA1 can promote the proliferation and cell cycle progression of glioma cells by upregulating cyclin D1 transcription. So UCA1 may serve as an independent prognostic indicator and a novel therapeutic target for glioma.

Triacetin-based acetate supplementation as a chemotherapeutic adjuvant therapy in glioma.

PubMed

Tsen, Andrew R; Long, Patrick M; Driscoll, Heather E; Davies, Matthew T; Teasdale, Benjamin A; Penar, Paul L; Pendlebury, William W; Spees, Jeffrey L; Lawler, Sean E; Viapiano, Mariano S; Jaworski, Diane M

2014-03-15

Cancer is associated with epigenetic (i.e., histone hypoacetylation) and metabolic (i.e., aerobic glycolysis) alterations. Levels of N-acetyl-L-aspartate (NAA), the primary storage form of acetate in the brain, and aspartoacylase (ASPA), the enzyme responsible for NAA catalysis to generate acetate, are reduced in glioma; yet, few studies have investigated acetate as a potential therapeutic agent. This preclinical study sought to test the efficacy of the food additive Triacetin (glyceryl triacetate, GTA) as a novel therapy to increase acetate bioavailability in glioma cells. The growth-inhibitory effects of GTA, compared to the histone deacetylase inhibitor Vorinostat (SAHA), were assessed in established human glioma cell lines (HOG and Hs683 oligodendroglioma, U87 and U251 glioblastoma) and primary tumor-derived glioma stem-like cells (GSCs), relative to an oligodendrocyte progenitor line (Oli-Neu), normal astrocytes, and neural stem cells (NSCs) in vitro. GTA was also tested as a chemotherapeutic adjuvant with temozolomide (TMZ) in orthotopically grafted GSCs. GTA-induced cytostatic growth arrest in vitro comparable to Vorinostat, but, unlike Vorinostat, GTA did not alter astrocyte growth and promoted NSC expansion. GTA alone increased survival of mice engrafted with glioblastoma GSCs and potentiated TMZ to extend survival longer than TMZ alone. GTA was most effective on GSCs with a mesenchymal cell phenotype. Given that GTA has been chronically administered safely to infants with Canavan disease, a leukodystrophy due to ASPA mutation, GTA-mediated acetate supplementation may provide a novel, safe chemotherapeutic adjuvant to reduce the growth of glioma tumors, most notably the more rapidly proliferating, glycolytic and hypoacetylated mesenchymal glioma tumors. © 2013 UICC.

Triacetin-based acetate supplementation as a chemotherapeutic adjuvant therapy in glioma

PubMed Central

Tsen, Andrew R.; Long, Patrick M.; Driscoll, Heather E.; Davies, Matthew T.; Teasdale, Benjamin A.; Penar, Paul L.; Pendlebury, William W.; Spees, Jeffrey L.; Lawler, Sean E.; Viapiano, Mariano S.; Jaworski, Diane M.

2013-01-01

Cancer is associated with epigenetic (i.e., histone hypoacetylation) and metabolic (i.e., aerobic glycolysis) alterations. Levels of N-acetyl-L-aspartate (NAA), the primary storage form of acetate in the brain, and aspartoacylase (ASPA), the enzyme responsible for NAA catalysis to generate acetate, are reduced in glioma; yet, few studies have investigated acetate as a potential therapeutic agent. This preclinical study sought to test the efficacy of the food additive Triacetin (glyceryl triacetate, GTA) as a novel therapy to increase acetate bioavailability in glioma cells. The growth-inhibitory effects of GTA, compared to the histone deacetylase inhibitor Vorinostat (SAHA), were assessed in established human glioma cell lines (HOG and Hs683 oligodendroglioma, U87 and U251 glioblastoma) and primary tumor-derived glioma stem-like cells (GSCs), relative to an oligodendrocyte progenitor line (Oli-Neu), normal astrocytes, and neural stem cells (NSCs) in vitro. GTA was also tested as a chemotherapeutic adjuvant with temozolomide (TMZ) in orthotopically grafted GSCs. GTA induced cytostatic growth arrest in vitro comparable to Vorinostat, but, unlike Vorinostat, GTA did not alter astrocyte growth and promoted NSC expansion. GTA alone increased survival of mice engrafted with glioblastoma GSCs and potentiated TMZ to extend survival longer than TMZ alone. GTA was most effective on GSCs with a mesenchymal cell phenotype. Given that GTA has been chronically administered safely to infants with Canavan disease, a leukodystrophy due to ASPA mutation, GTA-mediated acetate supplementation may provide a novel, safe chemotherapeutic adjuvant to reduce the growth of glioma tumors, most notably the more rapidly proliferating, glycolytic, and hypoacetylated mesenchymal glioma tumors. PMID:23996800

Effect of hypoxia on the expression of nuclear genes encoding mitochondrial proteins in U87 glioma cells.

PubMed

Minchenko, O H; Riabovol, O O; Tsymbal, D O; Minchenko, D O; Ratushna, O O

2016-01-01

We have studied the effect of hypoxia on the expression of nuclear genes encoding mitochondrial proteins in U87 glioma cells under the inhibition of IRE1 (inositol requiring enzyme-1), which controls cell proliferation and tumor growth as a central mediator of endoplasmic reticulum stress. It was shown that hypoxia down-regulated gene expression of malate dehydrogenase 2 (MDH2), malic enzyme 2 (ME2), mitochondrial aspartate aminotransferase (GOT2), and subunit B of succinate dehydrogenase (SDHB) in control (transfected by empty vector) glioma cells in a gene specific manner. At the same time, the expression level of mitochondrial NADP+-dependent isocitrate dehydrogenase 2 (IDH2) and subunit D of succinate dehydrogenase (SDHD) genes in these cells does not significantly change in hypoxic conditions. It was also shown that the inhibition of ІRE1 signaling enzyme function in U87 glioma cells decreases the effect of hypoxia on the expression of ME2, GOT2, and SDHB genes and introduces the sensitivity of IDH2 gene to hypoxia. Furthermore, the expression of all studied genes depends on IRE1-mediated endoplasmic reticulum stress signaling in gene specific manner, because ІRE1 knockdown significantly decreases their expression in normoxic conditions, except for IDH2 gene, which expression level is strongly up-regulated. Therefore, changes in the expression level of nuclear genes encoding ME2, MDH2, IDH2, SDHB, SDHD, and GOT2 proteins possibly reflect metabolic reprogramming of mitochondria by hypoxia and IRE1-mediated endoplasmic reticulum stress signaling and correlate with suppression of glioma cell proliferation under inhibition of the IRE1 enzyme function.

Inhibition of STAT3 and ErbB2 Suppresses Tumor Growth, Enhances Radiosensitivity, and Induces Mitochondria-Dependent Apoptosis in Glioma Cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gao Ling; Li Fengsheng; Dong Bo

2010-07-15

Purpose: Constitutively activated signal transducer and activator of transcription 3 (STAT3) and ErbB2 are involved in the pathogenesis of many tumors, including astrocytoma. Inactivation of these molecules is reported to result in radiosensitization. The purpose of this study was to investigate whether inhibition of STAT3, ErbB2, or both could enhance radiotherapy in the human glioma model (U251 and U87 cell lines). Methods and Materials: The RNAi plasmids targeting STAT3 or ErbB2 were constructed, and their downregulatory effects on target proteins were examined by immunoblotting. After combination treatment of RNAi with or without irradiation, the cell viability was determined using 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazoliummore » bromide (MTT) and clonogenic assays. The in vivo effect of combined treatment was determined using the U251 xenograft model. The apoptosis caused by the inhibition of STAT3 and ErbB2 was detected, and the mechanism involved in the apoptosis was investigated, including increases in caspase proteins, mitochondrial damage, and the expression of key modulating protein of different apoptosis pathways. Results: Transfection of U251 cells with STAT3 or ErbB2 siRNA plasmids specifically reduced their target gene expressions. Inhibition of STAT3 or ErbB2 greatly decreased glioma cell survival after 2, 4, or 6 Gy irradiation. Inhibition of STAT3 and ErbB2 also enhanced radiation-induced tumor growth inhibition in the U251 xenograft model. Furthermore, the suppression of either STAT3 or ErbB2 could induce U251 cell apoptosis, which was related primarily to the mitochondrial apoptotic pathway. Conclusions: These results indicated that simultaneous inhibition of STAT3 and ErbB2 expression can promote potent antitumor activity and radiosensitizing activity in human glioma.« less

Wnt/β-catenin signaling pathway inhibits the proliferation and apoptosis of U87 glioma cells via different mechanisms

PubMed Central

Gao, Liyang; Chen, Bing; Li, Jinhong; Yang, Fan; Cen, Xuecheng; Liao, Zhuangbing; Long, Xiao’ao

2017-01-01

The Wnt signaling pathway is necessary for the development of the central nervous system and is associated with tumorigenesis in various cancers. However, the mechanism of the Wnt signaling pathway in glioma cells has yet to be elucidated. Small-molecule Wnt modulators such as ICG-001 and AZD2858 were used to inhibit and stimulate the Wnt/β-catenin signaling pathway. Techniques including cell proliferation assay, colony formation assay, Matrigel cell invasion assay, cell cycle assay and Genechip microarray were used. Gene Ontology Enrichment Analysis and Gene Set Enrichment Analysis have enriched many biological processes and signaling pathways. Both the inhibiting and stimulating Wnt/β-catenin signaling pathways could influence the cell cycle, moreover, reduce the proliferation and survival of U87 glioma cells. However, Affymetrix expression microarray indicated that biological processes and networks of signaling pathways between stimulating and inhibiting the Wnt/β-catenin signaling pathway largely differ. We propose that Wnt/β-catenin signaling pathway might prove to be a valuable therapeutic target for glioma. PMID:28837560

Nutraceutical phycocyanin nanoformulation for efficient drug delivery of paclitaxel in human glioblastoma U87MG cell line

NASA Astrophysics Data System (ADS)

Agrawal, Madhunika; Yadav, Sanjeev Kumar; Agrawal, Satyam Kumar; Karmakar, Surajit

2017-08-01

To enhance the therapeutic efficacy of chemotherapy on glioblastoma U87MG cell line, paclitaxel-loaded phycocyanin nanoparticles (PTX-PcNPs) were prepared by modified desolvation process. PTX-PcNPs were characterised in terms of size, zeta potential, drug loading efficiency and drug release. Confocal laser scanning microscopy showed PTX-PcNPs could be internalised by U87MG cells. The anti-cancer activity was investigated in vitro by 3-(4,5-dimethylthizol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay with and without photodynamic therapy. It was observed that formulation could significantly inhibit growth of U87MG cells as compared to PTX alone and also induced apoptosis, which was evidenced by presence of apoptotic bodies and nuclear fragmentation in treated cells. The present study suggests that PTX-PcNPs can act as a promising drug delivery system for cancer treatment. [Figure not available: see fulltext.

PKC-ι promotes glioblastoma cell survival by phosphorylating and inhibiting BAD through a phosphatidylinositol 3-kinase pathway.

PubMed

Desai, S; Pillai, P; Win-Piazza, H; Acevedo-Duncan, M

2011-06-01

The focus of this research was to investigate the role of protein kinase C-iota (PKC-ι) in regulation of Bad, a pro-apoptotic BH3-only molecule of the Bcl-2 family in glioblastoma. Robust expression of PKC-ι is a hallmark of human glioma and benign and malignant meningiomas. The results were obtained from the two human glial tumor derived cell lines, T98G and U87MG. In these cells, PKC-ι co-localized and directly associated with Bad, as shown by immunofluorescence, immunoprecipitation, and Western blotting. Furthermore, in-vitro kinase activity assay showed that PKC-ι directly phosphorylated Bad at phospho specific residues, Ser-112, Ser-136 and Ser-155 which in turn induced inactivation of Bad and disruption of Bad/Bcl-XL dimer. Knockdown of PKC-ι by siRNA exhibited a corresponding reduction in Bad phosphorylation suggesting that PKC-ι may be a Bad kinase. PKC-ι knockdown also induced apoptosis in both the cell lines. Since, PKC-ι is an essential downstream mediator of the PI (3)-kinase, we hypothesize that glioma cell survival is mediated via a PI (3)-kinase/PDK1/PKC-ι/Bad pathway. Treatment with PI (3)-kinase inhibitors Wortmannin and LY294002, as well as PDK1 siRNA, inhibited PKC-ι activity and subsequent phosphorylation of Bad suggesting that PKC-ι regulates the activity of Bad in a PI (3)-kinase dependent manner. Thus, our data suggest that glioma cell survival occurs through a novel PI (3)-kinase/PDK1/PKC-ι/BAD mediated pathway. Published by Elsevier B.V.

Growth inhibition and chemosensitization of exogenous nitric oxide released from NONOates in glioma cells in vitro.

PubMed

Weyerbrock, Astrid; Baumer, Brunhilde; Papazoglou, Anna

2009-01-01

Exogenous nitric oxide (NO) from NO donors has cytotoxic, chemosensitizing, and radiosensitizing effects, and increases vascular permeability and blood flow in tumors. Yet little is known about whether these cytotoxic and chemosensitizing effects can be observed in glioma cells at doses that alter tumor physiological characteristics in vivo and whether these effects are tumor selective. The effect of NO released from proline NONOate, diethylamine NONOate, spermine NONOate, and sodium nitrite on cell proliferation, apoptosis, and chemosensitivity to carboplatin of cultured glioma cells was studied in C6, U87 glioma cells, human glioblastoma cells, and human astrocytes and fibroblasts. Although proline NONOate failed to induce cell death, the other NO donors induced growth arrest when present in high concentrations (10(-2) M) in all cell lines. Chemosensitization was observed after concomitant incubation with spermine NONOate and carboplatin in C6 and human glioblastoma cells. There is strong evidence that cell death occurs primarily by necrosis and to a lesser degree by apoptosis. The NO doses, which altered tumor physiology in vivo, were not cytotoxic, indicating that NO alters vascular permeability and cell viability in vivo by different mechanisms. The authors found that NO-generating agents at high concentrations are potent growth inhibitors and might also be useful as chemosensitizers in glioma cells. These data corroborate the theory that the use of NOgenerating agents may play a role in the multimodal treatment of malignant gliomas but that the NO release must be targeted more specifically to tumor cells to improve selectivity and efficacy.

Pharmacokinetics and antitumor efficacy of DSPE-PEG2000 polymeric liposomes loaded with quercetin and temozolomide: Analysis of their effectiveness in enhancing the chemosensitization of drug-resistant glioma cells

PubMed Central

HU, JUN; WANG, JUNJIE; WANG, GANG; YAO, ZHONGJUN; DANG, XIAOQIAN

2016-01-01

In the present study, a new type of DSPE-PEG2000 polymeric liposome for the brain-targeted delivery of poorly water-soluble anticancer drugs was successfully prepared and characterized. The nanoparticles were formed by the self-assembly of an amphiphilic polymer consisting of hydrophilic 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000] (DSPE-PEG2000). These nanoliposomes served as a safe delivery platform for the simultaneous delivery of quercetin (QUE) and temozolomide (TMZ) to rat brains. The 2-in-1 PEG2000-DSPE nanoliposomes containing QUE and TMZ (QUE/TMZ-NLs) were rapidly taken up by the U87 glioma cells in vitro, whereas at the same concentrations, the amounts of the free drugs taken up were minimal. The QUE/TMZ-NLs showed an enhanced potency in the U87 cells and the TMZ-resistant U87 cells (U87/TR cells), possibly due to the high intracellular drug concentration and the subsequent drug release. In vivo biodistribution experiments revealed a significant accumulation of QUE/TMZ-NLs in the brain, with significantly increased plasma concentrations of QUE and TMZ, as well as delayed clearance in our rat model of glioma. The results were not so significant for the QUE-loaded nanoliposomes (QUE-NLs) and free TMZ. The findings of our study establish the DSPE-PEG2000 polymeric liposome as a novel and effective nanocarrier for enhancing drug delivery to brain tumors. PMID:26782731

Resveratrol Targets AKT and p53 in Glioblastoma and Glioblastoma Stem-like Cells to Suppress Growth and Infiltration

PubMed Central

Clark, Paul A.; Bhattacharya, Saswati; Elmayan, Ardem; Darjatmoko, Soesiawati R.; Thuro, Bradley A.; Yan, Michael B.; van Ginkel, Paul R.; Polans, Arthur S.; Kuo, John S.

2016-01-01

Object Glioblastoma multiforme (GBM) is an aggressive brain cancer with median survival of less than two years with current treatment. GBM exhibits extensive intra-tumor and inter-patient heterogeneity, suggesting that successful therapies should exert broad anti-cancer activities. Therefore, the natural non-toxic pleiotropic agent, resveratrol, was studied for anti-tumorigenic effects against GBM. Methods Resveratrol’s effects on cell proliferation, sphere-forming ability, and invasion were tested using multiple patient-derived GBM stem-like cell (GSC) lines and established U87 glioma cells, and changes in oncogenic AKT and tumor suppressive p53 were analyzed. Resveratrol was also tested in vivo against U87 glioma flank xenografts using multiple delivery methods, including direct tumor injection. Finally, resveratrol was delivered directly to brain tissue to determine toxicity and achievable drug concentrations in the brain parenchyma. Results Resveratrol significantly inhibited proliferation in U87 glioma and multiple patient-derived GSC lines, demonstrating similar inhibitory concentrations across these phenotypically heterogeneous lines. Resveratrol also inhibited the sphere-forming ability of GSCs, suggesting anti-stem cell effects. Additionally, resveratrol blocked U87 glioma and GSC invasion in an in vitro Matrigel transwell assay at doses similar to those mediating anti-proliferative effects. In U87 glioma cells and GSCs, resveratrol reduced AKT phosphorylation and induced p53 expression and activation that led to transcription of downstream p53 target genes. Resveratrol administration via oral gavage or ad libitum in the water supply significantly suppressed GBM xenograft growth; intra-tumor or peri-tumor resveratrol injection further suppressed growth and approximating tumor regression. Intracranial resveratrol injection resulted in 100-fold higher local drug concentration compared to intravenous delivery, and with no apparent toxicity. Conclusions Resveratrol potently inhibited GBM and GBM stem-like cell growth and infiltration, acting partially via AKT deactivation and p53 induction, and suppressed glioblastoma growth in vivo. The ability of resveratrol to modulate AKT and p53, as well as reportedly many other anti-tumorigenic pathways, is attractive for therapy against a genetically heterogeneous tumor such as GBM. Although resveratrol exhibits low bioavailability when administered orally or intravenously, novel delivery methods such as direct injection (i.e. convection enhanced delivery) could potentially be used to achieve and maintain therapeutic doses in brain. Resveratrol’s non-toxic nature and broad anti-GBM effects make it a compelling candidate to supplement current GBM therapies. PMID:27419830

MiR-18a regulates the proliferation, migration and invasion of human glioblastoma cell by targeting neogenin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Song, Yichen, E-mail: jeff200064017@163.com; Wang, Ping, E-mail: pingwang8000@163.com; Institute of Pathology and Pathophysiology, China Medical University, Shenyang 110001

MiR-17-92 cluster has recently been reported as an oncogene in some tumors. However, the association of miR-18a, an important member of this cluster, with glioblastoma remains unknown. Therefore, this study aims to investigate the expression of miR-18a in glioblastoma and its role in biological behavior of U87 and U251 human glioblastoma cell lines. Quantitative RT-PCR results showed that miR-18a was highly expressed in glioblastoma tissues and U87 and U251 cell lines compared with that in human brain tissues and primary normal human astrocytes, and the expression levels were increased along with the rising pathological grades of glioblastoma. Neogenin was identifiedmore » as the target gene of miR-18a by dual-luciferase reporter assays. RT-PCR and western blot results showed that its expression levels were decreased along with the rising pathological grades of glioblastoma. Inhibition of miR-18a expression was established by transfecting exogenous miR-18a inhibitor into U87 and U251 cells, and its effects on the biological behavior of glioblastoma cells were studied using CCK-8 assay, transwell assay and flow cytometry. Inhibition of miR-18a expression in U87 and U251 cells significantly up-regulated neogenin, and dramatically suppressed the abilities of cell proliferation, migration and invasion, induced cell cycle arrest and promoted cellular apoptosis. Collectively, these results suggest that miR-18a may regulate biological behavior of human glioblastoma cells by targeting neogenin, and miR-18a can serve as a potential target in the treatment of glioblastoma. - Highlights: • MiR-18a was highly expressed in glioblastoma tissues and U87 and U251 cell lines. • Neogenin was identified as the target gene of miR-18a. • Neogenin expressions were decreased along with the rising pathological grades of glioblastoma. • Inhibition of miR-18a suppressed biological behavior of glioma cells by up-regulating neogenin.« less

Poly(ADP-ribose) polymerase-independent potentiation of nitrosourea cytotoxicity by 3-aminobenzamide in human malignant glioma cells.

PubMed

Winter, S; Weller, M

2000-06-16

Poly(ADP-ribose) polymerase is a zinc-finger DNA-binding protein that detects specifically DNA strand breaks generated by genotoxic agents and is thought to be involved in DNA repair. Here, we examined the effects of 3-aminobenzamide, a poly(ADP-ribose) polymerase inhibitor, on the chemosensitivity of human malignant glioma cells. 3-Aminobenzamide selectively potentiated the cytotoxicity of the nitrosoureas, nimustine, carmustine and lomustine in 10 of 12 human malignant glioma cell lines. In contrast, 3-aminobenzamide did not modulate the cytotoxic effects of doxorubicine, teniposide, vincristine, camptothecin or cytarabine. The nitrosoureas did not induce poly(ADP-ribose) polymerase activity in the glioma cells. Ectopic expression of truncated poly(ADP-ribose) polymerase containing the poly(ADP-ribose) polymerase DNA-binding domain, which acts as a dominant-negative mutant, in LN-18 or LN-229 cells did not alter the 3-aminobenzamide effect on nitrosourea-mediated cytotoxicity. Thus, 3-aminobenzamide may target another nicotinamide adenine dinucleotide (NAD)-requiring enzyme, but not poly(ADP-ribose) polymerase, when enhancing nitrosourea cytotoxicity in human malignant glioma cells. Carmustine cytotoxicity was associated with a G2/M arrest. Coexposure to carmustine and 3-aminobenzamide overcame this G2/M arrest in T98G cells, which are sensitized to carmustine by 3-aminobenzamide, but not in U251MG cells, which are refractory to 3-aminobenzamide-mediated sensitization to carmustine. Thus, 3-aminobenzamide-mediated sensitization to carmustine cytotoxicity may result from interference with the stable G2/M arrest response to carmustine in human glioma cells.

Epidermal Growth Factor Receptor Expression Modulates Antitumor Efficacy of Vandetanib or Cediranib Combined With Radiotherapy in Human Glioblastoma Xenografts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wachsberger, Phyllis R., E-mail: Phyllis.wachsberger@jeffersonhospital.org; Lawrence, Yaacov R.; Liu Yi

2012-01-01

Purpose: The purpose of this study was to determine the ability of radiation therapy (RT) combined with the tyrosine kinase inhibitors (TKI) vandetanib (antiepidermal growth factor receptor [EGFR] plus antivascular endothelial growth factor receptor [anti-VEGFR]) and cediranib (anti-VEGFR) to inhibit glioblastoma multiforme (GBM) growth. A secondary aim was to investigate how this regimen is modulated by tumor EGFR expression. Methods and Materials: Radiosensitivity was assessed by clonogenic cell survival assay. VEGF secretion was quantified by enzyme-linked immunosorbent assay. GBM (U87MG wild-type EGFR [wtEGFR] and U87MG EGFR-null) xenografts were treated with vandetanib, cediranib, and RT, alone or in combinations. Excised tumormore » sections were stained for proliferative and survival biomarkers. Results: In vitro, U87MG wtEGFR and U87 EGFR-null cells had similar growth kinetics. Neither TKI affected clonogenic cell survival following RT. However, in vivo, exogenous overexpression of wtEGFR decreased tumor doubling time (T2x) in U87MG xenografts (2.70 vs. 4.41 days for U87MG wtEGFR vs. U87MG vector, respectively). In U87MG EGFR-null cells, TKI combined with radiation was no better than radiation therapy alone. In U87MG wtEGFR, RT in combination with vandetanib (but not with cediranib) significantly increased tumor T2x compared with RT alone (T2x, 10.4 days vs. 4.8 days; p < 0.001). In vivo, growth delay correlated with suppression of pAkt, survivin, and Ki67 expression in tumor samples. The presence of EGFR augmented RT-stimulated VEGF release; this effect was inhibited by vandetanib. Conclusions: EGFR expression promoted tumor growth in vivo but not in vitro, suggesting a microenvironmental effect. GBM xenografts expressing EGFR exhibited greater sensitivity to both cediranib and vandetanib than EGFR-null tumors. Hence EGFR status plays a major role in determining a tumor's in vivo response to radiation combined with TKI, supporting a 'personalized' approach to GBM management.« less

Anti-SEMA3A Antibody: A Novel Therapeutic Agent to Suppress GBM Tumor Growth.

PubMed

Lee, Jaehyun; Shin, Yong Jae; Lee, Kyoungmin; Cho, Hee Jin; Sa, Jason K; Lee, Sang-Yun; Kim, Seok-Hyung; Lee, Jeongwu; Yoon, Yeup; Nam, Do-Hyun

2017-11-10

Glioblastoma (GBM) is classified as one of the most aggressive and lethal brain tumor. Great strides have been made in understanding the genomic and molecular underpinnings of GBM, which translated into development of new therapeutic approaches to combat such deadly disease. However, there are only few therapeutic agents that can effectively inhibit GBM invasion in a clinical framework. In an effort to address such challenges, we have generated anti-SEMA3A monoclonal antibody as a potential therapeutic antibody against GBM progression. We employed public glioma datasets, Repository of Molecular Brain Neoplasia Data and The Cancer Genome Atlas, to analyze SEMA3A mRNA expression in human GBM specimens. We also evaluated for protein expression level of SEMA3A via tissue microarray (TMA) analysis. Cell migration and proliferation kinetics were assessed in various GBM patient-derived cells (PDCs) and U87-MG cell-line for SEMA3A antibody efficacy. GBM patient-derived xenograft (PDX) models were generated to evaluate tumor inhibitory effect of anti-SEMA3A antibody in vivo. By combining bioinformatics and TMA analysis, we discovered that SEMA3A is highly expressed in human GBM specimens compared to non-neoplastic tissues. We developed three different anti-SEMA3A antibodies, in fully human IgG form, through screening phage-displayed synthetic antibody library using a classical panning method. Neutralization of SEMA3A significantly reduced migration and proliferation capabilities of PDCs and U87-MG cell-line in vitro. In PDX models, treatment with anti-SEMA3A antibody exhibited notable tumor inhibitory effect through down-regulation of cellular proliferative kinetics and tumor-associated macrophages recruitment. In present study, we demonstrated tumor inhibitory effect of SEMA3A antibody in GBM progression and present its potential relevance as a therapeutic agent in a clinical framework.

Preparation and characterization of teniposide PLGA nanoparticles and their uptake in human glioblastoma U87MG cells.

PubMed

Mo, Liqian; Hou, Lianbing; Guo, Dan; Xiao, Xiaoyan; Mao, Ping; Yang, Xixiao

2012-10-15

Many studies have demonstrated the uptake mechanisms of various nanoparticle delivery systems with different physicochemical properties in different cells. In this study, we report for the first time the preparation and characterization of teniposide (VM-26) poly(D,L-lactide-co-glycolide) (PLGA) nanoparticles (NPs) and their cellular uptake pathways in human glioblastoma U87MG cells. The nanoparticles prepared with oil-in-water (O/W) single-emulsion solvent evaporation method had a small particle size and spherical shape and provided effective protection against degradation of teniposide in PBS solution. Differential scanning calorimeter (DSC) thermograms concluded that VM-26 was dispersed as amorphous or disordered crystalline phase in the PLGA matrix. A cytotoxicity study revealed that, in a 24h period, blank PLGA NPs had no cytotoxicity, whereas teniposide-loaded PLGA NPs (VM-26-NPs) had U87MG cytotoxicity levels similar to free teniposide. Confocal laser scanning microscopy (CLSM) and transmission electron microscopy (TEM) images showed the distribution and degradation processes of nanoparticles in cells. An endocytosis inhibition test indicated that clathrin-mediated endocytosis and macropinocytosis were the primary modes of engulfment involved in the internalization of VM-26-NPs. Our findings suggest that PLGA nanoparticles containing a sustained release formula of teniposide may multiplex the therapeutic effect and ultimately degrade in lysosomal within human glioblastoma U87MG cells. Copyright © 2012 Elsevier B.V. All rights reserved.

Novel model of orthotopic U-87 MG glioblastoma resection in athymic nude mice.

PubMed

Bianco, John; Bastiancich, Chiara; Joudiou, Nicolas; Gallez, Bernard; des Rieux, Anne; Danhier, Fabienne

2017-06-01

In vitro and in vivo models of experimental glioma are useful tools to gain a better understanding of glioblastoma (GBM) and to investigate novel treatment strategies. However, the majority of preclinical models focus on treating solid intracranial tumours, despite surgical resection being the mainstay in the standard care of patients with GBM today. The lack of resection and recurrence models therefore has undermined efforts in finding a treatment for this disease. Here we present a novel orthotopic tumour resection and recurrence model that has potential for the investigation of local delivery strategies in the treatment of GBM. The model presented is simple to achieve through the use of a biopsy punch, is reproducible, does not require specific or expensive equipment, and results in a resection cavity suitable for local drug delivery systems, such as the implantation or injection of hydrogels. We show that tumour resection is well tolerated, does not induce deleterious neurological deficits, and significantly prolongs survival of mice bearing U-87 MG GBM tumours. In addition, the resulting cavity could accommodate adequate amounts of hydrogels for local delivery of chemotherapeutic agents to eliminate residual tumour cells that can induce tumour recurrence. Copyright © 2017 Elsevier B.V. All rights reserved.

Desmethylanhydroicaritin isolated from Sophora flavescens, shows antitumor activities in U87MG cells via inhibiting the proliferation, migration and invasion.

PubMed

Kang, Chang-Won; Kim, Nan-Hee; Jung, Huyn Ah; Choi, Hyung-Wook; Kang, Min-Jae; Choi, Jae-Sue; Kim, Gun-Do

2016-04-01

This study is the first report of the antitumor activities of desmethylanhydroicaritin (DMAI) isolated from Sophora flavescens on U87MG cells. Human glioblastoma is one of the most aggressive malignant type of brain tumors and highly diffuses to around normal brain tissues. DMAI showed anti-proliferation effects on U87MG cells at the concentration of 30μM, however did not affect to HEK-293 cells. DMAI induced anti-proliferation effects via ERK/MAPK, PI3K/Akt/mTOR signal pathway and G2/M phase cell cycle arrest. DMAI led to morphological change and inhibition of filapodia formation through regulation of Rac 1 and Cdc 42. In addition, migration and invasion of U87MG cells were inhibited by DMAI via down-regulation of matrix metalloproteinase (MMP) -2 and MMP -9 expressions and activities. Our results suggest that DMAI has a potential as a therapeutic agent against glioblastoma cells. Copyright © 2016 Elsevier B.V. All rights reserved.

The role of drebrin in glioma migration and invasion

DOE Office of Scientific and Technical Information (OSTI.GOV)

Terakawa, Yuzo; Department of Neurosurgery, Osaka City University Graduate School of Medicine, Osaka; Agnihotri, Sameer

Glioblastoma (GBM) is the most common primary brain tumor in adults. Despite current advances in therapy consisting of surgery followed by chemotherapy and radiation, the overall survival rate still remains poor. Therapeutic failures are partly attributable to the highly infiltrative nature of tumor adjacent to normal brain parenchyma. Recently, evidence is mounting to suggest that actin cytoskeleton dynamics are critical components of the cell invasion process. Drebrin is an actin-binding protein involved in the regulation of actin filament organization, and plays a significant role in cell motility; however, the role of drebrin in glioma cell invasiveness has not yet beenmore » fully elucidated. Therefore, this study was aimed to clarify the role of drebrin in glioma cell morphology and cell motility. Here we show that drebrin is expressed in glioma cell lines and in operative specimens of GBM. We demonstrate that stable overexpression of drebrin in U87 cells leads to alterations in cell morphology, and induces increased invasiveness in vitro while knockdown of drebrin in U87 cells by small interfering RNA (siRNA) decreases invasion and migration. In addition, we show that depletion of drebrin by siRNA alters glioma cell morphology in A172 GBM cell line. Our results suggest that drebrin contributes to the maintenance of cell shape, and may play an important role in glioma cell motility. - Highlights: ► Drebrin is an actin-binding protein aberrantly expressed in several cancers. ► Role of drebrin in glioma cell morphology and motility is previously unknown. ► We demonstrate that drebrin is expressed in 40% of glioblastoma specimens. ► Drebrin plays a significant role in modulating glioma cell migration and invasion.« less

TTBK2 circular RNA promotes glioma malignancy by regulating miR-217/HNF1β/Derlin-1 pathway.

PubMed

Zheng, Jian; Liu, Xiaobai; Xue, Yixue; Gong, Wei; Ma, Jun; Xi, Zhuo; Que, Zhongyou; Liu, Yunhui

2017-02-20

Circular RNAs are a subgroup of non-coding RNAs and generated by a mammalian genome. Herein, the expression and function of circular RNA circ-TTBK2 were investigated in human glioma cells. Fluorescence in situ hybridization and quantitative real-time PCR were conducted to profile the cell distribution and expression of circ-TTBK2 and microRNA-217 (miR-217) in glioma tissues and cells. Immunohistochemical and western blot were used to determine the expression of HNF1β and Derlin-1 in glioma tissues and cells. Stable knockdown of circ-TTBK2 or overexpression of miR-217 glioma cell lines (U87 and U251) were established to explore the function of circ-TTBK2 and miR-217 in glioma cells. Further, luciferase reports and RNA immunoprecipitation were used to investigate the correlation between circ-TTBK2 and miR-217. Cell Counting Kit-8, transwell assays, and flow cytometry were used to investigate circ-TTBK2 and miR-217 function including cell proliferation, migration and invasion, and apoptosis, respectively. ChIP assays were used to ascertain the correlations between HNF1β and Derlin-1. We found that circ-TTBK2 was upregulated in glioma tissues and cell lines, while linear TTBK2 was not dysregulated in glioma tissues and cells. Enhanced expression of circ-TTBK2 promoted cell proliferation, migration, and invasion, while inhibited apoptosis. MiR-217 was downregulated in glioma tissues and cell lines. We also found that circ-TTBK2, but not linear TTBK2, acted as miR-217 sponge in a sequence-specific manner. In addition, upregulated circ-TTBK2 decreased miR-217 expression and there was a reciprocal negative feedback between them in an Argonaute2-dependent manner. Moreover, reintroduction of miR-217 significantly reversed circ-TTBK2-mediated promotion of glioma progression. HNF1β was a direct target of miR-217, and played oncogenic role in glioma cells. Remarkably, circ-TTBK2 knockdown combined with miR-217 overexpression led to tumor regression in vivo. These results demonstrated a novel role circ-TTBK2 in the glioma progression.

Identification of a novel fusion gene HMGA2-EGFR in glioblastoma.

PubMed

Komuro, Akiyoshi; Raja, Erna; Iwata, Caname; Soda, Manabu; Isogaya, Kazunobu; Yuki, Keiko; Ino, Yasushi; Morikawa, Masato; Todo, Tomoki; Aburatani, Hiroyuki; Suzuki, Hiromichi; Ranjit, Melissa; Natsume, Atsushi; Mukasa, Akitake; Saito, Nobuhito; Okada, Hitoshi; Mano, Hiroyuki; Miyazono, Kohei; Koinuma, Daizo

2018-04-15

Glioblastoma is one of the most malignant forms of cancer, for which no effective targeted therapy has been found. Although The Cancer Genome Atlas has provided a list of fusion genes in glioblastoma, their role in progression of glioblastoma remains largely unknown. To search for novel fusion genes, we obtained RNA-seq data from TGS-01 human glioma-initiating cells, and identified a novel fusion gene (HMGA2-EGFR), encoding a protein comprising the N-terminal region of the high-mobility group AT-hook protein 2 (HMGA2) fused to the C-terminal region of epidermal growth factor receptor (EGFR), which retained the transmembrane and kinase domains of the EGFR. This fusion gene product showed transforming potential and a high tumor-forming capacity in cell culture and in vivo. Mechanistically, HMGA2-EGFR constitutively induced a higher level of phosphorylated STAT5B than EGFRvIII, an in-frame exon deletion product of the EGFR gene that is commonly found in primary glioblastoma. Forced expression of HMGA2-EGFR enhanced orthotopic tumor formation of the U87MG human glioma cell line. Furthermore, the EGFR kinase inhibitor erlotinib blocked sphere formation of TGS-01 cells in culture and inhibited tumor formation in vivo. These findings suggest that, in addition to gene amplification and in-frame exon deletion, EGFR signaling can also be activated by gene fusion, suggesting a possible avenue for treatment of glioblastoma. © 2017 UICC.

[MACF1 knockdown in glioblastoma multiforme cells increases temozolomide-induced cytotoxicity].

PubMed

Xie, Si-di; Chen, Zi-Yang; Wang, Hai; He, Min-Yi; Lu, Yun-Tao; Lei, Bing-Xi; Li, He-Zhen; Liu, Ya-Wei; Qi, Song-Tao

2017-09-20

To investigate the role of microtubule-actin crosslinking factor 1 (MACF1) in the response of glioma cells to temozolomide (TMZ). TMZ was applied to a human gliomablastoma cell line (U87) and changes in the protein expression and cellular localization were determined with Western blot, RT-PCR, and immunofluorescence. The responses of the cells with MACF1 expression knockdown by RNA interference to TMZ were assessed. TMZ-induced effects on MACF1 expression were also assessed by immunohistochemistry in a nude mouse model bearing human glioblastoma xenografts. TMZ resulted in significantly increased MACF1 expression (by about 2 folds) and changes in its localization in the gliomablastoma cells both in vitro and in vivo (P<0.01). Knockdown of MACF1 reduced the proliferation (by 45%) of human glioma cell lines treated with TMZ (P<0.01). TMZ-induced changes in MACF1 expression was accompanied by cytoskeletal rearrangement. MACF1 may be a potential therapeutic target for glioblastoma.

IRE1 inhibition affects the expression of insulin-like growth factor binding protein genes and modifies its sensitivity to glucose deprivation in U87 glioma cells.

PubMed

Minchenko, D O; Kharkova, A P; Tsymbal, D O; Karbovskyi, L L; Minchenko, O H

2015-10-01

The aim of the present study was to investigate the effect of inhibition of endoplasmic reticulum stress signaling mediated by IRE1/ERN1 (inositol-requiring enzyme 1/endoplasmic reticulum to nucleus signaling 1) on the expression of genes encoding different groups of insulin-like growth binding proteins (IGFBP6 and IGFBP7) and CCN family (IGFBP8/CTGF/CCN2, IGFBP9/NOV/CCN3, IGFBP10/CYR61/CCN1, WISP1/CCN4, and WISP2/CCN5) and its sensitivity to glucose deprivation in U87 glioma cells. The expression of IGFBP6, IGFBP7, IGFBP8, IGFBP9, IGFBP10, WISP1, and WISP2 genes was studied by qPCR in control U87 glioma cells (wild-type) and its subline with IRE1 signaling enzyme loss of function upon glucose deprivation. The expression of IGFBP8, IGFBP9, and WISP2 genes was up-regulated in control glioma cells upon glucose deprivation with most significant changes for IGFBP9 gene. At the same time, the expression of IGFBP6, IGFBP10, and WISP1 genes was resistant to glucose deprivation in these glioma cells, but the IGFBP7 gene expression was down-regulated. The inhibition of both enzymatic activities (kinase and endoribonuclease) of IRE1 in glioma cells modified the sensitivity of most studied gene expressions to glucose deprivation condition: introduced sensitivity of IGFBP10 and WISP1 genes to glucose deprivation, enhanced the effect of this deprivation on IGFBP7 and IGFBP9 gene expressions, and reduced this effect on WISP2 gene and induced suppressive effect of glucose deprivation on the expression of IGFBP8 gene. Furthermore, the inhibition of IRE1 strongly affected the expression of all studied genes in glioma cells upon regular growing condition in gene specific manner: up-regulated the expression levels of IGFBP7, IGFBP8, IGFBP10, WISP1, and WISP2 genes and down-regulated the IGFBP6 and IGFBP9 genes. The data of this investigation demonstrate that the expression of IGFBP7, IGFBP8, IGFBP9, and WISP2 genes are sensitive to glucose deprivation in U87 glioma cells and that inhibition of IRE1 signaling enzyme function may significantly affect the expression of all studied genes in the presence of glucose as well as modify the effect of glucose deprivation on the expression of most studied genes. These data also show that proteins encoded by these genes may participate in the regulation of metabolic and proliferative processes via IGF/INS receptors and possibly other signaling pathways as well, via IRE1 signaling, which is a central mediator of the unfolded protein response and an important component of the tumor growth and metabolic diseases.

Boron neutron capture therapy induces apoptosis of glioma cells through Bcl-2/Bax

PubMed Central

2010-01-01

Background Boron neutron capture therapy (BNCT) is an alternative treatment modality for patients with glioma. The aim of this study was to determine whether induction of apoptosis contributes to the main therapeutic efficacy of BNCT and to compare the relative biological effect (RBE) of BNCT, γ-ray and reactor neutron irradiation. Methods The neutron beam was obtained from the Xi'an Pulsed Reactor (XAPR) and γ-rays were obtained from [60Co] γ source of the Fourth Military Medical University (FMMU) in China. Human glioma cells (the U87, U251, and SHG44 cell lines) were irradiated by neutron beams at the XAPR or [60Co] γ-rays at the FMMU with different protocols: Group A included control nonirradiated cells; Group B included cells treated with 4 Gy of [60Co] γ-rays; Group C included cells treated with 8 Gy of [60Co] γ-rays; Group D included cells treated with 4 Gy BPA (p-borono-phenylalanine)-BNCT; Group E included cells treated with 8 Gy BPA-BNCT; Group F included cells irradiated in the reactor for the same treatment period as used for Group D; Group G included cells irradiated in the reactor for the same treatment period as used for Group E; Group H included cells irradiated with 4 Gy in the reactor; and Group I included cells irradiated with 8 Gy in the reactor. Cell survival was determined using the 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium (MTT) cytotoxicity assay. The morphology of cells was detected by Hoechst33342 staining and transmission electron microscope (TEM). The apoptosis rate was detected by flow cytometer (FCM). The level of Bcl-2 and Bax protein was measured by western blot analysis. Results Proliferation of U87, U251, and SHG44 cells was much more strongly inhibited by BPA-BNCT than by irradiation with [60Co] γ-rays (P < 0.01). Nuclear condensation was determined using both a fluorescence technique and electron microscopy in all cell lines treated with BPA-BNCT. Furthermore, the cellular apoptotic rates in Group D and Group E treated with BPA-BNCT were significantly higher than those in Group B and Group C irradiated by [60Co] γ-rays (P < 0.01). The clonogenicity of glioma cells was reduced by BPA-BNCT compared with cells treated in the reactor (Group F, G, H, I), and with the control cells (P < 0.01). Upon BPA-BNCT treatment, the Bax level increased in glioma cells, whereas Bcl-2 expression decreased. Conclusions Compared with γ-ray and reactor neutron irradiation, a higher RBE can be achieved upon treatment of glioma cells with BNCT. Glioma cell apoptosis induced by BNCT may be related to activation of Bax and downregulation of Bcl-2. PMID:21122152

Hypoxia Inducible Factor 1 (HIF1) Activation in U87 Glioma Cells Involves a Decrease in Reactive Oxygen Species Production and Protein Kinase C Activity

DTIC Science & Technology

1998-06-29

Curcumin DFX Desferrioxamine DNA Deoxyribonucleic Acid DPI Diphenyliodinium DPPD Diphenylphenylenediamine DTH Dithionite EMSA Electrophoretic mobility shift... neuroprotective effects (Fern et al., 1996, Morishita et al., 1 1997). The identification of a hypoxia inducible transcription factor known as HIF-1 (Semenza...derived EPO in the eNS neuroprotective response to hypoxia. Cloning of the human and murine EPO gene, the availability of a convenient EPa producing

IDH1 R132H Mutation Enhances Cell Migration by Activating AKT-mTOR Signaling Pathway, but Sensitizes Cells to 5-FU Treatment as NADPH and GSH Are Reduced

PubMed Central

Qiu, Jiangdong; Huang, Keting; Wu, Mindan; Xia, Chunlin

2017-01-01

Aim of study Mutations of isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2) gene were recently discovered in vast majority of World Health Organization (WHO) grade II/III gliomas. This study is to understand the effects of IDH1 R132H mutation in gliomagenesis and to develop new strategies to treat glioma with IDH1 R132H mutation. Materials and methods Over expression of IDH1 R132H in U87MG cells was done by transfecting cells with IDH1 R132H plasmid. MTT assay, scratch repair assay and western blot were performed to study effects of IDH1 R132H mutation on cell proliferation, migration, regulating AKT-mTOR signaling pathway and cell death respectively. NADP+/NADPH and GSH quantification assays were performed to evaluate effects of IDH1 R132H mutation on the production of antioxidant NADPH and GSH. Results We found that over expression of IDH1 R132H mutation decreased cell proliferation consistent with previous reports; however, it increased cell migration and enhanced AKT-mTOR signaling pathway activation. Mutations in isocitrate dehydrogenase (IDH) 1 also change the function of the enzymes and cause them to produce 2-hydroxyglutarate and not produce NADPH. We tested the level of NADPH and GSH and demonstrated that IDH1 R132H mutant stable cells had significantly low NADPH and GSH level compared to control or IDH1 wild type stable cells. The reduced antioxidants (NADPH and GSH) sensitized U87MG cells with IDH R132H mutant to 5-FU treatment. Conclusion Our study highlights the important role of IHD1 R132H mutant in up- regulating AKT-mTOR signaling pathway and enhancing cell migration. Furthermore, we demonstrate that IDH1 R132H mutation affects cellular redox status and sensitizes gliomas cells with IDH1 R132H mutation to 5FU treatment. PMID:28052098

HER2-targeted recombinant protein immuno-caspase-6 effectively induces apoptosis in HER2-overexpressing GBM cells in vitro and in vivo.

PubMed

Zhang, Leiming; Ren, Junlin; Zhang, Hangyu; Cheng, Gang; Xu, Yanming; Yang, Shuangwu; Dong, Chao; Fang, Dandong; Zhang, Jianning; Yang, Angang

2016-11-01

Glioblastoma multiforme (GBM), which is associated with a high rate of morbidity and mortality, is among the most malignant and treatment-refractory neoplasms in human adults. As GBM is highly resistant to conventional therapies, immunotherapies are a promising treatment candidate. HER2 is an attractive target for GBM immunotherapy, as its expression is highly associated with various types of GBM. We previously reported that a novel HER2-targeted recombinant protein e23sFv-Fdt-casp6 has an antitumor effect on HER2-positive gastric cancer cells. In this study, we established a genetically modified Chinese hamster ovary cell line, which produced and secreted e23sFv-Fdt-casp6 proteins. Following specific binding to and internalization into HER2-overexpressing tumor cells, the e23sFv-Fdt-casp6 protein induced tumor cell apoptosis and inhibited the proliferation of HER2-overexpressing A172 and U251MG cells in vitro, but not in U87MG cells with undetectable HER2. The e23sFv-Fdt-casp6 gene was introduced into severe combined immunodeficient mice bearing human glioblastoma xenografts by using intramuscular injections of a liposome-encapsulated vector. The recombinant protein e23sFv-Fdt-casp6 specifically targeted tumor cells and induced apoptosis, thereby leading to potent inhibition of tumor growth and prolonged the survival time of tumor-bearing mice. We concluded that e23sFv‑Fdt‑casp6 represents a promising HER2-targeted treatment option for human gliomas.

RNA-seq of 272 gliomas revealed a novel, recurrent PTPRZ1-MET fusion transcript in secondary glioblastomas.

PubMed

Bao, Zhao-Shi; Chen, Hui-Min; Yang, Ming-Yu; Zhang, Chuan-Bao; Yu, Kai; Ye, Wan-Lu; Hu, Bo-Qiang; Yan, Wei; Zhang, Wei; Akers, Johnny; Ramakrishnan, Valya; Li, Jie; Carter, Bob; Liu, Yan-Wei; Hu, Hui-Min; Wang, Zheng; Li, Ming-Yang; Yao, Kun; Qiu, Xiao-Guang; Kang, Chun-Sheng; You, Yong-Ping; Fan, Xiao-Long; Song, Wei Sonya; Li, Rui-Qiang; Su, Xiao-Dong; Chen, Clark C; Jiang, Tao

2014-11-01

Studies of gene rearrangements and the consequent oncogenic fusion proteins have laid the foundation for targeted cancer therapy. To identify oncogenic fusions associated with glioma progression, we catalogued fusion transcripts by RNA-seq of 272 gliomas. Fusion transcripts were more frequently found in high-grade gliomas, in the classical subtype of gliomas, and in gliomas treated with radiation/temozolomide. Sixty-seven in-frame fusion transcripts were identified, including three recurrent fusion transcripts: FGFR3-TACC3, RNF213-SLC26A11, and PTPRZ1-MET (ZM). Interestingly, the ZM fusion was found only in grade III astrocytomas (1/13; 7.7%) or secondary GBMs (sGBMs, 3/20; 15.0%). In an independent cohort of sGBMs, the ZM fusion was found in three of 20 (15%) specimens. Genomic analysis revealed that the fusion arose from translocation events involving introns 3 or 8 of PTPRZ and intron 1 of MET. ZM fusion transcripts were found in GBMs irrespective of isocitrate dehydrogenase 1 (IDH1) mutation status. sGBMs harboring ZM fusion showed higher expression of genes required for PIK3CA signaling and lowered expression of genes that suppressed RB1 or TP53 function. Expression of the ZM fusion was mutually exclusive with EGFR overexpression in sGBMs. Exogenous expression of the ZM fusion in the U87MG glioblastoma line enhanced cell migration and invasion. Clinically, patients afflicted with ZM fusion harboring glioblastomas survived poorly relative to those afflicted with non-ZM-harboring sGBMs (P < 0.001). Our study profiles the shifting RNA landscape of gliomas during progression and reveled ZM as a novel, recurrent fusion transcript in sGBMs. © 2014 Bao et al.; Published by Cold Spring Harbor Laboratory Press.

RNA-seq of 272 gliomas revealed a novel, recurrent PTPRZ1-MET fusion transcript in secondary glioblastomas

PubMed Central

Bao, Zhao-Shi; Yang, Ming-Yu; Zhang, Chuan-Bao; Yu, Kai; Ye, Wan-Lu; Hu, Bo-Qiang; Yan, Wei; Zhang, Wei; Akers, Johnny; Ramakrishnan, Valya; Li, Jie; Carter, Bob; Liu, Yan-Wei; Hu, Hui-Min; Wang, Zheng; Li, Ming-Yang; Yao, Kun; Qiu, Xiao-Guang; Kang, Chun-Sheng; You, Yong-Ping; Fan, Xiao-Long; Song, Wei Sonya; Li, Rui-Qiang

2014-01-01

Studies of gene rearrangements and the consequent oncogenic fusion proteins have laid the foundation for targeted cancer therapy. To identify oncogenic fusions associated with glioma progression, we catalogued fusion transcripts by RNA-seq of 272 gliomas. Fusion transcripts were more frequently found in high-grade gliomas, in the classical subtype of gliomas, and in gliomas treated with radiation/temozolomide. Sixty-seven in-frame fusion transcripts were identified, including three recurrent fusion transcripts: FGFR3-TACC3, RNF213-SLC26A11, and PTPRZ1-MET (ZM). Interestingly, the ZM fusion was found only in grade III astrocytomas (1/13; 7.7%) or secondary GBMs (sGBMs, 3/20; 15.0%). In an independent cohort of sGBMs, the ZM fusion was found in three of 20 (15%) specimens. Genomic analysis revealed that the fusion arose from translocation events involving introns 3 or 8 of PTPRZ and intron 1 of MET. ZM fusion transcripts were found in GBMs irrespective of isocitrate dehydrogenase 1 (IDH1) mutation status. sGBMs harboring ZM fusion showed higher expression of genes required for PIK3CA signaling and lowered expression of genes that suppressed RB1 or TP53 function. Expression of the ZM fusion was mutually exclusive with EGFR overexpression in sGBMs. Exogenous expression of the ZM fusion in the U87MG glioblastoma line enhanced cell migration and invasion. Clinically, patients afflicted with ZM fusion harboring glioblastomas survived poorly relative to those afflicted with non-ZM-harboring sGBMs (P < 0.001). Our study profiles the shifting RNA landscape of gliomas during progression and reveled ZM as a novel, recurrent fusion transcript in sGBMs. PMID:25135958

Density-Dependent Regulation of Glioma Cell Proliferation and Invasion Mediated by miR-9.

PubMed

Katakowski, Mark; Charteris, Nicholas; Chopp, Michael; Khain, Evgeniy

2016-12-01

The phenotypic axis of invasion and proliferation in malignant glioma cells is a well-documented phenomenon. Invasive glioma cells exhibit a decreased proliferation rate and a resistance to apoptosis, and invasive tumor cells dispersed in brain subsequently revert to proliferation and contribute to secondary tumor formation. One miRNA can affect dozens of mRNAs, and some miRNAs are potent oncogenes. Multiple miRNAs are implicated in glioma malignancy, and several of which have been identified to regulate tumor cell motility and division. Using rat 9 L gliosarcoma and human U87 glioblastoma cell lines, we investigated miRNAs associated with the switch between glioma cell invasion and proliferation. Using micro-dissection of 9 L glioma tumor xenografts in rat brain, we identified disparate expression of miR-9 between cells within the periphery of the primary tumor, and those comprising tumor islets within the invasive zone. Modifying miR-9 expression in in vitro assays, we report that miR-9 controls the axis of glioma cell invasion/proliferation, and that its contribution to invasion or proliferation is biphasic and dependent upon local tumor cell density. In addition, immunohistochemistry revealed elevated hypoxia inducible factor 1 alpha (HIF-1α) in the invasive zone as compared to the primary tumor periphery. We also found that hypoxia promotes miR-9 expression in glioma cells. Based upon these findings, we propose a hypothesis for the contribution of miR-9 to the dynamics glioma invasion and satellite tumor formation in brain adjacent to tumor.

Functionalized nano-graphene oxide particles for targeted fluorescence imaging and photothermy of glioma U251 cells.

PubMed

Li, Zhong-Jun; Li, Chao; Zheng, Mei-Guang; Pan, Jia-Dong; Zhang, Li-Ming; Deng, Yue-Fei

2015-01-01

This study was to prepare the functionalized nano-graphene oxide (nano-GO) particles, and observe targeted fluorescence imaging and photothermy of U251 glioma cells under near infrared (NIR) exposure. The functionalized nano-GO-Tf-FITC particles were prepared and then were incubated with U251 glioma cells. Estimation of CCK8 cell activity was adopted for measurement of cytotoxicity. The effect of fluorescein imaging was detected by fluorescence microscope with anti-CD71-FITC as a control. Finally, we detected the killing efficacy with flow cytometry after an 808 nm NIR exposure. Both nano-GO-Tf-FITC group and CD71-FITC group exhibited green-yellow fluorescence, while the control group without the target molecule nano-GO-FITC was negative. The nano-GO-Tf-FITC was incubated with U251 cells at 0.1 mg/ml, 1.0 mg/ml, 3.0 mg/ml and 5.0 mg/ml. After 48 h of incubation, the absorbance was 0.747 ± 0.031, 0.732 ± 0.043, 0.698 ± 0.051 and 0.682 ± 0.039, while the absorbance of control group is 0.759 ± 0.052. There is no significant difference between the nano-GO-FITC groups and control group. In addition, the apoptosis and death index of nano-GO-Tf-FITC group was significantly higher than that of nano-GO-FITC and blank control group (P < 0.05). The nano-GO-Tf-FITC particles with good biological compatibility and low cytotoxicity are successfully made, which have an observed effect of target imaging and photothermal therapy on glioma U251 cells.

An R132H Mutation in Isocitrate Dehydrogenase 1 Enhances p21 Expression and Inhibits Phosphorylation of Retinoblastoma Protein in Glioma Cells

PubMed Central

Miyata, Satsuki; Urabe, Masashi; Gomi, Akira; Nagai, Mutsumi; Yamaguchi, Takashi; Tsukahara, Tomonori; Mizukami, Hiroaki; Kume, Akihiro; Ozawa, Keiya; Watanabe, Eiju

2013-01-01

Cytosolic isocitrate dehydrogenase 1 (IDH1) with an R132H mutation in brain tumors loses its enzymatic activity for catalyzing isocitrate to α-ketoglutarate (α-KG) and acquires new activity whereby it converts α-KG to 2-hydroxyglutarate. The IDH1 mutation induces down-regulation of tricarboxylic acid cycle intermediates and up-regulation of lipid metabolism. Sterol regulatory element-binding proteins (SREBPs) regulate not only the synthesis of cholesterol and fatty acids but also acyclin-dependent kinase inhibitor p21 that halts the cell cycle at G1. Here we show that SREBPs were up-regulated in U87 human glioblastoma cells transfected with an IDH1R132H-expression plasmid. Small interfering ribonucleic acid (siRNA) for SREBP1 specifically decreased p21 messenger RNA (mRNA) levels independent of the p53 pathway. In IDH1R132H-expressing U87 cells, phosphorylation of Retinoblastoma (Rb) protein also decreased. We propose that metabolic changes induced by the IDH1 mutation enhance p21 expression via SREBP1 and inhibit phosphorylation of Rb, which slows progressionof the cell cycle and may be associated with non-aggressive features of gliomas with an IDH1 mutation. PMID:24077277

An R132H mutation in isocitrate dehydrogenase 1 enhances p21 expression and inhibits phosphorylation of retinoblastoma protein in glioma cells.

PubMed

Miyata, Satsuki; Urabe, Masashi; Gomi, Akira; Nagai, Mutsumi; Yamaguchi, Takashi; Tsukahara, Tomonori; Mizukami, Hiroaki; Kume, Akihiro; Ozawa, Keiya; Watanabe, Eiju

2013-01-01

Cytosolic isocitrate dehydrogenase 1 (IDH1) with an R132H mutation in brain tumors loses its enzymatic activity for catalyzing isocitrate to α-ketoglutarate (α-KG) and acquires new activity whereby it converts α-KG to 2-hydroxyglutarate. The IDH1 mutation induces down-regulation of tricarboxylic acid cycle intermediates and up-regulation of lipid metabolism. Sterol regulatory element-binding proteins (SREBPs) regulate not only the synthesis of cholesterol and fatty acids but also acyclin-dependent kinase inhibitor p21 that halts the cell cycle at G1. Here we show that SREBPs were up-regulated in U87 human glioblastoma cells transfected with an IDH1(R132H)-expression plasmid. Small interfering ribonucleic acid (siRNA) for SREBP1 specifically decreased p21 messenger RNA (mRNA) levels independent of the p53 pathway. In IDH1(R132H)-expressing U87 cells, phosphorylation of Retinoblastoma (Rb) protein also decreased. We propose that metabolic changes induced by the IDH1 mutation enhance p21 expression via SREBP1 and inhibit phosphorylation of Rb, which slows progression of the cell cycle and may be associated with non-aggressive features of gliomas with an IDH1 mutation.

Immunomodulation of glioma cells after gene therapy: induction of major histocompatibility complex class I but not class II antigen in vitro.

PubMed

Parsa, A T; Chi, J H; Hurley, P T; Jeyapalan, S A; Bruce, J N

2001-09-01

Acquired immunity has been demonstrated in Fischer rats bearing syngeneic 9L tumors after herpes simplex virus (HSV) thymidine kinase (TK) gene transfection and ganciclovir treatment. The nature of this immunity in rats and its relevance to the HSV TK/ganciclovir protocol for human subjects remain to be determined. In this study, levels of major histocompatibility complex (MHC) Class I and II antigen expression were measured before and after HSV TK transfection, in an effort to document immunomodulatory changes caused by gene therapy. Tumor cells from the 9L gliosarcoma cell line, three primary human glioma cultures, and the human glioma cell line U87 MG were transduced with HSV TK vector-containing supernatant from fibroblast-producing cells (titer of 5 x 10(6) colony-forming units/ml) and selected in G418 medium for neomycin resistance. Clones were pooled or individually selected for cell-killing assays with ganciclovir, to confirm TK expression (10(3) cells/well in a 96-well dish). Northern analyses using MHC Class I and Class II complementary deoxyribonucleic acid probes were performed on blots containing total ribonucleic acid from wild-type tumor cells and HSV TK transfectants. A beta-actin complementary deoxyribonucleic acid probe served as an internal control. Cell surface expression was confirmed with flow cytometry. The induction of MHC Class I was tested for cycloheximide and genistein sensitivity. All cell cultures exhibited increases in MHC Class I but not MHC Class II expression, as determined by Northern analysis densitometry and flow cytometry. Cycloheximide treatment did not diminish the up-regulation of MHC Class I after retroviral transfection, implicating a signal transduction pathway that does not require ongoing protein synthesis. Genistein pretreatment of cell cultures did diminish the up-regulation of MHC Class I, implicating a tyrosine kinase in the signaling cascade. Induction of MHC Class I in rat and human glioma cells after HSV TK retroviral gene therapy is a primary effect that is dependent on tyrosine kinase activity. Specific immune responses generated after transfection may represent an important general side effect of gene therapy protocols. Elucidation of the mechanism of immunomodulation after gene therapy will likely yield safer and more effective clinical protocols.

Impact of radiation therapy on the oncolytic adenovirus dl520: implications on the treatment of glioblastoma.

PubMed

Bieler, Alexa; Mantwill, Klaus; Holzmüller, Regina; Jürchott, Karsten; Kaszubiak, Alexander; Stärk, Sybille; Glockzin, Gabriel; Lage, Hermann; Grosu, Anca-Ligia; Gansbacher, Bernd; Holm, Per Sonne

2008-03-01

Viral oncolytic therapy is emerging as a new form of anticancer therapy and has shown promising preclinical results, especially in combination with radio- and chemotherapy. We recently reported that nuclear localization of the human transcription factor YB-1 in multidrug-resistant cells facilitates E1-independent adenoviral replication. The aim of this study was to evaluate the combined treatment of the conditionally-replicating adenovirus dl520 and radiotherapy in glioma cell lines in vitro and in human tumor xenografts. Furthermore, the dependency of YB-1 on dl520 replication was verified by shRNA directed down regulation of YB-1. Localization of YB-1 was determined by immunostaining. Glioma cell lines LN-18, U373 and U87 were infected with dl520. Induction of cytopathic effect (CPE), viral replication, viral yield and viral release were determined after viral infection, radiation therapy and the combination of both treatment modalities. The capacity of treatments alone or combined to induce tumor growth inhibition of subcutaneous U373 tumors was tested also in nude mice. Quantitative real-time PCR demonstrated that the shRNA-mediated down regulation of YB-1 is leading to a dramatic decrease in adenoviral replication of dl520. Immunostaining analysis showed that the YB-1 protein was predominantly located in the cytoplasm in the perinuclear space and less abundant in the nucleus. After irradiation we found an increase of nuclear YB-1. The addition of radiotherapy increased the oncolytic effect of dl520 with enhanced viral replication, viral yield and viral release. The oncolytic activity of dl520 plus radiation inhibited the growth of subcutaneous U373 tumors in a xenograft mouse model. Radiation mediated increase of nuclear YB-1 in glioma cells enhanced the oncolytic potential of adenovirus dl520.

IDH1 R132H decreases proliferation of glioma cell lines in vitro and in vivo.

PubMed

Bralten, Linda B C; Kloosterhof, Nanne K; Balvers, Rutger; Sacchetti, Andrea; Lapre, Lariesa; Lamfers, Martine; Leenstra, Sieger; de Jonge, Hugo; Kros, Johan M; Jansen, Erwin E W; Struys, Eduard A; Jakobs, Cornelis; Salomons, Gajja S; Diks, Sander H; Peppelenbosch, Maikel; Kremer, Andreas; Hoogenraad, Casper C; Smitt, Peter A E Sillevis; French, Pim J

2011-03-01

A high percentage of grade II and III gliomas have mutations in the gene encoding isocitrate dehydrogenase (IDH1). This mutation is always a heterozygous point mutation that affects the amino acid arginine at position 132 and results in loss of its native enzymatic activity and gain of alternative enzymatic activity (producing D-2-hydroxyglutarate). The objective of this study was to investigate the cellular effects of R132H mutations in IDH1. Functional consequences of IDH1(R132H) mutations were examined among others using fluorescence-activated cell sorting, kinome and expression arrays, biochemical assays, and intracranial injections on 3 different (glioma) cell lines with stable overexpression of IDH1(R132H) . IDH1(R132H) overexpression in established glioma cell lines in vitro resulted in a marked decrease in proliferation, decreased Akt phosphorylation, altered morphology, and a more contact-dependent cell migration. The reduced proliferation is related to accumulation of D-2-hydroxyglutarate that is produced by IDH1(R132H) . Mice injected with IDH1(R132H) U87 cells have prolonged survival compared to mice injected with IDH1(wt) or green fluorescent protein-expressing U87 cells. Our results demonstrate that IDH1(R132H) dominantly reduces aggressiveness of established glioma cell lines in vitro and in vivo. In addition, the IDH1(R132H) -IDH1(wt) heterodimer has higher enzymatic activity than the IDH1(R132H) -IDH1(R132H) homodimer. Our observations in model systems of glioma might lead to a better understanding of the biology of IDH1 mutant gliomas, which are typically low grade and often slow growing. Copyright © 2011 American Neurological Association.

Near-infrared optical imaging in glioblastoma xenograft with ligand targeting α3 integrin

PubMed Central

Xiao, Wenwu; Yao, Nianhuan; Peng, Li; Liu, Ruiwu; Lam, Kit S

2010-01-01

Purpose Patients with glioblastoma usually have a very poor prognosis. Even with a combination of radiotherapy plus temozolomide, the median survival of these patients is only 14.6 months. New treatment approaches to this cancer are needed. Our purpose is to develop new cell-surface binding ligands for glioblastoma cells, and use them as targeted imaging and therapeutic agents for this deadly disease. Methods One-bead one-compound combinatorial cyclic peptide libraries were screened with live human glioblastoma U-87MG cells. The binding affinity and targeting specificity of peptides identified were tested with in vitro experiments on cells and in vivo, and ex vivo experiments on U-87MG xegnograft mouse model. Results A cyclic peptide, LXY1, was identified and shown to be binding to the α3 integrin of U-87MG cells with moderately high affinity (Kd = 0.5+/−0.1 μM) and high specificity. Biotinylated LXY1, when complexed with streptavidin-Cy5.5 (SA-Cy5.5) conjugate, targeted both subcutaneous and orthotopic U-87MG xenograft implants in nude mice. The in vivo targeting specificity was further verified by strong inhibition of tumor uptake of LXY1-biotin-SA-Cy5.5 complex when intravenously injecting the animals with anti-α3 integrin antibody or excess unlabeled LXY1 prior to administrating the imaging probe. The smaller univalent LXY1-Cy5.5 conjugate (2279 Da) was found to have a faster accumulation in the U-87MG tumor and shorter retention time compared with the larger tetravalent LXY1-biotin-SA-Cy5.5 complex (~ 64 KDa). Conclusions Collectively, the data reveals that LXY1 has the potential to be developed into an effective imaging and therapeutic targeting agent for human glioblastoma. PMID:18712382

In vivo imaging of tumor vascular endothelial cells

NASA Astrophysics Data System (ADS)

Zhao, Dawen; Stafford, Jason H.; Zhou, Heling; Thorpe, Philip E.

2013-02-01

Phosphatidylserine (PS), normally restricted to the inner leaflet of the plasma membrane, becomes exposed on the outer surface of viable (non-apoptotic) endothelial cells in tumor blood vessels, probably in response to oxidative stresses present in the tumor microenvironment. In the present study, we optically imaged exposed PS on tumor vasculature in vivo using PGN635, a novel human monoclonal antibody that targets PS. PGN635 F(ab')2 was labeled with the near infrared (NIR) dye, IRDye 800CW. Human glioma U87 cells or breast cancer MDA-MB-231 cells were implanted subcutaneously or orthotopically into nude mice. When the tumors reached ~5 mm in diameter, 800CW- PGN635 was injected via a tail vein and in vivo dynamic NIR imaging was performed. For U87 gliomas, NIR imaging allowed clear detection of tumors as early as 4 h later, which improved over time to give a maximal tumor/normal ratio (TNR = 2.9 +/- 0.5) 24 h later. Similar results were observed for orthotopic MDA-MB-231 breast tumors. Localization of 800CW-PGN635 to tumors was antigen specific since 800CW-Aurexis, a control probe of irrelevant specificity, did not localize to the tumors, and pre-administration of unlabeled PGN635 blocked the uptake of 800CW-PGN635. Fluorescence microscopy confirmed that 800CW-PGN635 was binding to PS-positive tumor vascular endothelium. Our studies suggest that tumor vasculature can be successfully imaged in vivo to provide sensitive tumor detection.

The synthetic ligand of peroxisome proliferator-activated receptor-gamma ciglitazone affects human glioblastoma cell lines.

PubMed

Strakova, Nicol; Ehrmann, Jiri; Dzubak, Petr; Bouchal, Jan; Kolar, Zdenek

2004-06-01

Glioblastoma multiforme is the most common malignant brain tumor in adults, and it is among the most lethal of all cancers. Recent studies have shown that ligand activation of peroxisome proliferator-activated receptor (PPAR)-gamma can induce differentiation and inhibit proliferation of several cancer cells. In this study, we have investigated whether one PPARgamma ligand in particular, ciglitazone, inhibits cell viability and, additionally, whether it affects the cell cycle and apoptosis of human glioblastoma cell lines T98G, U-87 MG, A172, and U-118 MG. All glioblastoma cell lines were found to express PPARgamma protein, and following treatment with ciglitazone, localization was unchanged. Ciglitazone inhibited viability in a dose-dependent manner in all four tested glioblastoma cells after 24 h of treatment. Analysis of the cell cycle showed arrest in the G(1) phase and partial block in G(2)/M phase of the cell cycle. Cyclin D1 and cyclin B expression was decreased. Phosphorylation of Rb protein dropped as well. We found that ciglitazone was followed by increased expression of p27(Kip1) and p21(Waf1/Cip1). It also led to apoptosis induction: bax expression in T98G was elevated. Expression of the antiapoptotic protein bcl-2 was reduced in U-118 MG and U-87 MG and showed a slight decrease in A172 cells. Flow cytometry confirmed the induction of apoptosis. Moreover, PPARgamma ligand decreased telomerase activity in U-87 MG and U-118 MG cell lines. Our results demonstrate that ciglitazone inhibits the viability of human glioblastoma cell lines via induction of apoptosis; as a result, this ligand may offer potential new therapy for the treatment of central nervous system neoplasms.

Inhibition of the Rac1-WAVE2-Arp2/3 signaling pathway promotes radiosensitivity via downregulation of cofilin-1 in U251 human glioma cells.

PubMed

Zhou, Tao; Wang, Chen-Han; Yan, Hua; Zhang, Rui; Zhao, Jin-Bing; Qian, Chun-Fa; Xiao, Hong; Liu, Hong-Yi

2016-05-01

The Ras-related C3 botulinum toxin substrate 1 (Rac1)-WASP-family verprolin-homologous protein-2 (WAVE2)-actin-related protein 2/3 (Arp2/3) signaling pathway has been identified to be involved in cell migration and invasion in various types of cancer cell. Cofilin‑1 (CFL‑1), which is regulated by the Rac1‑WAVE2‑Arp2/3 signaling pathway, may promote radioresistance in glioma. Therefore, the present study aimed to investigate the potential role of the Rac1‑WAVE2‑Arp2/3 signaling pathway in radioresistance in U251 human glioma cells and elucidate its affect on CFL‑1 expression. Western blot analysis was performed to evaluate the protein expression of CFL‑1. In the present study, Rac1 was inhibited by NSC 23766, WAVE2 was inhibited by transfection with short hairpin (sh)RNA‑WAVE2 using Lipofectamine™ 2000 and Arp2/3 was inhibited by CK‑666. Cell viability was measured using the 3‑(4,5‑dimethylthiazol‑2‑yl)-2,5‑diphenyltetrazolium bromide assay, the cell migration ability was examined by a wound‑healing assay, and the cell invasion ability was assessed using a Transwell culture chamber system. The results showed that inhibition of the Rac1‑WAVE2‑Arp2/3 signaling pathway using NSC 23766, shRNA‑WAVE2 or CK‑666 reduced the cell viability, migration and invasion abilities in U251 human glioma cells, concordant with a reduced expression of CFL‑1. Furthermore, the expression of CFL‑1 was significantly increased in radioresistant U251 glioma cells when compared with normal U251 human glioma cells. These findings indicate that inhibition of the Rac1‑WAVE2‑Arp2/3 signaling pathway may promote radiosensitivity, which may partially result from the downregulation of CFL‑1 in U251 human glioma cells.

Sustained delivery of cytarabine-loaded vesicular phospholipid gels for treatment of xenografted glioma.

PubMed

Qi, Na; Cai, Cuifang; Zhang, Wei; Niu, Yantao; Yang, Jingyu; Wang, Lihui; Tian, Bin; Liu, Xiaona; Lin, Xia; Zhang, Yu; Zhang, Yan; He, Haibing; Chen, Kang; Tang, Xing

2014-09-10

This study described the development of vesicular phospholipid gels (VPGs) for sustained delivery of cytarabine (Ara-C) for the treatment of xenografted glioma. Ara-C-loaded VPGs in the state of a semisolid phospholipid dispersion looked like numerous vesicles tightly packing together under the freeze-fracture electron microscopy (FF-TEM), their release profiles displayed sustained drug release up to 384 h in vitro. The biodistribution of Ara-C in the rat brain showed that Ara-C-loaded VPGs could maintain therapeutic concentrations up to 5mm distance from the implantation site in brain tissue within 28 days. At the same time, fluorescence micrograph confirmed drug distribution in brain tissue visually. Furthermore, after single administration, Ara-C-loaded VPGs group significantly inhibited the U87-MG glioma growth in right flank in comparison with Ara-C solution (p<0.01). It was explained that the entrapped drug in VPGs could avoid degradation from cytidine deaminase and sustained release of drug from Ara-C-loaded VPGs could maintain the effective therapeutic levels for a long time around the tumor. In conclusion, Ara-C-loaded VPGs, with the properties of sustained release, high penetration capacity, nontoxicity and no shape restriction of the surgical cavity, are promising local delivery systems for post-surgical sustained chemotherapy against glioma. Copyright © 2014 Elsevier B.V. All rights reserved.

Antiglioma effects of N6-isopentenyladenosine, an endogenous isoprenoid end product, through the downregulation of epidermal growth factor receptor.

PubMed

Ciaglia, Elena; Abate, Mario; Laezza, Chiara; Pisanti, Simona; Vitale, Mario; Seneca, Vincenzo; Torelli, Giovanni; Franceschelli, Silvia; Catapano, Giuseppe; Gazzerro, Patrizia; Bifulco, Maurizio

2017-02-15

Malignant gliomas are highly dependent on the isoprenoid pathway for the synthesis of lipid moieties critical for cell proliferation. The isoprenoid derivative N6-isopentenyladenosine (iPA) displays pleiotropic biological effects, including a direct anti-tumor activity in several tumor models. The antiglioma effects of iPA was then explored in U87MG cells both in vitro and grafted in mice and the related molecular mechanism confirmed in primary derived patients' glioma cells. iPA powerfully inhibited tumor cell growth and induced caspase-dependent apoptosis through a mechanism involving a marked accumulation of the pro-apoptotic BIM protein and inhibition of EGFR. Indeed, activating AMPK following conversion into its iPAMP active form, iPA stimulated EGFR phosphorylation and ubiquitination along a proteasome-mediated pathway which was responsible for receptor degradation and its downstream signaling pathways inhibition, including the STAT3, ERK and AKT cascade. The inhibition of AMPK by compound C prevented iPA-mediated phosphorylation of EGFR, known to precede receptor loss. As expected the block of EGFR degradation, by exposure to the proteasome inhibitor MG132, significantly reduced iPA-induced cell death. Given the importance of receptor degradation in iPA-mediated cytotoxicity, we also documented that the EGFR expression levels in a panel of primary glioma cells confers them a high sensitivity to iPA treatment. In conclusion our study provides the first evidence of iPA antiglioma effect. Indeed, as glioma is driven by aberrant signaling of growth factor receptors, particularly the EGFR, iPA, alone or in association with EGFR targeted therapies, might be a promising therapeutic tool to achieve a potent anti-tumoral effect. © 2016 UICC.

Immunologic and gene expression profiles of spontaneous canine oligodendrogliomas.

PubMed

Filley, Anna; Henriquez, Mario; Bhowmik, Tanmoy; Tewari, Brij Nath; Rao, Xi; Wan, Jun; Miller, Margaret A; Liu, Yunlong; Bentley, R Timothy; Dey, Mahua

2018-05-01

Malignant glioma (MG), the most common primary brain tumor in adults, is extremely aggressive and uniformly fatal. Several treatment strategies have shown significant preclinical promise in murine models of glioma; however, none have produced meaningful clinical responses in human patients. We hypothesize that introduction of an additional preclinical animal model better approximating the complexity of human MG, particularly in interactions with host immune responses, will bridge the existing gap between these two stages of testing. Here, we characterize the immunologic landscape and gene expression profiles of spontaneous canine glioma and evaluate its potential for serving as such a translational model. RNA in situ hybridization, flowcytometry, and RNA sequencing were used to evaluate immune cell presence and gene expression in healthy and glioma-bearing canines. Similar to human MGs, canine gliomas demonstrated increased intratumoral immune cell infiltration (CD4+, CD8+ and CD4+Foxp3+ T cells). The peripheral blood of glioma-bearing dogs also contained a relatively greater proportion of CD4+Foxp3+ regulatory T cells and plasmacytoid dendritic cells. Tumors were strongly positive for PD-L1 expression and glioma-bearing animals also possessed a greater proportion of immune cells expressing the immune checkpoint receptors CTLA-4 and PD-1. Analysis of differentially expressed genes in our canine populations revealed several genetic changes paralleling those known to occur in human disease. Naturally occurring canine glioma has many characteristics closely resembling human disease, particularly with respect to genetic dysregulation and host immune responses to tumors, supporting its use as a translational model in the preclinical testing of prospective anti-glioma therapies proven successful in murine studies.

[Cell-ELA-based determination of binding affinity of DNA aptamer against U87-EGFRvIII cell].

PubMed

Tan, Yan; Liang, Huiyu; Wu, Xidong; Gao, Yubo; Zhang, Xingmei

2013-05-01

A15, a DNA aptamer with binding specificity for U87 glioma cells stably overexpressing the epidermal growth factor receptor variant III (U87-EGFRvIII), was generated by cell systematic evolution of ligands by exponential enrichment (cell-SELEX) using a random nucleotide library. Subsequently, we established a cell enzyme-linked assay (cell-ELA) to detect the affinity of A15 compared to an EGFR antibody. We used A15 as a detection probe and cultured U87-EGFRvIII cells as targets. Our data indicate that the equilibrium dissociation constants (K(d)) for A15 were below 100 nmol/L and had similar affinity compared to an EGFR antibody for U87-EGFRvIII. We demonstrated that the cell-ELA was a useful method to determine the equilibrium dissociation constants (K(d)) of aptamers generated by cell-SELEX.

Fascin-1 knock-down of human glioma cells reduces their microvilli/filopodia while improving their susceptibility to lymphocyte-mediated cytotoxicity

PubMed Central

Hoa, Neil T; Ge, Lisheng; Erickson, Kate L; Kruse, Carol A; Cornforth, Andrew N; Kuznetsov, Yurii; McPherson, Alex; Martini, Filippo; Jadus, Martin R

2015-01-01

Cancer cells derived from Glioblastoma multiforme possess membranous protrusions allowing these cells to infiltrate surrounding tissue, while resisting lymphocyte cytotoxicity. Microvilli and filopodia are supported by actin filaments cross-linked by fascin. Fascin-1 was genetically silenced within human U251 glioma cells; these knock-down glioma cells lost their microvilli/filopodia. The doubling time of these fascin-1 knock-down cells was doubled that of shRNA control U251 cells. Fascin-1 knock-down cells lost their transmigratory ability responding to interleukin-6 or insulin-like growth factor-1. Fascin-1 silenced U251 cells were more easily killed by cytolytic lymphocytes. Fascin-1 knock-down provides unique opportunities to augment glioma immunotherapy by simultaneously targeting several key glioma functions: like cell transmigration, cell division and resisting immune responses. PMID:25901196

Cannabidiol inhibits human glioma cell migration through a cannabinoid receptor-independent mechanism

PubMed Central

Vaccani, Angelo; Massi, Paola; Colombo, Arianna; Rubino, Tiziana; Parolaro, Daniela

2005-01-01

We evaluated the ability of cannabidiol (CBD) to impair the migration of tumor cells stimulated by conditioned medium. CBD caused concentration-dependent inhibition of the migration of U87 glioma cells, quantified in a Boyden chamber. Since these cells express both cannabinoid CB1 and CB2 receptors in the membrane, we also evaluated their engagement in the antimigratory effect of CBD. The inhibition of cell was not antagonized either by the selective cannabinoid receptor antagonists SR141716 (CB1) and SR144528 (CB2) or by pretreatment with pertussis toxin, indicating no involvement of classical cannabinoid receptors and/or receptors coupled to Gi/o proteins. These results reinforce the evidence of antitumoral properties of CBD, demonstrating its ability to limit tumor invasion, although the mechanism of its pharmacological effects remains to be clarified. PMID:15700028

Analysis of the activation status of Akt, NFkappaB, and Stat3 in human diffuse gliomas.

PubMed

Wang, Huamin; Wang, Hua; Zhang, Wei; Huang, Helen J; Liao, Warren S L; Fuller, Gregory N

2004-08-01

Loss of phosphatase and tensin homolog (PTEN) and amplification of the epidermal growth factor receptor (EGFR) gene contribute to the progression of gliomas. As downstream targets of the PTEN and EGFR signaling pathways, Akt, NFkappaB, and signal transducer and activator of transcription-3 (Stat3) have been shown to play important roles in the control of cell proliferation, apoptosis, and oncogenesis. We examined the activation status of Akt, NFkappaB, and Stat3 in 259 diffuse gliomas using tissue microarrays and immunohistochemistry, and evaluated their association with glioma grade. We observed significant positive correlations between the activation status of Akt and NFkappaB and glioma grade. In contrast, only focal immunoreactivity for phospho-Stat3 was observed in < 9% of high-grade gliomas. In addition, we observed a significant correlation between the activation of Akt and NFkappaB. Functional correlation between Akt activation and the activation of NFkappaB was confirmed in U251MG GBM cells in which inhibition of Akt activation either by stable expression of PTEN or by the PI3-kinase inhibitors, wortmannin and LY294002, led to a concomitant decrease in NFkappaB-binding activity. Thus, our results demonstrate that constitutive activation of Akt and NFkappaB, but not Stat3, contributes significantly to the progression of diffuse gliomas, and activation of Akt may lead to NFkappaB activation in high-grade gliomas.

[Elevated expression of B7-H6 in U87 cells-derived glioma stem like cells is associated with biological characteristics].

PubMed

Chen, Hanqing; Shi, Zhengpeng; Gao, Bing; Fu, Fengqing; Zhang, Xueguang

2016-09-01

Objective To investigate the expression and biological significance of costimulatory molecule B7-H6, a member of B7 family, in glioma stem like cells (GSLCs). Methods In virtue of the ability of forming neurospheres in vitro , GSLCs were isolated from U87 cells by cell sub-cloning. Real-time quantitative PCR and flow cytometry were performed to detect the expressions of stem cell related markers (c-myc, Sox2, CD133, nestin, and CXCR4), as well as the expressions of B7 family molecules. The different doses of adriamycin, carboplatin, cisplatin, were used to treat GSLCs for testing their chemotherapy-resistance. After the expression of B7-H6 in GSLCs was knockdown by siRNA, CCK-8 method was used to detect cell proliferation. Results GSLCs were successfully isolated from U87 cells, which formed neurospheres in vitro . The expressions of multiple stem cell markers were up-regulated and the GSLCs showed enhanced chemo therapy-resistance. B7 family members, B7-H1, B7-H3, B7-H4 and B7-H6 were expressed in GSLCs. Compared with primary U87 cells, GSLCs presented with a remarkably increased expression of B7-H6 on cell membrane. When B7-H6 was silenced by siRNA, cell proliferation was inhibited along with the decrease of c-myc expression. Conclusion The expression of B7-H6 is up-regulated in U87-derived GSLCs, which is associated with the biological characteristics of GSLCs.

CREB1 regulates glucose transport of glioma cell line U87 by targeting GLUT1.

PubMed

Chen, Jiaying; Zhang, Can; Mi, Yang; Chen, Fuxue; Du, Dongshu

2017-12-01

Glioma is stemmed from the glial cells in the brain, which is accounted for about 45% of all intracranial tumors. The characteristic of glioma is invasive growth, as well as there is no obvious boundary between normal brain tissue and glioma tissue, so it is difficult to resect completely with worst prognosis. The metabolism of glioma is following the Warburg effect. Previous researches have shown that GLUT1, as a glucose transporter carrier, affected the Warburg effect, but the molecular mechanism is not very clear. CREB1 (cAMP responsive element-binding protein1) is involved in various biological processes, and relevant studies confirmed that CREB1 protein regulated the expression of GLUT1, thus mediating glucose transport in cells. Our experiments mainly reveal that the CREB1 could affect glucose transport in glioma cells by regulating the expression of GLUT1, which controlled the metabolism of glioma and affected the progression of glioma.

Photochemical internalization-mediated nonviral gene transfection: polyamine core-shell nanoparticles as gene carrier

NASA Astrophysics Data System (ADS)

Zamora, Genesis; Wang, Frederick; Sun, Chung-Ho; Trinidad, Anthony; Kwon, Young Jik; Cho, Soo Kyung; Berg, Kristian; Madsen, Steen J.; Hirschberg, Henry

2014-10-01

The overall objective of the research was to investigate the utility of photochemical internalization (PCI) for the enhanced nonviral transfection of genes into glioma cells. The PCI-mediated introduction of the tumor suppressor gene phosphatase and tensin homolog (PTEN) or the cytosine deaminase (CD) pro-drug activating gene into U87 or U251 glioma cell monolayers and multicell tumor spheroids were evaluated. In the study reported here, polyamine-DNA gene polyplexes were encapsulated in a nanoparticle (NP) with an acid degradable polyketal outer shell. These NP synthetically mimic the roles of viral capsid and envelope, which transport and release the gene, respectively. The effects of PCI-mediated suppressor and suicide genes transfection efficiency employing either "naked" polyplex cores alone or as NP-shelled cores were compared. PCI was performed with the photosensitizer AlPcS2a and λ=670-nm laser irradiance. The results clearly demonstrated that the PCI can enhance the delivery of both the PTEN or CD genes in human glioma cell monolayers and multicell tumor spheroids. The transfection efficiency, as measured by cell survival and inhibition of spheroid growth, was found to be significantly greater at suboptimal light and DNA levels for shelled NPs compared with polyamine-DNA polyplexes alone.

HIF-2α mediates a marked increase in migration and stemness characteristics in a subset of glioma cells under hypoxia by activating an Oct-4/Sox-2-Mena (INV) axis.

PubMed

Bhagat, Mohita; Palanichamy, Jayanth Kumar; Ramalingam, Pradeep; Mudassir, Madeeha; Irshad, Khushboo; Chosdol, Kunzang; Sarkar, Chitra; Seth, Pankaj; Goswami, Sumanta; Sinha, Subrata; Chattopadhyay, Parthaprasad

2016-05-01

Hypoxia is a salient feature of most solid tumors and plays a central role in tumor progression owing to its multiple contributions to therapeutic resistance, metastasis, angiogenesis and stemness properties. Reports exist in literature about hypoxia increasing stemness characteristics and invasiveness potential of malignant cells. In order to delineate molecular crosstalk among factors driving glioma progression, we used knockdown and overexpression strategies. We have demonstrated that U87MG and A172 glioma cells inherently have a subset of cells with high migratory potential due to migration-inducing Mena transcripts. These cells also have elevated stemness markers (Sox-2 and Oct-4). There was a significant increase of number in this subset of migratory cells on exposure to hypoxia with corresponding elevation (over 1000 fold) in migration-inducing Mena transcripts. We were able to demonstrate that a HIF-2α-Sox-2/Oct-4-Mena (INV) axis that is strongly activated in hypoxia and markedly increases the migratory potential of the cells. Such cells also formed tumor spheres with greater efficiency. We have correlated our in-vitro results with human glioblastoma samples and found that hypoxia, invasiveness and stemness markers correlated well in native tumor samples. This study identifies a novel signaling mechanism mediated by HIF-2α in regulating invasiveness and stemness characteristics, suggesting that under hypoxic conditions, some tumor cells acquire more migratory potential by increased Pan Mena and Mena INV expression as a consequence of this HIF-2α mediated increase in Oct-4 and Sox-2. These properties would help the cells to form a new nidus after local invasion or metastasis. Copyright © 2016 Elsevier Ltd. All rights reserved.

EFFECT OF HYPOXIA ON THE EXPRESSION OF GENES THAT ENCODE SOME IGFBP AND CCN PROTEINS IN U87 GLIOMA CELLS DEPENDS ON IRE1 SIGNALING.

PubMed

Minchenko, O H; Kharkova, A P; Minchenko, D O; Karbovskyi, L L

2015-01-01

We have studied hypoxic regulation of the expression of different insulin-like growth factor binding protein genes in U87 glioma cells in relation to inhibition of IRE1 (inositol requiring enzyme-1), a central mediator of endoplasmic reticulum stress, which controls cell proliferation and tumor growth. We have demonstrated that hypoxia leads to up-regulation of the expression of IGFBP6, IGFBP7, IGFBP10/CYR61, WISP1, and WISP2 genes and down-regulation--of IGFBP9/NOV gene at the mRNA level in control glioma cells, being more signifcant changes for IGFBP10/CYR61 and WISP2 genes. At the same time, inhibition of IRE1 modifies the effect of hypoxia on the expression of all studied genes: eliminates sensitivity to hypoxia the expression of IGFBP7 and IGFBP9/NOV genes, suppresses effect of hypoxia on IGFBP6, IGFBP10/CYR61, and WISP2 genes, and slightly enhances hypoxic regulation of WISP1 gene expression in glioma cells. We have also demonstrated that the expression of all studied genes in glioma cells is regulated by IRE1 signaling enzyme upon normoxic condition, because inhibition of IRE1 significantly up-regulates IGFBP7, IGFBP10/CYR61, WISP1, and WISP2 genes and down-regulates IGFBP6 and IGFBP9/NOV genes as compared to control glioma cells. The present study demonstrates that hypoxia, which contributes to tumor growth, affects all studied IGFBP and WISP gene expressions and that inhibition of IRE1 preferentially abolishes or suppresses the hypoxic regulation of these gene expressions and thus possibly contributes to slower glioma growth. Moreover, inhibition of IRE1, which correlates with suppression of cell proliferation and glioma growth, is down-regulated expression of pro-proliferative IGFBP genes, attesting to the fact that endoplasmic reticulum stress is a necessary component of malignant tumor growth.

AshwaMAX and Withaferin A inhibits gliomas in cellular and murine orthotopic models

PubMed Central

Pohling, Christoph; Natarajan, Arutselvan; Witney, Timothy H.; Kaur, Jasdeep; Xu, Lingyun; Gowrishankar, Gayatri; D’Souza, Aloma L; Murty, Surya; Schick, Sophie; Chen, Liyin; Wu, Nicholas; Khaw, Phoo; Mischel, Paul; Abbasi, Taher; Usmani, Shahabuddin; Mallick, Parag

2017-01-01

Glioblastoma multiforme (GBM) is an aggressive, malignant cancer Johnson and O’Neill (J Neurooncol 107: 359–364, 2012). An extract from the winter cherry plant (Withania somnifera), AshwaMAX, is concentrated (4.3 %) for Withaferin A; a steroidal lactone that inhibits cancer cells Vanden Berghe et al. (Cancer Epidemiol Biomark Prev 23: 1985–1996, 2014). We hypothesized that AshwaMAX could treat GBM and that bioluminescence imaging (BLI) could track oral therapy in orthotopic murine models of glioblastoma. Human parietal-cortical glioblastoma cells (GBM2, GBM39) were isolated from primary tumors while U87-MG was obtained commercially. GBM2 was transduced with lentiviral vectors that express Green Fluorescent Protein (GFP)/firefly luciferase fusion proteins. Mutational, expression and proliferative status of GBMs were studied. Intracranial xenografts of glioblastomas were grown in the right frontal regions of female, nude mice (n = 3–5 per experiment). Tumor growth was followed through BLI. Neurosphere cultures (U87-MG, GBM2 and GBM39) were inhibited by AshwaMAX at IC50 of 1.4, 0.19 and 0.22 μM equivalent respectively and by Withaferin A with IC50 of 0.31, 0.28 and 0.25 μM respectively. Oral gavage, every other day, of AshwaMAX (40 mg/kg per day) significantly reduced bioluminescence signal (n = 3 mice, p < 0.02, four parameter non-linear regression analysis) in preclinical models. After 30 days of treatment, bioluminescent signal increased suggesting onset of resistance. BLI signal for control, vehicle-treated mice increased and then plateaued. Bioluminescent imaging revealed diffuse growth of GBM2 xenografts. With AshwaMAX, GBM neurospheres collapsed at nanomolar concentrations. Oral treatment studies on murine models confirmed that AshwaMAX is effective against orthotopic GBM. AshwaMAX is thus a promising candidate for future clinical translation in patients with GBM. PMID:26650066

AshwaMAX and Withaferin A inhibits gliomas in cellular and murine orthotopic models.

PubMed

Chang, Edwin; Pohling, Christoph; Natarajan, Arutselvan; Witney, Timothy H; Kaur, Jasdeep; Xu, Lingyun; Gowrishankar, Gayatri; D'Souza, Aloma L; Murty, Surya; Schick, Sophie; Chen, Liyin; Wu, Nicholas; Khaw, Phoo; Mischel, Paul; Abbasi, Taher; Usmani, Shahabuddin; Mallick, Parag; Gambhir, Sanjiv S

2016-01-01

Glioblastoma multiforme (GBM) is an aggressive, malignant cancer Johnson and O'Neill (J Neurooncol 107: 359-364, 2012). An extract from the winter cherry plant (Withania somnifera ), AshwaMAX, is concentrated (4.3 %) for Withaferin A; a steroidal lactone that inhibits cancer cells Vanden Berghe et al. (Cancer Epidemiol Biomark Prev 23: 1985-1996, 2014). We hypothesized that AshwaMAX could treat GBM and that bioluminescence imaging (BLI) could track oral therapy in orthotopic murine models of glioblastoma. Human parietal-cortical glioblastoma cells (GBM2, GBM39) were isolated from primary tumors while U87-MG was obtained commercially. GBM2 was transduced with lentiviral vectors that express Green Fluorescent Protein (GFP)/firefly luciferase fusion proteins. Mutational, expression and proliferative status of GBMs were studied. Intracranial xenografts of glioblastomas were grown in the right frontal regions of female, nude mice (n = 3-5 per experiment). Tumor growth was followed through BLI. Neurosphere cultures (U87-MG, GBM2 and GBM39) were inhibited by AshwaMAX at IC50 of 1.4, 0.19 and 0.22 µM equivalent respectively and by Withaferin A with IC50 of 0.31, 0.28 and 0.25 µM respectively. Oral gavage, every other day, of AshwaMAX (40 mg/kg per day) significantly reduced bioluminescence signal (n = 3 mice, p < 0.02, four parameter non-linear regression analysis) in preclinical models. After 30 days of treatment, bioluminescent signal increased suggesting onset of resistance. BLI signal for control, vehicle-treated mice increased and then plateaued. Bioluminescent imaging revealed diffuse growth of GBM2 xenografts. With AshwaMAX, GBM neurospheres collapsed at nanomolar concentrations. Oral treatment studies on murine models confirmed that AshwaMAX is effective against orthotopic GBM. AshwaMAX is thus a promising candidate for future clinical translation in patients with GBM.

Fluorescein-guided surgery for grade IV gliomas with a dedicated filter on the surgical microscope: preliminary results in 12 cases.

PubMed

Acerbi, Francesco; Broggi, Morgan; Eoli, Marica; Anghileri, Elena; Cuppini, Lucia; Pollo, Bianca; Schiariti, Marco; Visintini, Sergio; Orsi, Chiara; Franzini, Angelo; Broggi, Giovanni; Ferroli, Paolo

2013-07-01

Fluorescein is widely used as a fluorescent tracer for many applications. Its capability to accumulate in cerebral areas with blood-brain barrier damage makes it an ideal dye for intraoperative visualization of malignant gliomas (MG). We report our preliminary experience in fluorescein-guided removal of grade IV gliomas using a dedicated filter on the surgical microscope. In September 2011 we started a prospective phase II trial (FLUOGLIO) to evaluate the safety and obtain initial indications about the efficacy of fluorescein-guided surgery for MG. Patients with suspected MG amenable to complete resection of contrast-enhancing areas were eligible to participate in this study. This report is based on a preliminary analysis of the results of 12 patients with grade IV gliomas out of 15 consecutive cases (age range 48-72 years) enrolled since September 2011. Fluorescein was injected intravenously (i.v.) after intubation (5-10 mg/kg). The tumor was removed using a microsurgical technique and fluorescence visualization by BLU 400 or YELLOW 560 filters on a Pentero microscope (Carl Zeiss, Germany). The study was approved by our ethics committee and registered on the European Regulatory Authorities website (EudraCT no. 2011-002527-18). Histological analysis confirmed grade IV gliomas in 12/15 cases. Median preoperative tumor volume was 33.15 cm(3) (9.6-87.8 cm(3)). No adverse reaction related to the administration of fluorescein was registered. Contrast-enhanced tumor was completely removed in 75 % of the patients. This preliminary analysis suggested that the use of intravenous fluorescein during surgery on grade IV gliomas is safe and allows a high rate of complete resection of contrast-enhanced tumor at the early postoperative MRI.

Targeting delivery of etoposide to inhibit the growth of human glioblastoma multiforme using lactoferrin- and folic acid-grafted poly(lactide-co-glycolide) nanoparticles.

PubMed

Kuo, Yung-Chih; Chen, Yu-Chun

2015-02-01

Lactoferrin (Lf) and folic acid (FA) were crosslinked on poly(lactide-co-glycolide) (PLGA) nanoparticles (NPs) for transporting etoposide across the blood-brain barrier (BBB) and treating human brain malignant glioblastoma. Lf- and FA-grafted PLGA NPs (Lf/FA/PLGA NPs) were employed to permeate the monolayer of human brain-microvascular endothelial cells (HBMECs) regulated by human astrocytes and to inhibit the multiplication of U87MG cells. Lf/FA/PLGA NPs showed a satisfactory entrapment efficiency of etoposide and characteristics of sustained drug release. When compared with PLGA NPs, the permeability coefficient for etoposide across the BBB using Lf/FA/PLGA NPs increased about twofold. The antiproliferative efficacy against the growth of U87MG cells was in the following order: Lf/FA/PLGA NPs>FA/PLGA NPs>PLGA NPs>free etoposide solution. In addition, the targeting ability of Lf/FA/PLGA NPs was evidenced by immunostaining of Lf receptor on HBMECs and folate receptor on U87MG cells during endocytosis. Lf/FA/PLGA NPs with loaded etoposide can be a promising anticancer pharmacotherapy to enhance the delivery of etoposide to malignant brain tumors for preclinical trials. Copyright © 2014 Elsevier B.V. All rights reserved.

Gamma-glutamylcyclotransferase promotes the growth of human glioma cells by activating Notch-Akt signaling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shen, Shang-Hang; Yu, Ning; Liu, Xi-Yao

Glioma as an aggressive type tumor is rapidly growing and has become one of the leading cause of cancer-related death worldwide. γ-Glutamylcyclotransferase (GGCT) has been shown as a diagnostic marker in various cancers. To reveal whether there is a correlation between GGCT and human glioma, GGCT expression in human glioma tissues and cell lines was first determined. We found that GGCT expression was up-regulated in human glioma tissues and cell lines. Further, we demonstrate that GGCT knockdown inhibits glioma cell T98G and U251 proliferation and colony formation, whereas GGCT overexpression leads to oppose effects. GGCT overexpression promotes the expression ofmore » Notch receptors and activates Akt signaling in glioma cells, and Notch-Akt signaling is activated in glioma tissues with high expression of GGCT. Finally, we show that inhibition of Notch-Akt signaling with Notch inhibitor MK-0752 blocks the effects of GGCT on glioma proliferation and colony formation. In conclusion, GGCT plays a critical role in glioma cell proliferation and may be a potential cancer therapeutic target. - Highlights: • GGCT expression is up-regulated in human glioma tissues and cell lines. • GGCT promotes glioma cell growth and colony formation. • GGCT promotes the activation of Notch-Akt signaling in glioma cells and tissues. • Notch inhibition blocks the role of GGCT in human glioma cells.« less

Human IP10-scFv and DC-induced CTL synergistically inhibit the growth of glioma in a xenograft model.

PubMed

Wang, Xuan; Zhang, Fang-Cheng; Zhao, Hong-Yang; Lu, Xiao-Ling; Sun, Yun; Xiong, Zhi-Yong; Jiang, Xiao-Bing

2014-08-01

The epidermal growth factor receptor (EGFR) mutant of EGFRvIII is highly expressed in glioma cells, and the EGFRvIII-specific dendritic cell (DC)-induced tumor antigen-specific CD8(+) cytotoxic T lymphocytes (CTLs) may hold promise in cancer immunotherapy. Interferon (IFN)-γ-inducible protein (IP)-10 (IP-10) is a potent inhibitor of angiogenesis and can recruit CXCR3(+) T cells, including CD8(+) T cells, which are important for the control of tumor growth. In this study, we assessed if the combination of IP10-EGFRvIIIscFv with DC-induced CTLs would improve the therapeutic antitumor efficacy. IP10-scFv was generated by linking the human IP-10 gene with the DNA fragment for anti-EGFRvIIIscFv with a (Gly4Ser)3 flexible linker, purified by affinity chromatography, and characterized for its anti-EGFRvIII immunoreactivity and chemotactic activity. DCs were isolated from human peripheral blood monocyte cells and pulsed with EGFRvIII-peptide, then co-cultured with autologous CD8(+) T cells. BALB/c-nu mice were inoculated with human glioma U87-EGFRvIII cells in the brain and treated intracranially with IP10-scFv and/or intravenously with DC-induced CTLs for evaluating the therapeutic effect. Treatment with both IP10-scFv and EGFRvIII peptide-pulsed, DC-induced CTL synergistically inhibited the growth of glioma and prolonged the survival of tumor-bearing mice, which was accompanied by the inhibition of tumor angiogenesis and enhancement of cytotoxicity, thereby increasing the numbers of brain-infiltrating lymphocytes (BILs) and prolonging the residence time of CTLs in the tumor.

Ruta graveolens L. induces death of glioblastoma cells and neural progenitors, but not of neurons, via ERK 1/2 and AKT activation.

PubMed

Gentile, Maria Teresa; Ciniglia, Claudia; Reccia, Mafalda G; Volpicelli, Floriana; Gatti, Monica; Thellung, Stefano; Florio, Tullio; Melone, Mariarosa A B; Colucci-D'Amato, Luca

2015-01-01

Glioblastoma multiforme is a highly aggressive brain tumor whose prognosis is very poor. Due to early invasion of brain parenchyma, its complete surgical removal is nearly impossible, and even after aggressive combined treatment (association of surgery and chemo- and radio-therapy) five-year survival is only about 10%. Natural products are sources of novel compounds endowed with therapeutic properties in many human diseases, including cancer. Here, we report that the water extract of Ruta graveolens L., commonly known as rue, induces death in different glioblastoma cell lines (U87MG, C6 and U138) widely used to test novel drugs in preclinical studies. Ruta graveolens' effect was mediated by ERK1/2 and AKT activation, and the inhibition of these pathways, via PD98058 and wortmannin, reverted its antiproliferative activity. Rue extract also affects survival of neural precursor cells (A1) obtained from embryonic mouse CNS. As in the case of glioma cells, rue stimulates the activation of ERK1/2 and AKT in A1 cells, whereas their blockade by pharmacological inhibitors prevents cell death. Interestingly, upon induction of differentiation and cell cycle exit, A1 cells become resistant to rue's noxious effects but not to those of temozolomide and cisplatin, two alkylating agents widely used in glioblastoma therapy. Finally, rutin, a major component of the Ruta graveolens water extract, failed to cause cell death, suggesting that rutin by itself is not responsible for the observed effects. In conclusion, we report that rue extracts induce glioma cell death, discriminating between proliferating/undifferentiated and non-proliferating/differentiated neurons. Thus, it can be a promising tool to isolate novel drugs and also to discover targets for therapeutic intervention.

Hypoxic regulation of the expression of cell proliferation related genes in U87 glioma cells upon inhibition of ire1 signaling enzyme

PubMed

Minchenko, O H; Tsymbal, D O; Minchenko, D O; Riabovol, O O; Ratushna, O O; Karbovskyi, L L

2016-01-01

We have studied the effect of inhibition of IRE1 (inositol requiring enzyme 1), which is a central mediator of endoplasmic reticulum stress and a controller of cell proliferation and tumor growth, on hypoxic regulation of the expression of different proliferation related genes in U87 glioma cells. It was shown that hypoxia leads to up-regulation of the expression of IL13RA2, CD24, ING1, ING2, ENDOG, and POLG genes and to down-regulation – of KRT18, TRAPPC3, TSFM, and MTIF2 genes at the mRNA level in control glioma cells. Changes for ING1 and CD24 genes were more significant. At the same time, inhibition of IRE1 modifies the effect of hypoxia on the expression of all studied genes. In particular, it increases sensitivity to hypoxia of the expression of IL13RA2, TRAPPC3, ENDOG, and PLOG genes and suppresses the effect of hypoxia on the expression of ING1 gene. Additionally, it eliminates hypoxic regulation of KRT18, CD24, ING2, TSFM, and MTIF2 genes expressions and introduces sensitivity to hypoxia of the expression of BET1 gene in glioma cells. The present study demonstrates that hypoxia, which often contributes to tumor growth, affects the expression of almost all studied genes. Additionally, inhibition of IRE1 can both enhance and suppress the hypoxic regulation of these gene expressions in a gene specific manner and thus possibly contributes to slower glioma growth, but several aspects of this regulation must be further clarified.

[Overexpressed miRNA-134b inhibits proliferation and invasion of CD133+ U87 glioma stem cells].

PubMed

Liu, Yifeng; Zhang, Baochao; Wen, Changming; Wen, Gongling; Zhou, Guoping; Zhang, Jingwei; He, Haifa; Wang, Ning; Li, Wei

2017-05-01

Objective To investigate the role of microRNA-134b (miR-134b) in the tumorigenesis of glioma stem cells (GSCs) and the possible molecular mechanism. Methods Real-time quantitative PCR (qRT-PCR) was used to evalate the expression of miR-134b in CD133 + and CD133 - U87 GSCs. A lentiviral vector overexpressing miR-134b in U87 GSCs was constructed, and the effect of miR-134b overexpression on matrix metalloproteinase-2 (MMP-2), MMP-9 and MMP-12 expressions at both mRNA and protein levels were detected by qRT-PCR and Western blotting, respectively. Transwell TM assay was performed to determine the effect of miR-134b overexpression on GSCs invasion ability. Tumor xenograft models in nude mice were established to evaluate the effect of miR-134b overexpression on tumorgenesis in vivo. Results The qRT-PCR showed that, compared with CD133 - cells, miR-134b was significantly down-regulated in CD133 + cells. Cell line over-expressing miR-134b was successfully established, and miR-134b was up-regulated significantly compared with empty vector control. Overexpression of miR-134b remarkably inhibited the invasion of U87 GSCs and the expression of MMP-12. However, overexpression of miR-134b did not affect MMP-2 and MMP-9 expressions. miR-134b also suppressed U87 GSCs xenograft growth in vivo. Tumor volume in tumor xenograft model group was significantly lower than that in control group, and tumor weight decreased by 42% in the former group. Conclusion Overexpression of miR-134b inhibits the growth and invasion of CD133 + GSCs.

Cytotoxicity and apoptotic activities of alpha-, gamma- and delta-tocotrienol isomers on human cancer cells.

PubMed

Lim, Su-Wen; Loh, Hwei-San; Ting, Kang-Nee; Bradshaw, Tracey D; Zeenathul, Nazariah A

2014-12-06

Tocotrienols, especially the gamma isomer was discovered to possess cytotoxic effects associated with the induction of apoptosis in numerous cancers. Individual tocotrienol isomers are believed to induce dissimilar apoptotic mechanisms in different cancer types. This study was aimed to compare the cytotoxic potency of alpha-, gamma- and delta-tocotrienols, and to explore their resultant apoptotic mechanisms in human lung adenocarcinoma A549 and glioblastoma U87MG cells which are scarcely researched. The cytotoxic effects of alpha-, gamma- and delta-tocotrienols in both A549 and U87MG cancer cells were first determined at the cell viability and morphological aspects. DNA damage types were then identified by comet assay and flow cytometric study was carried out to support the incidence of apoptosis. The involvements of caspase-8, Bid, Bax and mitochondrial membrane permeability (MMP) in the execution of apoptosis were further expounded. All tocotrienols inhibited the growth of A549 and U87MG cancer cells in a concentration- and time-dependent manner. These treated cancer cells demonstrated some hallmarks of apoptotic morphologies, apoptosis was further confirmed by cell accumulation at the pre-G1 stage. All tocotrienols induced only double strand DNA breaks (DSBs) and no single strand DNA breaks (SSBs) in both treated cancer cells. Activation of caspase-8 leading to increased levels of Bid and Bax as well as cytochrome c release attributed by the disruption of mitochondrial membrane permeability in both A549 and U87MG cells were evident. This study has shown that delta-tocotrienol, in all experimental approaches, possessed a higher efficacy (shorter induction period) and effectiveness (higher induction rate) in the execution of apoptosis in both A549 and U87MG cancer cells as compared to alpha- and gamma-tocotrienols. Tocotrienols in particular the delta isomer can be an alternative chemotherapeutic agent for treating lung and brain cancers.

Antitumor Activity and Mechanism of a Reverse Transcriptase Inhibitor, Dapivirine, in Glioblastoma.

PubMed

Liu, Weiwen; Song, Xian-Lu; Zhao, Shan-Chao; He, Minyi; Wang, Hai; Chen, Ziyang; Xiang, Wei; Yi, Guozhong; Qi, Songtao; Liu, Yawei

2018-01-01

Dapivirine is one of reverse transcriptase inhibitors (RTIs). It is the prototype of diarylpyrimidines (DAPY), formerly known as TMC120 or DAPY R147681 (IUPAC name: 4- [[4-(2, 4, 6-trimethylphenyl) amino]-2-pyrimidinyl] amino]-benzonitrile; CAS no.244767-67-7). The purpose of this study is to investigate the antitumor activity of dapivirine, one of the RTIs, on U87 glioblastoma (GBM) cells in vitro and in vivo . U87 GBM cells were cultured and treated with or without dapivirine. Cell viability was evaluated by CCK-8 (Cell Counting Kit 8, CCK-8) assay; apoptosis was analyzed by flow cytometry; cell migration was evaluated by Boyden Chamber assay; Western blotting was performed to detect proteins related to apoptosis, epithelial-to-mesenchymal transition and autophagy. PathScan intracellular signaling array kit was used to detect important and well-characterized signaling molecules. Tumor xenograft model in nude mice was used to evaluate the antitumorigenic effect in vivo . Dapivirine weakened proliferation of glioma cells and induced the apoptosis of U87 glioblastoma cells. Furthermore, dapivirine regulated autophagy and induced Akt, Bad and SAPK/JNK activations. Moreover, the inhibition of glioma cell growth by dapivirine was also observed in nude mice in vivo . In summary, in our study dapivirine exposure induces stress, resulting in JNK and PI3K/Akt pathway activation through diminished inhibition of the apoptosis and autophagy cascade in U87 GBM cells, which inhibits cell growth in vitro and in vivo .

Saponin 1 Induces Apoptosis and Suppresses NF-κB-Mediated Survival Signaling in Glioblastoma Multiforme (GBM)

PubMed Central

Tang, Chi; Li, Bo; Wang, Yuangang; Gao, Zhenhui; Luo, Peng; Yin, Anan; Wang, Xiaoyang; Cheng, Guang; Fei, Zhou

2013-01-01

Saponin 1 is a triterpeniod saponin extracted from Anemone taipaiensis, a traditional Chinese medicine against rheumatism and phlebitis. It has also been shown to exhibit significant anti-tumor activity against human leukemia (HL-60 cells) and human hepatocellular carcinoma (Hep-G2 cells). Herein we investigated the effect of saponin 1 in human glioblastoma multiforme (GBM) U251MG and U87MG cells. Saponin 1 induced significant growth inhibition in both glioblastoma cell lines, with a 50% inhibitory concentration at 24 h of 7.4 µg/ml in U251MG cells and 8.6 µg/ml in U87MG cells, respectively. Nuclear fluorescent staining and electron microscopy showed that saponin 1 caused characteristic apoptotic morphological changes in the GBM cell lines. Saponin 1-induced apoptosis was also verified by DNA ladder electrophoresis and flow cytometry. Additionally, immunocytochemistry and western blotting analyses revealed a time-dependent decrease in the expression and nuclear location of NF-κB following saponin 1 treatment. Western blotting data indicated a significant decreased expression of inhibitors of apoptosis (IAP) family members,(e.g., survivin and XIAP) by saponin 1. Moreover, saponin 1 caused a decrease in the Bcl-2/Bax ratio and initiated apoptosis by activating caspase-9 and caspase-3 in the GBM cell lines. These findings indicate that saponin 1 inhibits cell growth of GBM cells at least partially by inducing apoptosis and inhibiting survival signaling mediated by NF-κB. In addition, in vivo study also demonstrated an obvious inhibition of saponin 1 treatment on the tumor growth of U251MG and U87MG cells-produced xenograft tumors in nude mice. Given the minimal toxicities of saponin 1 in non-neoplastic astrocytes, our results suggest that saponin 1 exhibits significant in vitro and in vivo anti-tumor efficacy and merits further investigation as a potential therapeutic agent for GBM. PMID:24278406

Inhibition of autophagy induced by proteasome inhibition increases cell death in human SHG-44 glioma cells.

PubMed

Ge, Peng-Fei; Zhang, Ji-Zhou; Wang, Xiao-Fei; Meng, Fan-Kai; Li, Wen-Chen; Luan, Yong-Xin; Ling, Feng; Luo, Yi-Nan

2009-07-01

The ubiquitin-proteasome system (UPS) and lysosome-dependent macroautophagy (autophagy) are two major intracellular pathways for protein degradation. Recent studies suggest that proteasome inhibitors may reduce tumor growth and activate autophagy. Due to the dual roles of autophagy in tumor cell survival and death, the effect of autophagy on the destiny of glioma cells remains unclear. In this study, we sought to investigate whether inhibition of the proteasome can induce autophagy and the effects of autophagy on the fate of human SHG-44 glioma cells. The proteasome inhibitor MG-132 was used to induce autophagy in SHG-44 glioma cells, and the effect of autophagy on the survival of SHG-44 glioma cells was investigated using an autophagy inhibitor 3-MA. Cell viability was measured by MTT assay. Apoptosis and cell cycle were detected by flow cytometry. The expression of autophagy related proteins was determined by Western blot. MG-132 inhibited cell proliferation, induced cell death and cell cycle arrest at G(2)/M phase, and activated autophagy in SHG-44 glioma cells. The expression of autophagy-related Beclin-1 and LC3-I was significantly up-regulated and part of LC3-I was converted into LC3-II. However, when SHG-44 glioma cells were co-treated with MG-132 and 3-MA, the cells became less viable, but cell death and cell numbers at G(2)/M phase increased. Moreover, the accumulation of acidic vesicular organelles was decreased, the expression of Beclin-1 and LC3 was significantly down-regulated and the conversion of LC3-II from LC3-I was also inhibited. Inhibition of the proteasome can induce autophagy in human SHG-44 glioma cells, and inhibition of autophagy increases cell death. This discovery may shed new light on the effect of autophagy on modulating the fate of SHG-44 glioma cells.Acta Pharmacologica Sinica (2009) 30: 1046-1052; doi: 10.1038/aps.2009.71.

KCTD2, an adaptor of Cullin3 E3 ubiquitin ligase, suppresses gliomagenesis by destabilizing c-Myc

PubMed Central

Kim, Eun-Jung; Kim, Sung-Hak; Jin, Xiong; Jin, Xun; Kim, Hyunggee

2017-01-01

Cullin3 E3 ubiquitin ligase ubiquitinates a wide range of substrates through substrate-specific adaptors Bric-a-brac, Tramtrack, and Broad complex (BTB) domain proteins. These E3 ubiquitin ligase complexes are involved in diverse cellular functions. Our recent study demonstrated that decreased Cullin3 expression induces glioma initiation and correlates with poor prognosis of patients with malignant glioma. However, the substrate recognition mechanism associated with tumorigenesis is not completely understood. Through yeast two-hybrid screening, we identified potassium channel tetramerization domain-containing 2 (KCTD2) as a BTB domain protein that binds to Cullin3. The interaction of Cullin3 and KCTD2 was verified using immunoprecipitation and immunofluorescence. Of interest, KCTD2 expression was markedly decreased in patient-derived glioma stem cells (GSCs) compared with non-stem glioma cells. Depletion of KCTD2 using a KCTD2-specific short-hairpin RNA in U87MG glioma cells and primary Ink4a/Arf-deficient murine astrocytes markedly increased self-renewal activity in addition with an increased expression of stem cell markers, and mouse in vivo intracranial tumor growth. As an underlying mechanism for these KCTD2-mediated phenotypic changes, we demonstrated that KCTD2 interacts with c-Myc, which is a key stem cell factor, and causes c-Myc protein degradation by ubiquitination. As a result, KCTD2 depletion acquires GSC features and affects aerobic glycolysis via expression changes in glycolysis-associated genes through c-Myc protein regulation. Of clinical significance was our finding that patients having a profile of KCTD2 mRNA-low and c-Myc gene signature-high, but not KCTD2 mRNA-low and c-Myc mRNA-high, are strongly associated with poor prognosis. This study describes a novel regulatory mode of c-Myc protein in malignant gliomas and provides a potential framework for glioma therapy by targeting c-Myc function. PMID:28060381

The delayed luminescence spectroscopy as tool to investigate the cytotoxic effect on human cancer cells of drug-loaded nanostructured lipid carrier

NASA Astrophysics Data System (ADS)

Grasso, R.; Gulino, M.; Scordino, A.; Musumeci, F.; Campisi, A.; Bonfanti, R.; Carbone, C.; Puglisi, G.

2016-05-01

The first results concerning the possibility to use Delayed Luminescence spectroscopy to evaluate the in vitro induction of cytotoxic effects on human glioblastoma cells of nanostructured lipid carrier and drug-loaded nanostructured lipid carrier are showed in this contribution. We tested the effects of nanostructured lipid carrier, ferulic acid and ferulic acidloaded nanostructured lipid carrier on U-87MG cell line. The study seems to confirm the ability of Delayed Luminescence to be sensible indicator of alterations induced on functionality of the mitochondrial respiratory chain complex I in U-87MG cancer cells when treated with nanostructured lipid carriers.

D-Amino acid oxidase-induced oxidative stress, 3-bromopyruvate and citrate inhibit angiogenesis, exhibiting potent anticancer effects.

PubMed

El Sayed, S M; El-Magd, R M Abou; Shishido, Y; Yorita, K; Chung, S P; Tran, D H; Sakai, T; Watanabe, H; Kagami, S; Fukui, K

2012-10-01

Angiogenesis is critical for cancer growth and metastasis. Steps of angiogenesis are energy consuming, while vascular endothelial cells are highly glycolytic. Glioblastoma multiforme (GBM) is a highly vascular tumor and this enhances its aggressiveness. D-amino acid oxidase (DAO) is a promising therapeutic protein that induces oxidative stress upon acting on its substrates. Oxidative stress-energy depletion (OSED) therapy was recently reported (El Sayed et al., Cancer Gene Ther, 19, 1-18, 2012). OSED combines DAO-induced oxidative stress with energy depletion caused by glycolytic inhibitors such as 3-bromopyruvate (3BP), a hexokinase II inhibitor that depleted ATP in cancer cells and induced production of hydrogen peroxide. 3BP disturbs the Warburg effect and antagonizes effects of lactate and pyruvate (El Sayed et al., J Bioenerg Biomembr, 44, 61-79, 2012). Citrate is a natural organic acid capable of inhibiting glycolysis by targeting phosphofructokinase. Here, we report that DAO, 3BP and citrate significantly inhibited angiogenesis, decreased the number of vascular branching points and shortened the length of vascular tubules. OSED delayed the growth of C6/DAO glioma cells. 3BP combined with citrate delayed the growth of C6 glioma cells and decreased significantly the number and size of C6 glioma colonies in soft agar. Human GBM cells (U373MG) were resistant to chemotherapy e.g. cisplatin and cytosine arabinoside, while 3BP was effective in decreasing the viability and disturbing the morphology of U373MG cells.

Polish Natural Bee Honeys Are Anti-Proliferative and Anti-Metastatic Agents in Human Glioblastoma multiforme U87MG Cell Line

PubMed Central

Moskwa, Justyna; Borawska, Maria H.; Markiewicz-Zukowska, Renata; Puscion-Jakubik, Anna; Naliwajko, Sylwia K.; Socha, Katarzyna; Soroczynska, Jolanta

2014-01-01

Honey has been used as food and a traditional medicament since ancient times. However, recently many scientists have been concentrating on the anti-oxidant, anti-proliferative, anti-inflammatory and other properties of honey. In this study, we investigated for the first time an anticancer effect of different honeys from Poland on tumor cell line - glioblastoma multiforme U87MG. Anti-proliferative activity of honeys and its interferences with temozolomide were determined by a cytotoxicity test and DNA binding by [H3]-thymidine incorporation. A gelatin zymography was used to conduct an evaluation of metalloproteinases (MMP-2 and MMP-9) expression in U87MG treatment with honey samples. The honeys were previously tested qualitatively (diastase activity, total phenolic content, lead and cadmium content). The data demonstrated that the examined honeys have a potent anti-proliferative effect on U87MG cell line in a time- and dose-dependent manner, being effective at concentrations as low as 0.5% (multifloral light honey - viability 53% after 72 h of incubation). We observed that after 48 h, combining honey with temozolomide showed a significantly higher inhibitory effect than the samples of honey alone. We observed a strong inhibition of MMP-2 and MMP-9 for the tested honeys (from 20 to 56% and from 5 to 58% compared to control, respectively). Our results suggest that Polish honeys have an anti-proliferative and anti-metastatic effect on U87MG cell line. Therefore, natural bee honey can be considered as a promising adjuvant treatment for brain tumors. PMID:24594866

Juglone reduces growth and migration of U251 glioblastoma cells and disrupts angiogenesis.

PubMed

Wang, Jian; Liu, Ke; Wang, Xiao-Feng; Sun, Dian-Jun

2017-10-01

Accumulating data show that prolylisomerase (Pin1) is overexpressed in human glioblastoma multiforme (GBM) specimens. Therefore, Pin1 inhibitors should be investigated as a new chemotherapeutic drug that may enhance the clinical management of human gliomas. Recently, juglone, a Pin1 inhibitor, was shown to exhibit potent anticancer activity in various tumor cells, but its role in human glioma cells remains unknown. In the present study, we determined if juglone exerts antitumor effects in the U251 human glioma cell line and investigated its potential underlying molecular mechanisms. Cell survival, apoptosis, migration, angiogenesis and molecular targets were identified with multiple detection techniques including the MTT cell proliferation assay, dual acridine orange/ethidium bromide staining, electron microscopy, transwell migration assay, chick chorioallantoic membrane assay, quantitative real-time polymerase chain reaction and immunoblotting. The results showed that 5-20 µM juglone markedly suppressed cell proliferation, induced apoptosis, and enhanced caspase-3 activity in U251 cells in a dose- and time-dependent manner. Moreover, juglone inhibited cell migration and the formation of new blood vessels. At the molecular level, juglone markedly suppressed Pin1 levels in a time-dependent manner. TGF-β1/Smad signaling, a critical upstream regulator of miR-21, was also suppressed by juglone. Moreover, the transient overexpression of Pin1 reversed its antitumor effects in U251 cells and inhibited juglone-mediated changes to the TGF-β1/miR-21 signaling pathway. These findings suggest that juglone inhibits cell growth by causing apoptosis, thereby inhibiting the migration of U251 glioma cells and disrupting angiogenesis; and that Pin1 is a critical target for juglone's antitumor activity. The present study provides evidence that juglone has in vitro efficacy against glioma. Therefore, additional studies are warranted to examine the clinical potential of juglone in human gliomas.

Suppression of STIM1 inhibits human glioblastoma cell proliferation and induces G0/G1 phase arrest

PubMed Central

2013-01-01

Background Depletion of calcium (Ca2+) from the endoplasmic reticulum (ER) activates the ubiquitous store-operated Ca2+ entry (SOCE) pathway which sustains long-term Ca2+ signals and is critical for cellular functions. Stromal interacting molecule 1 (STIM1) serves a dual role as an ER Ca2+ sensor and activator of SOCE. Aberrant expression of STIM1 could be observed in several human cancer cells. However, the role of STIM1 in regulating tumorigenesis of human glioblastoma still remains unclear. Methods Expression of STIM1 protein in a panel of human glioblastoma cell lines (U251, U87 and U373) in different transformation level were evaluated by Western blot method. STIM1 loss of function was performed on U251 cells, derived from grade IV astrocytomas-glioblastoma multiforme with a lentvirus-mediated short harpin RNA (shRNA) method. The biological impacts after knock down of STIM1 on glioblastoma cells were investigated in vitro and in vivo. Results We discovered that STIM1 protein was expressed in U251, U87 and U373 cells, and especially higher in U251 cells. RNA interference efficiently downregulated the expression of STIM1 in U251 cells at both mRNA and protein levels. Specific downregulation of STIM1 inhibited U251 cell proliferation by inducing cell cycle arrest in G0/G1 phase through regulation of cell cycle-related genes, such as p21Waf1/Cip1, cyclin D1 and cyclin-dependent kinase 4 (CDK4), and the antiproliferative effect of STIM1 silencing was also observed in U251 glioma xenograft tumor model. Conclusion Our findings confirm STIM1 as a rational therapeutic target in human glioblastoma, and also indicate that lentivirus-mediated STIM1 silencing is a promising therapeutic strategy for human glioblastoma. PMID:23578185

Suppression of STIM1 inhibits human glioblastoma cell proliferation and induces G0/G1 phase arrest.

PubMed

Li, Guilin; Zhang, Zhenxing; Wang, Renzhi; Ma, Wenbin; Yang, Ying; Wei, Junji; Wei, Yanping

2013-04-11

Depletion of calcium (Ca2+) from the endoplasmic reticulum (ER) activates the ubiquitous store-operated Ca2+ entry (SOCE) pathway which sustains long-term Ca2+ signals and is critical for cellular functions. Stromal interacting molecule 1 (STIM1) serves a dual role as an ER Ca2+ sensor and activator of SOCE. Aberrant expression of STIM1 could be observed in several human cancer cells. However, the role of STIM1 in regulating tumorigenesis of human glioblastoma still remains unclear. Expression of STIM1 protein in a panel of human glioblastoma cell lines (U251, U87 and U373) in different transformation level were evaluated by Western blot method. STIM1 loss of function was performed on U251 cells, derived from grade IV astrocytomas-glioblastoma multiforme with a lentvirus-mediated short harpin RNA (shRNA) method. The biological impacts after knock down of STIM1 on glioblastoma cells were investigated in vitro and in vivo. We discovered that STIM1 protein was expressed in U251, U87 and U373 cells, and especially higher in U251 cells. RNA interference efficiently downregulated the expression of STIM1 in U251 cells at both mRNA and protein levels. Specific downregulation of STIM1 inhibited U251 cell proliferation by inducing cell cycle arrest in G0/G1 phase through regulation of cell cycle-related genes, such as p21Waf1/Cip1, cyclin D1 and cyclin-dependent kinase 4 (CDK4), and the antiproliferative effect of STIM1 silencing was also observed in U251 glioma xenograft tumor model. Our findings confirm STIM1 as a rational therapeutic target in human glioblastoma, and also indicate that lentivirus-mediated STIM1 silencing is a promising therapeutic strategy for human glioblastoma.

Magnolol and honokiol exert a synergistic anti-tumor effect through autophagy and apoptosis in human glioblastomas

PubMed Central

Cheng, Yu-Chen; Hueng, Dueng-Yuan; Huang, Hua-Yin; Chen, Jang-Yi; Chen, Ying

2016-01-01

Glioblastoma (GBM) is a malignant brain tumor associated with a high mortality rate. The aim of this study is to investigate the synergistic effects of honokiol (Hono) and magnolol (Mag), extracted from Magnolia officinalis, on cytotoxicity and inhibition of human GBM tumor progression in cellular and animal models. In comparison with Hono or Mag alone, co-treatment with Hono and Mag (Hono-Mag) decreased cyclin A, D1 and cyclin-dependent kinase 2, 4, 6 significantly, leading to cell cycle arrest in U87MG and LN229 human glioma cells. In addition, phosphorylated phosphoinositide 3-kinase (p-PI3K), p-Akt, and Ki67 were decreased after Hono-Mag treatment, showing proliferation inhibition. Hono-Mag treatment also reduced p-p38 and p-JNK but elevated p-ERK expression. Besides, Hono-Mag treatment induced autophagy and intrinsic and extrinsic apoptosis. Both ERK and autophagy inhibitors enhanced Hono-Mag-induced apoptosis in LN229 cells, indicating a rescuer role of ERK. In human GBM orthotopic xenograft model, the Hono-Mag treatment inhibited the tumor progression and induced apoptosis more efficiently than Temozolomide, Hono, or Mag group. In conclusion, the Hono-Mag exerts a synergistic anti-tumor effect by inhibiting cell proliferation and inducing autophagy and apoptosis in human GBM cells. The Hono-Mag may be applied as an adjuvant therapy to improve the therapeutic efficacy of GBM treatment. PMID:27074557

Impact of meriolins, a new class of cyclin-dependent kinase inhibitors, on malignant glioma proliferation and neo-angiogenesis.

PubMed

Jarry, Marie; Lecointre, Céline; Malleval, Céline; Desrues, Laurence; Schouft, Marie-Thérèse; Lejoncour, Vadim; Liger, François; Lyvinec, Gildas; Joseph, Benoît; Loaëc, Nadège; Meijer, Laurent; Honnorat, Jérôme; Gandolfo, Pierrick; Castel, Hélène

2014-11-01

Glioblastomas are the most frequent and most aggressive primary brain tumors in adults. The median overall survival is limited to a few months despite surgery, radiotherapy, and chemotherapy. It is now clearly established that hyperactivity of cyclin-dependent kinases (CDKs) is one of the processes underlying hyperproliferation and tumoral growth. The marine natural products meridianins and variolins, characterized as CDK inhibitors, display a kinase-inhibitory activity associated with cytotoxic effects. In order to improve selectivity and efficiency of these CDK inhibitors, a series of hybrid compounds called meriolins have been synthesized. The potential antitumoral activity of meriolins was investigated in vitro on glioma cell lines (SW1088 and U87), native neural cells, and a human endothelial cell line (HUV-EC-C). The impact of intraperitoneal or intratumoral administrations of meriolin 15 was evaluated in vivo on 2 different nude mice-xenografted glioma models. Meriolins 3, 5, and 15 exhibited antiproliferative properties with nanomolar IC50 and induced cell-cycle arrest and CDK inhibition associated with apoptotic events in human glioma cell lines. These meriolins blocked the proliferation rate of HUV-EC-C through cell cycle arrest and apoptosis. In vivo, meriolin 15 provoked a robust reduction in tumor volume in spite of toxicity for highest doses, associated with inhibition of cell division, activation of caspase 3, reduction of CD133 cells, and modifications of the vascular architecture. Meriolins, and meriolin 15 in particular, exhibit antiproliferative and proapoptotic activities on both glioma and intratumoral endothelial cells, constituting key promising therapeutic lead compounds for the treatment of glioblastoma. © The Author(s) 2014. Published by Oxford University Press on behalf of the Society for Neuro-Oncology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Impact of meriolins, a new class of cyclin-dependent kinase inhibitors, on malignant glioma proliferation and neo-angiogenesis

PubMed Central

Jarry, Marie; Lecointre, Céline; Malleval, Céline; Desrues, Laurence; Schouft, Marie-Thérèse; Lejoncour, Vadim; Liger, François; Lyvinec, Gildas; Joseph, Benoît; Loaëc, Nadège; Meijer, Laurent; Honnorat, Jérôme; Gandolfo, Pierrick; Castel, Hélène

2014-01-01

Background Glioblastomas are the most frequent and most aggressive primary brain tumors in adults. The median overall survival is limited to a few months despite surgery, radiotherapy, and chemotherapy. It is now clearly established that hyperactivity of cyclin-dependent kinases (CDKs) is one of the processes underlying hyperproliferation and tumoral growth. The marine natural products meridianins and variolins, characterized as CDK inhibitors, display a kinase-inhibitory activity associated with cytotoxic effects. In order to improve selectivity and efficiency of these CDK inhibitors, a series of hybrid compounds called meriolins have been synthesized. Methods The potential antitumoral activity of meriolins was investigated in vitro on glioma cell lines (SW1088 and U87), native neural cells, and a human endothelial cell line (HUV-EC-C). The impact of intraperitoneal or intratumoral administrations of meriolin 15 was evaluated in vivo on 2 different nude mice-xenografted glioma models. Results Meriolins 3, 5, and 15 exhibited antiproliferative properties with nanomolar IC50 and induced cell-cycle arrest and CDK inhibition associated with apoptotic events in human glioma cell lines. These meriolins blocked the proliferation rate of HUV-EC-C through cell cycle arrest and apoptosis. In vivo, meriolin 15 provoked a robust reduction in tumor volume in spite of toxicity for highest doses, associated with inhibition of cell division, activation of caspase 3, reduction of CD133 cells, and modifications of the vascular architecture. Conclusion Meriolins, and meriolin 15 in particular, exhibit antiproliferative and proapoptotic activities on both glioma and intratumoral endothelial cells, constituting key promising therapeutic lead compounds for the treatment of glioblastoma. PMID:24891448

The sensitivity of glioma cells to pyropheophorbide-αmethyl ester-mediated photodynamic therapy is enhanced by inhibiting ABCG2.

PubMed

Pan, Li; Lin, Haidan; Tian, Si; Bai, Dingqun; Kong, Yuhan; Yu, Lehua

2017-09-01

To study the mechanisms of human glioblastoma cell resistance to methyl ester pyropheophorbide-a-mediated photodynamic therapy (MPPa-PDT) and the relationship between the cells and adenosine triphosphate-binding cassette superfamily G member 2 (ABCG2). The sensitivity of four human glioma cell lines (U87, A172, SHG-44, and U251) to MPPa-PDT was detected with a CCK-8 assay. Cell apoptosis, intracellular MPPa, and singlet oxygen were tested with flow cytometry. The mRNA and protein expression of ATP-binding cassette transporters (ABCG2, MRP1, and MDR1) were detected by PCR and Western blot, respectively. Both the sensitivity to MPPa-PDT and intracellular MPPa in A172 were the lowest among the four cell lines, while expression of ABCG2 mRNA and protein in A172 were the highest. The intracellular MPPa and ROS in A172 receiving MPPa-PDT significantly increased after using the ABCG2 inhibitor fumitremorgin C (FTC). Both cell viability and apoptosis in A172 cells undergoing MPPa-PDT were significantly improved with FTC. ABCG2 plays a significant role in the resistance of A172 to MPPa-PDT. Lasers Surg. Med. 49:719-726, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Release of tissue inhibitor of metalloproteinase-2 from alginate microcapsule encapsulating genetically engineered cells

PubMed Central

Kim, Yeon Seong; Jeong, Young-II; Jin, Shu-Guang; Pei, Jian; Wen, Min; Kim, In-Young; Moon, Kyung-Sub; Jung, Tae-Young; Ryu, Hyang-Hwa; Jung, Shin

2013-01-01

Background In this study, 293T cells were genetically engineered to secrete tissue inhibitor of metalloproteinase-2 (TIMP2) and encapsulated into alginate microcapsules to continuously release TIMP2 protein. Methods The anti-invasive potential of the microcapsules was studied in vitro using brain tumor cells. The TIMP2 gene was transfected to 293T cells, and genetically engineered 293TIMP2 cells were encapsulated into alginate microcapsules. Release of TIMP2 protein was detected with Western blot analysis and the anti-invasive potential against U87MG cells was tested using gelatin zymography and a Matrigel assay. Results Cell viability within the alginate microcapsules was maintained at a cell density of 5 × 106. Because polycationic polymers are helpful for maintaining the mechanical strength of microcapsules with good cell viability, the alginate microcapsules were reinforced with chitosan (0.1% w/v). Expression of TIMP2 protein in cell lysates and secretion of TIMP2 into the conditioned medium was confirmed by Western blot analysis. Alginate microcapsules encapsulating 293TIMP2 cells released TIMP2 protein into the medium efficiently, where the TIMP2 protein participated in degradation of the matrix metalloproteinase-2 enzyme and inhibited invasion of U87MG cells. Conclusion Alginate microcapsules encapsulating 293TIMP2 cells are promising candidates for anti-invasive treatment of glioma. PMID:24231999

Effective conversion of irinotecan to SN-38 after intratumoral drug delivery to an intracranial murine glioma model in vivo. Laboratory investigation.

PubMed

Wang, Weijun; Ghandi, Alex; Liebes, Leonard; Louie, Stan G; Hofman, Florence M; Schönthal, Axel H; Chen, Thomas C

2011-03-01

Irinotecan (CPT-11), a topoisomerase I inhibitor, is a cytotoxic agent with activity against malignant gliomas and other tumors. After systemic delivery, CPT-11 is converted to its active metabolite, SN-38, which displays significantly higher cytotoxic potency. However, the achievement of therapeutically effective plasma levels of CPT-11 and SN-38 is seriously complicated by variables that affect drug metabolism in the liver. Thus the capacity of CPT-11 to be converted to the active SN38 intratumorally in gliomas was addressed. For in vitro studies, 2 glioma cell lines, U87 and U251, were tested to determine the cytotoxic effects of CPT-11 and SN-38 in a dose-dependent manner. In vivo studies were performed by implanting U87 intracranially into athymic/nude mice. For a period of 2 weeks, SN-38, CPT-11, or vehicle was administered intratumorally by means of an osmotic minipump. One series of experiments measured the presence of SN-38 or CPT-11 in the tumor and surrounding brain tissues after 2 weeks' exposure to the drug. In a second series of experiments, after 2 weeks' exposure to the drug, the animals were maintained, in the absence of drug, until death. The survival curves were then calculated. The results show that the animals that had CPT-11 delivered intratumorally by the minipump expressed SN-38 in vivo. Furthermore, both CPT-11 and SN-38 accumulated at higher levels in tumor tissues compared with uninvolved brain. Intratumoral delivery of CPT-11 or SN-38 extended the average survival time of tumor-bearing animals from 22 days to 46 and 65 days, respectively. These results demonstrate that intratumorally administered CPT-11 can be effectively converted to SN-38 and this method of drug delivery is effective in extending the survival time of animals bearing malignant gliomas.

Characterization of Arginase Expression in Glioma-Associated Microglia and Macrophages

PubMed Central

Zhang, Leying; Gao, Hang; Song, Yanyan; Ren, Hui; Ouyang, Mao; Wu, Xiwei; D’Apuzzo, Massimo; Badie, Behnam

2016-01-01

Microglia (MG) and macrophages (MPs) represent a significant component of the inflammatory response to gliomas. When activated, MG/MP release a variety of pro-inflammatory cytokines, however, they also secrete anti-inflammatory factors that limit their cytotoxic function. The balance between pro and anti-inflammatory functions dictates their antitumor activity. To evaluate potential variations in MG and MP function in gliomas, we isolated these cells (and other Gr1+ cells) from intracranial GL261 murine gliomas by FACS and evaluated their gene expression profiles by microarray analysis. As expected, arginase 1 (Arg1, M2 marker) was highly expressed by tumor-associated Gr1+, MG and MP. However, in contrast to MP and Gr1+ cells that expressed Arg1 shortly after tumor trafficking, Arg1 expression in MG was delayed and occurred in larger tumors. Interestingly, depletion of MPs in tumors did not prevent MG polarization, suggesting direct influence of tumor-specific factors on MG Arg1 upregulation. Finally, Arg1 expression was confirmed in human GBM samples, but most Arg1+ cells were neutrophils and not MPs. These findings confirm variations in tumor MG and MP polarization states and its dependency on tumor microenvironmental factors. PMID:27936099

Characterization of Arginase Expression in Glioma-Associated Microglia and Macrophages.

PubMed

Zhang, Ian; Alizadeh, Darya; Liang, Junling; Zhang, Leying; Gao, Hang; Song, Yanyan; Ren, Hui; Ouyang, Mao; Wu, Xiwei; D'Apuzzo, Massimo; Badie, Behnam

2016-01-01

Microglia (MG) and macrophages (MPs) represent a significant component of the inflammatory response to gliomas. When activated, MG/MP release a variety of pro-inflammatory cytokines, however, they also secrete anti-inflammatory factors that limit their cytotoxic function. The balance between pro and anti-inflammatory functions dictates their antitumor activity. To evaluate potential variations in MG and MP function in gliomas, we isolated these cells (and other Gr1+ cells) from intracranial GL261 murine gliomas by FACS and evaluated their gene expression profiles by microarray analysis. As expected, arginase 1 (Arg1, M2 marker) was highly expressed by tumor-associated Gr1+, MG and MP. However, in contrast to MP and Gr1+ cells that expressed Arg1 shortly after tumor trafficking, Arg1 expression in MG was delayed and occurred in larger tumors. Interestingly, depletion of MPs in tumors did not prevent MG polarization, suggesting direct influence of tumor-specific factors on MG Arg1 upregulation. Finally, Arg1 expression was confirmed in human GBM samples, but most Arg1+ cells were neutrophils and not MPs. These findings confirm variations in tumor MG and MP polarization states and its dependency on tumor microenvironmental factors.

A nutrient mixture inhibits glioblastoma xenograft U-87 MG growth in male nude mice.

PubMed

Roomi, M W; Kalinovsky, T; Rath, M; Niedzwiecki, A

2016-03-01

Brain tumors are highly aggressive tumors characterized by secretions of high levels of matrix metalloproteinase-2 and -9, leading to tumor growth, invasion and metastasis by digesting the basement membrane and extracellular matrix components. We previously demonstrated the effectiveness of a nutrient mixture (NM) containing ascorbic acid, lysine, proline, and green tea extract in vitro: on activity of urokinase plasminogen activator, matrix metalloproteinases and TIMPs in various human glioblastoma (LN-18, T-98G and A-172) cell lines and on glioblastoma A-172 cell proliferation and Matrigel invasion. Our main objective in this study was to investigate the effect of the NM in vivo on human glioblastoma U-87 MG cell line. Athymic male nude mice inoculated with 3·10(6) U-87 MG cells subcutaneously and were fed a regular diet or a regular diet supplemented with 0.5% NM. Four weeks later, the mice were sacrificed, the tumors were weighed and measured. The samples were studied histologically. NM inhibited tumor weight and tumor burden by 53% (p = 0.015) and 48% (p = 0.010), respectively. These results suggest the therapeutic potential of NM as an adjuvant in the treatment of glioblastoma.

INHIBITION OF ERN1 SIGNALING ENZYME AFFECTS HYPOXIC REGULATION OF THE EXPRESSION OF E2F8, EPAS1, HOXC6, ATF3, TBX3 AND FOXF1 GENES IN U87 GLIOMA CELLS.

PubMed

Minchenko, O H; Tsymbal, D O; Minchenko, D O; Kovalevska, O V; Karbovskyi, L L; Bikfalvi, A

2015-01-01

Hypoxia as well as the endoplasmic reticulum stress are important factors of malignant tumor growth and control of the expression of genes, which regulate numerous metabolic processes and cell proliferation. Furthermore, blockade of ERN1 (endoplasmic reticulum to nucleus 1) suppresses cell proliferation and tumor growth. We studied the effect of hypoxia on the expression of genes encoding the transcription factors such as E2F8 (E2F transcription factor 8), EPAS1 (endothelial PAS domain protein 1), TBX3 (T-box 3), ATF3 (activating transcription factor 3), FOXF1 (forkhead box F), and HOXC6 (homeobox C6) in U87 glioma cells with and without ERN1 signaling enzyme function. We have established that hypoxia enhances the expression of HOXC6, E2F8, ATF3, and EPAS1 genes but does not change TBX3 and FOXF1 gene expression in glioma cells with ERNI function. At the same time, the expression level of all studied genes is strongly decreased, except for TBX3 gene, in glioma cells without ERN1 function. Moreover, the inhibition of ERN1 signaling enzyme function significantly modifies the effect of hypoxia on the expression of these transcription factor genes. removes or introduces this regulation as well as changes a direction or magnitude of hypoxic regulation. Present study demonstrates that fine-tuning of the expression of proliferation related genes depends upon hypoxia and ERN1-mediated endoplasmic reticulum stress signaling and correlates with slower proliferation rate of glioma cells without ERN1 function.

Antitumor Activity and Mechanism of a Reverse Transcriptase Inhibitor, Dapivirine, in Glioblastoma

PubMed Central

Liu, Weiwen; Song, Xian-lu; Zhao, Shan-chao; He, Minyi; Wang, Hai; Chen, Ziyang; Xiang, Wei; Yi, Guozhong; Qi, Songtao; Liu, Yawei

2018-01-01

Ethnopharmacological relevance: Dapivirine is one of reverse transcriptase inhibitors (RTIs). It is the prototype of diarylpyrimidines (DAPY), formerly known as TMC120 or DAPY R147681 (IUPAC name: 4- [[4-(2, 4, 6-trimethylphenyl) amino]-2-pyrimidinyl] amino]-benzonitrile; CAS no.244767-67-7). Aim: The purpose of this study is to investigate the antitumor activity of dapivirine, one of the RTIs, on U87 glioblastoma (GBM) cells in vitro and in vivo. Materials and Methods: U87 GBM cells were cultured and treated with or without dapivirine. Cell viability was evaluated by CCK-8 (Cell Counting Kit 8, CCK-8) assay; apoptosis was analyzed by flow cytometry; cell migration was evaluated by Boyden Chamber assay; Western blotting was performed to detect proteins related to apoptosis, epithelial-to-mesenchymal transition and autophagy. PathScan intracellular signaling array kit was used to detect important and well-characterized signaling molecules. Tumor xenograft model in nude mice was used to evaluate the antitumorigenic effect in vivo. Results: Dapivirine weakened proliferation of glioma cells and induced the apoptosis of U87 glioblastoma cells. Furthermore, dapivirine regulated autophagy and induced Akt, Bad and SAPK/JNK activations. Moreover, the inhibition of glioma cell growth by dapivirine was also observed in nude mice in vivo. Conclusion: In summary, in our study dapivirine exposure induces stress, resulting in JNK and PI3K/Akt pathway activation through diminished inhibition of the apoptosis and autophagy cascade in U87 GBM cells, which inhibits cell growth in vitro and in vivo. PMID:29290776

Overexpression of ceramide synthase 1 increases C18-ceramide and leads to lethal autophagy in human glioma

PubMed Central

Wang, Zheng; Wen, Lijun; Zhu, Fei; Wang, Yanping; Xie, Qing; Chen, Zijun; Li, Yunsen

2017-01-01

Ceramide synthase 1 (CERS1) is the most highly expressed CERS in the central nervous system, and ceramide with an 18-carbon–containing fatty acid chain (C18-ceramide) in the brain plays important roles in signaling and sphingolipid development. However, the roles of CERS1 and C18-ceramide in glioma are largely unknown. In the present study, measured by electrospray ionization linear ion trap mass spectrometry, C18-ceramide was significantly lower in glioma tumor tissues compared with controls (P < 0.001), indicating that C18-ceramide might have a role in glioma. These roles were examined by reconstitution of C18-ceramide in U251 and A172 glioma cells via addition of exogenous C18-ceramide or overexpression of CERS1, which has been shown to specifically induce the generation of C18-ceramide. Overexpression of CERS1 or adding exogenous C18-ceramide inhibited cell viability and induced cell death by activating endoplasmic reticulum stress, which induced lethal autophagy and inhibited PI3K/AKT signal pathway in U251 and A172 glioma cells. Moreover, overexpression of CERS1 or adding exogenous C18-ceramide increased the sensitivity of U251 and A172 glioma cells to teniposide (VM-26). Thus, the combined therapy of CERS1/C18-ceramide and VM-26 may be a novel therapeutic strategy for the treatment of human glioma. PMID:29262618

MAP30 promotes apoptosis of U251 and U87 cells by suppressing the LGR5 and Wnt/β-catenin signaling pathway, and enhancing Smac expression

PubMed Central

Jiang, Yilin; Miao, Junjie; Wang, Dongliang; Zhou, Jingru; Liu, Bo; Jiao, Feng; Liang, Jiangfeng; Wang, Yangshuo; Fan, Cungang; Zhang, Qingjun

2018-01-01

Significant antitumor activity of Momordica anti-human immunodeficiency virus protein of 30 kDa (MAP30) purified from Momordica charantia has been the subject of previous research. However, the effective mechanism of MAP30 on malignant glioma cells has not yet been clarified. The aim of the present study was to investigate the effects and mechanism of MAP30 on U87 and U251 cell lines. A Cell Counting Kit-8 assay, wound healing assay and Transwell assay were used to detect the effects on U87 and U251 cells treated with different concentrations of MAP30 (0.5, 1, 2, 4, 8 and 16 µM) over different periods of time. Proliferation, migration and invasion of each cell line were markedly inhibited by MAP30 in a dose- and time-dependent manner. Flow cytometry and fluorescence staining demonstrated that apoptosis increased and the cell cycle was arrested in S-phase in the two investigated cell lines following MAP30 treatment. Western blot analysis demonstrated that leucine-rich-repeat-containing G-protein-coupled receptor 5 (LGR5) expression and key proteins in the Wnt/β-catenin signaling pathway were apparently decreased, whereas second mitochondria-derived activator of caspase (Smac) protein expression significantly increased with MAP30 treatment in the same manner. These results suggest that MAP30 markedly induces apoptosis in U87 and U251 cell lines by suppressing LGR5 and the Wnt/β-catenin signaling pathway, and enhancing Smac expression in a dose- and time-dependent manner. PMID:29556310

Mobile phone specific electromagnetic fields induce transient DNA damage and nucleotide excision repair in serum-deprived human glioblastoma cells.

PubMed

Al-Serori, Halh; Ferk, Franziska; Kundi, Michael; Bileck, Andrea; Gerner, Christopher; Mišík, Miroslav; Nersesyan, Armen; Waldherr, Monika; Murbach, Manuel; Lah, Tamara T; Herold-Mende, Christel; Collins, Andrew R; Knasmüller, Siegfried

2018-01-01

Some epidemiological studies indicate that the use of mobile phones causes cancer in humans (in particular glioblastomas). It is known that DNA damage plays a key role in malignant transformation; therefore, we investigated the impact of the UMTS signal which is widely used in mobile telecommunications, on DNA stability in ten different human cell lines (six brain derived cell lines, lymphocytes, fibroblasts, liver and buccal tissue derived cells) under conditions relevant for users (SAR 0.25 to 1.00 W/kg). We found no evidence for induction of damage in single cell gel electrophoresis assays when the cells were cultivated with serum. However, clear positive effects were seen in a p53 proficient glioblastoma line (U87) when the cells were grown under serum free conditions, while no effects were found in p53 deficient glioblastoma cells (U251). Further experiments showed that the damage disappears rapidly in U87 and that exposure induced nucleotide excision repair (NER) and does not cause double strand breaks (DSBs). The observation of NER induction is supported by results of a proteome analysis indicating that several proteins involved in NER are up-regulated after exposure to UMTS; additionally, we found limited evidence for the activation of the γ-interferon pathway. The present findings show that the signal causes transient genetic instability in glioma derived cells and activates cellular defense systems.

Contact-independent cell death of human microglial cells due to pathogenic Naegleria fowleri trophozoites.

PubMed

Kim, Jong-Hyun; Kim, Daesik; Shin, Ho-Joon

2008-12-01

Free-living Naegleria fowleri leads to a fatal infection known as primary amebic meningoencephalitis in humans. Previously, the target cell death could be induced by phagocytic activity of N. fowleri as a contact-dependent mechanism. However, in this study we investigated the target cell death under a non-contact system using a tissue-culture insert. The human microglial cells, U87MG cells, co-cultured with N. fowleri trophozoites for 30 min in a non-contact system showed morphological changes such as the cell membrane destruction and a reduction in the number. By fluorescence-activated cell sorter (FACS) analysis, U87MG cells co-cultured with N. fowleri trophozoites in a non-contact system showed a significant increase of apoptotic cells (16%) in comparison with that of the control or N. fowleri lysate. When U87MG cells were co-cultured with N. fowleri trophozoites in a non-contact system for 30 min, 2 hr, and 4 hr, the cytotoxicity of amebae against target cells was 40.5, 44.2, and 45.6%, respectively. By contrast, the cytotoxicity of non-pathogenic N. gruberi trophozoites was 10.2, 12.4, and 13.2%, respectively. These results suggest that the molecules released from N. fowleri in a contact-independent manner as well as phagocytosis in a contact-dependent manner may induce the host cell death.

Contact-Independent Cell Death of Human Microglial Cells due to Pathogenic Naegleria fowleri Trophozoites

PubMed Central

Kim, Jong-Hyun

2008-01-01

Free-living Naegleria fowleri leads to a fatal infection known as primary amebic meningoencephalitis in humans. Previously, the target cell death could be induced by phagocytic activity of N. fowleri as a contact-dependent mechanism. However, in this study we investigated the target cell death under a non-contact system using a tissue-culture insert. The human microglial cells, U87MG cells, co-cultured with N. fowleri trophozoites for 30 min in a non-contact system showed morphological changes such as the cell membrane destruction and a reduction in the number. By fluorescence-activated cell sorter (FACS) analysis, U87MG cells co-cultured with N. fowleri trophozoites in a non-contact system showed a significant increasse of apoptotic cells (16%) in comparison with that of the control or N. fowleri lysate. When U87MG cells were co-cultured with N. fowleri trophozoites in a non-contact system for 30 min, 2 hr, and 4 hr, the cytotoxicity of amebae against target cells was 40.5, 44.2, and 45.6%, respectively. By contrast, the cytotoxicity of non-pathogenic N. gruberi trophozoites was 10.2, 12.4, and 13.2%, respectively. These results suggest that the molecules released from N. fowleri in a contact-independent manner as well as phagocytosis in a contact-dependent manner may induce the host cell death. PMID:19127326

Lebbeckoside C, a new triterpenoid saponin from the stem barks of Albizia lebbeck inhibits the growth of human glioblastoma cells.

PubMed

Noté, Olivier Placide; Ngo Mbing, Joséphine; Kilhoffer, Marie-Claude; Pegnyemb, Dieudonné Emmanuel; Lobstein, Annelise

2018-02-19

One new acacic acid-type saponin, named lebbeckoside C (1), was isolated from the stem barks of Albizia lebbeck. Its structure was established on the basis of extensive analysis of 1D and 2D NMR ( 1 H, 13 C NMR, DEPT, COSY, TOCSY, ROESY, HSQC and HMBC) experiments, HRESIMS studies, and by chemical evidence as 3-O-[β-d-xylopyranosyl-(l→2)-β-d-fucopyranosyl-(1→6)-[β-d-glucopyranosyl(1→2)]-β-d-glucopyranosyl]-21-O-{(2E,6S)-6-O-{4-O-[(2E,6S)-2,6-dimethyl-6-O-(β-d-quinovopyranosyl)octa-2,7-dienoyl]-4-O-[(2E,6S)-2,6-dimethyl-6-O-(β-d-quinovopyranosyl)octa-2,7-dienoyl]-β-d-quinovopyranosyl}-2,6-dimethylocta-2,7-dienoyl}acacic acid 28 O-[β-d-quinovopyranosyl-(l→3)-[α-l-arabinofuranosyl-(l→4)]-α-l-rhamnopyranosyl-(l→2)-β-d-glucopyranosyl] ester. The isolated saponin (1) displayed significant cytotoxic activity against the human glioblastoma cell line U-87 MG and TG1 stem-like glioma cells isolated from a patient tumor with IC 50 values of 1.69 and 1.44 μM, respectively.

Decreasing GSH and increasing ROS in chemosensitivity gliomas with IDH1 mutation.

PubMed

Shi, Jinlong; Sun, Baolan; Shi, Wei; Zuo, Hao; Cui, Daming; Ni, Lanchun; Chen, Jian

2015-02-01

Gliomas are the most malignant and aggressive primary brain tumor in adults. Despite concerted efforts to improve therapies, their prognosis remains very poor. Isocitrate dehydrogenase 1 (IDH1) mutations have been discovered frequently in glioma patients and are strongly correlated with improved survival. However, the effect of IDH1 mutations on the chemosensitivity of gliomas remains unclear. In this study, we generated clonal U87 and U251 glioma cell lines overexpressing the R132H mutant protein (IDH1-R132H). Compared with control cells and cells overexpressing IDH wild type (IDH1-WT), both types of IDH1-R132H cells were more sensitive to temozolomide (TMZ) and cis-diamminedichloroplatinum (CDDP) in a time- and dose-dependent manner. The IDH1-R132H-induced higher chemosensitivity was associated with nicotine adenine disphosphonucleotide (NADPH), glutathione (GSH) depletion, and reactive oxygen species (ROS) generation. Accordingly, this IDH1-R132H-induced growth inhibition was effectively abrogated by GSH in vitro and in vivo. Our study provides direct evidence that the improved survival in patients with IDH1-R132H tumors may partly result from the effects of the IDH1-R132H protein on chemosensitivity. The primary cellular events associated with improved survival are the GSH depletion and increased ROS generation.

Down-regulation of MicroRNA-133 predicts poor overall survival and regulates the growth and invasive abilities in glioma.

PubMed

Liu, Yu; Han, Lili; Bai, Yahui; Du, Wei; Yang, Bo

2018-02-01

miRNAs were reported as oncogene or tumour suppressors in various cancers and played important roles in tumour development and progression. Dysregulated miR-133 has been reported in several cancers, however, the expression and biological function of miR-133 in glioma remained unclear. In this study, we found that miR-133 expression level was significantly decreased in glioma tissues and cell lines by RT-qPCR. Then miR-133 mimics were used to evaluate the effects of miR-133 on cell proliferation and invasion in vitro. We found that overexpressed miR-133 could significantly suppress cell growth, and invasion in U87 cells. Additionally, we found that forkhead box C1 (FOXC1) was overexpressed in glioma tissue and it was directly regulated by miR-133. Overall, this study is the first proof to demonstrate that miR-133 function as tumour suppressor in glioma and inhibit cell proliferation and invasioned by directly targeting FOXC1, implying miR-133 as a potential therapeutic target for glioma.

PAQR3 inhibits the proliferation, migration and invasion in human glioma cells.

PubMed

Tang, Shi-Lei; Gao, Yuan-Lin; Hu, Wen-Zhong

2017-08-01

Progestin and AdipoQ Receptor 3 (PAQR3), a member of the PAQR family, is down-regulated in several types of cancers and has been closely associated with tumor progression and development. However, little is known about the functions of PAQR3 in the tumorigenesis of human glioma. Therefore, in this report, we investigated the role of PAQR3 in human glioma. Our results showed that the expression of PAQR3 was significantly reduced in human glioma tissues and cell lines. PAQR3 overexpression inhibited the proliferation of glioma cells in vitro and attenuated tumor xenograft growth in vivo. In addition, PAQR3 overexpression suppressed the migration and invasion of glioma cells, as well as prevented the EMT process. Mechanistic studies demonstrated that PAQR3 overexpression significantly down-regulated the levels of phosphorylated PI3K and Akt in U251 cells. In conclusion, these results demonstrated that PAQR3 inhibited the proliferation, migration and invasion in glioma cells, at least in part, through the inactivation of PI3K/Akt signaling pathway. Therefore, PAQR3 may be a therapeutic target for the treatment of glioma. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

An optical assessment of the effects of glioma growth on resting state networks in mice (Conference Presentation)

NASA Astrophysics Data System (ADS)

Orukari, Inema E.; Bauer, Adam Q.; Baxter, Grant A.; Rubin, Joshua B.; Culver, Joseph P.

2017-02-01

Gliomas are known to cause significant changes in normal brain function that lead to cognitive deficits. Disruptions in resting state networks (RSNs) are thought to underlie these changes. However, investigating the effects of glioma growth on RSNs in humans is complicated by the heterogeneity in lesion size, type, and location across subjects. In this study, we evaluated the effects of tumor growth on RSNs over time in a controlled mouse model of glioma growth. Methods: Glioma cells (5x104-105 U87s) were stereotactically injected into the forepaw somatosensory cortex of adult nude mice (n=5). Disruptions in RSNs were evaluated weekly with functional connectivity optical intrinsic signal imaging (fcOIS). Tumor growth was monitored with MRI and weekly bioluminescence imaging (BLI). In order to characterize how tumor growth affected different RSNs over time, we calculated a number of functional connectivity (fc) metrics, including homotopic (bilateral) connectivity, spatial similarity, and node degree. Results: Deficits in fc initiate near the lesion, and over a period of several weeks, extend more globally. The reductions in spatial similarity were found to strongly correlate with the BLI signal indicating that increased tumor size is associated with increased RSN disruption. Conclusions: We have shown that fcOIS is capable of detecting alterations in mouse RSNs due to brain tumor growth. A better understanding of how RSN disruption contributes to the development of cognitive deficits in brain tumor patients may lead to better patient risk stratification and consequently improved cognitive outcomes.

Arginylglycylaspartic Acid-Surface-Functionalized Doxorubicin-Loaded Lipid-Core Nanocapsules as a Strategy to Target Alpha(V) Beta(3) Integrin Expressed on Tumor Cells

PubMed Central

Antonow, Michelli B.; Franco, Camila; Prado, Willian; Beckenkamp, Aline; Silveira, Gustavo P.; Buffon, Andréia; Guterres, Sílvia S.

2017-01-01

Doxorubicin (Dox) clinical use is limited by dose-related cardiomyopathy, becoming more prevalent with increasing cumulative doses. Previously, we developed Dox-loaded lipid-core nanocapsules (Dox-LNC) and, in this study, we hypothesized that self-assembling and interfacial reactions could be used to obtain arginylglycylaspartic acid (RGD)-surface-functionalized-Dox-LNC, which could target tumoral cells overexpressing αvβ3 integrin. Human breast adenocarcinoma cell line (MCF-7) and human glioblastoma astrocytoma (U87MG) expressing different levels of αvβ3 integrin were studied. RGD-functionalized Dox-LNC were prepared with Dox at 100 and 500 mg·mL−1 (RGD-MCMN (Dox100) and RGD-MCMN (Dox500)). Blank formulation (RGD-MCMN) had z-average diameter of 162 ± 6 nm, polydispersity index of 0.11 ± 0.04, zeta potential of +13.2 ± 1.9 mV and (6.2 ± 1.1) × 1011 particles mL−1, while RGD-MCMN (Dox100) and RGD-MCMN (Dox500) showed respectively 146 ± 20 and 215 ± 25 nm, 0.10 ± 0.01 and 0.09 ± 0.03, +13.8 ± 2.3 and +16.4 ± 1.5 mV and (6.9 ± 0.6) × 1011 and (6.1 ± 1.0) × 1011 particles mL−1. RGD complexation was 7.73 × 104 molecules per nanocapsule and Dox loading were 1.51 × 104 and 7.64 × 104 molecules per nanocapsule, respectively. RGD-functionalized nanocapsules had an improved uptake capacity by U87MG cells. Pareto chart showed that the cell viability was mainly affected by the Dox concentration and the period of treatment in both MCF-7 and U87MG. The influence of RGD-functionalization on cell viability was a determinant factor exclusively to U87MG. PMID:29271920

Biological properties of potent inhibitors of class I phosphatidylinositide 3-kinases: from PI-103 through PI-540, PI-620 to the oral agent GDC-0941.

PubMed

Raynaud, Florence I; Eccles, Suzanne A; Patel, Sonal; Alix, Sonia; Box, Gary; Chuckowree, Irina; Folkes, Adrian; Gowan, Sharon; De Haven Brandon, Alexis; Di Stefano, Francesca; Hayes, Angela; Henley, Alan T; Lensun, Letitia; Pergl-Wilson, Giles; Robson, Anthony; Saghir, Nahid; Zhyvoloup, Alexander; McDonald, Edward; Sheldrake, Peter; Shuttleworth, Stephen; Valenti, Melanie; Wan, Nan Chi; Clarke, Paul A; Workman, Paul

2009-07-01

The phosphatidylinositide 3-kinase pathway is frequently deregulated in human cancers and inhibitors offer considerable therapeutic potential. We previously described the promising tricyclic pyridofuropyrimidine lead and chemical tool compound PI-103. We now report the properties of the pharmaceutically optimized bicyclic thienopyrimidine derivatives PI-540 and PI-620 and the resulting clinical development candidate GDC-0941. All four compounds inhibited phosphatidylinositide 3-kinase p110alpha with IC(50) < or = 10 nmol/L. Despite some differences in isoform selectivity, these agents exhibited similar in vitro antiproliferative properties to PI-103 in a panel of human cancer cell lines, with submicromolar potency in PTEN-negative U87MG human glioblastoma cells and comparable phosphatidylinositide 3-kinase pathway modulation. PI-540 and PI-620 exhibited improvements in solubility and metabolism with high tissue distribution in mice. Both compounds gave improved antitumor efficacy over PI-103, following i.p. dosing in U87MG glioblastoma tumor xenografts in athymic mice, with treated/control values of 34% (66% inhibition) and 27% (73% inhibition) for PI-540 (50 mg/kg b.i.d.) and PI-620 (25 mg/kg b.i.d.), respectively. GDC-0941 showed comparable in vitro antitumor activity to PI-103, PI-540, and PI-620 and exhibited 78% oral bioavailability in mice, with tumor exposure above 50% antiproliferative concentrations for >8 hours following 150 mg/kg p.o. and sustained phosphatidylinositide 3-kinase pathway inhibition. These properties led to excellent dose-dependent oral antitumor activity, with daily p.o. dosing at 150 mg/kg achieving 98% and 80% growth inhibition of U87MG glioblastoma and IGROV-1 ovarian cancer xenografts, respectively. Together, these data support the development of GDC-0941 as a potent, orally bioavailable inhibitor of phosphatidylinositide 3-kinase. GDC-0941 has recently entered phase I clinical trials.

Biological properties of potent inhibitors of class I phosphatidylinositide 3-kinases: from PI-103 through PI-540, PI-620 to the oral agent GDC-0941

PubMed Central

Raynaud, Florence I.; Eccles, Suzanne A.; Patel, Sonal; Alix, Sonia; Box, Gary; Chuckowree, Irina; Folkes, Adrian; Gowan, Sharon; De Haven Brandon, Alexis; Di Stefano, Francesca; Hayes, Angela; Henley, Alan T.; Lensun, Letitia; Pergl-Wilson, Giles; Robson, Anthony; Saghir, Nahid; Zhyvoloup, Alexander; McDonald, Edward; Sheldrake, Peter; Shuttleworth, Stephen; Valenti, Melanie; Wan, Nan Chi; Clarke, Paul A.; Workman, Paul

2009-01-01

The phosphatidylinositide 3-kinase pathway is frequently deregulated in human cancers and inhibitors offer considerable therapeutic potential. We previously described the promising tricyclic pyridofuropyrimidine lead and chemical tool compound PI-103. We now report the properties of the pharmaceutically optimized bicyclic thienopyrimidine derivatives PI-540 and PI-620 and the resulting clinical development candidate GDC-0941. All four compounds inhibited phosphatidylinositide 3-kinase p110α with IC50 ≤ 10 nmol/L. Despite some differences in isoform selectivity, these agents exhibited similar in vitro antiproliferative properties to PI-103 in a panel of human cancer cell lines, with submicromolar potency in PTEN-negative U87MG human glioblastoma cells and comparable phosphatidylinositide 3-kinase pathway modulation. PI-540 and PI-620 exhibited improvements in solubility and metabolism with high tissue distribution in mice. Both compounds gave improved antitumor efficacy over PI-103, following i.p. dosing in U87MG glioblastoma tumor xenografts in athymic mice, with treated/control values of 34% (66% inhibition) and 27% (73% inhibition) for PI-540 (50 mg/kg b.i.d.) and PI-620 (25 mg/kg b.i.d.), respectively. GDC-0941 showed comparable in vitro antitumor activity to PI-103, PI-540, and PI-620 and exhibited 78% oral bioavailability in mice, with tumor exposure above 50% anti-proliferative concentrations for >8 hours following 150 mg/kg p.o. and sustained phosphatidylinositide 3-kinase pathway inhibition. These properties led to excellent dose-dependent oral antitumor activity, with daily p.o. dosing at 150 mg/kg achieving 98% and 80% growth inhibition of U87MG glioblastoma and IGROV-1 ovarian cancer xenografts, respectively. Together, these data support the development of GDC-0941 as a potent, orally bioavailable inhibitor of phosphatidylinositide 3-kinase. GDC-0941 has recently entered phase I clinical trials. PMID:19584227

3-Bromopyruvate antagonizes effects of lactate and pyruvate, synergizes with citrate and exerts novel anti-glioma effects.

PubMed

El Sayed, S M; El-Magd, R M Abou; Shishido, Y; Chung, S P; Diem, T H; Sakai, T; Watanabe, H; Kagami, S; Fukui, K

2012-02-01

Oxidative stress-energy depletion therapy using oxidative stress induced by D-amino acid oxidase (DAO) and energy depletion induced by 3-bromopyruvate (3BP) was reported recently (El Sayed et al., Cancer Gene Ther., 19, 1-18, 2012). Even in the presence of oxygen, cancer cells oxidize glucose preferentially to produce lactate (Warburg effect) which seems vital for cancer microenvironment and progression. 3BP is a closely related structure to lactate and pyruvate and may antagonize their effects as a novel mechanism of its action. Pyruvate exerted a potent H(2)O(2) scavenging effect to exogenous H(2)O(2), while lactate had no scavenging effect. 3BP induced H(2)O(2) production. Pyruvate protected against H(2)O(2)-induced C6 glioma cell death, 3BP-induced C6 glioma cell death but not against DAO/D-serine-induced cell death, while lactate had no protecting effect. Lactate and pyruvate protected against 3BP-induced C6 glioma cell death and energy depletion which were overcome with higher doses of 3BP. Lactate and pyruvate enhanced migratory power of C6 glioma which was blocked by 3BP. Pyruvate and lactate did not protect against C6 glioma cell death induced by other glycolytic inhibitors e.g. citrate (inhibitor of phosphofructokinase) and sodium fluoride (inhibitor of enolase). Serial doses of 3BP were synergistic with citrate in decreasing viability of C6 glioma cells and spheroids. Glycolysis subjected to double inhibition using 3BP with citrate depleted ATP, clonogenic power and migratory power of C6 glioma cells. 3BP induced a caspase-dependent cell death in C6 glioma. 3BP was powerful in decreasing viability of human glioblastoma multiforme cells (U373MG) and C6 glioma in a dose- and time-dependent manner.

Protein kinase D2 regulates migration and invasion of U87MG glioblastoma cells in vitro

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bernhart, Eva; Damm, Sabine; Wintersperger, Andrea

Glioblastoma multiforme (GBM) is the most common malignant brain tumor, which, despite combined modality treatment, reoccurs and is invariably fatal for affected patients. Recently, a member of the serine/threonine protein kinase D (PRKD) family, PRKD2, was shown to be a potent mediator of glioblastoma growth. Here we studied the role of PRKD2 in U87MG glioblastoma cell migration and invasion in response to sphingosine-1-phosphate (S1P), an activator of PRKD2 and a GBM mitogen. Time-lapse microscopy demonstrated that random cell migration was significantly diminished in response to PRKD2 silencing. The pharmacological PRKD family inhibitor CRT0066101 decreased chemotactic migration and invasion across uncoatedmore » or matrigel-coated Transwell inserts. Silencing of PRKD2 attenuated migration and invasion of U87MG cells even more effectively. In terms of downstream signaling, CRT0066101 prevented PRKD2 autophosphorylation and inhibited p44/42 MAPK and to a smaller extent p54/46 JNK and p38 MAPK activation. PRKD2 silencing impaired activation of p44/42 MAPK and p54/46 JNK, downregulated nuclear c-Jun protein levels and decreased c-Jun{sup S73} phosphorylation without affecting the NFκB pathway. Finally, qPCR array analyses revealed that silencing of PRKD2 downregulates mRNA levels of integrin alpha-2 and -4 (ITGA2 and -4), plasminogen activator urokinase (PLAU), plasminogen activator urokinase receptor (PLAUR), and matrix metallopeptidase 1 (MMP1). Findings of the present study identify PRKD2 as a potential target to interfere with glioblastoma cell migration and invasion, two major determinants contributing to recurrence of glioblastoma after multimodality treatment. Highlights: • Sphingosine-1-phosphate induces glioma cell migration and invasion. • Part of the effects is mediated by protein kinase D2 (PRKD2) activation. • Inactivation of PRKD2 attenuates glioblastoma cell migration and invasion. • Both, RNAi and pharmacological inhibition of PRKD2 inhibits MAPK signaling. • PRKD2 regulates transcription of gene products implicated in migration and invasion.« less

The CHAC1-inhibited Notch3 pathway is involved in temozolomide-induced glioma cytotoxicity.

PubMed

Chen, Peng-Hsu; Shen, Wan-Lin; Shih, Chwen-Ming; Ho, Kuo-Hao; Cheng, Chia-Hsiung; Lin, Cheng-Wei; Lee, Chin-Cheng; Liu, Ann-Jeng; Chen, Ku-Chung

2017-04-01

Glioblastoma multiforme (GBM) is the high-grade primary glioma in adults. Temozolomide (TMZ), an alkylating agent of the imidazotetrazine series, is a first-line chemotherapeutic drug for clinical therapy. However, the expense of TMZ therapy and increasing drug resistance to TMZ decreases its therapeutic effects. Therefore, our aim was to investigate the detailed molecular mechanisms of TMZ-mediated cytotoxicity to enhance the efficacy of TMZ in clinical GBM therapy. First, TMZ-mediated gene expression profiles and networks in U87-MG cells were identified by transcriptome microarray and bioinformatic analyses. Cation transport regulator-like protein 1 (CHAC1) was the most highly TMZ-upregulated gene. Overexpression and knockdown of CHAC1 expression significantly influenced TMZ-mediated cell viability, apoptosis, caspase-3 activation, and poly(ADP ribose) polymerase (PARP) degradation. The c-Jun N-terminal kinase (JNK)1/c-JUN pathway was identified to participate in TMZ-upregulated CHAC1 expression via transcriptional control. Furthermore, CHAC1 levels were significantly decreased in GBM cell lines, TCGA array data, and tumor tissues. Overexpression of CHAC1 enhanced glioma apoptotic death via caspase-3/9 activation, PARP degradation, autophagy formation, reactive oxygen species generation, increased intracellular calcium, and loss of the mitochondria membrane potential. Finally, we also identified that TMZ significantly reduced Notch3 levels, which are upregulated in gliomas. TMZ also induced CHAC1 to bind to the Notch3 protein and inhibit Notch3 activation, resulting in attenuation of Notch3-mediated downstream signaling pathways. These results emphasize that CHAC1-inhibited Notch3 signaling can influence TMZ-mediated cytotoxicity. Our findings may provide novel therapeutic strategies for future glioblastoma therapy. Copyright © 2016 Elsevier Ltd. All rights reserved.

Critical roles of chemokine receptor CCR5 in regulating glioblastoma proliferation and invasion.

PubMed

Zhao, Lanfu; Wang, Yuan; Xue, Yafei; Lv, Wenhai; Zhang, Yufu; He, Shiming

2015-11-01

Glioblastoma (GBM) is the most prevalent malignant primary brain tumor in adults and exhibits a spectrum of aberrantly aggressive phenotype. Tumor cell proliferation and invasion are critically regulated by chemokines and their receptors. Recent studies have shown that the chemokine CCL5 and its receptor CCR5 play important roles in tumor invasion and metastasis. Nonetheless, the roles of the CCR5 in GBM still remain unclear. The present study provides the evidence that the chemokine receptor CCR5 is highly expressed and associated with poor prognosis in human GBM. Mechanistically, CCL5-CCR5 mediates activation of Akt, and subsequently induces proliferation and invasive responses in U87 and U251 cells. Moreover, down-regulation of CCR5 significantly inhibited the growth of glioma in U87 tumor xenograft mouse model. Finally, high CCR5 expression in GBM is correlated with increased p-Akt expression in patient samples. Together, these findings suggest that the CCR5 is a critical molecular event associated with gliomagenesis. © The Author 2015. Published by ABBS Editorial Office in association with Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences.

Ex vivo cultures of glioblastoma in three-dimensional hydrogel maintain the original tumor growth behavior and are suitable for preclinical drug and radiation sensitivity screening

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jiguet Jiglaire, Carine, E-mail: carine.jiguet-jiglaire@univ-amu.fr; CRO2, UMR 911, Faculté de Médecine de la Timone, 27 boulevard Jean Moulin, 13284 Marseille Cedex; INSERM, U911, 13005 Marseille

Identification of new drugs and predicting drug response are major challenges in oncology, especially for brain tumors, because total surgical resection is difficult and radiation therapy or chemotherapy is often ineffective. With the aim of developing a culture system close to in vivo conditions for testing new drugs, we characterized an ex vivo three-dimensional culture system based on a hyaluronic acid-rich hydrogel and compared it with classical two-dimensional culture conditions. U87-MG glioblastoma cells and seven primary cell cultures of human glioblastomas were subjected to radiation therapy and chemotherapy drugs. It appears that 3D hydrogel preserves the original cancer growth behaviormore » and enables assessment of the sensitivity of malignant gliomas to radiation and drugs with regard to inter-tumoral heterogeneity of therapeutic response. It could be used for preclinical assessment of new therapies. - Highlights: • We have compared primary glioblastoma cell culture in a 2D versus 3D-matrix system. • In 3D morphology, organization and markers better recapitulate the original tumor. • 3D-matrix culture might represent a relevant system for more accurate drug screening.« less

Enhanced In Vivo Tumor Detection by Active Tumor Cell Targeting Using Multiple Tumor Receptor-Binding Peptides Presented on Genetically Engineered Human Ferritin Nanoparticles.

PubMed

Kwon, Koo Chul; Ko, Ho Kyung; Lee, Jiyun; Lee, Eun Jung; Kim, Kwangmeyung; Lee, Jeewon

2016-08-01

Human ferritin heavy-chain nanoparticle (hFTH) is genetically engineered to present tumor receptor-binding peptides (affibody and/or RGD-derived cyclic peptides, named 4CRGD here) on its surface. The affibody and 4CRGD specifically and strongly binds to human epidermal growth factor receptor I (EGFR) and human integrin αvβ3, respectively, which are overexpressed on various tumor cells. Through in vitro culture of EGFR-overexpressing adenocarcinoma (MDA-MB-468) and integrin-overexpressing glioblastoma cells (U87MG), it is clarified that specific interactions between receptors on tumor cells and receptor-binding peptides on engineered hFTH is critical in active tumor cell targeting. After labeling with the near-infrared fluorescence dye (Cy5.5) and intravenouse injection into MDA-MB-468 or U87MG tumor-bearing mice, the recombinant hFTHs presenting either peptide or both of affibody and 4CRGD are successfully delivered to and retained in the tumor for a prolonged period of time. In particular, the recombinant hFTH presenting both affibody and 4CRGD notably enhances in vivo detection of U87MG tumors that express heterogeneous receptors, integrin and EGFR, compared to the other recombinant hFTHs presenting either affibody or 4CRGD only. Like affibody and 4CRGD used in this study, other multiple tumor receptor-binding peptides can be also genetically introduced to the hFTH surface for actively targeting of in vivo tumors with heterogenous receptors. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Feasibility of Using Bevacizumab With Radiation Therapy and Temozolomide in Newly Diagnosed High-Grade Glioma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Narayana, Ashwatha; Golfinos, John G.; Fischer, Ingeborg

2008-10-01

Introduction: Bevacizumab, a monoclonal antibody against vascular endothelial growth factor (VEGF), has shown promise in the treatment of patients with recurrent high-grade glioma. The purpose of this study is to test the feasibility of using bevacizumab with chemoradiation in the primary management of high-grade glioma. Methods and Materials: Fifteen patients with high-grade glioma were treated with involved field radiation therapy to a dose of 59.4 Gy at 1.8 Gy/fraction with bevacizumab 10 mg/kg on Days 14 and 28 and temozolomide 75 mg/m{sup 2}. Subsequently, bevacizumab 10 mg/kg was continued every 2 weeks with temozolomide 150 mg/m{sup 2} for 12 months.more » Changes in relative cerebral blood volume, perfusion-permeability index, and tumor volume measurement were measured to assess the therapeutic response. Immunohistochemistry for phosphorylated VEGF receptor 2 (pVEGFR2) was performed. Results: Thirteen patients (86.6%) completed the planned bevacizumab and chemoradiation therapy. Four Grade III/IV nonhematologic toxicities were seen. Radiographic responses were noted in 13 of 14 assessable patients (92.8%). The pVEGFR2 staining was seen in 7 of 8 patients (87.5%) at the time of initial diagnosis. Six patients have experienced relapse, 3 at the primary site and 3 as diffuse disease. One patient showed loss of pVEGFR2 expression at relapse. One-year progression-free survival and overall survival rates were 59.3% and 86.7%, respectively. Conclusion: Use of antiangiogenic therapy with radiation and temozolomide in the primary management of high-grade glioma is feasible. Perfusion imaging with relative cerebral blood volume, perfusion-permeability index, and pVEGFR2 expression may be used as a potential predictor of therapeutic response. Toxicities and patterns of relapse need to be monitored closely.« less

Identification of EGFRvIII-derived CTL epitopes restricted by HLA A0201 for dendritic cell based immunotherapy of gliomas.

PubMed

Wu, An-hua; Xiao, Jing; Anker, Lars; Hall, Walter A; Gregerson, Dale S; Cavenee, Webster K; Chen, Wei; Low, Walter C

2006-01-01

The type III variant of the epidermal growth factor receptor (EGFRvIII) mutation is present in 20-25% of patients with glioblastoma multiforme (GBM). EGFRvIII is not expressed in normal tissue and is therefore a suitable candidate antigen for dendritic cell (DC) based immunotherapy of GBM. To identify the antigenic epitope(s) that may serve as targets for EGFRvIII-specific cytotoxic T lymphocytes (CTLs), the peptide sequence of EGFRvIII was screened with two software programs to predict candidate epitopes restricted by the major histocompatibility complex class I subtype HLA-A0201, which is the predominant subtype in most ethnic groups. Three predicted peptides were constructed and loaded to mature human DCs generated from peripheral blood monocytes. Autologous CD8+ T cells were stimulated in vitro with the EGFRvIII peptide-pulsed DCs. One of the three peptides was found to induce EGFRvIII-specific CTLs as demonstrated by IFN-gamma production and cytotoxicity against HLA-A0201+ EGFRvIII transfected U87 glioma cells. These results suggest that vaccination with EGFRvIII peptide-pulsed DCs or adoptive transfer of in vitro elicited EGFRvIII-specific CTLs by EGFRvIII peptide-pulsed DCs are potential approaches to the treatment of glioma patients.

Propolis changes the anticancer activity of temozolomide in U87MG human glioblastoma cell line.

PubMed

Markiewicz-Żukowska, Renata; Borawska, Maria H; Fiedorowicz, Anna; Naliwajko, Sylwia K; Sawicka, Diana; Car, Halina

2013-02-27

Propolis is a honey bee product which contains many active compounds, such as CAPE or chrysin, and has many beneficial activities. Recently, its anti-tumor properties have been discussed. We have tested whether the ethanolic extract of propolis (EEP) interferes with temozolomide (TMZ) to inhibit U87MG cell line growth. The U87MG glioblastoma cell line was exposed to TMZ (10-100 μM), EEP (10-100 μg/ml) or a mixture of TMZ and EEP during 24, 48 or 72 hours. The cell division was examined by the H3-thymidine incorporation, while the western blot method was used for detection of p65 subunit of NF-κB and ELISA test to measure the concentration of its p50 subunit in the nucleus. We have found that both, TMZ and EEP administrated alone, had a dose- and time-dependent inhibitory effect on the U87MG cell line growth, which was manifested by gradual reduction of cell viability and alterations in proliferation rate. The anti-tumor effect of TMZ (20 μM) was enhanced by EEP, which was especially well observed after a short time of exposition, where simultaneous usage of TMZ and EEP resulted in a higher degree of growth inhibition than each biological factor used separately. In addition, cells treated with TMZ presented no changes in NF-κB activity in prolonged time of treatment and EEP only slightly reduced the nuclear translocation of this transcription factor. In turn, the combined incubation with TMZ and EEP led to an approximately double reduction of NF-κB nuclear localization. We conclude that EEP presents cytotoxic properties and may cooperate with TMZ synergistically enhancing its growth inhibiting activity against glioblastoma U87MG cell line. This phenomenon may be at least partially mediated by a reduced activity of NF-κB.

Propolis changes the anticancer activity of temozolomide in U87MG human glioblastoma cell line

PubMed Central

2013-01-01

Background Propolis is a honey bee product which contains many active compounds, such as CAPE or chrysin, and has many beneficial activities. Recently, its anti-tumor properties have been discussed. We have tested whether the ethanolic extract of propolis (EEP) interferes with temozolomide (TMZ) to inhibit U87MG cell line growth. Methods The U87MG glioblastoma cell line was exposed to TMZ (10-100 μM), EEP (10-100 μg/ml) or a mixture of TMZ and EEP during 24, 48 or 72 hours. The cell division was examined by the H3-thymidine incorporation, while the western blot method was used for detection of p65 subunit of NF-κB and ELISA test to measure the concentration of its p50 subunit in the nucleus. Results We have found that both, TMZ and EEP administrated alone, had a dose- and time-dependent inhibitory effect on the U87MG cell line growth, which was manifested by gradual reduction of cell viability and alterations in proliferation rate. The anti-tumor effect of TMZ (20 μM) was enhanced by EEP, which was especially well observed after a short time of exposition, where simultaneous usage of TMZ and EEP resulted in a higher degree of growth inhibition than each biological factor used separately. In addition, cells treated with TMZ presented no changes in NF-κB activity in prolonged time of treatment and EEP only slightly reduced the nuclear translocation of this transcription factor. In turn, the combined incubation with TMZ and EEP led to an approximately double reduction of NF-κB nuclear localization. Conclusions We conclude that EEP presents cytotoxic properties and may cooperate with TMZ synergistically enhancing its growth inhibiting activity against glioblastoma U87MG cell line. This phenomenon may be at least partially mediated by a reduced activity of NF-κB. PMID:23445763

Anticancer activity of flavonoids isolated from Achyrocline satureioides in gliomas cell lines.

PubMed

Souza, Priscila Oliveira de; Bianchi, Sara Elis; Figueiró, Fabrício; Heimfarth, Luana; Moresco, Karla Suzana; Gonçalves, Rosângela Mayer; Hoppe, Juliana Bender; Klein, Caroline Peres; Salbego, Christianne Gazzana; Gelain, Daniel Pens; Bassani, Valquíria Linck; Zanotto Filho, Alfeu; Moreira, José Claudio Fonseca

2018-05-04

Achyrocline satureioides, popularly known as "marcela", is a medicinal plant found in South America. This plant is rich in flavonoids, which have been reported to exert numerous biological activities. The aim of this study was to purify, identify and evaluate the mechanisms underlining anticancer activity of A. satureioides flavonoids in glioma cell lines (U87, U251 and C6) as well as their comparative toxicity in normal brain cells (primary astrocytes, neurons and organotypic hippocampal cultures). The main flavonoids present in A. satureioides are luteolin, quercetin, 3-O-methyl-quercetin and achyrobichalcone, the later a very unique metabolite present in this plant. Isolated flavonoids as well as A. satureioides extracts reduced proliferation and clonogenic survival, and induced apoptosis of glioma cell lines. In addition, A. satureioides flavonoids potentiated the cytotoxic effect and apoptosis induction by the glioma chemotherapeutic temozolomide (TMZ). Importantly, A. satureioides flavonoids were less cytotoxic to astrocytes, neuron:astrocytes co-cultures and hippocampal cultures if compared to gliomas. Investigation of 10 cancer-related pathways showed a reduced activation of MYC and the Map kinases ERK and JNK by A. satureioides flavonoid-enriched extract, an effect not observed when individual flavonoids were evaluated. Altogether, the herein presented results show that A. satureioides extract possesses a combination of flavonoids, some unique for this plant, which have synergistic anticancer activity and potential for further studies in vivo. Copyright © 2018 Elsevier Ltd. All rights reserved.

microPET Imaging of Glioma Integrin (alpha-v, beta-3) Expression Using Cu-64-Labeled Tetrameric RGD Peptide

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wu, Yun; Zhang, , Xianzhong; Xiong, , Zhengming

2005-10-01

Integrins ?v?3 and ?v?5 play a critical role in tumor-induced angiogenesis and metastasis, and have become promising diagnostic indicators and therapeutic targets of tumors. Radiolabeled RGD peptides that are integrin-specific may be used for non-invasive imaging of integrin expression level as well as for integrin-targeted radionuclide therapy. We previously conjugated a series of mono- and dimeric RGD peptides with 1,4,7,10-tetraazacyclododecane-N, N?,N??,N???-tetraacetic acid (DOTA) and labeled these with copper-64 for microPET imaging in various mouse xenograft models. The copper-64 tracers showed ?v?3-selective tumor uptake, but the magnitude of tumor uptake was relatively low, the tumor washout was rapid, and non-target organ/tissuemore » retention was high. In this study we developed a tetrameric RGD peptide tracer 64Cu-DOTA-E{l_brace}E[c(RGDfK)]2{r_brace}2 for positron emission tomography (PET) imaging of integrin ?v?3 expression in a subcutaneous U87MG glioma xenograft model in female athymic nude mice. The RGD tetramer showed significantly higher integrin binding affinity than the corresponding mono- and dimeric RGD analogs, most likely due to polyvalency effect. The radiolabeled peptide showed rapid blood clearance (0.61 ? 0.01%ID/g at 30 min and 0.21 ? 0.01 %ID/g at 4 h postinjection (p.i.), respectively) and predominantly renal excretion. Tumor uptake was rapid and high and the tumor washout was slow (9.93 ? 1.05 %ID/g at 30 min p.i. and 4.56 ? 0.51 %ID/g at 24 h post-injection). The metabolic stability of 64Cu-DOTA-E{l_brace}E[c(RGDfK)]2{r_brace}2 was determined in mouse blood, urine, and liver and kidney homogenates at different times after tracer injection. The average fractions of intact tracer in these organs at 1 h were approximately 70, 58, 51 and 26 percent, respectively. Non-invasive microPET imaging studies showed significant tumor uptake and good contrast in the subcutaneous tumor-bearing mice, which agreed well with the biodistribution results. Integrin ?v?3 specificity was demonstrated by successful blocking of tumor uptake of 64Cu-DOTA-E{l_brace}E[c(RGDfK)]2{r_brace}2 in the presence of excess amount of c(RGDyK) at 1 h postinjection. The highest absorbed radiation doses determined for the human reference adult were received by the urinary bladder wall (0.263 mGy/MBq), kidneys (0.0298 mGy/MBq), and liver (0.0244 mGy/MBq). Assuming 0.5-g U87MG glioma tumors in man, we calculated an absorbed dose of 65.3 mGy/MBq (242 rad/mCi) following a single injection of 64Cu-DOTA-E{l_brace}E[c(RGDfK)]2{r_brace}2. In conclusion, the high integrin avidity and favorable biokinetics make 64Cu-DOTA-E{l_brace}E[c(RGDfK)]2{r_brace}2 a promising agent for peptide receptor radionuclide imaging therapy of integrin-positive tumors.« less

The prosurvival role of autophagy in Resveratrol-induced cytotoxicity in human U251 glioma cells

PubMed Central

2009-01-01

Background Previous study reported that resveratrol has anti-tumor activity. In this study, we investigated the involvement of autophagy in the resveratrol-induced apoptotic death of human U251 glioma cells. Methods The growth inhibition of U251 cells induced by resveratrol was assessed with methyl thiazolyl tetrazolium (MTT). The activation of autophagy and proapoptotic effect were characterized by monodansylcadaverine labeling and Hoechst stain, respectively. Mitochondrialtransmembrane potential (ΔΨm) was measured as a function of drug treatment using 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide (JC-1). The role of autophagy and apoptosis in the resveratrol-induced death of U251 cells was assessed using autophagic and caspase inhibitors. Immunofluorescence, flow cytometry, and Western blot analysis were used to study the apoptotic and autophagic mechanisms. Results Methyl thiazolyl tetrazolium (MTT) assays indicated that resveratrol decreased the viability of U251 cells in a dose- and time-dependent manner. Flow cytometry analysis indicated that resveratrol increased cell population at sub-G1 phase, an index of apoptosis. Furthermore, resveratrol-induced cell death was associated with a collapse of the mitochondrial membrane potential. The pan-caspase inhibitor Z-VAD-fmk suppressed resveratrol-induced U251 cell death. Resveratrol stimulated autophagy was evidenced by punctuate monodansylcadaverine(MDC) staining and microtubule-associated protein light chain 3 (LC3) immunoreactivty. Resveratrol also increased protein levels of beclin 1 and membrane form LC3 (LC3-II). Autophagy inhibitors 3-methylademine (3-MA) and bafilomycin A1 sensitized the cytotoxicity of resveratrol. Conclusion Together, these findings indicate that resveratrol induces autophagy in human U251 glioma cells and autophagy suppressed resveratrol-induced apoptosis. This study thus suggests that autophagy inhibitors can increase the cytotoxicity of resveratrol to glioma cells. PMID:19566920

Genetic modification of neurons to express bevacizumab for local anti-angiogenesis treatment of glioblastoma.

PubMed

Hicks, Martin J; Funato, Kosuke; Wang, Lan; Aronowitz, Eric; Dyke, Jonathan P; Ballon, Douglas J; Havlicek, David F; Frenk, Esther Z; De, Bishnu P; Chiuchiolo, Maria J; Sondhi, Dolan; Hackett, Neil R; Kaminsky, Stephen M; Tabar, Viviane; Crystal, Ronald G

2015-01-01

The median survival of glioblastoma multiforme (GBM) is approximately 1 year. Following surgical removal, systemic therapies are limited by the blood-brain barrier. To circumvent this, we developed a method to modify neurons with the genetic sequence for therapeutic monoclonal antibodies using adeno-associated virus (AAV) gene transfer vectors, directing persistent, local expression in the tumor milieu. The human U87MG GBM cell line or patient-derived early passage GBM cells were administered to the striatum of NOD/SCID immunodeficient mice. AAVrh.10BevMab, an AAVrh.10-based vector coding for bevacizumab (Avastin), an anti-human vascular endothelial growth factor (VEGF) monoclonal antibody, was delivered to the area of the GBM xenograft. Localized expression of bevacizumab was demonstrated by quantitative PCR, ELISA and western blotting. Immunohistochemistry showed that bevacizumab was expressed in neurons. Concurrent administration of AAVrh.10BevMab with the U87MG tumor reduced tumor blood vessel density and tumor volume, and increased survival. Administration of AAVrh.10BevMab 1 week after U87MG xenograft reduced growth and increased survival. Studies with patient-derived early passage GBM primary cells showed a reduction in primary tumor burden with an increased survival. These data support the strategy of AAV-mediated central nervous system gene therapy to treat GBM, overcoming the blood-brain barrier through local, persistent delivery of an anti-angiogenesis monoclonal antibody.

A pilot study of peptide vaccines for VEGF receptor 1 and 2 in patients with recurrent/progressive high grade glioma.

PubMed

Shibao, Shunsuke; Ueda, Ryo; Saito, Katsuya; Kikuchi, Ryogo; Nagashima, Hideaki; Kojima, Atsuhiro; Kagami, Hiroshi; Pareira, Eriel Sandika; Sasaki, Hikaru; Noji, Shinobu; Kawakami, Yutaka; Yoshida, Kazunari; Toda, Masahiro

2018-04-20

Early-phase clinical studies of glioma vaccines have shown feasibility and encouraging preliminary clinical activity. A vaccine that targets tumor angiogenesis factors in glioma microenvironment has not been reported. Therefore, we performed a pilot study to evaluate the safety and immunogenicity of a novel vaccination targeting tumor angiogenesis with synthetic peptides for vascular endothelial growth factor (VEGF) receptor epitopes in patients with recurrent/progressive high grade gliomas. Eight patients received intranodal vaccinations weekly at a dose of 2mg/kg bodyweight 8 times. T-lymphocyte responses against VEGF receptor (VEGFR) epitopes were assessed by enzyme linked immunosorbent spot assays. This treatment was well-tolerated in patients. The first four vaccines induced positive immune responses against at least one of the targeted VEGFR epitopes in the peripheral blood mononuclear cells in 87.5% of patients. The median overall survival time in all patients was 15.9 months. Two achieved progression-free status lasting at least 6 months. Two patients with recurrent GBM demonstrated stable disease. Plasma IL-8 level was negatively correlated with overall survival. These data demonstrate the safety and immunogenicity of VEGFR peptide vaccines targeting tumor vasculatures in high grade gliomas.

Carbon Ion Irradiation Inhibits Glioma Cell Migration Through Downregulation of Integrin Expression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rieken, Stefan, E-mail: Stefan.Rieken@med.uni-heidelberg.de; Habermehl, Daniel; Wuerth, Lena

2012-05-01

Purpose: To investigate the effect of carbon ion irradiation on glioma cell migration. Methods and Materials: U87 and Ln229 glioma cells were irradiated with photons and carbon ions. Migration was analyzed 24 h after irradiation. Fluorescence-activated cell sorting analysis was performed in order to quantify surface expression of integrins. Results: Single photon doses of 2 Gy and 10 Gy enhanced {alpha}{sub {nu}}{beta}{sub 3} and {alpha}{sub {nu}}{beta}{sub 5} integrin expression and caused tumor cell hypermigration on both vitronectin (Vn) and fibronectin (Fn). Compared to integrin expression in unirradiated cells, carbon ion irradiation caused decreased integrin expression and inhibited cell migration onmore » both Vn and Fn. Conclusion: Photon radiotherapy (RT) enhances the risk of tumor cell migration and subsequently promotes locoregional spread via photon induction of integrin expression. In contrast to photon RT, carbon ion RT causes decreased integrin expression and suppresses glioma cell migration on both Vn and Fn, thus promising improved local control.« less

Polysaccharide peptide isolated from grass-cultured Ganoderma lucidum induces anti-proliferative and pro-apoptotic effects in the human U251 glioma cell line

PubMed Central

Wang, Chunhua; Lin, Dongmei; Chen, Quan; Lin, Shuqian; Shi, Songsheng; Chen, Chunmei

2018-01-01

The Ganoderma lucidum (G. lucidum) mushroom is one of the most extensively studied functional foods, known for its numerous health benefits, including the inhibition of tumor cell growth. The present study assessed the anti-proliferative and pro-apoptotic activity of a novel G. lucidum polysaccharide peptide (GL-PP) in human glioma U251 cells, which was purified from grass-cultured G. lucidum. GL-PP is a glycopeptide with an average molecular weight of 42,635 Da and a polysaccharide-to-peptide ratio of 88.70:11.30. The polysaccharides were composed of l-arabinose, d-mannose and d-glucose at a molar ratio of 1.329:0.372:2.953 and a total of 17 amino acids were detected. The results of the current study demonstrated that GL-PP significantly inhibited U251 cellular proliferation. The proportion of G0/G1 phase cells and sub-G1 phase cells significantly increased as the concentration of GL-PP increased, as did the activity of caspase-3. These results indicate that GL-PP directly inhibited human glioma U251 proliferation by inducing cell cycle arrest and promoting apoptosis. PMID:29541200

Polysaccharide peptide isolated from grass-cultured Ganoderma lucidum induces anti-proliferative and pro-apoptotic effects in the human U251 glioma cell line.

PubMed

Wang, Chunhua; Lin, Dongmei; Chen, Quan; Lin, Shuqian; Shi, Songsheng; Chen, Chunmei

2018-04-01

The Ganoderma lucidum ( G. lucidum ) mushroom is one of the most extensively studied functional foods, known for its numerous health benefits, including the inhibition of tumor cell growth. The present study assessed the anti-proliferative and pro-apoptotic activity of a novel G. lucidum polysaccharide peptide (GL-PP) in human glioma U251 cells, which was purified from grass-cultured G. lucidum . GL-PP is a glycopeptide with an average molecular weight of 42,635 Da and a polysaccharide-to-peptide ratio of 88.70:11.30. The polysaccharides were composed of l-arabinose, d-mannose and d-glucose at a molar ratio of 1.329:0.372:2.953 and a total of 17 amino acids were detected. The results of the current study demonstrated that GL-PP significantly inhibited U251 cellular proliferation. The proportion of G 0 /G 1 phase cells and sub-G 1 phase cells significantly increased as the concentration of GL-PP increased, as did the activity of caspase-3. These results indicate that GL-PP directly inhibited human glioma U251 proliferation by inducing cell cycle arrest and promoting apoptosis.

The novel agent phospho-glycerol-ibuprofen-amide (MDC-330) inhibits glioblastoma growth in mice: an effect mediated by cyclin D1

PubMed Central

Bartels, Lauren E.; Mattheolabakis, George; Vaeth, Brandon M.; LaComb, Joseph F.; Wang, Ruixue; Zhi, Jizu; Komninou, Despina; Rigas, Basil; Mackenzie, Gerardo G.

2016-01-01

Given that glioblastoma multiforme (GBM) is associated with poor prognosis, new agents are urgently needed. We developed phospho-glycerol-ibuprofen-amide (PGIA), a novel ibuprofen derivative, and evaluated its safety and efficacy in preclinical models of GBM, and its mechanism of action using human GBM cells and animal tumor models. Furthermore, we explored whether formulating PGIA in polymeric nanoparticles could enhance its levels in the brain. PGIA was 3.7- to 5.1-fold more potent than ibuprofen in suppressing the growth of human GBM cell lines. PGIA 0.75× IC50 inhibited cell proliferation by 91 and 87% in human LN-229 and U87-MG GBM cells, respectively, and induced strong G1/S arrest. In vivo, compared with control, PGIA reduced U118-MG and U87-MG xenograft growth by 77 and 56%, respectively (P < 0.05), and was >2-fold more efficacious than ibuprofen. Normal human astrocytes were resistant to PGIA, indicating selectivity. Mechanistically, PGIA reduced cyclin D1 levels in a time- and concentration-dependent manner in GBM cells and in xenografts. PGIA induced cyclin D1 degradation via the proteasome pathway and induced dephosphorylation of GSK3β, which was required for cyclin D1 turnover. Furthermore, cyclin D1 overexpression rescued GBM cells from the cell growth inhibition by PGIA. Moreover, the formulation of PGIA in poly-(l)-lactic acid poly(ethylene glycol) polymeric nanoparticles improved its pharmacokinetics in mice, delivering PGIA to the brain. PGIA displays strong efficacy against GBM, crosses the blood-brain barrier when properly formulated, reaching the target tissue, and establishes cyclin D1 as an important molecular target. Thus, PGIA merits further evaluation as a potential therapeutic option for GBM. PMID:26905586

Cn-AMP2 from green coconut water is an anionic anticancer peptide.

PubMed

Prabhu, Saurabh; Dennison, Sarah R; Mura, Manuela; Lea, Robert W; Snape, Timothy J; Harris, Frederick

2014-12-01

Globally, death due to cancers is likely to rise to over 20 million by 2030, which has created an urgent need for novel approaches to anticancer therapies such as the development of host defence peptides. Cn-AMP2 (TESYFVFSVGM), an anionic host defence peptide from green coconut water of the plant Cocos nucifera, showed anti-proliferative activity against the 1321N1 and U87MG human glioma cell lines with IC50 values of 1.25 and 1.85 mM, respectively. The membrane interactive form of the peptide was found to be an extended conformation, which primarily included β-type structures (levels > 45%) and random coil architecture (levels > 45%). On the basis of these and other data, it is suggested that the short anionic N-terminal sequence (TES) of Cn-AMP2 interacts with positively charged moieties in the cancer cell membrane. Concomitantly, the long hydrophobic C-terminal sequence (YFVFSVGM) of the peptide penetrates the membrane core region, thereby driving the translocation of Cn-AMP2 across the cancer cell membrane to attack intracellular targets and induce anti-proliferative mechanisms. This work is the first to demonstrate that anionic host defence peptides have activity against human glioblastoma, which potentially provides an untapped source of lead compounds for development as novel agents in the treatment of these and other cancers. Copyright © 2014 European Peptide Society and John Wiley & Sons, Ltd.

Vitamin K3-2,3-epoxide induction of apoptosis with activation of ROS-dependent ERK and JNK protein phosphorylation in human glioma cells.

PubMed

Wu, Jender; Chien, Chih-Chiang; Yang, Liang-Yo; Huang, Guan-Cheng; Cheng, Min-Chi; Lin, Che-Tong; Shen, Shing-Chuan; Chen, Yen-Chou

2011-08-15

2-Methyl-1,4-naphthoquinone (menadione or vitamin K3; EPO) and K3-2,3-epoxide (EPO1), but not vitamin K3-3-OH (EPO2), exhibited cytotoxicity that caused DNA fragmentation and chromatin condensation in U87 and C6 cells. EPO1 showed more-potent cytotoxicity than EPO, and the IC(50) values of EPO and EPO1 in U87 cells were 37.5 and 15.7μM, respectively. Activation of caspase 3 enzyme activity with cleavage of caspase 3 protein was detected in EPO1-treated U87 and C6 cells, and the addition of the caspase 3 peptidyl inhibitor, DEVD-FMK, reduced the cytotoxic effect of EPO1. An increase in the intracellular ROS level by EPO1 was observed in the DCHF-DA analysis, and EPO1-induced apoptosis and caspase 3 protein cleavage were prevented by adding the antioxidant, N-acetyl-cysteine (NAC), with decreased ROS production elicited by EPO1. Activation of ERK and JNK, but not p38, via phosphorylation induction was identified in EPO1- but not EPO- or EPO2-treated U87 and C6 cells, and this was blocked by adding NAC. However, the ERK inhibitor, PD98059, and the JNK inhibitor, SP600125, showed no effect on EPO1-induced cytotoxicity in either cell type. Our findings demonstrate that 2,3-epoxide substitution significantly potentiates the apoptotic effect of vitamin K3 via stimulating ROS production, which may be useful in the chemotherapy of glioblastoma cells. Copyright © 2011. Published by Elsevier Ireland Ltd.

BCNU wafer placement with temozolomide (TMZ) in the immediate postoperative period after tumor resection followed by radiation therapy with TMZ in patients with newly diagnosed high grade glioma: final results of a prospective, multi-institutional, phase II trial.

PubMed

Burri, Stuart H; Prabhu, Roshan S; Sumrall, Ashley L; Brick, Wendy; Blaker, Brian D; Heideman, Brent E; Boltes, Peggy; Kelly, Renee; Symanowski, James T; Wiggins, Walter F; Ashby, Lynn; Norton, H James; Judy, Kevin; Asher, Anthony L

2015-06-01

Temozolomide (TMZ) and BCNU have demonstrated anti-glioma synergism in preclinical models. We report final data from a prospective, multi-institutional study of BCNU wafers and early TMZ followed by radiation therapy with TMZ in patients with newly diagnosed malignant glioma. 65 patients were consented in 4 institutions, and 46 patients (43 GBM, 3 AA) were eligible for analysis. After resection and BCNU wafer placement, TMZ began on day four postoperatively. Radiation and TMZ (RT/TMZ) were then administered, followed by monthly TMZ at 200 mg/m2 for the first 26 patients, which was reduced to 150 mg/m2 for the remaining 20 patients. Non-hematologic toxicities were minimal. Nine of 27 patients (33 %) who received 200 mg/m2 TMZ, but only 1 of 20 (5 %) who received 150 mg/m2, experienced grade 3/4 thrombocytopenia. Median progression free survival (PFS) and overall survival (OS) period was 8.5 and 18 months, respectively. The 1-year OS rate was 76 %, which is a significant improvement compared with the historical control 1-year OS rate of 59 % (p = 0.023). However, there was no difference in 1-year OS compared with standard RT/TMZ (p = 0.12) or BCNU wafer followed by RT/TMZ (p = 0.87) in post hoc analyses. Early post-operative TMZ can be safely administered with BCNU wafers following resection of malignant glioma at the 150 mg/m2 dose level. Although there was an OS benefit compared to historical control, there was no indication of benefit for BCNU wafers and early TMZ in addition to standard RT/TMZ or early TMZ in addition to regimens of BCNU wafers followed by RT/TMZ.

Cytotoxic Effects of Temozolomide and Radiation are Additive- and Schedule-Dependent

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chalmers, Anthony J., E-mail: a.j.chalmers@sussex.ac.u; Genome Damage and Stability Centre, University of Sussex, Falmer; Ruff, Elliot M.

2009-12-01

Purpose: Despite aggressive therapy comprising radical radiation and temozolomide (TMZ) chemotherapy, the prognosis for patients with glioblastoma multiforme (GBM) remains poor, particularly if tumors express O{sup 6}-methylguanine-DNA-methyltransferase (MGMT). The interactions between radiation and TMZ remain unclear and have important implications for scheduling and for developing strategies to improve outcomes. Methods and Materials: Factors determining the effects of combination therapy on clonogenic survival, cell-cycle checkpoint signaling and DNA repair were investigated in four human glioma cell lines (T98G, U373-MG, UVW, U87-MG). Results: Combining TMZ and radiation yielded additive cytotoxicity, but only when TMZ was delivered 72 h before radiation. Radiosensitization wasmore » not observed. TMZ induced G2/M cell-cycle arrest at 48-72 h, coincident with phosphorylation of Chk1 and Chk2. Additive G2/M arrest and Chk1/Chk2 phosphorylation was only observed when TMZ preceded radiation by 72 h. The ataxia-telangiectasia mutated (ATM) inhibitor KU-55933 increased radiation sensitivity and delayed repair of radiation-induced DNA breaks, but did not influence TMZ effects. The multiple kinase inhibitor caffeine enhanced the cytotoxicity of chemoradiation and exacerbated DNA damage. Conclusions: TMZ is not a radiosensitizing agent but yields additive cytotoxicity in combination with radiation. Our data indicate that TMZ treatment should commence at least 3 days before radiation to achieve maximum benefit. Activation of G2/M checkpoint signaling by TMZ and radiation has a cytoprotective effect that can be overcome by dual inhibition of ATM and ATR. More specific inhibition of checkpoint signaling will be required to increase treatment efficacy without exacerbating toxicity.« less

Investigation of imatinib loaded surface decorated biodegradable nanocarriers against glioblastoma cell lines: Intracellular uptake and cytotoxicity studies.

PubMed

Khan, Abrar M; Ahmad, Farhan Jalees; Panda, Amulya K; Talegaonkar, Sushama

2016-06-30

Overexpression of P-glycoprotein (P-gp) efflux transporter in glioma cells thwarts the build-up of therapeutic concentration of drugs usually resulting into poor therapeutic outcome. To surmount aforesaid challenge, Imatinib (IMM) loaded Poly-lactide-co-glycolic acid nanoparticles (IMM-PLGA-NPs) were developed and optimized by Box Behnken Design as a new treatment stratagem in malignant glioma. Optimized NPs were functionalized with Pluronic(®) P84, P-gp inhibitor (IMM-PLGA-P84-NPs) which showed size, PDI, zeta potential, drug loading, 182.63±13.56nm, 0.196±0.021, -15.2±1.49mV, 40.63±2.04μg/mg, respectively. Intracellular uptake study conducted on A172, U251MG and C6 glioma cells demonstrated significantly high uptake of IMM through NPs when compared with IMM solution (IMM-S), p<0.001. IMM-PLGA-P84-NPs showed better uptake in P-gp expressing cell line (U251MG and C6) while uncoated NPs showed higher uptake in non-P-gp expressing cell line (A-172). Cytotoxicity studies demonstrated significantly low IC50 for both IMM-PLGA-NPs and IMM-PLGA-P84-NPs when compared with IC50 of IMM-S. IMM-PLGA-P84-NPs showed a significantly low IC50 against P-gp overexpressing cell lines when compared with IC50 of IMM-PLGA-NPs. In contrary, IMM-PLGA-NPs showed lower IC50 against non P-gp expressing cell line. This study demonstrated the feasibility of targeting surface decorated NPs to multidrug resistant gliomas. However, to address its clinical utility extensive in vivo studies are required. Copyright © 2016 Elsevier B.V. All rights reserved.

Hardware, software, and scanning issues encountered during small animal imaging of photodynamic therapy in the athymic nude rat

NASA Astrophysics Data System (ADS)

Cross, Nathan; Sharma, Rahul; Varghai, Davood; Spring-Robinson, Chandra; Oleinick, Nancy L.; Muzic, Raymond F., Jr.; Dean, David

2007-02-01

Small animal imaging devices are now commonly used to study gene activation and model the effects of potential therapies. We are attempting to develop a protocol that non-invasively tracks the affect of Pc 4-mediated photodynamic therapy (PDT) in a human glioma model using structural image data from micro-CT and/or micro-MR scanning and functional data from 18F-fluorodeoxy-glucose (18F-FDG) micro-PET imaging. Methods: Athymic nude rat U87-derived glioma was imaged by micro-PET and either micro-CT or micro-MR prior to Pc 4-PDT. Difficulty insuring animal anesthesia and anatomic position during the micro-PET, micro-CT, and micro-MR scans required adaptation of the scanning bed hardware. Following Pc 4-PDT the animals were again 18F-FDG micro-PET scanned, euthanized one day later, and their brains were explanted and prepared for H&E histology. Histology provided the gold standard for tumor location and necrosis. The tumor and surrounding brain functional and structural image data were then isolated and coregistered. Results: Surprisingly, both the non-PDT and PDT groups showed an increase in tumor functional activity when we expected this signal to disappear in the group receiving PDT. Co-registration of the functional and structural image data was done manually. Discussion: As expected, micro-MR imaging provided better structural discrimination of the brain tumor than micro-CT. Contrary to expectations, in our preliminary analysis 18F-FDG micro-PET imaging does not readily discriminate the U87 tumors that received Pc 4-PDT. We continue to investigate the utility of micro-PET and other methods of functional imaging to remotely detect the specificity and sensitivity of Pc 4-PDT in deeply placed tumors.

Nuclear Motility in Glioma Cells Reveals a Cell-Line Dependent Role of Various Cytoskeletal Components

PubMed Central

Kiss, Alexa; Horvath, Peter; Rothballer, Andrea; Kutay, Ulrike; Csucs, Gabor

2014-01-01

Nuclear migration is a general term for the movement of the nucleus towards a specific site in the cell. These movements are involved in a number of fundamental biological processes, such as fertilization, cell division, and embryonic development. Despite of its importance, the mechanism of nuclear migration is still poorly understood in mammalian cells. In order to shed light on the mechanical processes underlying nuclear movements, we adapted a micro-patterning based assay. C6 rat and U87 human glioma cells seeded on fibronectin patterns - thereby forced into a bipolar morphology - displayed oscillatory movements of the nucleus or the whole cell, respectively. We found that both the actomyosin system and microtubules are involved in the nuclear/cellular movements of both cell lines, but their contributions are cell-/migration-type specific. Dynein activity was necessary for nuclear migration of C6 cells but active myosin-II was dispensable. On the other hand, coupled nuclear and cellular movements of U87 cells were driven by actomyosin contraction. We explain these cell-line dependent effects by the intrinsic differences in the overall mechanical tension due to the various cytoskeletal elements inside the cell. Our observations showed that the movements of the nucleus and the centrosome are strongly correlated and display large variation, indicating a tight but flexible coupling between them. The data also indicate that the forces responsible for nuclear movements are not acting directly via the centrosome. Based on our observations, we propose a new model for nuclear oscillations in C6 cells in which dynein and microtubule dynamics are the main drivers of nuclear movements. This mechanism is similar to the meiotic nuclear oscillations of Schizosaccharomyces pombe and may be evolutionary conserved. PMID:24691067

TLR9-ERK-mTOR signaling is critical for autophagic cell death induced by CpG oligodeoxynucleotide 107 combined with irradiation in glioma cells

PubMed Central

Li, Xiaoli; Cen, Yanyan; Cai, Yongqing; Liu, Tao; Liu, Huan; Cao, Guanqun; Liu, Dan; Li, Bin; Peng, Wei; Zou, Jintao; Pang, Xueli; Zheng, Jiang; Zhou, Hong

2016-01-01

Synthetic oligodeoxynucleotides containing unmethylated CpG dinucleotides (CpG ODN) function as potential radiosensitizers for glioma treatment, although the underlying mechanism is unclear. It was observed that CpG ODN107, when combined with irradiation, did not induce apoptosis. Herein, the effect of CpG ODN107 + irradiation on autophagy and the related signaling pathways was investigated. In vitro, CpG ODN107 + irradiation induced autophagosome formation, increased the ratio of LC3 II/LC3 I, beclin 1 and decreased p62 expression in U87 cells. Meanwhile, CpG ODN107 also increased LC3 II/LC3 I expression in U251 and CHG-5 cells. In vivo, CpG ODN107 combined with local radiotherapy induced autophagosome formation in orthotopic transplantation tumor. Investigation of the molecular mechanisms demonstrated that CpG ODN107 + irradiation increased the levels of TLR9 and p-ERK, and decreased the level of p-mTOR in glioma cells. Further, TLR9-specific siRNA could affect the expressions of p-ERK and autophagy-related proteins in glioma cells. Taken together, CpG ODN107 combined with irradiation could induce autophagic cell death, and this effect was closely related to the TLR9-ERK-mTOR signaling pathway in glioma cells, providing new insights into the investigation mechanism of CpG ODN. PMID:27251306

Preparation and characterization of Fe3O4@Au-C225 composite targeted nanoparticles for MRI of human glioma.

PubMed

Ge, Yaoqi; Zhong, Yuejiao; Ji, Guozhong; Lu, Qianling; Dai, Xinyu; Guo, Zhirui; Zhang, Peng; Peng, Gang; Zhang, Kangzhen; Li, Yuntao

2018-01-01

To study the characterization of Fe3O4@Au-C225 composite targeted MNPs. Fe3O4@Au-C225 was prepared by the absorption method. The immunosorbent assay was used to evaluate its absorption efficiency at C225 Fc. ZETA SIZER3000 laser particle size analyzer, ultraviolet photometer and its characteristics were analyzed by VSM. the targeting effect of Fe3O4@Au-C225 composite targeted MNPs on U251 cells in vitro were detected by 7.0 Tesla Micro-MR; and subcutaneous transplanted human glioma in nude mice were performed the targeting effect in vivo after tail vein injection of Fe3O4@Au-C225 composite targeted MNPs by MRI. The self-prepared Fe3O4@Au composite MNPs can adsorb C225 with high efficiency of adsorption so that Fe3O4@Au-C225 composite targeted MNPs were prepared successfully. Fe3O4@Au-C225 composite targeted MNPs favorably targeted human glioma cell line U251 in vitro; Fe3O4@Au-C225 composite targeted MNPs have good targeting ability to xenografted glioma on nude mice in vivo, and can be traced by MRI. The Fe3O4@Au-C225 composite targeted MNPs have the potential to be used as a tracer for glioma in vivo.

Dynamic contrast enhanced-magnetic resonance imaging (DCE-MRI) for the assessment of Pc 4-sensitized photodynamic therapy of a U87-derived glioma model in the athymic nude rat

NASA Astrophysics Data System (ADS)

Anka, Ali; Thompson, Paul; Mott, Eric; Sharma, Rahul; Zhang, Ruozhen; Cross, Nathan; Sun, Jiayang; Flask, Chris A.; Oleinick, Nancy L.; Dean, David

2010-02-01

Introduction: Dynamic Contrast-Enhanced-Magnetic Resonance Imaging (DCE-MRI) may provide a means of tracking the outcome of Pc 4-sensitized photodynamic therapy (PDT) in deeply placed lesions (e.g., brain tumors). We previously determined that 150 μL of gadolinium (Gd-DTPA) produces optimal enhancement of U87-derived intracerebral tumors in an athymic nude rat glioma model. We wish to determine how consistently DCE-MRI enhancement will detect an increase in Gd-enhancement of these tumors following Pc 4-PDT. Methods: We injected 2.5 x 105 U87 cells into the brains of 6 athymic nude rats. After 7-8 days pre-Pc 4 PDT peri-tumor DCE-MRI images were acquired on a 7.0T microMRI scanner before and after administration of 150 μL Gd. DCE-MRI scans were repeated on Days 11, 12, and 13 following Pc 4-PDT (Day 8 or 9). Results: Useful DCE-MRI data were obtained for these animals before and after Pc 4- PDT. In the pre-Pc 4-PDT DCE-MRI scans an average normalized peak Gd enhancement was observed in tumor tissue that was 1.297 times greater than baseline (0.035 Standard Error [SE]). The average normalized peak Gd enhancement in the tumor tissue in the scan following PDT (Day 11) was 1.537 times greater than baseline (0.036 SE), a statistically significant increase in enhancement (p = 0.00584) over the pre-PDT level. Discussion: A 150 μL Gd dose appears to provide an unambiguous increase in signal indicating Pc 4-PDT-induced necrosis of the U87-derived tumor. Our DCEMRI protocol may allow the development of a clinically robust, unambiguous, non-invasive technique for the assessment of PDT outcome.

A Hypoxia-Targeted Boron Neutron Capture Therapy Agent for the Treatment of Glioma.

PubMed

Luderer, Micah John; Muz, Barbara; de la Puente, Pilar; Chavalmane, Sanmathi; Kapoor, Vaishali; Marcelo, Raymundo; Biswas, Pratim; Thotala, Dinesh; Rogers, Buck; Azab, Abdel Kareem

2016-10-01

Boron neutron capture therapy (BNCT) has the potential to become a viable cancer treatment modality, but its clinical translation has been limited by the poor tumor selectivity of agents. To address this unmet need, a boronated 2-nitroimidazole derivative (B-381) was synthesized and evaluated for its capability of targeting hypoxic glioma cells. B-381 has been synthesized from a 1-step reaction. Using D54 and U87 glioma cell lines, the in vitro cytotoxicity and cellular accumulation of B-381 has been evaluated under normoxic and hypoxic conditions compared to L-boronophenylalanine (BPA). Furthermore, tumor retention of B-381 was evaluated in vivo. B-381 had low cytotoxicity in normal and cancer cells. Unlike BPA, B-381 illustrated preferential retention in hypoxic glioma cells compared to normoxic glioma cells and normal tissues in vitro. In vivo, B-381 illustrated significantly higher long-term tumor retention compared to BPA, with 9.5-fold and 6.5-fold higher boron levels at 24 and 48 h, respectively. B-381 represents a new class of BNCT agents in which their selectivity to tumors is based on hypoxic tumor metabolism. Further studies are warranted to evaluate B-381 and similar compounds as preclinical candidates for future BNCT clinical trials for the treatment of glioma.

Development of induced glioblastoma by implantation of a human xenograft in Yucatan minipig as a large animal model.

PubMed

Khoshnevis, Mehrdad; Carozzo, Claude; Bonnefont-Rebeix, Catherine; Belluco, Sara; Leveneur, Olivia; Chuzel, Thomas; Pillet-Michelland, Elodie; Dreyfus, Matthieu; Roger, Thierry; Berger, François; Ponce, Frédérique

2017-04-15

Glioblastoma is the most common and deadliest primary brain tumor for humans. Despite many efforts toward the improvement of therapeutic methods, prognosis is poor and the disease remains incurable with a median survival of 12-14.5 months after an optimal treatment. To develop novel treatment modalities for this fatal disease, new devices must be tested on an ideal animal model before performing clinical trials in humans. A new model of induced glioblastoma in Yucatan minipigs was developed. Nine immunosuppressed minipigs were implanted with the U87 human glioblastoma cell line in both the left and right hemispheres. Computed tomography (CT) acquisitions were performed once a week to monitor tumor growth. Among the 9 implanted animals, 8 minipigs showed significant macroscopic tumors on CT acquisitions. Histological examination of the brain after euthanasia confirmed the CT imaging findings with the presence of an undifferentiated glioma. Yucatan minipig, given its brain size and anatomy (gyrencephalic structure) which are comparable to humans, provides a reliable brain tumor model for preclinical studies of different therapeutic METHODS: in realistic conditions. Moreover, the short development time, the lower cyclosporine and caring cost and the compatibility with the size of commercialized stereotactic frames make it an affordable and practical animal model, especially in comparison with large breed pigs. This reproducible glioma model could simulate human anatomical conditions in preclinical studies and facilitate the improvement of novel therapeutic devices, designed at the human scale from the outset. Copyright © 2017 Elsevier B.V. All rights reserved.

Optimal gadolinium dose level for magnetic resonance imaging (MRI) contrast enhancement of U87-derived tumors in athymic nude rats for the assessment of photodynamic therapy

NASA Astrophysics Data System (ADS)

Cross, Nathan; Varghai, Davood; Flask, Chris A.; Feyes, Denise K.; Oleinick, Nancy L.; Dean, David

2009-02-01

This study aims to determine the effect of varying gadopentetate dimeglumine (Gd-DTPA) dose on Dynamic Contrast Enhanced-Magnetic Resonance Imaging (DCE-MRI) tracking of brain tumor photodynamic therapy (PDT) outcome. Methods: We injected 2.5 x 105 U87 cells (derived from human malignant glioma) into the brains of six athymic nude rats. After 9, 12, and 13 days DCE-MRI images were acquired on a 9.4 T micro-MRI scanner before and after administration of 100, 150, or 200 μL of Gd-DTPA. Results: Tumor region normalized DCE-MRI scan enhancement at peak was: 1.217 over baseline (0.018 Standard Error [SE]) at the 100 μL dose, 1.339 (0.013 SE) at the 150 μL dose, and 1.287 (0.014 SE) at the 200 μL dose. DCE-MRI peak tumor enhancement at the 150 μL dose was significantly greater than both the 100 μL dose (p < 3.323E-08) and 200 μL dose (p < 0.0007396). Discussion: In this preliminary study, the 150 μL Gd-DTPA dose provided the greatest T1 weighted contrast enhancement, while minimizing negative T2* effects, in DCE-MRI scans of U87-derived tumors. Maximizing Gd-DTPA enhancement in DCE-MRI scans may assist development of a clinically robust (i.e., unambiguous) technique for PDT outcome assessment.

Targeted Antiepidermal Growth Factor Receptor (Cetuximab) Immunoliposomes Enhance Cellular Uptake In Vitro and Exhibit Increased Accumulation in an Intracranial Model of Glioblastoma Multiforme

PubMed Central

Mortensen, Joachim Høg

2013-01-01

Therapeutic advances do not circumvent the devastating fact that the survival rate in glioblastoma multiforme (GBM) is less than 5%. Nanoparticles consisting of liposome-based therapeutics are provided against a variety of cancer types including GBM, but available liposomal formulations are provided without targeting moieties, which increases the dosing demands to reach therapeutic concentrations with risks of side effects. We prepared PEGylated immunoliposomes (ILs) conjugated with anti-human epidermal growth factor receptor (EGFR) antibodies Cetuximab (α-hEGFR-ILs). The affinity of the α-hEGFR-ILs for the EGF receptor was evaluated in vitro using U87 mg and U251 mg cells and in vivo using an intracranial U87 mg xenograft model. The xenograft model was additionally analyzed with respect to permeability to endogenous albumin, tumor size, and vascularization. The in vitro studies revealed significantly higher binding of α-hEGFR-ILs when compared with liposomes conjugated with isotypic nonimmune immunoglobulin. The uptake and internalization of the α-hEGFR-ILs by U87 mg cells were further confirmed by 3D deconvolution analyses. In vivo, the α-hEGFR-ILs accumulated to a higher extent inside the tumor when compared to nonimmune liposomes. The data show that α-hEGFR-ILs significantly enhance the uptake and accumulation of liposomes in this experimental model of GBM suggestive of improved specific nanoparticle-based delivery. PMID:24175095

Role of a second chemotherapy in recurrent malignant glioma patients who progress on bevacizumab.

PubMed

Quant, Eudocia C; Norden, Andrew D; Drappatz, Jan; Muzikansky, Alona; Doherty, Lisa; Lafrankie, Debra; Ciampa, Abigail; Kesari, Santosh; Wen, Patrick Y

2009-10-01

Bevacizumab is a humanized monoclonal antibody against vascular endothelial growth factor (VEGF) that has efficacy in recurrent malignant gliomas, particularly in combination with irinotecan. However, responses are rarely durable. Continuation of bevacizumab in combination with another chemotherapeutic agent may demonstrate some activity. In this article we present a retrospective review of 54 patients with recurrent malignant gliomas who progressed on a bevacizumab-containing regimen and were then treated with an alternate bevacizumab-containing regimen. All patients received intravenous bevacizumab (5-10 mg/kg) every 2 weeks alone or in combination with an additional chemotherapeutic agent, such as irinotecan. There was no limit on the number of prior therapies. Clinical characteristics and outcomes were reviewed. Tumor progression was determined by a combination of clinical status and radiographic changes. Patients were 33 men, 21 women (median age, 50 years; range, 23-72 years) with a median KPS score of 80 prior to the first bevacizumab-containing regimen and 70 prior to the second regimen; median prior chemotherapy regimens including the first bevacizumab-containing regimen was 3 (range, 2-5). Median progression-free survival (PFS) on the first bevacizumab-containing regimen was 124 days (95% confidence interval [CI], 87-154 days); 6-month (6M)-PFS was 33%. Median PFS on the second bevacizumab-containing regimen was 37.5 days (95% CI, 34-42 days); 6M-PFS was 2%. Ten patients on the first regimen and 12 patients on the second regimen suffered grade 3/4 toxicities. Those patients with malignant gliomas who progressed despite a bevacizumab-containing regimen rarely responded to the second bevacizumab-containing chemotherapeutic regimen. In such patients, alternate therapies should be considered.

Role of a second chemotherapy in recurrent malignant glioma patients who progress on bevacizumab

PubMed Central

Quant, Eudocia C.; Norden, Andrew D.; Drappatz, Jan; Muzikansky, Alona; Doherty, Lisa; LaFrankie, Debra; Ciampa, Abigail; Kesari, Santosh; Wen, Patrick Y.

2009-01-01

Bevacizumab is a humanized monoclonal antibody against vascular endothelial growth factor (VEGF) that has efficacy in recurrent malignant gliomas, particularly in combination with irinotecan. However, responses are rarely durable. Continuation of bevacizumab in combination with another chemotherapeutic agent may demonstrate some activity. In this article we present a retrospective review of 54 patients with recurrent malignant gliomas who progressed on a bevacizumab-containing regimen and were then treated with an alternate bevacizumab-containing regimen. All patients received intravenous bevacizumab (5–10 mg/kg) every 2 weeks alone or in combination with an additional chemotherapeutic agent, such as irinotecan. There was no limit on the number of prior therapies. Clinical characteristics and outcomes were reviewed. Tumor progression was determined by a combination of clinical status and radiographic changes. Patients were 33 men, 21 women (median age, 50 years; range, 23–72 years) with a median KPS score of 80 prior to the first bevacizumab-containing regimen and 70 prior to the second regimen; median prior chemotherapy regimens including the first bevacizumab-containing regimen was 3 (range, 2–5). Median progression-free survival (PFS) on the first bevacizumab-containing regimen was 124 days (95% confidence interval [CI], 87–154 days); 6-month (6M)-PFS was 33%. Median PFS on the second bevacizumab-containing regimen was 37.5 days (95% CI, 34–42 days); 6M-PFS was 2%. Ten patients on the first regimen and 12 patients on the second regimen suffered grade 3/4 toxicities. Those patients with malignant gliomas who progressed despite a bevacizumab-containing regimen rarely responded to the second bevacizumab-containing chemotherapeutic regimen. In such patients, alternate therapies should be considered. PMID:19332770

A pilot study of peptide vaccines for VEGF receptor 1 and 2 in patients with recurrent/progressive high grade glioma

PubMed Central

Shibao, Shunsuke; Ueda, Ryo; Saito, Katsuya; Kikuchi, Ryogo; Nagashima, Hideaki; Kojima, Atsuhiro; Kagami, Hiroshi; Pareira, Eriel Sandika; Sasaki, Hikaru; Noji, Shinobu; Kawakami, Yutaka; Yoshida, Kazunari; Toda, Masahiro

2018-01-01

Object Early-phase clinical studies of glioma vaccines have shown feasibility and encouraging preliminary clinical activity. A vaccine that targets tumor angiogenesis factors in glioma microenvironment has not been reported. Therefore, we performed a pilot study to evaluate the safety and immunogenicity of a novel vaccination targeting tumor angiogenesis with synthetic peptides for vascular endothelial growth factor (VEGF) receptor epitopes in patients with recurrent/progressive high grade gliomas. Methods Eight patients received intranodal vaccinations weekly at a dose of 2mg/kg bodyweight 8 times. T-lymphocyte responses against VEGF receptor (VEGFR) epitopes were assessed by enzyme linked immunosorbent spot assays. Results This treatment was well-tolerated in patients. The first four vaccines induced positive immune responses against at least one of the targeted VEGFR epitopes in the peripheral blood mononuclear cells in 87.5% of patients. The median overall survival time in all patients was 15.9 months. Two achieved progression-free status lasting at least 6 months. Two patients with recurrent GBM demonstrated stable disease. Plasma IL-8 level was negatively correlated with overall survival. Conclusion These data demonstrate the safety and immunogenicity of VEGFR peptide vaccines targeting tumor vasculatures in high grade gliomas. PMID:29765561

Nanomelatonin triggers superior anticancer functionality in a human malignant glioblastoma cell line

NASA Astrophysics Data System (ADS)

Yadav, Sanjeev Kumar; Srivastava, Anup Kumar; Dev, Atul; Kaundal, Babita; Choudhury, Subhasree Roy; Karmakar, Surajit

2017-09-01

Melatonin (MEL) has promising medicinal value as an anticancer agent in a variety of malignancies, but there are difficulties in achieving a therapeutic dose due to its short half-life, low bioavailability, poor solubility and extensive first-pass metabolism. In this study chitosan/tripolyphosphate (TPP) nanoparticles were prepared by an ionic gelation method to overcome the therapeutic challenges of melatonin and to improve its anticancer efficacy. Characterization of the melatonin-loaded chitosan (MEL-CS) nanoformulation was performed using transmission and scanning electron microscopies, dynamic light scattering, Fourier transform infrared spectroscopy, Raman spectroscopy and x-ray diffraction. In vitro release, cellular uptake and efficacy studies were tested for their enhanced anticancer potential in human U87MG glioblastoma cells. Confocal studies revealed higher cellular uptake of MEL-CS nanoparticles and enhanced anticancer efficacy in human malignant glioblastoma cancer cells than in healthy non-malignant human HEK293T cells in mono- and co-culture models. Our study has shown for the first time that MEL-CS nanocomposites are therapeutically more effective as compared to free MEL at inducing functional anticancer efficacy in the human brain tumour U87MG cell line.

Effects of targeting moiety, linker, bifunctional chelator, and molecular charge on biological properties of 64Cu-labeled triphenylphosphonium cations.

PubMed

Kim, Young-Seung; Yang, Chang-Tong; Wang, Jianjun; Wang, Lijun; Li, Zi-Bo; Chen, Xiaoyuan; Liu, Shuang

2008-05-22

In this report, we present the synthesis and evaluation of six new 64Cu-labeled triphenylphosphonium (TPP) cations. Biodistribution studies were performed using the athymic nude mice bearing U87MG human glioma xenografts to explore the impact of TPP moieties, linkers, bifunctional chelators (BFCs), and molecular charge on biological properties of 64Cu radiotracers. On the basis of the results from this study, it is concluded that (1) mTPP (tris(4-methoxyphenyl)phosphonium) is a better mitochondrion-targeting molecule than TPP and 3mTPP (tris(2,4,6-trimethoxyphenyl)phosphonium); (2) DO3A (1,4,7,10-tetraazacyclododecane-4,7,10-triacetic acid) and DO2A (1,4,7,10-tetraazacyclododecane-4,7-diacetic acid) are suitable BFCs for the 64Cu-labeling of TPP cations; (3) NOTA-Bn ( S-2-(4-thioureidobenzyl)-1,4,7-triazacyclononane-1,4,7-triacetic acid) has a significant adverse effect on the radiotracer tumor uptake and tumor-to-background ratios; and (4) monoanionic BFCs should be avoided to ensure that 64Cu chelate has a neutral or negative charge. Considering the tumor uptake and tumor/liver ratios, 64Cu(DO2A-xy-TPP)+ is the best candidate for more extensive evaluations in different tumor-bearing animal models.

64Cu-Labeled triphenylphosphonium and triphenylarsonium cations as highly tumor-selective imaging agents.

PubMed

Wang, Jianjun; Yang, Chang-Tong; Kim, Young-Seung; Sreerama, Subramanya G; Cao, Qizhen; Li, Zi-Bo; He, Zhengjie; Chen, Xiaoyuan; Liu, Shuang

2007-10-18

This report presents synthesis and evaluation of the 64Cu-labeled triphenylphosphonium (TPP) cations as new radiotracers for imaging tumors by positron emission tomography. Biodistribution properties of 64Cu-L1, 64Cu-L2, 64Cu-L3, and 99mTc-Sestamibi were evaluated in athymic nude mice bearing U87MG human glioma xenografts. The most striking difference is that 64Cu-L1, 64Cu-L2, and 64Cu-L3 have much lower heart uptake (<0.6% ID/g) than 99mTc-Sestamibi ( approximately 18% ID/g) at >30 min p.i. Their tumor/heart ratios increase steadily from approximately 1 at 5 min p.i. to approximately 5 at 120 min p.i. The tumor/heart ratio of 64Cu-L3 is approximately 40 times better than that of 99mTc-Sestamibi at 120 min postinjection. Results from in vitro assays show that 64Cu-L1 is able to localize in tumor mitochondria. The tumor is clearly visualized in the tumor-bearing mice administered with 64Cu-L1 as 30 min postinjection. The 64Cu-labeled TPP/TPA cations are very selective radiotracers that are able to provide the information of mitochondrial bioenergetic function in tumors by monitoring mitochondrial potential in a noninvasive fashion.

Inhibitors of GLUT/SLC2A Enhance the Action of BCNU and Temozolomide against High-Grade Gliomas.

PubMed

Azzalin, Alberto; Nato, Giulia; Parmigiani, Elena; Garello, Francesca; Buffo, Annalisa; Magrassi, Lorenzo

2017-04-01

Glucose transport across glioblastoma membranes plays a crucial role in maintaining the enhanced glycolysis typical of high-grade gliomas and glioblastoma. We tested the ability of two inhibitors of the glucose transporters GLUT/SLC2A superfamily, indinavir (IDV) and ritonavir (RTV), and of one inhibitor of the Na/glucose antiporter type 2 (SGLT2/SLC5A2) superfamily, phlorizin (PHZ), in decreasing glucose consumption and cell proliferation of human and murine glioblastoma cells. We found in vitro that RTV, active on at least three different GLUT/SLC2A transporters, was more effective than IDV, a specific inhibitor of GLUT4/SLC2A4, both in decreasing glucose consumption and lactate production and in inhibiting growth of U87MG and Hu197 human glioblastoma cell lines and primary cultures of human glioblastoma. PHZ was inactive on the same cells. Similar results were obtained when cells were grown in adherence or as 3D multicellular tumor spheroids. RTV treatment but not IDV treatment induced AMP-activated protein kinase (AMPKα) phosphorylation that paralleled the decrease in glycolytic activity and cell growth. IDV, but not RTV, induced an increase in GLUT1/SLC2A1 whose activity could compensate for the inhibition of GLUT4/SLC2A4 by IDV. RTV and IDV pass poorly the blood brain barrier and are unlikely to reach sufficient liquoral concentrations in vivo to inhibit glioblastoma growth as single agents. Isobologram analysis of the association of RTV or IDV and 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) or 4-methyl-5-oxo-2,3,4,6,8-pentazabicyclo[4.3.0]nona-2,7,9-triene-9-carboxamide (TMZ) indicated synergy only with RTV on inhibition of glioblastoma cells. Finally, we tested in vivo the combination of RTV and BCNU on established GL261 tumors. This drug combination increased the overall survival and allowed a five-fold reduction in the dose of BCNU. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Triterpenoid saponins from Albizia lebbeck (L.) Benth and their inhibitory effect on the survival of high grade human brain tumor cells.

PubMed

Noté, Olivier Placide; Jihu, Dong; Antheaume, Cyril; Zeniou, Maria; Pegnyemb, Dieudonné Emmanuel; Guillaume, Dominique; Chneiwess, Hervé; Kilhoffer, Marie Claude; Lobstein, Annelise

2015-03-02

As part of our search of new bioactive triterpenoid saponins from Cameroonian Mimosaceae plants, phytochemical investigation of the roots of Albizia lebbeck led to the isolation of two new oleanane-type saponins, named lebbeckosides A-B (1-2). Their structures were established on the basis of extensive 1D and 2D NMR ((1)H, (13)C NMR, DEPT, COSY, TOCSY, ROESY, HSQC, and HMBC) and HRESIMS studies, and by chemical evidence. Compounds 1-2 were evaluated for their inhibitory effect on the metabolism of high grade human brain tumor cells, the human glioblastoma U-87 MG cell lines and the glioblastoma stem-like TG1 cells isolated from a patient tumor, and known to be particularly resistant to standard therapies. The isolated saponins showed significant cytotoxic activity against U-87 MG and TG1 cancer cells with IC50 values of 3.46 μM and 1.36 μM for 1, and 2.10 μM and 2.24 μM for 2, respectively. Copyright © 2014 Elsevier Ltd. All rights reserved.

Preparation and characterization of Fe3O4@Au-C225 composite targeted nanoparticles for MRI of human glioma

PubMed Central

Ge, Yaoqi; Zhong, Yuejiao; Ji, Guozhong; Lu, Qianling; Dai, Xinyu; Guo, Zhirui; Zhang, Peng; Peng, Gang; Zhang, Kangzhen; Li, Yuntao

2018-01-01

Objective To study the characterization of Fe3O4@Au-C225 composite targeted MNPs. Methods Fe3O4@Au-C225 was prepared by the absorption method. The immunosorbent assay was used to evaluate its absorption efficiency at C225 Fc. ZETA SIZER3000 laser particle size analyzer, ultraviolet photometer and its characteristics were analyzed by VSM. the targeting effect of Fe3O4@Au-C225 composite targeted MNPs on U251 cells in vitro were detected by 7.0 Tesla Micro-MR; and subcutaneous transplanted human glioma in nude mice were performed the targeting effect in vivo after tail vein injection of Fe3O4@Au-C225 composite targeted MNPs by MRI. Results The self-prepared Fe3O4@Au composite MNPs can adsorb C225 with high efficiency of adsorption so that Fe3O4@Au-C225 composite targeted MNPs were prepared successfully. Fe3O4@Au-C225 composite targeted MNPs favorably targeted human glioma cell line U251 in vitro; Fe3O4@Au-C225 composite targeted MNPs have good targeting ability to xenografted glioma on nude mice in vivo, and can be traced by MRI. Conclusion The Fe3O4@Au-C225 composite targeted MNPs have the potential to be used as a tracer for glioma in vivo. PMID:29652919

68Ga-labeling and in vivo evaluation of a uPAR binding DOTA- and NODAGA-conjugated peptide for PET imaging of invasive cancers.

PubMed

Persson, Morten; Madsen, Jacob; Østergaard, Søren; Ploug, Michael; Kjaer, Andreas

2012-05-01

The urokinase-type plasminogen activator receptor (uPAR) is a well-established biomarker for tumor aggressiveness and metastatic potential. DOTA-AE105 and DOTA-AE105-NH(2) labeled with (64)Cu have previously been demonstrated to be able to noninvasively monitor uPAR expression using positron emission tomography (PET) in human cancer xenograft mice models. Here we introduce (68)Ga-DOTA-AE105-NH(2) and (68)Ga-NODAGA-AE105-NH(2) and evaluate their imaging properties using small-animal PET. Synthesis of DOTA-AE105-NH(2) and NODAGA-AE105-NH(2) was based on solid-phase peptide synthesis protocols using the Fmoc strategy. (68)GaCl(3) was eluted from a (68)Ge/(68)Ga generator. The eluate was either concentrated on a cation-exchange column or fractionated and used directly for labeling. For in vitro characterization of both tracers, partition coefficient, buffer and plasma stability, uPAR binding affinity and cell uptake were determined. To characterize the in vivo properties, dynamic microPET imaging was carried out in nude mice bearing human glioma U87MG tumor xenograft. In vitro experiments revealed uPAR binding affinities in the lower nM range for both conjugated peptides and identical to AE105. Labeling of DOTA-AE105-NH(2) and NODAGA-AE105-NH(2) with (68)Ga was done at 95°C and room temperature, respectively. The highest radiochemical yield and purity were obtained using fractionated elution, whereas a negative effect of acetone on labeling efficiency for NODAGA-AE105-NH(2) was observed. Good stability in phosphate-buffered saline and mouse plasma was observed. High cell uptake was found for both tracers in U87MG tumor cells. Dynamic microPET imaging demonstrated good tumor-to-background ratio for both tracers. Tumor uptake was 2.1% ID/g and 1.3% ID/g 30 min postinjection and 2.0% ID/g and 1.1% ID/g 60 min postinjection for (68)Ga-NODAGA-AE105-NH(2) and (68)Ga-DOTA-AE105-NH(2), respectively. A significantly higher tumor-to-muscle ratio (P<.05) was found for (68)Ga-NODAGA-AE105-NH(2) 60 min postinjection. The use of (68)Ga-DOTA-AE105-NH(2) and (68)Ga-NODAGA-AE105-NH(2) as the first gallium-68 labeled uPAR radiotracers for noninvasive PET imaging is reported, which combine versatility with good imaging properties. These new tracers thus constitute an interesting alternative to the (64)Cu-labeled version ((64)Cu-DOTA-AE105 and 64Cu-DOTA-AE105-NH(2)) for detecting uPAR expression in tumor tissue. In our hands, the fractionated elution approach was superior for labeling of peptides, and (68)Ga-NODAGA-AE105-NH(2) is the favored tracer as it provides the highest tumor-to-background ratio. Copyright © 2012 Elsevier Inc. All rights reserved.

Chimeric antigen receptor containing ICOS signaling domain mediates specific and efficient antitumor effect of T cells against EGFRvIII expressing glioma

PubMed Central

2013-01-01

Background Adoptive transfer of chimeric antigen receptor (CAR)-modified T cells appears to be a promising immunotherapeutic strategy. CAR combines the specificity of antibody and cytotoxicity of cytotoxic T lymphocytes, enhancing T cells’ ability to specifically target antigens and to effectively kill cancer cells. Recent efforts have been made to integrate the costimulatory signals in the CAR to improve the antitumor efficacy. Epidermal growth factor receptor variant III (EGFRvIII) is an attractive therapeutic target as it frequently expresses in glioma and many other types of cancers. Our current study aimed to investigate the specific and efficient antitumor effect of T cells modified with CAR containing inducible costimulator (ICOS) signaling domain. Methods A second generation of EGFRvIII/CAR was generated and it contained the EGFRvIII single chain variable fragment, ICOS signaling domain and CD3ζ chain. Lentiviral EGFRvIII/CAR was prepared and human CD3+ T cells were infected by lentivirus encoding EGFRvIII/CAR. The expression of EGFRvIII/CAR on CD3+ T cells was confirmed by flow cytometry and Western blot. The functions of EGFRvIII/CAR+ T cells were evaluated using in vitro and in vivo methods including cytotoxicity assay, cytokine release assay and xenograft tumor mouse model. Results Chimeric EGFRvIIIscFv-ICOS-CD3ζ (EGFRvIII/CAR) was constructed and lentiviral EGFRvIII/CAR were made to titer of 106 TU/ml. The transduction efficiency of lentiviral EGFRvIII/CAR on T cells reached around 70% and expression of EGFRvIII/CAR protein was verified by immunoblotting as a band of about 57 kDa. Four hour 51Cr release assays demonstrated specific and efficient cytotoxicity of EGFRvIII/CAR+ T cells against EGFRvIII expressing U87 cells. A robust increase in the IFN-γ secretion was detected in the co-culture supernatant of the EGFRvIII/CAR+ T cells and the EGFRvIII expressing U87 cells. Intravenous and intratumor injection of EGFRvIII/CAR+ T cells inhibited the in vivo growth of the EGFRvIII expressing glioma cells. Conclusions Our study demonstrates that the EGFRvIII/CAR-modified T cells can destroy glioma cells efficiently in an EGFRvIII specific manner and release IFN-γ in an antigen dependent manner. The specific recognition and effective killing activity of the EGFRvIII-directed T cells with ICOS signaling domain lays a foundation for us to employ such approach in future cancer treatment. PMID:23656794

Chimeric antigen receptor containing ICOS signaling domain mediates specific and efficient antitumor effect of T cells against EGFRvIII expressing glioma.

PubMed

Shen, Chan-Juan; Yang, Yu-Xiu; Han, Ethan Q; Cao, Na; Wang, Yun-Fei; Wang, Yi; Zhao, Ying-Ying; Zhao, Li-Ming; Cui, Jian; Gupta, Puja; Wong, Albert J; Han, Shuang-Yin

2013-05-09

Adoptive transfer of chimeric antigen receptor (CAR)-modified T cells appears to be a promising immunotherapeutic strategy. CAR combines the specificity of antibody and cytotoxicity of cytotoxic T lymphocytes, enhancing T cells' ability to specifically target antigens and to effectively kill cancer cells. Recent efforts have been made to integrate the costimulatory signals in the CAR to improve the antitumor efficacy. Epidermal growth factor receptor variant III (EGFRvIII) is an attractive therapeutic target as it frequently expresses in glioma and many other types of cancers. Our current study aimed to investigate the specific and efficient antitumor effect of T cells modified with CAR containing inducible costimulator (ICOS) signaling domain. A second generation of EGFRvIII/CAR was generated and it contained the EGFRvIII single chain variable fragment, ICOS signaling domain and CD3ζ chain. Lentiviral EGFRvIII/CAR was prepared and human CD3+ T cells were infected by lentivirus encoding EGFRvIII/CAR. The expression of EGFRvIII/CAR on CD3+ T cells was confirmed by flow cytometry and Western blot. The functions of EGFRvIII/CAR+ T cells were evaluated using in vitro and in vivo methods including cytotoxicity assay, cytokine release assay and xenograft tumor mouse model. Chimeric EGFRvIIIscFv-ICOS-CD3ζ (EGFRvIII/CAR) was constructed and lentiviral EGFRvIII/CAR were made to titer of 106 TU/ml. The transduction efficiency of lentiviral EGFRvIII/CAR on T cells reached around 70% and expression of EGFRvIII/CAR protein was verified by immunoblotting as a band of about 57 kDa. Four hour 51Cr release assays demonstrated specific and efficient cytotoxicity of EGFRvIII/CAR+ T cells against EGFRvIII expressing U87 cells. A robust increase in the IFN-γ secretion was detected in the co-culture supernatant of the EGFRvIII/CAR+ T cells and the EGFRvIII expressing U87 cells. Intravenous and intratumor injection of EGFRvIII/CAR+ T cells inhibited the in vivo growth of the EGFRvIII expressing glioma cells. Our study demonstrates that the EGFRvIII/CAR-modified T cells can destroy glioma cells efficiently in an EGFRvIII specific manner and release IFN-γ in an antigen dependent manner. The specific recognition and effective killing activity of the EGFRvIII-directed T cells with ICOS signaling domain lays a foundation for us to employ such approach in future cancer treatment.

Angelica polysaccharides inhibit the growth and promote the apoptosis of U251 glioma cells in vitro and in vivo.

PubMed

Zhang, Wen-Feng; Yang, Yan; Li, Xin; Xu, Da-Yan; Yan, Yu-Li; Gao, Qiao; Jia, Ai-Ling; Duan, Ming-Hua

2017-09-15

Angelica sinensis (Oliv) Diels (Apiaceae) is a traditional medicine that has been used for more than 2000 years in China. It exhibits various therapeutic effects including neuroprotective, anti-oxidant, anti-inflammatory, and immunomodulatory activities. Angelica polysaccharides (APs), bioactive constituents of Angelica have been shown to be responsible for these effects; however, the utility of APs for the treatment of glioma and their mechanism of action remain to be elucidated. In this study, we investigated the inhibitory effects of APs on a glioma cell line and their molecular mechanism of action. U251 cells were utilized to confirm the effects of APs on glioma. The human glioblastoma cell line U251 was utilized for both in vitro and in vivo models, in which we tested the effects of APs. Flow cytometry, gene expression analysis, western blotting, and MTT assays were used to elucidate the effects of APs on cell proliferation, cell cycle, and apoptosis. The results demonstrated that APs significantly inhibited the growth and proliferation of U251 cells and induced their apoptosis. Furthermore, APs effectively reduced the expression of several cell cycle regulators: cyclins D1, B, and E. The apoptosis suppressor protein Bcl-2 was also downregulated, and the expression of pro-apoptotic proteins Bax and cleaved-caspase-3 increased. Additionally, APs inhibited the transforming growth factor (TGF)-β signaling pathway and stimulated the expression of E-cadherin, thus prohibiting cell growth. In conclusion, the results indicate that APs attenuate the tumorigenicity of glioma cells and promote their apoptosis by suppressing the TGF-β signaling pathway. The present study therefore provides evidence of the inhibitory effects of APs against glioma progression, and proposes their potential application as alternative therapeutic agents for glioma. Copyright © 2017 Elsevier GmbH. All rights reserved.

A Dual Tracer 18F-FCH/18F-FDG PET Imaging of an Orthotopic Brain Tumor Xenograft Model.

PubMed

Fu, Yilong; Ong, Lai-Chun; Ranganath, Sudhir H; Zheng, Lin; Kee, Irene; Zhan, Wenbo; Yu, Sidney; Chow, Pierce K H; Wang, Chi-Hwa

2016-01-01

Early diagnosis of low grade glioma has been a challenge to clinicians. Positron Emission Tomography (PET) using 18F-FDG as a radio-tracer has limited utility in this area because of the high background in normal brain tissue. Other radiotracers such as 18F-Fluorocholine (18F-FCH) could provide better contrast between tumor and normal brain tissue but with high incidence of false positives. In this study, the potential application of a dual tracer 18F-FCH/18F-FDG-PET is investigated in order to improve the sensitivity of PET imaging for low grade glioma diagnosis based on a mouse orthotopic xenograft model. BALB/c nude mice with and without orthotopic glioma xenografts from U87 MG-luc2 glioma cell line are used for the study. The animals are subjected to 18F-FCH and 18F-FDG PET imaging, and images acquired from two separate scans are superimposed for analysis. The 18F-FCH counts are subtracted from the merged images to identify the tumor. Micro-CT, bioluminescence imaging (BLI), histology and measurement of the tumor diameter are also conducted for comparison. Results show that there is a significant contrast in 18F-FCH uptake between tumor and normal brain tissue (2.65 ± 0.98), but with a high false positive rate of 28.6%. The difficulty of identifying the tumor by 18F-FDG only is also proved in this study. All the tumors can be detected based on the dual tracer technique of 18F-FCH/18F-FDG-PET imaging in this study, while the false-positive caused by 18F-FCH can be eliminated. Dual tracer 18F-FCH/18F-FDG PET imaging has the potential to improve the visualization of low grade glioma. 18F-FCH delineates tumor areas and the tumor can be identified by subtracting the 18F-FCH counts. The sensitivity was over 95%. Further studies are required to evaluate the possibility of applying this technique in clinical trials.

A Dual Tracer 18F-FCH/18F-FDG PET Imaging of an Orthotopic Brain Tumor Xenograft Model

PubMed Central

Ranganath, Sudhir H.; Zheng, Lin; Kee, Irene; Zhan, Wenbo; Yu, Sidney; Chow, Pierce K. H.; Wang, Chi-Hwa

2016-01-01

Early diagnosis of low grade glioma has been a challenge to clinicians. Positron Emission Tomography (PET) using 18F-FDG as a radio-tracer has limited utility in this area because of the high background in normal brain tissue. Other radiotracers such as 18F-Fluorocholine (18F-FCH) could provide better contrast between tumor and normal brain tissue but with high incidence of false positives. In this study, the potential application of a dual tracer 18F-FCH/18F-FDG-PET is investigated in order to improve the sensitivity of PET imaging for low grade glioma diagnosis based on a mouse orthotopic xenograft model. BALB/c nude mice with and without orthotopic glioma xenografts from U87 MG-luc2 glioma cell line are used for the study. The animals are subjected to 18F-FCH and 18F-FDG PET imaging, and images acquired from two separate scans are superimposed for analysis. The 18F-FCH counts are subtracted from the merged images to identify the tumor. Micro-CT, bioluminescence imaging (BLI), histology and measurement of the tumor diameter are also conducted for comparison. Results show that there is a significant contrast in 18F-FCH uptake between tumor and normal brain tissue (2.65 ± 0.98), but with a high false positive rate of 28.6%. The difficulty of identifying the tumor by 18F-FDG only is also proved in this study. All the tumors can be detected based on the dual tracer technique of 18F-FCH/ 18F-FDG-PET imaging in this study, while the false-positive caused by 18F-FCH can be eliminated. Dual tracer 18F-FCH/18F-FDG PET imaging has the potential to improve the visualization of low grade glioma. 18F-FCH delineates tumor areas and the tumor can be identified by subtracting the 18F-FCH counts. The sensitivity was over 95%. Further studies are required to evaluate the possibility of applying this technique in clinical trials. PMID:26844770

Knockdown of HDAC1 expression suppresses invasion and induces apoptosis in glioma cells.

PubMed

Wang, Xiao-Qiang; Bai, Hong-Min; Li, Shi-Ting; Sun, Hui; Min, Ling-Zhao; Tao, Bang-Bao; Zhong, Jun; Li, Bin

2017-07-18

Glioma is the most common malignant tumor of the central nervous system, with a low survival rate of five years worldwide. Although high expression and prognostic value of histone deacetylase 1 (HDAC1) have been recently reported in various types of human tumors, the molecular mechanism underlying the biological function of HDAC1 in glioma is still unclear. We found that HDAC1 was elevated in glioma tissues and cell lines. HDAC1 expression was closely related with pathological grade and overall survival of patients with gliomas. Downregulation of HDAC1 inhibited cell proliferation, prevented invasion of glioma cell lines, and induced cell apoptosis. The expression of apoptosis and metastasis related molecules were detected by RT-PCR and Western blot, respectively, in U251 and T98G cells with HDAC1 knockdown. We found that HDAC1 knockdown upregulated expression of BIM, BAX, cleaved CASPASE3 and E-CADHERIN, and decreased expression of TWIST1, SNAIL and MMP9 in U251 and T98G cells with HDAC1 knockdown. In vivo data showed that knockdown of HDAC1 inhibited tumor growth in nude mice. In summary, HDAC1 may therefore be considered an unfavorable progression indicator for glioma patients, and may also serve as a potential therapeutic target.

Phosphatidylserine-targeted liposome for enhanced glioma-selective imaging.

PubMed

Zhang, Liang; Habib, Amyn A; Zhao, Dawen

2016-06-21

Phosphatidylserine (PS), which is normally intracellular, becomes exposed on the outer surface of viable endothelial cells (ECs) of tumor vasculature. Utilizing a PS-targeting antibody, we have recently established a PS-targeted liposomal (PS-L) nanoplatform that has demonstrated to be highly tumor-selective. Because of the vascular lumen-exposed PS that is immediately accessible without a need to penetrate the intact blood brain barrier (BBB), we hypothesize that the systemically administered PS-L binds specifically to tumor vascular ECs, becomes subsequently internalized into the cells and then enables its cargos to be efficiently delivered to glioma parenchyma. To test this, we exploited the dual MRI/optical imaging contrast agents-loaded PS-L and injected it intravenously into mice bearing intracranial U87 glioma. At 24 h, both in vivo optical imaging and MRI depicted enhanced tumor contrast, distinct from the surrounding normal brain. Intriguingly, longitudinal MRI revealed temporal and spatial intratumoral distribution of the PS-L by following MRI contrast changes, which appeared punctate in tumor periphery at an earlier time point (4 h), but became clustering and disseminated throughout the tumor at 24 h post injection. Importantly, glioma-targeting specificity of the PS-L was antigen specific, since a control probe of irrelevant specificity showed minimal accumulation in the glioma. Together, these results indicate that the PS-L nanoplatform enables the enhanced, glioma-targeted delivery of imaging contrast agents by crossing the tumor BBB efficiently, which may also serve as a useful nanoplatform for anti-glioma drugs.

A Hypoxia-Targeted Boron Neutron Capture Therapy Agent for the Treatment of Glioma

PubMed Central

Luderer, Micah John; Muz, Barbara; de la Puente, Pilar; Chavalmane, Sanmathi; Kapoor, Vaishali; Marcelo, Raymundo; Biswas, Pratim; Thotala, Dinesh; Rogers, Buck; Azab, Abdel Kareem

2016-01-01

Purpose Boron neutron capture therapy (BNCT) has the potential to become a viable cancer treatment modality, but its clinical translation has been limited by the poor tumor selectivity of agents. To address this unmet need, a boronated 2-nitroimidazole derivative (B-381) was synthesized and evaluated for its capability of targeting hypoxic glioma cells. Methods B-381 has been synthesized from a 1-step reaction. Using D54 and U87 glioma cell lines, the in vitro cytotoxicity and cellular accumulation of B-381 has been evaluated under normoxic and hypoxic conditions compared to L-boronophenylalanine (BPA). Furthermore, tumor retention of B-381 was evaluated in vivo. Results B-381 had low cytotoxicity in normal and cancer cells. Unlike BPA, B-381 illustrated preferential retention in hypoxic glioma cells compared to normoxic glioma cells and normal tissues in vitro. In vivo, B-381 illustrated significantly higher long-term tumor retention compared to BPA, with 9.5-fold and 6.5-fold higher boron levels at 24 and 48 h, respectively. Conclusions B-381 represents a new class of BNCT agents in which their selectivity to tumors is based on tumor hypoxic metabolism, and further studies are warranted to evaluate this compound and similar compounds as preclinical candidates for future BNCT clinical trials for the treatment of glioma. PMID:27401411

Monitoring oxygen levels in orthotopic human glioma xenograft following carbogen inhalation and chemotherapy by implantable resonator-based oximetry.

PubMed

Hou, Huagang; Krishnamurthy Nemani, Venkata; Du, Gaixin; Montano, Ryan; Song, Rui; Gimi, Barjor; Swartz, Harold M; Eastman, Alan; Khan, Nadeem

2015-04-01

Hypoxia is a critical hallmark of glioma, and significantly compromises treatment efficacy. Unfortunately, techniques for monitoring glioma pO2 to facilitate translational research are lacking. Furthermore, poor prognosis of patients with malignant glioma, in particular glioblastoma multiforme, warrant effective strategies that can inhibit hypoxia and improve treatment outcome. EPR oximetry using implantable resonators was implemented for monitoring pO2 in normal cerebral tissue and U251 glioma in mice. Breathing carbogen (95% O2 + 5% CO2 ) was tested for hyperoxia in the normal brain and glioma xenografts. A new strategy to inhibit glioma growth by rationally combining gemcitabine and MK-8776, a cell cycle checkpoint inhibitor, was also investigated. The mean pO2 of left and right hemisphere were ∼56-69 mmHg in the normal cerebral tissue of mice. The mean baseline pO2 of U251 glioma on the first and fifth day of measurement was 21.9 ± 3.7 and 14.1 ± 2.4 mmHg, respectively. The mean brain pO2 including glioma increased by at least 100% on carbogen inhalation, although the response varied between the animals over days. Treatment with gemcitabine + MK-8776 significantly increased pO2 and inhibited glioma growth assessed by MRI. In conclusion, EPR oximetry with implantable resonators can be used to monitor the efficacy of carbogen inhalation and chemotherapy on orthotopic glioma in mice. The increase in glioma pO2 of mice breathing carbogen can be used to improve treatment outcome. The treatment with gemcitabine + MK-8776 is a promising strategy that warrants further investigation. © 2014 UICC.

S100B Attenuates Microglia Activation in Gliomas: Possible Role of STAT3 Pathway

PubMed Central

Zhang, Leying; Liu, Wei; Alizadeh, Darya; Zhao, Dongchang; Farrukh, Omar; Lin, Jeffrey; Badie, Sam A.; Badie, Behnam

2010-01-01

Despite significant infiltration into tumors, the effector function of macrophages (MPs) and microglia (MG) appears to be suppressed in gliomas. Although STAT3 pathway is thought to play a role in this process, the exact mechanism by which gliomas induce STAT3 activation in MPs and MG is not known. Because activation of receptor for advanced glycation end products (RAGE) can induce STAT3, and because gliomas express high levels of S100B, a RAGE ligand, we hypothesized that MP/MG STAT3 activity may be modulated through S100B-RAGE interaction. Exposure of N9 MG and bone marrow-derived monocytes (BMM) to GL261 glioma condition medium (GCM) and low (nM) levels of S100B increased RAGE expression, induced STAT3 and suppressed MG function in vitro. Furthermore, neutralization of S100B in GCM, partially reversed IL-1β suppression in BMM, suggesting that the inhibitory effect of GCM to be in part due to S100B. Finally, blockage of S100B-RAGE interaction inhibited STAT3 activation in N9 MG and in glioma MG/MP in vivo. These findings suggest that the RAGE pathway may play an important role in STAT3 induction in glioma-associated MG/MPs, and that this process may be mediated through S100B. PMID:21264954

S100B attenuates microglia activation in gliomas: possible role of STAT3 pathway.

PubMed

Zhang, Leying; Liu, Wei; Alizadeh, Darya; Zhao, Dongchang; Farrukh, Omar; Lin, Jeffrey; Badie, Sam A; Badie, Behnam

2011-03-01

Despite significant infiltration into tumors, the effector function of macrophages (MPs) and microglia (MG) appears to be suppressed in gliomas. Although STAT3 pathway is thought to play a role in this process, the exact mechanism by which gliomas induce STAT3 activation in MPs and MG is not known. Because activation of receptor for advanced glycation end products (RAGE) can induce STAT3, and because gliomas express high levels of S100B, a RAGE ligand, we hypothesized that MP/MG STAT3 activity may be modulated through S100B-RAGE interaction. Exposure of N9 MG and bone marrow-derived monocytes (BMM) to GL261 glioma condition medium (GCM) and low (nM) levels of S100B increased RAGE expression, induced STAT3 and suppressed MG function in vitro. Furthermore, neutralization of S100B in GCM, partially reversed IL-1β suppression in BMM, suggesting that the inhibitory effect of GCM to be in part due to S100B. Finally, blockage of S100B-RAGE interaction inhibited STAT3 activation in N9 MG and in glioma MG/MP in vivo. These findings suggest that the RAGE pathway may play an important role in STAT3 induction in glioma-associated MG/MPs, and that this process may be mediated through S100B. Copyright © 2010 Wiley-Liss, Inc.

Low Dose of Doxorubicin Potentiates the Effect of Temozolomide in Glioblastoma Cells.

PubMed

Villodre, Emilly Schlee; Kipper, Franciele Cristina; Silva, Andrew Oliveira; Lenz, Guido; Lopez, Patrícia Luciana da Costa

2018-05-01

Glioblastoma (GBM) is an aggressive brain tumor with temozolomide (TMZ)-based chemotherapy as the main therapeutic strategy. Doxorubicin (DOX) is not used in gliomas due to its low bioavailability in the brain; however, new delivery strategies and low doses may be effective in the long term, especially as part of a drug cocktail. Our aim was to evaluate the chronic effects of low doses of DOX and TMZ in GBM. Human U87-ATCC cells and a primary GBM culture were chronically treated with TMZ (5 μM) and DOX (1 and 10 nM) alone or combined. DOX resulted in a reduction in the number of cells over a period of 35 days and delayed the cell regrowth. In addition, DOX induced cell senescence and reduced tumor sphere formation and the proportion of NANOG- and OCT4-positive cells after 7 days. Low doses of TMZ potentiated the effects of DOX on senescence and sphere formation. This combined response using low doses of DOX may pave the way for its use in glioma therapy, with new technologies to overcome its low blood-brain barrier permeability.

Resveratrol induces cellular senescence with attenuated mono-ubiquitination of histone H2B in glioma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gao, Zhen; Xu, Michael S.; Barnett, Tamara L.

2011-04-08

Research highlights: {yields} Resveratrol induces cellular senescence in glioma cell. {yields} Resveratrol inhibits mono-ubiquitination of histone H2B at K120. {yields} Depletion of RNF20, phenocopies the inhibitory effects of resveratrol. {yields} Mono-ubiquitination of histone H2B at K120 is a novel target of resveratrol. {yields} RNF20 inhibits cellular senescence in proliferating glioma cells. -- Abstract: Resveratrol (3,4',5-trihydroxy-trans-stilbene), a polyphenol naturally occurring in grapes and other plants, has cancer chemo-preventive effects and therapeutic potential. Although resveratrol modulates multiple pathways in tumor cells, how resveratrol or its affected pathways converge on chromatin to mediate its effects is not known. Using glioma cells as amore » model, we showed here that resveratrol inhibited cell proliferation and induced cellular hypertrophy by transforming spindle-shaped cells to enlarged, irregular and flatten-shaped ones. We further showed that resveratrol-induced hypertrophic cells expressed senescence-associated-{beta}-galactosidase, suggesting that resveratrol-induced cellular senescence in glioma cells. Consistent with these observations, we demonstrated that resveratrol inhibited clonogenic efficiencies in vitro and tumor growth in a xenograft model. Furthermore, we found that acute treatment of resveratrol inhibited mono-ubiquitination of histone H2B at K120 (uH2B) in breast, prostate, pancreatic, lung, brain tumor cells as well as primary human cells. Chronic treatment with low doses of resveratrol also inhibited uH2B in the resveratrol-induced senescent glioma cells. Moreover, we showed that depletion of RNF20, a ubiquitin ligase of histone H2B, inhibited uH2B and induced cellular senescence in glioma cells in vitro, thereby recapitulated the effects of resveratrol. Taken together, our results suggest that uH2B is a novel direct or indirect chromatin target of resveratrol and RNF20 plays an important role in inhibiting cellular senescence programs that are intact in glioma cells.« less

ERGO: A pilot study of ketogenic diet in recurrent glioblastoma

PubMed Central

RIEGER, JOHANNES; BÄHR, OLIVER; MAURER, GABRIELE D.; HATTINGEN, ELKE; FRANZ, KEA; BRUCKER, DANIEL; WALENTA, STEFAN; KÄMMERER, ULRIKE; COY, JOHANNES F.; WELLER, MICHAEL; STEINBACH, JOACHIM P.

2014-01-01

Limiting dietary carbohydrates inhibits glioma growth in preclinical models. Therefore, the ERGO trial (NCT00575146) examined feasibility of a ketogenic diet in 20 patients with recurrent glioblastoma. Patients were put on a low-carbohydrate, ketogenic diet containing plant oils. Feasibility was the primary endpoint, secondary endpoints included the percentage of patients reaching urinary ketosis, progression-free survival (PFS) and overall survival. The effects of a ketogenic diet alone or in combination with bevacizumab was also explored in an orthotopic U87MG glioblastoma model in nude mice. Three patients (15%) discontinued the diet for poor tolerability. No serious adverse events attributed to the diet were observed. Urine ketosis was achieved at least once in 12 of 13 (92%) evaluable patients. One patient achieved a minor response and two patients had stable disease after 6 weeks. Median PFS of all patients was 5 (range, 3–13) weeks, median survival from enrollment was 32 weeks. The trial allowed to continue the diet beyond progression. Six of 7 (86%) patients treated with bevacizumab and diet experienced an objective response, and median PFS on bevacizumab was 20.1 (range, 12–124) weeks, for a PFS at 6 months of 43%. In the mouse glioma model, ketogenic diet alone had no effect on median survival, but increased that of bevacizumab-treated mice from 52 to 58 days (p<0.05). In conclusion, a ketogenic diet is feasible and safe but probably has no significant clinical activity when used as single agent in recurrent glioma. Further clinical trials are necessary to clarify whether calorie restriction or the combination with other therapeutic modalities, such as radiotherapy or anti-angiogenic treatments, could enhance the efficacy of the ketogenic diet. PMID:24728273

Human Fibronectin Extra-Domain B (EDB)-Specific Aptide (APTEDB) Radiolabelling with Technetium-99m as a Potent Targeted Tumour-Imaging Agent.

PubMed

Mohammadgholi, Mohsen; Sadeghzadeh, Nourollah; Erfani, Mostafa; Abediankenari, Saeid; Abedi, Seyed Mohammad; Emrarian, Iman; Jafari, Narjes; Behzadi, Ramezan

2018-01-01

Human fibronectin extra-domain B (EDB) is particularly expressed during angiogenesis progression. It is, thus, a promising marker of tumour growth. Aptides are a novel class of peptides with high-affinity binding to specific protein targets. APTEDB is an antagonist-like ligand that especially interacts with human fibronectin EDB. This study was the first attempt in which the hydrazinonicotinamide (HYNIC)-conjugated APTEDB was labelled with technetium-99m (99mTc) as an appropriate radiotracer and tricine/EDDA exchange labeling. Radiochemical purity, normal saline, and serum stability were evaluated by HPLC and radio-isotope TLC scanner. Other examinations, such as protein-binding calculation, dissociation radioligand binding assay, and partition coefficient constant determination, were also carried out. The cellular-specific binding of 99mTc- HYNIC-conjugated APTEDB was assessed in two EDB-positive (U87MG) and EDB-negative (U373MG) cell lines. Bio-distribution was investigated in normal mice as well as in U87MG and U373MG tumour-bearing mice. Eventually, the radiolabelled APTEDB was used for tumour imaging using planar SPECT. Radiolabelling was achieved with high purity (up to 97%) and accompanied by high solution (over 90% after overnight) and serum (80% after 2 hours) stability. The obtained cellular-specific binding ratio was greater than nine-fold. In-vivo experiments showed rapid blood clearance with mainly renal excretion and tumour uptake specificity (0.48±0.03% ID/g after 1h). The results of the imaging also confirmed considerable tumour uptake for EDB-positive cell line compared with the EDB-negative one. Aptides are considered to be a potent candidate for biopharmaceutical applications. They can be modified with imaging or therapeutic agents. This report shows the capability of 99mTc-HYNIC-APTEDB for human EDB-expressing tumours detection. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Time- and spectrally resolved characteristics of flavin fluorescence in U87MG cancer cells in culture

NASA Astrophysics Data System (ADS)

Horilova, Julia; Cunderlikova, Beata; Marcek Chorvatova, Alzbeta

2015-05-01

Early detection of cancer is crucial for the successful diagnostics of its presence and its subsequent treatment. To improve cancer detection, we tested the progressive multimodal optical imaging of U87MG cells in culture. A combination of steady-state spectroscopic methods with the time-resolved approach provides a new insight into the native metabolism when focused on endogenous tissue fluorescence. In this contribution, we evaluated the metabolic state of living U87MG cancer cells in culture by means of endogenous flavin fluorescence. Confocal microscopy and time-resolved fluorescence imaging were employed to gather spectrally and time-resolved images of the flavin fluorescence. We observed that flavin fluorescence in U87MG cells was predominantly localized outside the cell nucleus in mitochondria, while exhibiting a spectral maximum under 500 nm and fluorescence lifetimes under 1.4 ns, suggesting the presence of bound flavins. In some cells, flavin fluorescence was also detected inside the cell nuclei in the nucleoli, exhibiting longer fluorescence lifetimes and a red-shifted spectral maximum, pointing to the presence of free flavin. Extra-nuclear flavin fluorescence was diminished by 2-deoxyglucose, but failed to increase with 2,4-dinitrophenol, the uncoupler of oxidative phosphorylation, indicating that the cells use glycolysis, rather than oxidative phosphorylation for functioning. These gathered data are the first step toward monitoring the metabolic state of U87MG cancer cells.

Convection-enhanced delivery of a topoisomerase I inhibitor (nanoliposomal topotecan) and a topoisomerase II inhibitor (pegylated liposomal doxorubicin) in intracranial brain tumor xenografts1

PubMed Central

Yamashita, Yoji; Krauze, Michal T.; Kawaguchi, Tomohiro; Noble, Charles O.; Drummond, Daryl C.; Park, John W.; Bankiewicz, Krystof S.

2007-01-01

Despite multimodal treatment options, the response and survival rates for patients with malignant gliomas remain dismal. Clinical trials with convection-enhanced delivery (CED) have recently opened a new window in neuro-oncology to the direct delivery of chemotherapeutics to the CNS, circumventing the blood-brain barrier and reducing systemic side effects. Our previous CED studies with liposomal chemotherapeutics have shown promising antitumor activity in rodent brain tumor models. In this study, we evaluated a combination of nanoliposomal topotecan (nLs-TPT) and pegylated liposomal doxorubicin (PLD) to enhance efficacy in our brain tumor models, and to establish a CED treatment capable of improving survival from malignant brain tumors. Both liposomal drugs decreased key enzymes involved in tumor cell replication in vitro. Synergistic effects of nLs-TPT and PLD on U87MG cell death were found. The combination displayed excellent efficacy in a CED-based survival study 10 days after tumor cell implantation. Animals in the control group and those in single-agent groups had a median survival of less than 30 days, whereas the combination group experienced a median survival of more than 90 days. We conclude that CED of two liposomal chemotherapeutics (nLs-TPT and PLD) may be an effective treatment option for malignant gliomas. PMID:17018695

miR-204 reverses temozolomide resistance and inhibits cancer initiating cells phenotypes by degrading FAP-α in glioblastoma.

PubMed

Yang, Yun-Na; Zhang, Xiang-Hua; Wang, Yan-Ming; Zhang, Xi; Gu, Zheng

2018-05-01

Malignant gliomas are treated with temozolomide (TMZ) at present, but often exhibit resistance to this agent. Cancer-initiating cells (CICs) have been suggested to lead to TMZ resistance. The mechanisms underlying CICs-based TMZ resistance are not fully understood. MicroRNAs (miRNAs) have been demonstrated to serve important roles in tumorigenesis and TMZ resistance. In the present study, a sphere forming assay and western blot analysis were performed to detect the formation of CICs and fibroblast activation protein α (FAP-α) protein expression. It was revealed that TMZ resistance promoted the formation of CICs and upregulated FAP-α expression in glioblastoma cells. Over-expressing FAP-α was also demonstrated to promote TMZ resistance and induce the formation of CICs in U251MG cells. In addition, using a reverse transcription-quantitative polymerase chain reaction, it was observed that miR-204 was downregulated in U251MG-resistant (-R) cells. miR-204 expression negatively correlated with the FAP-α levels in human glioblastoma tissues, and it may inhibit the formation of CICs and reverse TMZ resistance in U251MG-R cells. Therefore, it was concluded that miR-204 reversed temozolomide resistance and inhibited CICs phenotypes by degrading FAP-α in glioblastoma.

miR-204 reverses temozolomide resistance and inhibits cancer initiating cells phenotypes by degrading FAP-α in glioblastoma

PubMed Central

Yang, Yun-Na; Zhang, Xiang-Hua; Wang, Yan-Ming; Zhang, Xi; Gu, Zheng

2018-01-01

Malignant gliomas are treated with temozolomide (TMZ) at present, but often exhibit resistance to this agent. Cancer-initiating cells (CICs) have been suggested to lead to TMZ resistance. The mechanisms underlying CICs-based TMZ resistance are not fully understood. MicroRNAs (miRNAs) have been demonstrated to serve important roles in tumorigenesis and TMZ resistance. In the present study, a sphere forming assay and western blot analysis were performed to detect the formation of CICs and fibroblast activation protein α (FAP-α) protein expression. It was revealed that TMZ resistance promoted the formation of CICs and upregulated FAP-α expression in glioblastoma cells. Over-expressing FAP-α was also demonstrated to promote TMZ resistance and induce the formation of CICs in U251MG cells. In addition, using a reverse transcription-quantitative polymerase chain reaction, it was observed that miR-204 was downregulated in U251MG-resistant (-R) cells. miR-204 expression negatively correlated with the FAP-α levels in human glioblastoma tissues, and it may inhibit the formation of CICs and reverse TMZ resistance in U251MG-R cells. Therefore, it was concluded that miR-204 reversed temozolomide resistance and inhibited CICs phenotypes by degrading FAP-α in glioblastoma. PMID:29725461

The effects of CD147 on the cell proliferation, apoptosis, invasion, and angiogenesis in glioma.

PubMed

Yin, Haoyuan; Shao, Ying; Chen, Xuan

2017-01-01

To analyze the effects of extracellular matrix metalloproteinase inducer (CD147) on glioma proliferation, apoptosis, invasion, and angiogenesis. Tissue samples were obtained from 101 glioma cases while normal brain tissues were obtained from 30 brain injury cases. Immunohistochemical assay was performed to detect the expressions of CD147, CD34, and VEGF in tissue samples. QRT-PCR was performed to detect the relative expression of CD147 mRNA in human glioma cell lines. CD147 siRNA was transfected into glioma cell line U251. Cell proliferation, apoptosis, invasion, and angiogenesis were tested by MTT, flow cytometry, Transwell assay, and vasculogenic mimicry assay, respectively. Expressions of relative proteins were analyzed with western blot. CD147 was positively expressed with the percentage of 0, 37.5, 44.8, 67.9, and 85.7 % in normal tissues and glioma tissues with WHO grades I-IV, respectively, and the scores of MVDand VEGF were associated with the expression of CD147. CD147 was significantly upregulated in the human glioma cell lines (P < 0.05). Downregulated the expression of CD147 suppressed cell proliferation, blocked cell cycle, induced apoptosis, inhibited cell invasion and angiogenesis in glioma cells in vitro. The expression of CD147 was significantly associated with WHO tumor grade and angiogenesis; silencing of CD147 contributed to inhibition of glioma proliferation, invasion, and angiogenesis. Our study provided firm evidence that CD 147 is a potential glioma target for anti-angiogenic therapies.

DW10075, a novel selective and small-molecule inhibitor of VEGFR, exhibits antitumor activities both in vitro and in vivo

PubMed Central

Li, Meng-yuan; Lv, Yong-cong; Tong, Lin-jiang; Peng, Ting; Qu, Rong; Zhang, Tao; Sun, Yi-ming; Chen, Yi; Wei, Li-xin; Geng, Mei-yu; Duan, Wen-hu; Xie, Hua; Ding, Jian

2016-01-01

Aim: Targeting the VEGF/VEGF receptor (VEGFR) pathway has proved to be an effective antiangiogenic approach for cancer treatment. Here, we identified 6-((2-((3-acetamidophenyl)amino)pyrimidin-4-yl)oxy)-N-phenyl-1-naphthamide (designated herein as DW10075) as a novel and highly selective inhibitor of VEGFRs. Methods: In vitro tyrosine kinase activity was measured using ELISA, and intracellular signaling pathway proteins were detected by Western blot analysis. Endothelial cell proliferation was examined with CCK-8 assays, and tumor cell proliferation was determined with SRB assays. Cell migration, tube formation and rat aortic ring assays were used to detect antiangiogenic activity. Antitumor efficacy was further evaluated in U87-MG human glioblastoma xenograft tumors in nude mice receiving DW10075 (500 mg·kg−1·d−1, po) for two weeks. Results: Among a panel of 21 kinases tested, DW10075 selectively inhibited VEGFR-1, VEGFR-2 and VEGFR-3 (the IC50 values were 6.4, 0.69 and 5.5 nmol/L, respectively), but did not affect 18 other kinases including FGFR and PDGFR at 10 μmol/L. DW10075 significantly blocked VEGF-induced activation of VEGFR and its downstream signaling transduction in primary human umbilical vein endothelial cells (HUVECs), thus inhibited VEGF-induced HUVEC proliferation. DW10075 (1–100 nmol/L) dose-dependently inhibited VEGF-induced HUVEC migration and tube formation and suppressed angiogenesis in both the rat aortic ring model and the chicken chorioallantoic membrane model. Furthermore, DW10075 exhibited anti-proliferative activity against 22 different human cancer cell lines with IC50 values ranging from 2.2 μmol/L (for U87-MG human glioblastoma cells) to 22.2 μmol/L (for A375 melanoma cells). In U87-MG xenograft tumors in nude mice, oral administration of DW10075 significantly suppressed tumor growth, and reduced the expression of CD31 and Ki67 in the tumor tissues. Conclusion: DW10075 is a potent and highly selective inhibitor of VEGFR that deserves further development. PMID:26806300

Dynamic glucose enhanced (DGE) MRI for combined imaging of blood-brain barrier break down and increased blood volume in brain cancer.

PubMed

Xu, Xiang; Chan, Kannie W Y; Knutsson, Linda; Artemov, Dmitri; Xu, Jiadi; Liu, Guanshu; Kato, Yoshinori; Lal, Bachchu; Laterra, John; McMahon, Michael T; van Zijl, Peter C M

2015-12-01

Recently, natural d-glucose was suggested as a potential biodegradable contrast agent. The feasibility of using d-glucose for dynamic perfusion imaging was explored to detect malignant brain tumors based on blood brain barrier breakdown. Mice were inoculated orthotopically with human U87-EGFRvIII glioma cells. Time-resolved glucose signal changes were detected using chemical exchange saturation transfer (glucoCEST) MRI. Dynamic glucose enhanced (DGE) MRI was used to measure tissue response to an intravenous bolus of d-glucose. DGE images of mouse brains bearing human glioma showed two times higher and persistent changes in tumor compared with contralateral brain. Area-under-curve (AUC) analysis of DGE delineated blood vessels and tumor and had contrast comparable to the AUC determined using dynamic contrast enhanced (DCE) MRI with GdDTPA, both showing a significantly higher AUC in tumor than in brain (P < 0.005). Both CEST and relaxation effects contribute to the signal change. DGE MRI is a feasible technique for studying brain tumor enhancement reflecting differences in tumor blood volume and permeability with respect to normal brain. We expect DGE will provide a low-risk and less expensive alternative to DCE MRI for imaging cancer in vulnerable populations, such as children and patients with renal impairment. © 2015 Wiley Periodicals, Inc.

Dynamic Glucose Enhanced (DGE) MRI for Combined Imaging of Blood Brain Barrier Break Down and Increased Blood Volume in Brain Cancer

PubMed Central

Xu, Xiang; Chan, Kannie WY; Knutsson, Linda; Artemov, Dmitri; Xu, Jiadi; Liu, Guanshu; Kato, Yoshinori; Lal, Bachchu; Laterra, John; McMahon, Michael T.; van Zijl, Peter C.M.

2015-01-01

Purpose Recently, natural d-glucose was suggested as a potential biodegradable contrast agent. The feasibility of using d-glucose for dynamic perfusion imaging was explored to detect malignant brain tumors based on blood brain barrier breakdown. Methods Mice were inoculated orthotopically with human U87-EGFRvIII glioma cells. Time-resolved glucose signal changes were detected using chemical exchange saturation transfer (glucoCEST) MRI. Dynamic glucose enhanced (DGE) MRI was used to measure tissue response to an intravenous bolus of d-glucose. Results DGE images of mouse brains bearing human glioma showed two times higher and persistent changes in tumor compared to contralateral brain. Area-under-curve (AUC) analysis of DGE delineated blood vessels and tumor and had contrast comparable to the AUC determined using dynamic contrast enhanced (DCE) MRI with GdDTPA, both showing a significantly higher AUC in tumor than in brain (p<0.005). Both CEST and relaxation effects contribute to the signal change. Conclusion DGE MRI is a feasible technique for studying brain tumor enhancement reflecting differences in tumor blood volume and permeability with respect to normal brain. We expect DGE will provide a low-risk and less expensive alternative to DCE MRI for imaging cancer in vulnerable populations, such as children and patients with renal impairment. PMID:26404120

The inhibitory effect of CIL-102 on the growth of human astrocytoma cells is mediated by the generation of reactive oxygen species and induction of ERK1/2 MAPK

DOE Office of Scientific and Technical Information (OSTI.GOV)

Teng, Chih-Chuan; Institute of Basic Medicine Science, National Cheng Kung University, Tainan, Taiwan; Kuo, Hsing-Chun

2012-08-15

CIL-102 (1-[4-(furo[2,3-b]quinolin-4-ylamino)phenyl]ethanone) is the major active agent of the alkaloid derivative of Camptotheca acuminata, with multiple pharmacological activities, including anticancer effects and promotion of apoptosis. The mechanism by which CIL-102 inhibits growth remains poorly understood in human astrocytoma cells. Herein, we investigated the molecular mechanisms by which CIL-102 affects the generation of reactive oxygen species (ROS) and cell cycle G2/M arrest in glioma cells. Treatment of U87 cells with 1.0 μM CIL-102 resulted in phosphorylation of extracellular signal-related kinase (ERK1/2), downregulation of cell cycle-related proteins (cyclin A, cyclin B, cyclin D1, and cdk1), and phosphorylation of cdk1Tyr{sup 15} and Cdc25cSer{supmore » 216}. Furthermore, treatment with the ERK1/2 inhibitor PD98059 abolished CIL-102-induced Cdc25cSer{sup 216} expression and reversed CIL-102-inhibited cdk1 activation. In addition, N-acetyl cysteine (NAC), an ROS scavenger, blocked cell cycle G2/M arrest and phosphorylation of ERK1/2 and Cdc25cSer{sup 216} in U87 cells. CIL-102-mediated ERK1/2 and ROS production, and cell cycle arrest were blocked by treatment with specific inhibitors. In conclusion, we have identified a novel CIL-102-inhibited proliferation in U87 cells by activating the ERK1/2 and Cdc25cSer{sup 216} cell cycle-related proteins and inducing ROS production; this might be a new mechanism in human astrocytoma cells. -- Highlights: ► We show the effects of CIL-102 on the G2/M arrest of human astrocytoma cells. ► ROS and the Ras/ERK1/2 triggering pathways are involved in the CIL-102 treatment. ► CIL-102 induces sustained activation of ERK1/2 and Cdc25c and ROS are required.« less

NOX4-mediated ROS production induces apoptotic cell death via down-regulation of c-FLIP and Mcl-1 expression in combined treatment with thioridazine and curcumin.

PubMed

Seo, Seung Un; Kim, Tae Hwan; Kim, Dong Eun; Min, Kyoung-Jin; Kwon, Taeg Kyu

2017-10-01

Thioridazine is known to have anti-tumor effects by inhibiting PI3K/Akt signaling, which is an important signaling pathway in cell survival. However, thioridazine alone does not induce apoptosis in head and neck squamous cell carcinoma (AMC-HN4), human breast carcinoma (MDA-MB231), and human glioma (U87MG) cells. Therefore, we investigated whether combined treatment with thioridazine and curcumin induces apoptosis. Combined treatment with thioridazine and curcumin markedly induced apoptosis in cancer cells without inducing apoptosis in human normal mesangial cells and human normal umbilical vein cells (EA.hy926). We found that combined treatment with thioridazine and curcumin had synergistic effects in AMC-HN4 cells. Among apoptosis-related proteins, thioridazine plus curcumin induced down-regulation of c-FLIP and Mcl-1 expression at the post-translational levels in a proteasome-dependent manner. Augmentation of proteasome activity was related to the up-regulation of proteasome subunit alpha 5 (PSMA5) expression in curcumin plus thioridazine-treated cells. Combined treatment with curcumin and thioridazine produced intracellular ROS in a NOX4-dependent manner, and ROS-mediated activation of Nrf2/ARE signaling played a critical role in the up-regulation of PSMA5 expression. Furthermore, ectopic expression of c-FLIP and Mcl-1 inhibited apoptosis in thioridazine and curcumin-treated cells. Therefore, we demonstrated that thioridazine plus curcumin induces proteasome activity by up-regulating PSMA5 expression via NOX4-mediated ROS production and that down-regulation of c-FLIP and Mcl-1 expression post-translationally is involved in apoptosis. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Antitumor efficacy of PKI-587, a highly potent dual PI3K/mTOR kinase inhibitor.

PubMed

Mallon, Robert; Feldberg, Larry R; Lucas, Judy; Chaudhary, Inder; Dehnhardt, Christoph; Santos, Efren Delos; Chen, Zecheng; dos Santos, Osvaldo; Ayral-Kaloustian, Semiramis; Venkatesan, Aranapakam; Hollander, Irwin

2011-05-15

The aim of this study was to show preclinical efficacy and clinical development potential of PKI-587, a dual phosphoinositide 3-kinase (PI3K)/mTOR inhibitor. In vitro class 1 PI3K enzyme and human tumor cell growth inhibition assays and in vivo five tumor xenograft models were used to show efficacy. In vitro, PKI-587 potently inhibited class I PI3Ks (IC(50) vs. PI3K-α = 0.4 nmol/L), PI3K-α mutants, and mTOR. PKI-587 inhibited growth of 50 diverse human tumor cell lines at IC(50) values of less than 100 nmol/L. PKI-587 suppressed phosphorylation of PI3K/mTOR effectors (e.g., Akt), and induced apoptosis in human tumor cell lines with elevated PI3K/mTOR signaling. MDA-MB-361 [breast; HER2(+), PIK3CA mutant (E545K)] was particularly sensitive to this effect, with cleaved PARP, an apoptosis marker, induced by 30 nmol/L PKI-587 at 4 hours. In vivo, PKI-587 inhibited tumor growth in breast (MDA-MB-361, BT474), colon (HCT116), lung (H1975), and glioma (U87MG) xenograft models. In MDA-MB-361 tumors, PKI-587 (25 mg/kg, single dose i.v.) suppressed Akt phosphorylation [at threonine(T)308 and serine(S)473] for up to 36 hours, with cleaved PARP (cPARP) evident up to 18 hours. PKI-587 at 25 mg/kg (once weekly) shrank large (∼1,000 mm(3)) MDA-MB-361 tumors and suppressed tumor regrowth. Tumor regression correlated with suppression of phosphorylated Akt in the MDA-MB-361 model. PKI-587 also caused regression in other tumor models, and efficacy was enhanced when given in combination with PD0325901 (MEK 1/2 inhibitor), irinotecan (topoisomerase I inhibitor), or HKI-272 (neratinib, HER2 inhibitor). Significant antitumor efficacy and a favorable pharmacokinetic/safety profile justified phase 1 clinical evaluation of PKI-587. ©2011 AACR.

Sustained Radiosensitization of Hypoxic Glioma Cells after Oxygen Pretreatment in an Animal Model of Glioblastoma and In Vitro Models of Tumor Hypoxia

PubMed Central

Clarke, Ryon H.; Moosa, Shayan; Anzivino, Matthew; Wang, Yi; Floyd, Desiree Hunt; Purow, Benjamin W.; Lee, Kevin S.

2014-01-01

Glioblastoma multiforme (GBM) is the most common and lethal form of brain cancer and these tumors are highly resistant to chemo- and radiotherapy. Radioresistance is thought to result from a paucity of molecular oxygen in hypoxic tumor regions, resulting in reduced DNA damage and enhanced cellular defense mechanisms. Efforts to counteract tumor hypoxia during radiotherapy are limited by an attendant increase in the sensitivity of healthy brain tissue to radiation. However, the presence of heightened levels of molecular oxygen during radiotherapy, while conventionally deemed critical for adjuvant oxygen therapy to sensitize hypoxic tumor tissue, might not actually be necessary. We evaluated the concept that pre-treating tumor tissue by transiently elevating tissue oxygenation prior to radiation exposure could increase the efficacy of radiotherapy, even when radiotherapy is administered after the return of tumor tissue oxygen to hypoxic baseline levels. Using nude mice bearing intracranial U87-luciferase xenografts, and in vitro models of tumor hypoxia, the efficacy of oxygen pretreatment for producing radiosensitization was tested. Oxygen-induced radiosensitization of tumor tissue was observed in GBM xenografts, as seen by suppression of tumor growth and increased survival. Additionally, rodent and human glioma cells, and human glioma stem cells, exhibited prolonged enhanced vulnerability to radiation after oxygen pretreatment in vitro, even when radiation was delivered under hypoxic conditions. Over-expression of HIF-1α reduced this radiosensitization, indicating that this effect is mediated, in part, via a change in HIF-1-dependent mechanisms. Importantly, an identical duration of transient hyperoxic exposure does not sensitize normal human astrocytes to radiation in vitro. Taken together, these results indicate that briefly pre-treating tumors with elevated levels of oxygen prior to radiotherapy may represent a means for selectively targeting radiation-resistant hypoxic cancer cells, and could serve as a safe and effective adjuvant to radiation therapy for patients with GBM. PMID:25350400

The Fanconi anemia (FA) pathway confers glioma resistance to DNA alkylating agents.

PubMed

Chen, Clark C; Taniguchi, Toshiyasu; D'Andrea, Alan

2007-05-01

DNA alkylating agents including temozolomide (TMZ) and 1,3-bis[2-chloroethyl]-1-nitroso-urea (BCNU) are the most common form of chemotherapy in the treatment of gliomas. Despite their frequent use, the therapeutic efficacy of these agents is limited by the development of resistance. Previous studies suggest that the mechanism of this resistance is complex and involves multiple DNA repair pathways. To better define the pathways contributing to the mechanisms underlying glioma resistance, we tested the contribution of the Fanconi anemia (FA) DNA repair pathway. TMZ and BCNU treatment of FA-proficient cell lines led to a dose- and time-dependent increase in FANCD2 mono-ubiquitination and FANCD2 nuclear foci formation, both hallmarks of FA pathway activation. The FA-deficient cells were more sensitive to TMZ/BCNU relative to their corrected, isogenic counterparts. To test whether these observations were pertinent to glioma biology, we screened a panel of glioma cell lines and identified one (HT16) that was deficient in the FA repair pathway. This cell line exhibited increased sensitivity to TMZ and BCNU relative to the FA-proficient glioma cell lines. Moreover, inhibition of FA pathway activation by a small molecule inhibitor (curcumin) or by small interference RNA suppression caused increased sensitivity to TMZ/BCNU in the U87 glioma cell line. The BCNU sensitizing effect of FA inhibition appeared additive to that of methyl-guanine methyl transferase inhibition. The results presented in this paper underscore the complexity of cellular resistance to DNA alkylating agents and implicate the FA repair pathway as a determinant of this resistance.

Oleanane-type saponins from Anemone taipaiensis and their cytotoxic activities.

PubMed

Wang, Xiaoyang; Zhang, Wei; Gao, Kai; Lu, Yunyang; Tang, Haifeng; Sun, Xiaoli

2013-09-01

Phytochemical investigation of the n-BuOH extract of the rhizomes of Anemone taipaiensis led to the isolation of three new oleanane-type triterpenoid saponins (1-3), together with four known saponins (4-7). Their structures were elucidated on the basis of spectroscopic analysis and chemical derivatization. All the compounds were isolated for the first time from A. taipaiensis. The cytotoxicity of these compounds was evaluated in five human cancer cell lines including A549 (lung carcinoma), HeLa (cervical carcinoma), HepG2 (hepatocellular carcinoma), HL-60 (promyelocytic leukemia), and U87MG (glioblastoma). The monodesmosidic saponin 4 exhibited cytotoxic activity toward all cancer cell lines, with IC50 values ranging from 6.42 to 18.16 μM. In addition, the bisdesmosidic saponins 1 and 7 showed selective cytotoxicity against the U87MG cells. Copyright © 2013 Elsevier B.V. All rights reserved.

Asiatic acid induces endoplasmic reticulum stress and apoptotic death in glioblastoma multiforme cells both in vitro and in vivo

PubMed Central

Kavitha, Chandagirikoppal V.; Jain, Anil K.; Agarwal, Chapla; Pierce, Angela; Keating, Amy; Huber, Kendra M.; Serkova, Natalie J.; Wempe, Michael F.; Agarwal, Rajesh; Deep, Gagan

2014-01-01

Glioblastoma multiforme (GBM) is an untreatable malignancy. Existing therapeutic options are insufficient, and adversely affect functional and non-cancerous cells in the brain impairing different functions of the body. Therefore, there is an urgent need for additional preventive and therapeutic non-toxic drugs against GBM. Asiatic acid (AsA; 2,3,23-trihydroxy-12-ursen-28-oic acid, C30H48O5) is a natural small molecule widely used to treat various neurological disorders, and the present research investigates AsA’s efficacy against GBM both in vitro and in vivo. Results showed that AsA treatment (10–100 μM) decreased the human GBM cell (LN18, U87MG, and U118MG) viability, with better efficacy than temozolomide at equimolar doses. Orally administered AsA (30 mg/kg/day) strongly decreased tumor volume in mice when administered immediately after ectopic U87MG xenograft implantation (54% decrease, p≤0.05) or in mice with established xenografts (48% decrease, p≤0.05) without any apparent toxicity. Importantly, AsA feeding (30 mg/kg/twice a day) also decreased the orthotopic U87MG xenografts growth in nude mice as measured by magnetic resonance imaging. Using LC/MS-MS methods, AsA was detected in mice plasma and brain tissue, confirming that AsA crosses blood-brain barrier. Mechanistic studies showed that AsA induces apoptotic death by modulating the protein expression of several apoptosis regulators (caspases, Bcl2 family members, and survivin) in GBM cells. Furthermore, AsA induced ER stress (increased GRP78 and Calpain, and decreased Calnexin and IRE1α expression), enhanced free intra-cellular calcium, and damaged cellular organization in GBM cells. These experimental results demonstrate that AsA is effective against GBM, and advocate further pre-clinical and clinical evaluations of AsA against GBM. PMID:25252179

Tumor-associated macrophages are predominant carriers of cyclodextrin-based nanoparticles into gliomas.

PubMed

Alizadeh, Darya; Zhang, Leying; Hwang, Jungyeon; Schluep, Thomas; Badie, Behnam

2010-04-01

The goal of this study was to evaluate the mechanism of cyclodextrin-based nanoparticle (CDP-NP) uptake into a murine glioma model. Using mixed in vitro culture systems, we demonstrated that CDP-NPs were preferentially taken up by BV2 and N9 microglia (MG) cells compared with GL261 glioma cells. Fluorescent microscopy and flow cytometry analysis of intracranial GL261 gliomas confirmed these findings and demonstrated a predominant CDP-NP uptake by macrophages (MPs) and MG within and around the tumor site. Notably, in mice bearing bilateral intracranial tumor, MG and MPs carrying CDP-NPs were able to migrate to the contralateral tumors. In conclusion, these studies better characterize the cellular distribution of CDP-NPs in intracranial tumors and demonstrate that MPs and MG could potentially be used as nanoparticle drug carriers into malignant brain tumors. The goal of this study was to evaluate the mechanism of cyclodextrin-based nanoparticle (CDP-NP) uptake into a murine glioma model. CDP-NP was preferentially taken up microglia (MG) cells as compared to glioma cells. A predominant CDP-NP uptake by macrophages and MG was also shown in and around the tumor site. Macrophages and MG could potentially be used as nanoparticle drug carriers into malignant brain tumors. Copyright 2010 Elsevier Inc. All rights reserved.

Overexpression of Large-Conductance Calcium-Activated Potassium Channels in Human Glioblastoma Stem-Like Cells and Their Role in Cell Migration.

PubMed

Rosa, Paolo; Sforna, Luigi; Carlomagno, Silvia; Mangino, Giorgio; Miscusi, Massimo; Pessia, Mauro; Franciolini, Fabio; Calogero, Antonella; Catacuzzeno, Luigi

2017-09-01

Glioblastomas (GBMs) are brain tumors characterized by diffuse invasion of cancer cells into the healthy brain parenchyma, and establishment of secondary foci. GBM cells abundantly express large-conductance, calcium-activated potassium (BK) channels that are thought to promote cell invasion. Recent evidence suggests that the GBM high invasive potential mainly originates from a pool of stem-like cells, but the expression and function of BK channels in this cell subpopulation have not been studied. We investigated the expression of BK channels in GBM stem-like cells using electrophysiological and immunochemical techniques, and assessed their involvement in the migratory process of this important cell subpopulation. In U87-MG cells, BK channel expression and function were markedly upregulated by growth conditions that enriched the culture in GBM stem-like cells (U87-NS). Cytofluorimetric analysis further confirmed the appearance of a cell subpopulation that co-expressed high levels of BK channels and CD133, as well as other stem cell markers. A similar association was also found in cells derived from freshly resected GBM biopsies. Finally, transwell migration tests showed that U87-NS cells migration was much more sensitive to BK channel block than U87-MG cells. Our data show that BK channels are highly expressed in GBM stem-like cells, and participate to their high migratory activity. J. Cell. Physiol. 232: 2478-2488, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Alkaloid extracts of Ficus species and palm oil-derived tocotrienols synergistically inhibit proliferation of human cancer cells.

PubMed

Abubakar, Ibrahim Babangida; Lim, Kuan-Hon; Loh, Hwei-San

2015-01-01

Tocotrienols have been reported to possess anticancer effects other than anti-inflammatory and antioxidant activities. This study explored the potential synergism of antiproliferative effects induced by individual alkaloid extracts of Ficus fistulosa, Ficus hispida and Ficus schwarzii combined with δ- and γ-tocotrienols against human brain glioblastoma (U87MG), lung adenocarcinoma (A549) and colorectal adenocarcinoma (HT-29) cells. Cell viability and morphological results demonstrated that extracts containing a mixture of alkaloids from the leaves and bark of F. schwarzii inhibited the proliferation of HT-29 cells, whereas the alkaloid extracts of F. fistulosa inhibited the proliferation of both U87MG and HT-29 cells and showed synergism in combined treatments with either δ- or γ-tocotrienol resulting in 2.2-34.7 fold of reduction in IC50 values of tocotrienols. The observed apoptotic cell characteristics in conjunction with the synergistic antiproliferative effects of Ficus species-derived alkaloids and tocotrienols assuredly warrant future investigations towards the development of a value-added chemotherapeutic regimen against cancers.

Zn{sup 2+} induces apoptosis in human highly metastatic SHG-44 glioma cells, through inhibiting activity of the voltage-gated proton channel Hv1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Yifan; Zhang, Shangrong; Li, Shu Jie, E-mail: shujieli@nankai.edu.cn

Highlights: •Hv1 is expressed in highly metastatic glioma cell. •Zn{sup 2+} ions induces apoptosis in highly metastatic glioma cells. •Zn{sup 2+} ions markedly inhibit proton secretion. •Zn{sup 2+} ions reduce the gelatinase activity. •Inhibition of Hv1 activity via Zn{sup 2+} ions can effectively retard the cancer growth. -- Abstract: In contrast to the voltage-gated K{sup +} channels, the voltage-gated proton channel Hv1 contains a voltage-sensor domain but lacks a pore domain. Here, we showed that Hv1 is expressed in the highly metastatic glioma cell SHG-44, but lowly in the poorly metastatic glioma cell U-251. Inhibition of Hv1 activity by 140more » μM zinc chloride induces apoptosis in the human highly metastatic glioma cells. Zn{sup 2+} ions markedly inhibit proton secretion, and reduce the gelatinase activity in the highly metastatic glioma cells. In vivo, the glioma tumor sizes of the implantation of the SHG-44 xenografts in nude mice that were injected zinc chloride solution, were dramatically smaller than that in the controlled groups. The results demonstrated that the inhibition of Hv1 activity via Zn{sup 2+} ions can effectively retard the cancer growth and suppress the cancer metastasis by the decrease of proton extrusion and the down-regulation of gelatinase activity. Our results suggest that Zn{sup 2+} ions may be used as a potential anti-glioma drug for glioma therapy.« less

Adult, embryonic and fetal hemoglobin are expressed in human glioblastoma cells.

PubMed

Emara, Marwan; Turner, A Robert; Allalunis-Turner, Joan

2014-02-01

Hemoglobin is a hemoprotein, produced mainly in erythrocytes circulating in the blood. However, non-erythroid hemoglobins have been previously reported in other cell types including human and rodent neurons of embryonic and adult brain, but not astrocytes and oligodendrocytes. Human glioblastoma multiforme (GBM) is the most aggressive tumor among gliomas. However, despite extensive basic and clinical research studies on GBM cells, little is known about glial defence mechanisms that allow these cells to survive and resist various types of treatment. We have shown previously that the newest members of vertebrate globin family, neuroglobin (Ngb) and cytoglobin (Cygb), are expressed in human GBM cells. In this study, we sought to determine whether hemoglobin is also expressed in GBM cells. Conventional RT-PCR, DNA sequencing, western blot analysis, mass spectrometry and fluorescence microscopy were used to investigate globin expression in GBM cell lines (M006x, M059J, M059K, M010b, U87R and U87T) that have unique characteristics in terms of tumor invasion and response to radiotherapy and hypoxia. The data showed that α, β, γ, δ, ζ and ε globins are expressed in all tested GBM cell lines. To our knowledge, we are the first to report expression of fetal, embryonic and adult hemoglobin in GBM cells under normal physiological conditions that may suggest an undefined function of those expressed hemoglobins. Together with our previous reports on globins (Ngb and Cygb) expression in GBM cells, the expression of different hemoglobins may constitute a part of series of active defence mechanisms supporting these cells to resist various types of treatments including chemotherapy and radiotherapy.

Tangeretin induces cell cycle arrest and apoptosis through upregulation of PTEN expression in glioma cells.

PubMed

Ma, Li-Li; Wang, Da-Wei; Yu, Xu-Dong; Zhou, Yan-Ling

2016-07-01

Tangeretin (TANG), present in peel of citrus fruits, has been shown to various medicinal properties such as chemopreventive and neuroprotective. However, the chemopreventive effect of TANG on glioblastoma cells has not been examined. The present study was designed to explore the anticancer potential of TANG in glioblastoma cells and to investigate the related mechanism. Human glioblastoma U-87MG and LN-18 cells were treated with 45μM concentration of TANG and cell growth was measured by MTT assay. The cell cycle distribution and cell death were measured by flow cytometry. The expression of cell cycle and apoptosis related genes were analyzed by quantitative RT-PCR and western blot. The cells treated with TANG were significantly increased cell growth suppression and cell death effects than vehicle treated cells. Further, TANG treatment increases G2/M arrest and apoptosis by modulating PTEN and cell-cycle regulated genes such as cyclin-D and cdc-2 mRNA and protein expressions. Moreover, the ability of TANG to decrease cell growth and to induce cell death was compromised when PTEN was knockdown by siRNA. Taken together, the chemopreventive effect of TANG is associated with regulation of cell-cycle and apoptosis in glioblastoma, thereby attenuating glioblastoma cell growth. Hence, the present findings suggest that TANG may be a therapeutic agent for glioblastoma treatment. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Early versus late GD-DTPA MRI enhancement in experimental glioblastomas.

PubMed

Farace, Paolo; Tambalo, Stefano; Fiorini, Silvia; Merigo, Flavia; Daducci, Alessandro; Nicolato, Elena; Conti, Giamaica; Degrassi, Anna; Sbarbati, Andrea; Marzola, Pasquina

2011-03-01

To compare early versus late enhancement in two glioblastoma models characterized by different infiltrative/edematous patterns. Three weeks after inoculation into nude mice of U87MG and U251 cells, T1-weighted images were acquired early (10.5 min), intermediate (21 min) and late (30.5 min) after a bolus injection of Gd-DTPA at 300 μ mol/kg dosage. EARLY(TH) and LATE(TH) were the corresponding volumes with an enhancement higher than a threshold TH, defined by the mean (μ) and standard deviation (σ) on a contralateral healthy area. ADD(TH) was the enhancing volume found in LATE(TH) but not in EARLY(TH). T2 imaging of both tumors was performed, and T2 mapping of U251. In all tumors, LATE(TH) was significantly higher than EARLY(TH) for TH ranging from μ+σ to μ+5σ. The ADD(TH) /EARLY(TH) ratio was not significantly different when U251 and U87MG tumors were compared. In the U87MG tumors, some enhancement was observed outside the regularly demarcated T2-hyperintense area. In the U251 tumors, irregularly T2 demarcated, a large portion of ADD(μ+3σ) had normal T2 values. At histology, U251 showed a higher infiltrative pattern than U87MG. In these models, the increase over time in the enhancing volume did not depend on the different infiltrative/edematous patterns and was not closely related with edema. Copyright © 2011 Wiley-Liss, Inc.

Inhibition of CYP4A by a novel flavonoid FLA-16 prolongs survival and normalizes tumor vasculature in glioma.

PubMed

Wang, Chenlong; Li, Ying; Chen, Honglei; Zhang, Jie; Zhang, Jing; Qin, Tian; Duan, Chenfan; Chen, Xuewei; Liu, Yanzhuo; Zhou, Xiaoyang; Yang, Jing

2017-08-28

Glioblastomas rapidly become refractory to anti-VEGF therapies. We previously showed that cytochrome P450 (CYP) 4A-derived 20-hydroxyeicosatetraenoic acid (20-HETE) promotes angiogenesis. Here, we tested whether a novel flavonoid (FLA-16) prolongs survival and normalizes tumor vasculature in glioma through CYP4A inhibition. FLA-16 improved survival, reduced tumor burden, and normalized vasculature, accompanied with the decreased secretion of 20-HETE, VEGF and TGF-β in tumor-associated macrophages (TAMs) and endothelial progenitor cells (EPCs) in C6 and U87 gliomas. FLA-16 attenuated vascular abnormalization induced by co-implantation of GL261 glioma cells with CYP4A10 high macrophages or EPCs. Mechanistically, the conditional medium from TAMs and EPCs treated with FLA-16 enhanced the migration of pericyte cells, and decreased the proliferation and migration of endothelial cells, which were reversed by CYP4A overexpression or exogenous addition of 20-HETE, VEGF and TGF-β. Furthermore, FLA-16 prevented crosstalk between TAMs and EPCs during angiogenesis. These results suggest that CYP4A inhibition by FLA-16 prolongs survival and normalizes vasculature in glioma through decreasing production of TAMs and EPCs-derived VEGF and TGF-β. This may represent a potential therapeutic strategy to overcome resistance to anti-VEGF treatment by effects on vessels and immune cells. Copyright © 2017 Elsevier B.V. All rights reserved.

Involvement of nitric oxide synthase in matrix metalloproteinase-9- and/or urokinase plasminogen activator receptor-mediated glioma cell migration

PubMed Central

2013-01-01

Background Src tyrosine kinase activates inducible nitric oxide synthase (iNOS) and, in turn, nitric oxide production as a means to transduce cell migration. Src tyrosine kinase plays a key proximal role to control α9β1 signaling. Our recent studies have clearly demonstrated the role of α9β1 integrin in matrix metalloproteinase-9 (MMP-9) and/or urokinase plasminogen activator receptor (uPAR)-mediated glioma cell migration. In the present study, we evaluated the involvement of α9β1 integrin-iNOS pathway in MMP-9- and/or uPAR-mediated glioma cell migration. Methods MMP-9 and uPAR shRNAs and overexpressing plasmids were used to downregulate and upregulate these molecules, respectively in U251 glioma cells and 5310 glioma xenograft cells. The effect of treatments on migration and invasion potential of these glioma cells were assessed by spheroid migration, wound healing, and Matrigel invasion assays. In order to attain the other objectives we also performed immunocytochemical, immunohistochemical, RT-PCR, Western blot and fluorescence-activated cell sorting (FACS) analysis. Results Immunohistochemical analysis revealed the prominent association of iNOS with glioblastoma multiforme (GBM). Immunofluorescence analysis showed prominent expression of iNOS in glioma cells. MMP-9 and/or uPAR knockdown by respective shRNAs reduced iNOS expression in these glioma cells. RT-PCR analysis revealed elevated iNOS mRNA expression in either MMP-9 or uPAR overexpressed glioma cells. The migration potential of MMP-9- and/or uPAR-overexpressed U251 glioma cells was significantly inhibited after treatment with L-NAME, an inhibitor of iNOS. Similarly, a significant inhibition of the invasion potential of the control or MMP-9/uPAR-overexpressed glioma cells was noticed after L-NAME treatment. A prominent reduction of iNOS expression was observed in the tumor regions of nude mice brains, which were injected with 5310 glioma cells, after MMP-9 and/or uPAR knockdown. Protein expressions of cSrc, phosphoSrc and p130Cas were reduced with simultaneous knockdown of both MMP-9 and uPAR. Conclusions Taken together, our results from the present and earlier studies clearly demonstrate that α9β1 integrin-mediated cell migration utilizes the iNOS pathway, and inhibition of the migratory potential of glioma cells by simultaneous knockdown of MMP-9 and uPAR could be attributed to the reduced α9β1 integrin and iNOS levels. PMID:24325546

[18F]-FLT Positron Emission Tomography can be used to image the response of sensitive tumors to PI3-Kinase inhibition with the novel agent GDC-0941

PubMed Central

Cawthorne, Christopher; Burrows, Natalie; Gieling, Roben G.; Morrow, Christopher J; Forster, Duncan; Gregory, Jamil; Radigois, Marc; Smigova, Alison; Babur, Muhammad; Simpson, Kathryn; Hodgkinson, Cassandra; Brown, Gavin; McMahon, Adam; Dive, Caroline; Hiscock, Duncan; Wilson, Ian; Williams, Kaye J

2013-01-01

The Phosphatidylinositide 3-kinase (PI3-K) pathway is deregulated in a range of cancers, and several targeted inhibitors are entering the clinic. This study aimed to investigate whether the PET tracer 3′-Deoxy-3′-[18F]fluorothymidine ([18F]-FLT) is suitable to mark the effect of the novel PI-3K inhibitor GDC-0941 which has entered phase II clinical trial. CBA nude mice bearing U87 glioma and HCT116 colorectal xenografts were imaged at baseline with [18F]-FLT and at acute (18h) and chronic (186h) timepoints after twice-daily administration of GDC-0941 (50mg/kg) or vehicle. Tumor uptake normalized to blood pool was calculated, and tissue was analyzed at sacrifice for PI3-K pathway inhibition and thymidine kinase (TK1) expression. Uptake of [18F]-FLT was also assessed in tumors inducibly overexpressing a dominant-negative form of the PI3-K p85 subunit Δp85α, as well as HCT116 liver metastases after GDC-0941 therapy. GDC-0941 treatment induced tumor stasis in U87 xenografts, whereas inhibition of HCT116 tumors was more variable. Tumor uptake of [18F]-FLT was significantly reduced following GDC-0941 dosing in responsive tumors at the acute timepoint, and correlated with pharmacodynamic markers of PI3-K signaling inhibition and significant reduction in TK1 expression in U87, but not HCT116, tumors. Reduction of PI3-K signaling via expression of Δp85α significantly reduced tumor growth and [18F]-FLT uptake, as did treatment of HCT116 liver metastases with GDC-0941. These results indicate that [18F]-FLT is a strong candidate for the non-invasive measurement of GDC-0941 action. PMID:23427298

[18F]-FLT positron emission tomography can be used to image the response of sensitive tumors to PI3-kinase inhibition with the novel agent GDC-0941.

PubMed

Cawthorne, Christopher; Burrows, Natalie; Gieling, Roben G; Morrow, Christopher J; Forster, Duncan; Gregory, Jamil; Radigois, Marc; Smigova, Alison; Babur, Muhammad; Simpson, Kathryn; Hodgkinson, Cassandra; Brown, Gavin; McMahon, Adam; Dive, Caroline; Hiscock, Duncan; Wilson, Ian; Williams, Kaye J

2013-05-01

The phosphoinositide 3-kinase (PI3K) pathway is deregulated in a range of cancers, and several targeted inhibitors are entering the clinic. This study aimed to investigate whether the positron emission tomography tracer 3'-deoxy-3'-[(18)F]fluorothymidine ([(18)F]-FLT) is suitable to mark the effect of the novel PI3K inhibitor GDC-0941, which has entered phase II clinical trial. CBA nude mice bearing U87 glioma and HCT116 colorectal xenografts were imaged at baseline with [(18)F]-FLT and at acute (18 hours) and chronic (186 hours) time points after twice-daily administration of GDC-0941 (50 mg/kg) or vehicle. Tumor uptake normalized to blood pool was calculated, and tissue was analyzed at sacrifice for PI3K pathway inhibition and thymidine kinase (TK1) expression. Uptake of [(18)F]-FLT was also assessed in tumors inducibly overexpressing a dominant-negative form of the PI3K p85 subunit p85α, as well as HCT116 liver metastases after GDC-0941 therapy. GDC-0941 treatment induced tumor stasis in U87 xenografts, whereas inhibition of HCT116 tumors was more variable. Tumor uptake of [(18)F]-FLT was significantly reduced following GDC-0941 dosing in responsive tumors at the acute time point and correlated with pharmacodynamic markers of PI3K signaling inhibition and significant reduction in TK1 expression in U87, but not HCT116, tumors. Reduction of PI3K signaling via expression of Δp85α significantly reduced tumor growth and [(18)F]-FLT uptake, as did treatment of HCT116 liver metastases with GDC-0941. These results indicate that [(18)F]-FLT is a strong candidate for the noninvasive measurement of GDC-0941 action. ©2013 AACR

Potential involvement of F0F1-ATP(synth)ase and reactive oxygen species in apoptosis induction by the antineoplastic agent erucylphosphohomocholine in glioblastoma cell lines : a mechanism for induction of apoptosis via the 18 kDa mitochondrial translocator protein.

PubMed

Veenman, Leo; Alten, Julia; Linnemannstöns, Karen; Shandalov, Yulia; Zeno, Sivan; Lakomek, Max; Gavish, Moshe; Kugler, Wilfried

2010-07-01

Erucylphosphohomocholine (ErPC3, Erufosine) was reported previously to induce apoptosis in otherwise highly apoptosis-resistant malignant glioma cell lines while sparing their non-tumorigenic counterparts. We also previously found that the mitochondrial 18 kDa Translocator Protein (TSPO) is required for apoptosis induction by ErPC3. These previous studies also suggested involvement of reactive oxygen species (ROS). In the present study we further investigated the potential involvement of ROS generation, the participation of the mitochondrial respiration chain, and the role of the mitochondrial F(O)F(1)-ATP(synth)ase in the pro-apoptotic effects of ErPC3 on U87MG and U118MG human glioblastoma cell lines. For this purpose, cells were treated with the ROS chelator butylated hydroxyanisole (BHA), the mitochondrial respiration chain inhibitors rotenone, antimycin A, myxothiazol, and the uncoupler CCCP. Also oligomycin and piceatannol were studied as inhibitors of the F(O) and F(1) subunits of the mitochondrial F(O)F(1)-ATP(synth)ase, respectively. BHA was able to attenuate apoptosis induction by ErPC3, including mitochondrial ROS generation as determined with cardiolipin oxidation, as well as collapse of the mitochondrial membrane potential (Deltapsi(m)). Similarly, we found that oligomycin attenuated apoptosis and collapse of the Deltapsi(m), normally induced by ErPC3, including the accompanying reductions in cellular ATP levels. Other inhibitors of the mitochondrial respiration chain, as well as piceatannol, did not show such effects. Consequently, our findings strongly point to a role for the F(O) subunit of the mitochondrial F(O)F(1)-ATP(synth)ase in ErPC3-induced apoptosis and dissipation of Deltapsi(m) as well as ROS generation by ErPC3 and TSPO.

Dexamethasone Alleviates Tumor-Associated Brain Damage and Angiogenesis

PubMed Central

Fan, Zheng; Sehm, Tina; Rauh, Manfred; Buchfelder, Michael

2014-01-01

Children and adults with the most aggressive form of brain cancer, malignant gliomas or glioblastoma, often develop cerebral edema as a life-threatening complication. This complication is routinely treated with dexamethasone (DEXA), a steroidal anti-inflammatory drug with pleiotropic action profile. Here we show that dexamethasone reduces murine and rodent glioma tumor growth in a concentration-dependent manner. Low concentrations of DEXA are already capable of inhibiting glioma cell proliferation and at higher levels induce cell death. Further, the expression of the glutamate antiporter xCT (system Xc −; SLC7a11) and VEGFA is up-regulated after DEXA treatment indicating early cellular stress responses. However, in human gliomas DEXA exerts differential cytotoxic effects, with some human glioma cells (U251, T98G) resistant to DEXA, a finding corroborated by clinical data of dexamethasone non-responders. Moreover, DEXA-resistant gliomas did not show any xCT alterations, indicating that these gene expressions are associated with DEXA-induced cellular stress. Hence, siRNA-mediated xCT knockdown in glioma cells increased the susceptibility to DEXA. Interestingly, cell viability of primary human astrocytes and primary rodent neurons is not affected by DEXA. We further tested the pharmacological effects of DEXA on brain tissue and showed that DEXA reduces tumor-induced disturbances of the microenvironment such as neuronal cell death and tumor-induced angiogenesis. In conclusion, we demonstrate that DEXA inhibits glioma cell growth in a concentration and species-dependent manner. Further, DEXA executes neuroprotective effects in brains and reduces tumor-induced angiogenesis. Thus, our investigations reveal that DEXA acts pleiotropically and impacts tumor growth, tumor vasculature and tumor-associated brain damage. PMID:24714627

Knockdown of NF-E2-related factor 2 inhibits the proliferation and growth of U251MG human glioma cells in a mouse xenograft model.

PubMed

Ji, Xiang-Jun; Chen, Sui-Hua; Zhu, Lin; Pan, Hao; Zhou, Yuan; Li, Wei; You, Wan-Chun; Gao, Chao-Chao; Zhu, Jian-Hong; Jiang, Kuan; Wang, Han-Dong

2013-07-01

NF-E2-related factor 2 (Nrf2) is a pivotal transcription factor of cellular responses to oxidative stress and recent evidence suggests that Nrf2 plays an important role in cancer pathobiology. However, the underlying mechanism has yet to be elucidated, particularly in glioma. In the present study, we investigated the role of Nrf2 in the clinical prognosis, cell proliferation and tumor growth of human glioblastoma multiforme (GBM). We detected overexpression of Nrf2 protein levels in GBM compared to normal brain tissues. Notably, higher protein levels of Nrf2 were significantly associated with poorer overall survival and 1-year survival for GBM patients. Furthermore, we constructed the plasmid Si-Nrf2 and transduced it into U251MG cells to downregulate the expression of Nrf2 and established stable Nrf2 knockdown cells. The downregulation of Nrf2 suppressed cell proliferation in vitro and tumor growth in mouse xenograft models. We performed immunohistochemistry staining to detect the protein levels of Nrf2, Ki-67, caspase-3 and CD31 in the xenograft tumors and found that the expression levels of Nrf2 and Ki-67 were much lower in the Si-Nrf2 group compared to the Si-control group. In addition, the number of caspase-3-positive cells was significantly increased in the Si-Nrf2 group. By analysis of microvessel density (MVD) assessed by CD31, the MVD value in the Si-Nrf2 group decreased significantly compared to the Si-control group. These findings indicate that the knockdown of Nrf2 may suppress tumor growth by inhibiting cell proliferation, increasing cell apoptosis and inhibiting angiogenesis. These results highlight the potential of Nrf2 as a candidate molecular target to control GBM cell proliferation and tumor growth.

Stat3 inhibition activates tumor macrophages and abrogates glioma growth in mice.

PubMed

Zhang, Leying; Alizadeh, Darya; Van Handel, Michelle; Kortylewski, Marcin; Yu, Hua; Badie, Behnam

2009-10-01

As the main effector-cell population of the central nervous system, microglia (MG) are considered to play an important immunoregulatory function in a number of pathological conditions such as inflammation, trauma, degenerative disease, and brain tumors. Recent studies, however, have suggested that the anti-neoplastic function of MG may be suppressed in malignant brain tumors. Considering the proposed suppressive role of signal transducers and activators of transcription 3 (Stat3) in antitumor immunity, we evaluated the role of Stat3 inhibition on MG and macrophage (MP) activation and tumor growth in a murine glioma model. N9 MG cells were exposed to GL261 glioma conditioned medium (GL261-CM) and evaluated for Stat3 activity and cytokine expression. Furthermore, the role of Stat3 inhibition on MG and MP activation was studied both in vitro and in vivo. Finally, the effect of Stat3 inhibition on tumor growth was assessed in intracranial GL261 gliomas. GL261-CM increased Stat3 activity in N9 cells in vitro and resulted in overexpression of IL-10 and IL-6, and downregulation of IL1-beta, a pro-inflammatory cytokine. Inhibition of Stat3 by CPA-7 or siRNA reversed glioma-induced cytokine expression profile in N9 cells. Furthermore, inactivation of Stat3 in intracranial GL261 tumors by siRNA resulted in MG/MP activation and tumor growth inhibition. Glioma-induced MG and MP suppression may be mediated thorough Stat3. Inhibition of Stat3 function in tumor MG/MP may result in their activation and can potentially be used as an adjunct immunotherapy approach for gliomas.

WE-H-207A-08: Characterization of a Broad-Spectrum Cancer Targeted MRI Contrast Agent

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brunnquell, C; Zhang, R; Pinchuk, A

Purpose: To characterize the relaxation properties and tumor targeting capabilities of a novel alkylphosphocholine (APC) analog MR contrast agent, Gd-DO3A-404. Methods: Relaxivities were measured via T1 and T2 mapping of Gd-DO3A-404 with inversion recovery and spin echo pulse sequences, respectively. Uptake was characterized in flank xenograft models of non-small cell lung cancer (A549) and glioma (U87) and compared with uptake of Dotarem. Mice (N=3 per model per agent) were delivered 2.34 moles contrast intravenously. T1-weighted MRI and T1 maps were acquired pre-contrast and at multiple time points up to seven days post-contrast. For Dotarem imaging, T1-weighted MRI was performed atmore » multiple time points from one minute to one day. Results: Relaxivities of Gd-DO3A-404 in plasma were r1=5.74 and r2=20.4 s-1/mm at 4.7T, comparing favorably to clinical contrast agent Dotarem (r1=3.3, r2=4.7). Specific, sustained uptake of Gd-DO3A-404 was observed in U87 and A549. The ratio of tumor:muscle T1-weighted signal increased from 1.24 pre-contrast to 2.12 twenty-four hours post-contrast in U87 and from 1.14 to 2.16 (same time points) in A549. Significant signal enhancement was maintained until 7 and 4 days post-contrast in U87 and A549, respectively. In comparison, uptake and washout of Dotarem in U87 occurred over the course of fifteen minutes. The ratio of tumor:muscle T1-weighted signal increased only 59% as much as Gd-DO3A-404, ranging from 1.15 pre-contrast to a maximum of 1.67 five minutes post-contrast. Significant signal enhancement from Dotarem was not sustained beyond one hour post-contrast. Conclusion: These results indicate that with favorable relaxation characteristics and sustained signal-enhancing uptake in multiple tumor models, Gd-DO3A-404 has great potential as a tumor-targeting MR contrast agent. As part of a library of APC analogs labeled with PET/optical tracers and therapeutic radionuclides, Gd-DO3A-404 further expands theranostic capabilities. Future work will investigate applications in orthotopic glioma imaging, simultaneous PET/MR, and neutron capture therapy.« less

Betulinic acid derivatives NVX-207 and B10 for treatment of glioblastoma--an in vitro study of cytotoxicity and radiosensitization.

PubMed

Bache, Matthias; Bernhardt, Stephan; Passin, Sarina; Wichmann, Henri; Hein, Anja; Zschornak, Martin; Kappler, Matthias; Taubert, Helge; Paschke, Reinhard; Vordermark, Dirk

2014-10-30

Betulinic acid (BA), a pentacyclic triterpene, represents a new therapeutic substance that has potential benefits for treating glioblastoma. Recently, new strategies for producing BA derivatives with improved properties have evolved. However, few studies have examined the combination of BA or BA derivatives using radiotherapy. The effects of two BA derivatives, NVX-207 and B10, on cellular and radiobiological behavior were analyzed using glioblastoma cell lines (U251MG, U343MG and LN229). Based on IC50 values under normoxic conditions, we detected a 1.3-2.9-fold higher cytotoxicity of the BA derivatives B10 and NVX-207, respectively, compared to BA. Incubation using both BA derivatives led to decreased cell migration, cleavage of PARP and decreased protein expression levels of Survivin. Weak radiation sensitivity enhancement was observed in U251MG cells after treatment with both BA derivatives. The enhancement factors at an irradiation dose of 6 Gy after treatment with 5 µM NVX-207 and 5 µM B10 were 1.32 (p=0.029) and 1.55 (p=0.002), respectively. In contrast to BA, neither NVX-207 nor B10 had additional effects under hypoxic conditions. Our results suggest that the BA derivatives NVX-207 and B10 improve the effects of radiotherapy on human malignant glioma cells, particularly under normoxic conditions.

Betulinic Acid Derivatives NVX-207 and B10 for Treatment of Glioblastoma—An in Vitro Study of Cytotoxicity and Radiosensitization

PubMed Central

Bache, Matthias; Bernhardt, Stephan; Passin, Sarina; Wichmann, Henri; Hein, Anja; Zschornak, Martin; Kappler, Matthias; Taubert, Helge; Paschke, Reinhard; Vordermark, Dirk

2014-01-01

Betulinic acid (BA), a pentacyclic triterpene, represents a new therapeutic substance that has potential benefits for treating glioblastoma. Recently, new strategies for producing BA derivatives with improved properties have evolved. However, few studies have examined the combination of BA or BA derivatives using radiotherapy. The effects of two BA derivatives, NVX-207 and B10, on cellular and radiobiological behavior were analyzed using glioblastoma cell lines (U251MG, U343MG and LN229). Based on IC50 values under normoxic conditions, we detected a 1.3–2.9-fold higher cytotoxicity of the BA derivatives B10 and NVX-207, respectively, compared to BA. Incubation using both BA derivatives led to decreased cell migration, cleavage of PARP and decreased protein expression levels of Survivin. Weak radiation sensitivity enhancement was observed in U251MG cells after treatment with both BA derivatives. The enhancement factors at an irradiation dose of 6 Gy after treatment with 5 µM NVX-207 and 5 µM B10 were 1.32 (p = 0.029) and 1.55 (p = 0.002), respectively. In contrast to BA, neither NVX-207 nor B10 had additional effects under hypoxic conditions. Our results suggest that the BA derivatives NVX-207 and B10 improve the effects of radiotherapy on human malignant glioma cells, particularly under normoxic conditions. PMID:25361208

Suppression of autophagy augments the radiosensitizing effects of STAT3 inhibition on human glioma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yuan, Xiaopeng; Du, Jie; Hua, Song

Radiotherapy is an essential component of the standard therapy for newly diagnosed glioblastoma. To increase the radiosensitivity of glioma cells is a feasible solution to improve the therapeutic effects. It has been suggested that inhibition of signal transducer and activator of transcription 3 (STAT3) can radiosensitize glioma cells, probably via the activation of mitochondrial apoptotic pathway. In this study, human malignant glioma cells, U251 and A172, were treated with an STAT3 inhibitor, WP1066, or a short hairpin RNA plasmid targeting STAT3 to suppress the activation of STAT3 signaling. The radiosensitizing effects of STAT3 inhibition were confirmed in glioma cells. Intriguingly,more » combination of ionizing radiation exposure and STAT3 inhibition triggered a pronounced increase of autophagy flux. To explore the role of autophagy, glioma cells were treated with 3-methyladenine or siRNA for autophagy-related gene 5, and it was demonstrated that inhibition of autophagy further strengthened the radiosensitizing effects of STAT3 inhibition. Accordingly, more apoptotic cells were induced by the dual inhibition of autophagy and STAT3 signaling. In conclusion, our data revealed a protective role of autophagy in the radiosensitizing effects of STAT3 inhibition, and inhibition of both autophagy and STAT3 might be a potential therapeutic strategy to increase the radiosensitivity of glioma cells. - Highlights: • Inactivation of STAT3 signaling radiosensitizes malignant glioma cells. • STAT3 inhibition triggers a significant increase of autophagy flux induced by ionizing radiation in glioma cells. • Suppression of autophagy further strengthens the radiosensitizing effects of STAT3 inhibition in glioma cells. • Dual inhibition of autophagy and STAT3 induce massive apoptotic cells upon exposure to ionizing radiation.« less

Multifunctional targeted liposomal drug delivery for efficient glioblastoma treatment

PubMed Central

Belhadj, Zakia; Zhan, Changyou; Ying, Man; Wei, Xiaoli; Xie, Cao; Yan, Zhiqiang; Lu, Weiyue

2017-01-01

Glioblastoma multiforme (GBM) has been considered to be the most malignant brain tumors. Due to the existence of various barriers including the blood–brain barrier (BBB) and blood–brain tumor barrier (BBTB) greatly hinder the accumulation and deep penetration of chemotherapeutics, the treatment of glioma remains to be the most challenging task in clinic. In order to circumvent these hurdles, we developed a multifunctional liposomal glioma-targeted drug delivery system (c(RGDyK)/pHA-LS) modified with cyclic RGD (c(RGDyK)) and p-hydroxybenzoic acid (pHA) in which c(RGDyK) could target integrin αvβ3 overexpressed on the BBTB and glioma cells and pHA could target dopamine receptors on the BBB. In vitro, c(RGDyK)/pHA-LS could target glioblastoma cells (U87), brain capillary endothelial cells (bEnd.3) and umbilical vein endothelial cells (HUVECs) through a comprehensive pathway. Besides, c(RGDyK)/pHA-LS could also increase the cytotoxicity of doxorubicin encapsulated in liposomes on glioblastoma cells, and was able to penetrate inside the glioma spheroids after traversing the in vitro BBB and BBTB. In vivo, we demonstrated the targeting ability of c(RGDyK)/pHA-LS to intracranial glioma. As expected, c(RGDyK)/pHA-LS/DOX showed a median survival time of 35 days, which was 2.31-, 1.76- and 1.5-fold higher than that of LS/DOX, c(RGDyK)-LS/DOX, and pHA-LS/DOX, respectively. The findings here suggested that the multifunctional glioma-targeted drug delivery system modified with both c(RGDyK) and pHA displayed strong antiglioma efficiency in vitro and in vivo, representing a promising platform for glioma therapy. PMID:28978003

γ-Glutamyl transferase 7 is a novel regulator of glioblastoma growth.

PubMed

Bui, Timothy T; Nitta, Ryan T; Kahn, Suzana A; Razavi, Seyed-Mostafa; Agarwal, Maya; Aujla, Parvir; Gholamin, Sharareh; Recht, Lawrence; Li, Gordon

2015-04-07

Glioblastoma (GBM) is the most malignant primary brain tumor in adults, with a median survival time of one and a half years. Traditional treatments, including radiation, chemotherapy, and surgery, are not curative, making it imperative to find more effective treatments for this lethal disease. γ-Glutamyl transferase (GGT) is a family of enzymes that was shown to control crucial redox-sensitive functions and to regulate the balance between proliferation and apoptosis. GGT7 is a novel GGT family member that is highly expressed in brain and was previously shown to have decreased expression in gliomas. Since other members of the GGT family were found to be altered in a variety of cancers, we hypothesized that GGT7 could regulate GBM growth and formation. To determine if GGT7 is involved in GBM tumorigenesis, we modulated GGT7 expression in two GBM cell lines (U87-MG and U138) and monitored changes in tumorigenicity in vitro and in vivo. We demonstrated for the first time that GBM patients with low GGT7 expression had a worse prognosis and that 87% (7/8) of primary GBM tissue samples showed a 2-fold decrease in GGT7 expression compared to normal brain samples. Exogenous expression of GGT7 resulted in a 2- to 3-fold reduction in proliferation and anchorage-independent growth under minimal growth conditions (1% serum). Decreasing GGT7 expression using either short interfering RNA or short hairpin RNA consistently increased proliferation 1.5- to 2-fold. In addition, intracranial injections of U87-MG cells with reduced GGT7 expression increased tumor growth in mice approximately 2-fold, and decreased mouse survival. To elucidate the mechanism by which GGT7 regulates GBM growth, we analyzed reactive oxygen species (ROS) levels in GBM cells with modulated GGT7 expression. We found that enhanced GGT7 expression reduced ROS levels by 11-33%. Our study demonstrates that GGT7 is a novel player in GBM growth and that GGT7 can play a critical role in tumorigenesis by regulating anti-oxidative damage. Loss of GGT7 may increase the cellular ROS levels, inducing GBM occurrence and growth. Our findings suggest that GGT7 can be a promising biomarker and a potential therapeutic target for GBM.

CRISPR/Cas9-Mediated Knockin Application in Cell Therapy: A Non-viral Procedure for Bystander Treatment of Glioma in Mice.

PubMed

Meca-Cortés, Oscar; Guerra-Rebollo, Marta; Garrido, Cristina; Borrós, Salvador; Rubio, Nuria; Blanco, Jeronimo

2017-09-15

The use of non-viral procedures, together with CRISPR/Cas9 genome-editing technology, allows the insertion of single-copy therapeutic genes at pre-determined genomic sites, overcoming safety limitations resulting from random gene insertions of viral vectors with potential for genome damage. In this study, we demonstrate that combination of non-viral gene delivery and CRISPR/Cas9-mediated knockin via homology-directed repair can replace the use of viral vectors for the generation of genetically modified therapeutic cells. We custom-modified human adipose mesenchymal stem cells (hAMSCs), using electroporation as a transfection method and CRISPR/Cas9-mediated knockin for the introduction and stable expression of a 3 kb DNA fragment including the eGFP (selectable marker) and a variant of the herpes simplex virus 1 thymidine kinase genes (therapeutic gene), under the control of the human elongation factor 1 alpha promoter in exon 5 of the endogenous thymidine kinase 2 gene. Using a U87 glioma model in SCID mice, we show that the therapeutic capacity of the new CRISPR/Cas9-engineered hAMSCs is equivalent to that of therapeutic hAMSCs generated by introduction of the same therapeutic gene by transduction with a lentiviral vector previously published by our group. This strategy should be of general use to other applications requiring genetic modification of therapeutic cells. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Evaluation of Nutritional Composition of The Dried Seaweed Ulva lactuca from Pameungpeuk Waters, Indonesia.

PubMed

Rasyid, Abdullah

2017-07-01

The nutritional composition of the dried seaweed Ulva lactuca from Pameungpeuk waters, including proximate, vitamins, minerals, dietary fibre and heavy metal has been carried out. The objective of this present study is to know the nutritional composition of the dried seaweed U. lactuca for utilisation in human nutrition in the future. Results show that carbohydrate was the major component in the proximate analysis of U. lactuca in the present study. The carbohydrate content was 58.1%. Moisture, ash, protein and fat content were 16.9%, 11.2%, 13.6% and 0.19% respectively, while dietary fibre was 28.4%. The vitamin A content was examined in this study less than 0.5 IU/100 mg while vitamin B1 (thiamine) and vitamin B2 (riboflavin) were 4.87 mg/kg and 0.86 mg/kg respectively. The calcium content was 1828 mg/100 g higher than other minerals. The heavy metal content examined in this study were lower than the limit of the quality criteria applied to edible seaweeds sold in Indonesia. Based on the results of this study show that U. lactuca has potential to be developed as an alternative source of a healthy food for human in the future.

β-Asarone promotes Temozolomide's entry into glioma cells and decreases the expression of P-glycoprotein and MDR1.

PubMed

Wang, Nanbu; Zhang, Qinxin; Ning, Baile; Luo, Laiyu; Fang, Yongqi

2017-06-01

Glioma is the most common primary brain tumor and has an undesirable prognosis due to the blood-brain barrier (BBB) and drug resistance. A thorough investigation of the changes in intracellular drug concentrations is important to observe therapeutic effects and cell resistance. P-glycoprotein (P-gp) is an essential protein of Multi-drug resistance 1 (MDR1). The over-expression of P-gp and MDR1 is associated with poor prognosis and drug-resistance in glioma. However, β-asarone can pass through the BBB easily and increase the drug concentration in the rat brain. Our aim is to study the effect of β-asarone on promoting the entry of temozolomide (TMZ) into human glioma U251 cells. The cells were divided into three groups: model group, TMZ group (300μM) and co-administration group (360μM β-asarone; 300μM TMZ). We further detected P-gp and MDR1 expression in U251 and rat glioma C6 cells in four groups: model group (U251/C6), TMZ group (U251 300μM, C6 420μM), β-asarone group (U251 360μM, C6 450μM) and co-administration group (β-asarone 360μM, TMZ 300μM for U251; β-asarone 450μM, TMZ 420μM for C6). Then, high performance liquid chromatography was used to determine the intracellular and extracellular levels of TMZ. Morphological changes in both cells were observed by the microscope. The Counting Kit-8 assay was used to measure the cell proliferation and toxicity. Cell immunohistochemistry/immunofluorescence, flowcytometry and western blot were synchronously used to examine the expression of P-gp. We also determined the levels of MDR1 mRNA by RT-PCR. The results showed that β-asarone could promote the entry of TMZ into U251 cells through the membrane. The co-administration of β-asarone and TMZ also decreased cell proliferation and the expression of P-gp and MDR1 better than single medication in U251 and C6 cells. All of the data suggest that β-asarone might contribute to treatment by promoting TMZ's entry into glioma cells, thereby contributing to anti-cancer growth and inhibiting P-gp and MDR1 expression. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Engineered Modular Recombinant Transporters: Application of New Platform for Targeted Radiotherapeutic Agents to {alpha}-Particle Emitting {sup 211}At

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rosenkranz, Andrey A.; Department of Biophysics, Biological Faculty, Moscow State University, Moscow; Vaidyanathan, Ganesan

2008-09-01

Purpose: To generate and evaluate a modular recombinant transporter (MRT) for targeting {sup 211}At to cancer cells overexpressing the epidermal growth factor receptor (EGFR). Methods and Materials: The MRT was produced with four functional modules: (1) human epidermal growth factor as the internalizable ligand, (2) the optimized nuclear localization sequence of simian vacuolating virus 40 (SV40) large T-antigen, (3) a translocation domain of diphtheria toxin as an endosomolytic module, and (4) the Escherichia coli hemoglobin-like protein (HMP) as a carrier module. MRT was labeled using N-succinimidyl 3-[{sup 211}At]astato-5-guanidinomethylbenzoate (SAGMB), its {sup 125}I analogue SGMIB, or with {sup 131}I using Iodogen.more » Binding, internalization, and clonogenic assays were performed with EGFR-expressing A431, D247 MG, and U87MG.wtEGFR human cancer cell lines. Results: The affinity of SGMIB-MRT binding to A431 cells, determined by Scatchard analysis, was 22 nM, comparable to that measured before labeling. The binding of SGMIB-MRT and its internalization by A431 cancer cells was 96% and 99% EGFR specific, respectively. Paired label assays demonstrated that compared with Iodogen-labeled MRT, SGMIB-MRT and SAGMB-MRT exhibited more than threefold greater peak levels and durations of intracellular retention of activity. SAGMB-MRT was 10-20 times more cytotoxic than [{sup 211}At]astatide for all three cell lines. Conclusion: The results of this study have demonstrated the initial proof of principle for the MRT approach for designing targeted {alpha}-particle emitting radiotherapeutic agents. The high cytotoxicity of SAGMB-MRT for cancer cells overexpressing EGFR suggests that this {sup 211}At-labeled conjugate has promise for the treatment of malignancies, such as glioma, which overexpress this receptor.« less

Imaging of platelet-derived growth factor receptor β expression in glioblastoma xenografts using affibody molecule 111In-DOTA-Z09591.

PubMed

Tolmachev, Vladimir; Varasteh, Zohreh; Honarvar, Hadis; Hosseinimehr, Seyed Jalal; Eriksson, Olof; Jonasson, Per; Frejd, Fredrik Y; Abrahmsen, Lars; Orlova, Anna

2014-02-01

The overexpression and excessive signaling of platelet-derived growth factor receptor β (PDGFRβ) has been detected in cancers, atherosclerosis, and a variety of fibrotic diseases. Radionuclide in vivo visualization of PDGFRβ expression might help to select PDGFRβ targeting treatment for these diseases. The goal of this study was to evaluate the feasibility of in vivo radionuclide imaging of PDGFRβ expression using an Affibody molecule, a small nonimmunoglobulin affinity protein. The PDGFRβ-binding Z09591 Affibody molecule was site-specifically conjugated with a maleimido derivative of DOTA and labeled with (111)In. Targeting of the PDGFRβ-expressing U-87 MG glioblastoma cell line using (111)In-DOTA-Z09591 was evaluated in vitro and in vivo. DOTA-Z09591 was stably labeled with (111)In with preserved specific binding to PDGFRβ-expressing cells in vitro. The dissociation constant for (111)In-DOTA-Z09591 binding to U-87 MG cells was determined to be 92 ± 10 pM. In mice bearing U-87 MG xenografts, the tumor uptake of (111)In-DOTA-Z09591 was 7.2 ± 2.4 percentage injected dose per gram and the tumor-to-blood ratio was 28 ± 14 at 2 h after injection. In vivo receptor saturation experiments demonstrated that targeting of U-87 MG xenografts in mice was PDGFRβ-specific. U-87 MG xenografts were clearly visualized using small-animal SPECT/CT at 3 h after injection. This study demonstrates the feasibility of in vivo visualization of PDGFRβ-expressing xenografts using an Affibody molecule. Further development of radiolabeled Affibody molecules might provide a useful clinical imaging tool for PDGFRβ expression during various pathologic conditions.

Transferrin receptor-targeted theranostic gold nanoparticles for photosensitizer delivery in brain tumors

NASA Astrophysics Data System (ADS)

Dixit, Suraj; Novak, Thomas; Miller, Kayla; Zhu, Yun; Kenney, Malcolm E.; Broome, Ann-Marie

2015-01-01

Therapeutic drug delivery across the blood-brain barrier (BBB) is not only inefficient, but also nonspecific to brain stroma. These are major limitations in the effective treatment of brain cancer. Transferrin peptide (Tfpep) targeted gold nanoparticles (Tfpep-Au NPs) loaded with the photodynamic pro-drug, Pc 4, have been designed and compared with untargeted Au NPs for delivery of the photosensitizer to brain cancer cell lines. In vitro studies of human glioma cancer lines (LN229 and U87) overexpressing the transferrin receptor (TfR) show a significant increase in cellular uptake for targeted conjugates as compared to untargeted particles. Pc 4 delivered from Tfpep-Au NPs clusters within vesicles after targeting with the Tfpep. Pc 4 continues to accumulate over a 4 hour period. Our work suggests that TfR-targeted Au NPs may have important therapeutic implications for delivering brain tumor therapies and/or providing a platform for noninvasive imaging.

Transferrin receptor-targeted theranostic gold nanoparticles for photosensitizer delivery in brain tumors

PubMed Central

Dixit, Suraj; Novak, Thomas; Miller, Kayla; Zhu, Yun; Kenney, Malcolm E.

2015-01-01

Therapeutic drug delivery across the blood-brain barrier (BBB) is not only inefficient, but also nonspecific to brain stroma. These are major limitations in the effective treatment of brain cancer. Transferrin peptide (Tfpep) targeted gold nanoparticles (Tfpep-Au NPs) loaded with the photodynamic pro-drug, Pc 4, have been designed and compared with untargeted Au NPs for delivery of the photosensitizer to brain cancer cell lines. In vitro studies of human glioma cancer lines (LN229 and U87) overexpressing the transferrin receptor (TfR) show a significant increase in cellular uptake for targeted conjugates as compared to un-targeted particles. Pc 4 delivered from Tfpep-Au NPs clusters within vesicles after targeting with the Tfpep. Pc 4 continues to accumulate over a 4 hour period. Our work suggests that TfR-targeted Au NPs may have important therapeutic implications for delivering brain tumor therapies and/or providing a platform for noninvasive imaging. PMID:25519743

Preclinical Efficacy of the Anti-Hepatocyte Growth Factor Antibody Ficlatuzumab in a Mouse Brain Orthotopic Glioma Model Evaluated by Bioluminescence, PET, and MRI

PubMed Central

Mittra, Erik S.; Fan-Minogue, Hua; Lin, Frank I.; Karamchandani, Jason; Sriram, Venkataraman; Han, May; Gambhir, Sanjiv S.

2016-01-01

Purpose Ficlatuzumab is a novel therapeutic agent targeting the hepatocyte growth factor (HGF)/c-MET pathway. We summarize extensive preclinical work using this agent in a mouse brain orthotopic model of glioblastoma. Experimental Design Sequential experiments were done using eight- to nine-week-old nude mice injected with 3 × 105 U87 MG (glioblastoma) cells into the brain. Evaluation of ficlatuzumab dose response for this brain tumor model and comparison of its response to ficlatuzumab and to temozolamide were conducted first. Subsequently, various small-animal imaging modalities, including bioluminescence imaging (BLI), positron emission tomography (PET), and MRI, were used with a U87 MG-Luc 2 stable cell line, with and without the use of ficlatuzumab, to evaluate the ability to non-invasively assess tumor growth and response to therapy. ANOVA was conducted to evaluate for significant differences in the response. Results There was a survival benefit with ficlatuzumab alone or in combination with temozolamide. BLI was more sensitive than PET in detecting tumor cells. Fluoro-D-thymidine (FLT) PET provided a better signal-to-background ratio than 2[18F]fluoro-2-deoxy-D-glucose (FDG) PET. In addition, both BLI and FLT PET showed significant changes over time in the control group as well as with response to therapy. MRI does not disclose any time-dependent change. Also, the MRI results showed a temporal delay in comparison to the BLI and FLT PET findings, showing similar results one drug cycle later. Conclusions Targeting the HGF/c-MET pathway with the novel agent ficlatuzumab appears promising for the treatment of glioblastoma. Various clinically applicable imaging modalities including FLT, PET, and MRI provide reliable ways of assessing tumor growth and response to therapy. Given the clinical applicability of these findings, future studies on patients with glioblastoma may be appropriate. PMID:23983258

Brain Exposure of Two Selective Dual CDK4 and CDK6 Inhibitors and the Antitumor Activity of CDK4 and CDK6 Inhibition in Combination with Temozolomide in an Intracranial Glioblastoma Xenograft.

PubMed

Raub, Thomas J; Wishart, Graham N; Kulanthaivel, Palaniappan; Staton, Brian A; Ajamie, Rose T; Sawada, Geri A; Gelbert, Lawrence M; Shannon, Harlan E; Sanchez-Martinez, Concepcion; De Dios, Alfonso

2015-09-01

Effective treatments for primary brain tumors and brain metastases represent a major unmet medical need. Targeting the CDK4/CDK6-cyclin D1-Rb-p16/ink4a pathway using a potent CDK4 and CDK6 kinase inhibitor has potential for treating primary central nervous system tumors such as glioblastoma and some peripheral tumors with high incidence of brain metastases. We compared central nervous system exposures of two orally bioavailable CDK4 and CDK6 inhibitors: abemaciclib, which is currently in advanced clinical development, and palbociclib (IBRANCE; Pfizer), which was recently approved by the U.S. Food and Drug Administration. Abemaciclib antitumor activity was assessed in subcutaneous and orthotopic glioma models alone and in combination with standard of care temozolomide (TMZ). Both inhibitors were substrates for xenobiotic efflux transporters P-glycoprotein and breast cancer resistant protein expressed at the blood-brain barrier. Brain Kp,uu values were less than 0.2 after an equimolar intravenous dose indicative of active efflux but were approximately 10-fold greater for abemaciclib than palbociclib. Kp,uu increased 2.8- and 21-fold, respectively, when similarly dosed in P-gp-deficient mice. Abemaciclib had brain area under the curve (0-24 hours) Kp,uu values of 0.03 in mice and 0.11 in rats after a 30 mg/kg p.o. dose. Orally dosed abemaciclib significantly increased survival in a rat orthotopic U87MG xenograft model compared with vehicle-treated animals, and efficacy coincided with a dose-dependent increase in unbound plasma and brain exposures in excess of the CDK4 and CDK6 Ki values. Abemaciclib increased survival time of intracranial U87MG tumor-bearing rats similar to TMZ, and the combination of abemaciclib and TMZ was additive or greater than additive. These data show that abemaciclib crosses the blood-brain barrier and confirm that both CDK4 and CDK6 inhibitors reach unbound brain levels in rodents that are expected to produce enzyme inhibition; however, abemaciclib brain levels are reached more efficiently at presumably lower doses than palbociclib and are potentially on target for a longer period of time. Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.

A chitosan/beta-glycerophosphate thermo-sensitive gel for the delivery of ellagic acid for the treatment of brain cancer.

PubMed

Kim, Sungwoo; Nishimoto, Satoru K; Bumgardner, Joel D; Haggard, Warren O; Gaber, M Waleed; Yang, Yunzhi

2010-05-01

We report here the development of a chitosan/beta-glycerophosphate(Ch/beta-GP) thermo-sensitive gel to deliver ellagic acid (EA) for cancer treatment. The properties of the Ch/beta-GP gels were characterized regarding chemical structure, surface morphology, and viscoelasticity. In vitro EA release rate from the EA loaded Ch/beta-GP gel and chitosan degradation rate were investigated. The anti-tumor effect of the EA loaded Ch/beta-GP gel on brain cancer cells (human U87 glioblastomas and rat C6 glioma cells) was evaluated by examining cell viability. Cell number and activity were monitored by the MTS assay. The Ch/beta-GP solution formed a heat-induced gel at body temperature, and the gelation temperature and time were affected by the final pH of the Ch/beta-GP solution. The lysozyme increased the EA release rate by 2.5 times higher than that in the absence of lysozyme. Dialyzed chitosan solution with final pH 6.3 greatly reduced the beta-GP needed for gelation, thereby significantly improving the biocompatibility of gel (p < 0.001). The chitosan gels containing 1% (w/v) of ellagic acid significantly reduced viability of U87 cells and C6 cells compared with the chitosan gels at 3 days incubation (p < 0.01, and p < 0.001, respectively). Copyright 2010 Elsevier Ltd. All rights reserved.

Combinatorial therapy with adenoviral-mediated PTEN and a PI3K inhibitor suppresses malignant glioma cell growth in vitro and in vivo by regulating the PI3K/AKT signaling pathway.

PubMed

Nan, Yang; Guo, Liyun; Song, Yunpeng; Wang, Le; Yu, Kai; Huang, Qiang; Zhong, Yue

2017-08-01

Glioblastoma is a highly invasive and challenging tumor of the central nervous system. The mutation/deletion of the tumor suppressor phosphatase and tensin homolog (PTEN) gene is the main genetic change identified in glioblastomas. PTEN plays a critical role in tumorigenesis and has been shown to be an important therapeutic target. The phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 is commonly used to inhibit glioma cell growth via regulation of the PI3K/AKT signaling pathway. In this study, we examined the growth inhibitory effects of a combinatorial therapy of adenoviral-mediated PTEN (Ad-PTEN) and LY294002 on LN229 and U251 glioma cells in vitro and on tumor xenografts in vivo. In vitro, LN229 and U251 glioma cells were treated by combinatorial therapy with Ad-PTEN and LY294002. The growth ability was determined by MTT assay. The cell cycle distribution was analyzed by flow cytometry. Cell invasive ability was analyzed by transwell invasion assay and cell apoptosis analysis via FITC-Annexin V analysis. In vivo, U251 subcutaneous glioblastoma xenograft was used to assay anti-tumor effect of combinatorial therapy with Ad-PTEN and LY294002 by mean volume of tumors, immunohistochemistry and TUNEL method. The combinatorial treatment clearly suppressed cell proliferation, arrested the cell cycle, reduced cell invasion and promoted cell apoptosis compared with the Ad-PTEN or LY294002 treatment alone. The treatment worked by inhibiting the PI3K/AKT pathway. In addition, the growth of U251 glioma xenografts treated with the combination of Ad-PTEN and LY294002 was significantly inhibited compared with those treated with Ad-PTEN or LY294002 alone. Our data indicated that the combination of Ad-PTEN and LY294002 effectively suppressed the malignant growth of human glioma cells in vitro and in tumor xenografts, suggesting a promising new approach for glioma gene therapy that warrants further investigation.

Regulation of DNA repair mechanism in human glioma xenograft cells both in vitro and in vivo in nude mice.

PubMed

Ponnala, Shivani; Veeravalli, Krishna Kumar; Chetty, Chandramu; Dinh, Dzung H; Rao, Jasti S

2011-01-01

Glioblastoma Multiforme (GBM) is the most lethal form of brain tumor. Efficient DNA repair and anti-apoptotic mechanisms are making glioma treatment difficult. Proteases such as MMP9, cathepsin B and urokinase plasminogen activator receptor (uPAR) are over expressed in gliomas and contribute to enhanced cancer cell proliferation. Non-homologous end joining (NHEJ) repair mechanism plays a major role in double strand break (DSB) repair in mammalian cells. Here we show that silencing MMP9 in combination with uPAR/cathepsin B effects NHEJ repair machinery. Expression of DNA PKcs and Ku70/80 at both mRNA and protein levels in MMP9-uPAR (pMU) and MMP9-cathepsin B (pMC) shRNA-treated glioma xenograft cells were reduced. FACS analysis showed an increase in apoptotic peak and proliferation assays revealed a significant reduction in the cell population in pMU- and pMC-treated cells compared to untreated cells. We hypothesized that reduced NHEJ repair led to DSBs accumulation in pMU- and pMC-treated cells, thereby initiating cell death. This hypothesis was confirmed by reduced Ku70/Ku80 protein binding to DSB, increased comet tail length and elevated γH2AX expression in treated cells compared to control. Immunoprecipitation analysis showed that EGFR-mediated lowered DNA PK activity in treated cells compared to controls. Treatment with pMU and pMC shRNA reduced the expression of DNA PKcs and ATM, and elevated γH2AX levels in xenograft implanted nude mice. Glioma cells exposed to hypoxia and irradiation showed DSB accumulation and apoptosis after pMU and pMC treatments compared to respective controls. Our results suggest that pMU and pMC shRNA reduce glioma proliferation by DSB accumulation and increase apoptosis under normoxia, hypoxia and in combination with irradiation. Considering the radio- and chemo-resistant cancers favored by hypoxia, our study provides important therapeutic potential of MMP9, uPAR and cathepsin B shRNA in the treatment of glioma from clinical stand point.

Regulation of DNA Repair Mechanism in Human Glioma Xenograft Cells both In Vitro and In Vivo in Nude Mice

PubMed Central

Ponnala, Shivani; Veeravalli, Krishna Kumar; Chetty, Chandramu; Dinh, Dzung H.; Rao, Jasti S.

2011-01-01

Background Glioblastoma Multiforme (GBM) is the most lethal form of brain tumor. Efficient DNA repair and anti-apoptotic mechanisms are making glioma treatment difficult. Proteases such as MMP9, cathepsin B and urokinase plasminogen activator receptor (uPAR) are over expressed in gliomas and contribute to enhanced cancer cell proliferation. Non-homologous end joining (NHEJ) repair mechanism plays a major role in double strand break (DSB) repair in mammalian cells. Methodology/Principal Findings Here we show that silencing MMP9 in combination with uPAR/cathepsin B effects NHEJ repair machinery. Expression of DNA PKcs and Ku70/80 at both mRNA and protein levels in MMP9-uPAR (pMU) and MMP9-cathepsin B (pMC) shRNA-treated glioma xenograft cells were reduced. FACS analysis showed an increase in apoptotic peak and proliferation assays revealed a significant reduction in the cell population in pMU- and pMC-treated cells compared to untreated cells. We hypothesized that reduced NHEJ repair led to DSBs accumulation in pMU- and pMC-treated cells, thereby initiating cell death. This hypothesis was confirmed by reduced Ku70/Ku80 protein binding to DSB, increased comet tail length and elevated γH2AX expression in treated cells compared to control. Immunoprecipitation analysis showed that EGFR-mediated lowered DNA PK activity in treated cells compared to controls. Treatment with pMU and pMC shRNA reduced the expression of DNA PKcs and ATM, and elevated γH2AX levels in xenograft implanted nude mice. Glioma cells exposed to hypoxia and irradiation showed DSB accumulation and apoptosis after pMU and pMC treatments compared to respective controls. Conclusion/Significance Our results suggest that pMU and pMC shRNA reduce glioma proliferation by DSB accumulation and increase apoptosis under normoxia, hypoxia and in combination with irradiation. Considering the radio- and chemo-resistant cancers favored by hypoxia, our study provides important therapeutic potential of MMP9, uPAR and cathepsin B shRNA in the treatment of glioma from clinical stand point. PMID:22022560

[Vitamin K3-induced activation of molecular oxygen in glioma cells].

PubMed

Krylova, N G; Kulagova, T A; Semenkova, G N; Cherenkevich, S N

2009-01-01

It has been shown by the method of fluorescent analysis that the rate of hydrogen peroxide generation in human U251 glioma cells under the effect of lipophilic (menadione) or hydrophilic (vikasol) analogues of vitamin K3 was different. Analyzing experimental data we can conclude that menadione underwent one- and two-electron reduction by intracellular reductases in glioma cells. Reduced forms of menadione interact with molecular oxygen leading to reactive oxygen species (ROS) generation. The theoretical model of ROS generation including two competitive processes of one- and two-electron reduction of menadione has been proposed. Rate constants of ROS generation mediated by one-electron reduction process have been estimated.

Vitamin K1 enhances sorafenib-induced growth inhibition and apoptosis of human malignant glioma cells by blocking the Raf/MEK/ERK pathway

PubMed Central

2012-01-01

Background The combined effects of anticancer drugs with nutritional factors against tumor cells have been reported previously. This study characterized the efficacy and possible mechanisms of the combination of sorafenib and vitamin K1 (VK1) on glioma cell lines. Methods We examined the effects of sorafenib, VK1 or their combination on the proliferation and apoptosis of human malignant glioma cell lines (BT325 and U251) by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay, flow cytometry and 4′,6-diamidino-2-phenylindole (DAPI) assay. The signaling pathway changes were detected by western blotting. Results Sorafenib, as a single agent, showed antitumor activity in a dose-dependent manner in glioma cells, but the effects were more pronounced when used in combination with VK1 treatment. Sorafenib in combination with VK1 treatment produced marked potentiation of growth inhibition and apoptosis, and reduced expression of phospho-mitogen-activated protein kinase kinase (MEK) and phospho-extracellular signal-regulated kinase (ERK). Furthermore, the expression levels of antiapoptotic proteins Bcl-2 and Mcl-1 were significantly reduced. Conclusions Our findings indicated that VK1 enhanced the cytotoxicity effect of sorafenib through inhibiting the Raf/MEK/ERK signaling pathway in glioma cells, and suggested that sorafenib in combination with VK1 maybe a new therapeutic option for patients with gliomas. PMID:22520038

circ-SHKBP1 Regulates the Angiogenesis of U87 Glioma-Exposed Endothelial Cells through miR-544a/FOXP1 and miR-379/FOXP2 Pathways.

PubMed

He, Qianru; Zhao, Lini; Liu, Yunhui; Liu, Xiaobai; Zheng, Jian; Yu, Hai; Cai, Heng; Ma, Jun; Liu, Libo; Wang, Ping; Li, Zhen; Xue, Yixue

2018-03-02

Circular RNAs (circRNAs) are a type of endogenous non-coding RNAs, which have been considered to mediate diverse tumorigenesis including angiogenesis. The present study aims to elucidate the potential role and molecular mechanism of circ-SHKBP1 in regulating the angiogenesis of U87 glioma-exposed endothelial cells (GECs). The expression of circ-SHKBP1, but not linear SHKBP1, was significantly upregulated in GECs compared with astrocyte-exposed endothelial cells (AECs). circ-SHKBP1 knockdown inhibited the viability, migration, and tube formation of GECs dramatically. The expressions of miR-379/miR-544a were downregulated in GECs, and circ-SHKBP1 functionally targeted miR-544a/miR-379 in an RNA-induced silencing complex (RISC) manner. Dual-luciferase reporter assay demonstrated that forkhead box P1/P2 (FOXP1/FOXP2) were targets of miR-544a/miR-379. The expressions of FOXP1/FOXP2 were upregulated in GECs, and silencing of FOXP1/FOXP2 inhibited the viability, migration, and tube formation of GECs. Meanwhile, FOXP1/FOXP2 promoted angiogenic factor with G patch and FHA domains 1 (AGGF1) expression at the transcriptional level. Furthermore, knockdown of AGGF1 suppressed the viability, migration, and tube formation of GECs via phosphatidylinositol 3-kinase (PI3K)/AKT and extracellular signal-regulated kinase (ERK)1/2 pathways. Taken together, the present study demonstrated that circ-SHKBP1 regulated the angiogenesis of GECs through miR-544a/FOXP1 and miR-379/FOXP2 pathways, and these findings might provide a potential target and effective strategy for combined therapy of gliomas. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Propofol depresses cisplatin cytotoxicity via the inhibition of gap junctions.

PubMed

Zhang, Yuan; Wang, Xiyan; Wang, Qin; Ge, Hui; Tao, Liang

2016-06-01

The general anesthetic, propofol, affects chemotherapeutic activity, however, the mechanism underlying its effects remains to be fully elucidated. Our previous study showed that tramadol and flurbiprofen depressed the cytotoxicity of cisplatin via the inhibition of gap junction (GJ) intercellular communication (GJIC) in connexin (Cx)32 HeLa cells. The present study investigated whether the effects of propofol on the cytotoxicity of cisplatin were mediated by GJ in U87 glioma cells and Cx26‑transfected HeLa cells. Standard colony formation assay was used to determine the cytotoxicity of cisplatin. Parachute dye coupling assay was used to measure GJ function, and western blot analysis was used to determine the expression levels of Cx32. The results revealed that exposure of the U87 glioma cells and the Cx26-transfected HeLa cells to cisplatin for 1 h reduced clonogenic survival in low density cultures (without GJs) and high density cultures (with GJs). However, the toxic effect was higher in the high density culture. In addition, pretreatment of the cells with propofol significantly reduced cisplatin‑induced cytotoxicity, but only in the presence of functional GJs. Furthermore, propofol significantly inhibited dye coupling through junctional channels, and a long duration of exposure of the cells to propofol downregulated the expression levels of Cx43 and Cx26. These results demonstrated that the inhibition of GJIC by propofol affected the therapeutic efficacy of chemotherapeutic drugs. The present study provides evidence of a novel mechanism underlying the effects of analgesics in counteracting chemotherapeutic efficiency.

A Novel Ideal Radionuclide Imaging System for Non-invasively Cell Monitoring built on Baculovirus Backbone by Introducing Sleeping Beauty Transposon

PubMed Central

Lv, Jing; Pan, Yu; Ju, Huijun; Zhou, Jinxin; Cheng, Dengfeng; Shi, Hongcheng; Zhang, Yifan

2017-01-01

Sleeping Beauty (SB) transposon is an attractive tool in stable transgene integration both in vitro and in vivo; and we introduced SB transposon into recombinant sodium-iodide symporter baculovirus system (Bac-NIS system) to facilitate long-term expression of recombinant sodium-iodide symporter. In our study, two hybrid baculovirus systems (Bac-eGFP-SB-NeoR and Bac-NIS-SB-NeoR) were successfully constructed and used to infect U87 glioma cells. After G418 selection screening, the Bac-eGFP-SB-NeoR-U87 cells remained eGFP positive, at the 18th and 196th day post transfection (96.03 ± 0.21% and 97.43 ± 0.81%), while eGFP positive population declined significantly at 18 days in cells transfected with unmodified baculovirus construct. NIS gene expression by Bac-NIS-SB-NeoR-U87 cells was also maintained for 28 weeks as determined by radioiodine uptake assay, reverse transcription-polymerase chain reaction (RT-PCR) and Western Blot (WB) assay. When transplanted in mice, Bac-NIS-SB-NeoR-U87 cells also expressed NIS gene stably as monitored by SPECT imaging for 43 days until the tumor-bearing mice were sacrificed. Herein, we showed that incorporation of SB in Bac-NIS system (hybrid Bac-NIS-SB-NeoR) can achieve a long-term transgene expression and can improve radionuclide imaging in cell tracking and monitoring in vivo. PMID:28262785

Radiosensitization of human glioma cells by tamoxifen is associated with the inhibition of PKC-ι activity in vitro.

PubMed

Yang, Lei; Yuan, Xiaopeng; Wang, Jie; Gu, Cheng; Zhang, Haowen; Yu, Jiahua; Liu, Fenju

2015-07-01

The present study aimed to investigate the radiosensitizing effects of tamoxifen (TAM), a non-steroidal anti-estrogen drug, in human glioma A172 and U251 cells in vitro . A colony-forming assay revealed that TAM enhances radiosensitivity in A172 and U251 cells. Treatment with TAM also increased the percentage of apoptotic cells subsequent to ionizing radiation, and increased the expression of apoptotic markers, including cleaved caspase-3 and poly(ADP-ribose) polymerase. Ionizing radiation induced G2/M phase arrest, which was alleviated within 24 h when the radiation-induced DNA damage was repaired. However, flow cytometry analysis revealed that TAM treatment delayed the recovery of cell cycle progression. Additional examination demonstrated that TAM-mediated protein kinase C-ι (PKC-ι) inhibition may lead to the activation of pro-apoptotic B-cell lymphoma 2-associated death promoter, and the dephosphorylation of cyclin-dependent kinase 7, resulting in increased cell apoptosis and sustained G2/M phase arrest following exposure to radiation. The present data indicate that the radiosensitizing effects of TAM on glioma cells are partly due to the inhibition of PKC-ι activity in vitro .

Near-infrared light-activated red-emitting upconverting nanoplatform for T1-weighted magnetic resonance imaging and photodynamic therapy.

PubMed

Tang, Xiang-Long; Wu, Jun; Lin, Ben-Lan; Cui, Sheng; Liu, Hong-Mei; Yu, Ru-Tong; Shen, Xiao-Dong; Wang, Ting-Wei; Xia, Wei

2018-05-12

Photodynamic therapy (PDT) has increasingly become an efficient and attractive cancer treatment modality based on reactive oxygen species (ROS) that can induce tumor death after irradiation with ultraviolet or visible light. Herein, to overcome the limited tissue penetration in traditional PDT, a novel near-infrared (NIR) light-activated NaScF 4 : 40% Yb, 2% Er@CaF 2 upconversion nanoparticle (rUCNP) is successfully designed and synthesized. Chlorin e6, a photosensitizer and a chelating agent for Mn 2+ , is loaded into human serum albumin (HSA) that further conjugates onto rUCNPs. To increase the ability to target glioma tumor, an acyclic Arg-Gly-Asp peptide (cRGDyK) is linked to rUCNPs@HSA(Ce6-Mn). This nanoplatform enables efficient adsorption and conversion of NIR light (980 nm) into bright red emission (660 nm), which can trigger the photosensitizer Ce6-Mn complex for PDT and T 1 -weighted magnetic resonance imaging (T 1 -weighted MRI) for glioma diagnosis. Our in vitro and in vivo experiments demonstrate that NIR light-activated and glioma tumor-targeted PDT can generate large amounts of intracellular ROS that induce U87 cell apoptosis and suppress glioma tumor growth owing to the deep tissue penetration of irradiated light and excellent tumor-targeting ability. Thus, this nanoplatform holds potential for applications in T 1 -weighted MRI diagnosis and PDT of glioma for antitumor therapy. A near-infrared (NIR) light-activated nanoplatform for photodynamic therapy (PDT) was designed and synthesized. The Red-to-Green (R/G) ratio of NaScF 4 : 40% Yb, 2% Er almost reached 9, a value that was much higher than that of a traditional Yb/Er-codoped upconversion nanoparticle (rUCNP). By depositing a CaF 2 shell, the red-emission intensities of the rUCNPs were seven times strong as that of NaScF 4 : 40% Yb, 2% Er. The enhanced red-emitting rUCNPs could be applied in many fields such as bioimaging, controlled release, and real-time diagnosis. The nanoplatform had a strong active glioma-targeting ability, and all results achieved on subcutaneous glioma demonstrated that our NIR light-activated red-emitting upconverting nanoplatform was efficient for PDT. By loading Ce6-Mn complex into rUCNPs@HSA-RGD, the nanoplatform could be used as a T 1 -weighted magnetic resonance imaging agent for tumor diagnosis. Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

IDH1 R132H mutation regulates glioma chemosensitivity through Nrf2 pathway.

PubMed

Li, Kaishu; Ouyang, Leping; He, Mingliang; Luo, Ming; Cai, Wangqing; Tu, Yalin; Pi, Rongbiao; Liu, Anmin

2017-04-25

Numerous studies have reported that glioma patients with isocitrate dehydrogenase 1(IDH1) R132H mutation are sensitive to temozolomide treatment. However, the mechanism of IDH1 mutations on the chemosensitivity of glioma remains unclear. In this study, we investigated the role and the potential mechanism of Nrf2 in IDH1 R132H-mediated drug resistance. Wild type IDH1 (R132H-WT) and mutant IDH1 (R132H) plasmids were constructed. Stable U87 cells and U251 cells overexpressing IDH1 were generated. Phenotypic differences between IDH1-WT and IDH1 R132H overexpressing cells were evaluated using MTT, cell colony formation assay, scratch test assay and flow cytometry. Expression of IDH1 and its associated targets, nuclear factor-erythroid 2-related factor 2 (Nrf2), NAD(P)H quinine oxidoreductase 1 (NQO1), multidrug resistant protein 1 (MRP1) and p53 were analyzed. The IDH1 R132H overexpressing cells were more sensitive to temozolomide than WT and the control, and Nrf2 was significantly decreased in IDH1 R132H overexpressing cells. We found that knocking down Nrf2 could decrease resistance to temozolomide. The nuclear translocation of Nrf2 in IDH1 R132H overexpressing cells was lower than the WT and the control groups after temozolomide treatment. When compared with WT cells, NQO1 expression was reduced in IDH1 R132H cells, especially after temozolomide treatment. P53 was involved in the resistance mechanism of temozolomide mediated by Nrf2 and NQO1. Nrf2 played an important role in IDH1 R132H-mediated drug resistance. The present study provides new insight for glioma chemotherapy with temozolomide.

IDH1 R132H mutation regulates glioma chemosensitivity through Nrf2 pathway

PubMed Central

Luo, Ming; Cai, Wangqing; Tu, Yalin; Pi, Rongbiao; Liu, Anmin

2017-01-01

Purpose Numerous studies have reported that glioma patients with isocitrate dehydrogenase 1(IDH1) R132H mutation are sensitive to temozolomide treatment. However, the mechanism of IDH1 mutations on the chemosensitivity of glioma remains unclear. In this study, we investigated the role and the potential mechanism of Nrf2 in IDH1 R132H-mediated drug resistance. Methods Wild type IDH1 (R132H-WT) and mutant IDH1 (R132H) plasmids were constructed. Stable U87 cells and U251 cells overexpressing IDH1 were generated. Phenotypic differences between IDH1-WT and IDH1 R132H overexpressing cells were evaluated using MTT, cell colony formation assay, scratch test assay and flow cytometry. Expression of IDH1 and its associated targets, nuclear factor-erythroid 2-related factor 2 (Nrf2), NAD(P)H quinine oxidoreductase 1 (NQO1), multidrug resistant protein 1 (MRP1) and p53 were analyzed. Results The IDH1 R132H overexpressing cells were more sensitive to temozolomide than WT and the control, and Nrf2 was significantly decreased in IDH1 R132H overexpressing cells. We found that knocking down Nrf2 could decrease resistance to temozolomide. The nuclear translocation of Nrf2 in IDH1 R132H overexpressing cells was lower than the WT and the control groups after temozolomide treatment. When compared with WT cells, NQO1 expression was reduced in IDH1 R132H cells, especially after temozolomide treatment. P53 was involved in the resistance mechanism of temozolomide mediated by Nrf2 and NQO1. Conclusions Nrf2 played an important role in IDH1 R132H-mediated drug resistance. The present study provides new insight for glioma chemotherapy with temozolomide. PMID:28427200

Human Atg8-cardiolipin interactions in mitophagy: Specific properties of LC3B, GABARAPL2 and GABARAP.

PubMed

Antón, Zuriñe; Landajuela, Ane; Hervás, Javier H; Montes, L Ruth; Hernández-Tiedra, Sonia; Velasco, Guillermo; Goñi, Felix M; Alonso, Alicia

2016-12-01

The phospholipid cardiolipin (CL) has been proposed to play a role in selective mitochondrial autophagy, or mitophagy. CL externalization to the outer mitochondrial membrane would act as a signal for the human Atg8 ortholog subfamily, MAP1LC3 (LC3). The latter would mediate both mitochondrial recognition and autophagosome formation, ultimately leading to removal of damaged mitochondria. We have applied quantitative biophysical techniques to the study of CL interaction with various Atg8 human orthologs, namely LC3B, GABARAPL2 and GABARAP. We have found that LC3B interacts preferentially with CL over other di-anionic lipids, that CL-LC3B binding occurs with positive cooperativity, and that the CL-LC3B interaction relies only partially on electrostatic forces. CL-induced increased membrane fluidity appears also as an important factor helping LC3B to bind CL. The LC3B C terminus remains exposed to the hydrophilic environment after protein binding to CL-enriched membranes. In intact U87MG human glioblastoma cells rotenone-induced autophagy leads to LC3B translocation to mitochondria and subsequent delivery of mitochondria to lysosomes. We have also observed that GABARAP, but not GABARAPL2, interacts with CL in vitro. However neither GABARAP nor GABARAPL2 were translocated to mitochondria in rotenone-treated U87MG cells. Thus the various human Atg8 orthologs might play specific roles in different autophagic processes.

Matched rabbit monoclonal antibodies against αv-series integrins reveal a novel αvβ3-LIBS epitope, and permit routine staining of archival paraffin samples of human tumors

PubMed Central

Goodman, Simon L.; Grote, Hans Juergen; Wilm, Claudia

2012-01-01

Summary The relationship between integrin expression and function in pathologies is often contentious as comparisons between human pathological expression and expression in cell lines is difficult. In addition, the expression of even integrins αvβ6 and αvβ8 in tumor cell lines is not comprehensively documented. Here, we describe rabbit monoclonal antibodies (RabMabs) against the extracellular domains of αv integrins that react with both native integrins and formalin fixed, paraffin embedded (FFPE) human tissues. These RabMabs, against αvβ3 (EM22703), αvβ5 (EM09902), αvβ6 (EM05201), αvβ8 (EM13309), and pan-αv (EM01309), recognize individual integrin chains in Western blots and in flow cytometry. EM22703 detected a ligand-induced binding site (LIBS), reporting an epitope enhanced by the binding of an RGD-peptide to αvβ3. αvβ8 was rarely expressed in human tumor specimens, and weakly expressed in non-small-cell lung carcinoma (NSCLC). However, ovarian carcinoma cell lines expressed αvβ8, as did some melanoma cells, whereas U87MG glioma lacked αvβ8 expression. We observed an unexpected strong expression of αvβ6 in tumor samples of invasive ductal breast adenoma, colorectal carcinoma (CRC), and NSCLC. αvβ3 was strongly expressed in some invasive NSCLC cohorts. Interestingly, PC3 prostate cell and human prostate tumors did not express αvβ3. The RabMabs stained plasma membranes in FFPE-immunohistochemistry (IHC) samples of tumor cell lines from lung, ovary, colon, prostate, squamous cell carcinoma of head and neck (SCCHN), breast, and pancreas carcinomas. The RabMabs are unique tools for probing αv integrin biology, and suggest that especially αvβ6 and αvβ8 biologies still have much to reveal. PMID:23213423

Near-Infrared Fluorescent Deoxyglucose Analog for Tumor Optical Imaging in Cell Culture and in Living Mice

PubMed Central

Cheng, Zhen; Levi, Jelena; Xiong, Zhengming; Gheysens, Olivier; Keren, Shay; Chen, Xiaoyuan; Gambhir, Sanjiv Sam

2011-01-01

2-deoxy-2-[18F]fluoro-d-glucose ([18F]FDG) has extensively been used for clinical diagnosis, staging and therapy monitoring of cancer and other diseases. Non-radioactive glucose analogs enabling the screening of the glucose metabolic rate of tumors are of particular interest for anticancer drug development. A non-radioactive fluorescent deoxyglucose analog may have many applications for both imaging of tumors and monitoring therapeutic efficacy of drugs in living animals and may eventually translate to clinical applications. We found that a fluorescent 2-deoxyglucose analog, 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxy-d-glucose (2-NBDG) can be delivered in several tumor cells via the glucose transporters (GLUTs). We therefore conjugated d-glucosamine with a near-infrared (NIR) fluorphor Cy5.5 and tested the feasibility of Cy5.5-d-glucosamine conjugate (Cy5.5-2DG) for NIR fluorescence imaging of tumors in a pre-clinical xenograft animal model. Cy5.5-2DG was prepared by conjugating Cy5.5 monofunctional N-hydroxysuccinimide ester (Cy5.5-NHS) and d-glucosamine followed by high-performance liquid chromatography purification. The accumulation of Cy5.5-2DG and Cy5.5-NHS in different tumor cell lines at 37 °C and 4 °C were imaged using a fluorescence microscope. Tumor targeting and retention of Cy5.5-2DG and Cy5.5-NHS in a subcutaneous U87MG glioma and A375M melanoma tumor model were evaluated and quantified by a Xenogen IVIS 200 optical cooled charged-coupled device system. Fluorescence microscopy imaging shows that Cy5.5-2DG and Cy5.5-NHS are taken up and trapped by a variety of tumor cell lines at 37 °C incubation, while they exhibit marginal uptake at 4 °C. The tumor cell uptake of Cy5.5-2DG can not be blocked by the 50 mM d-glucose, suggesting that Cy5.5-2DG may not be delivered in tumor cells by GLUTs. U87MG and A375M tumor localization were clearly visualized in living mice with both NIR fluorescent probes. Tumor/muscle contrast was clearly visible as early as 30 min post-injection, and the highest U87MG tumor/muscle ratio of 2.81 ± 0.10, 3.34 ± 0.23 were achieved 24 hours post-injection for Cy5.5-2DG and Cy5.5-NHS, respectively. While as a comparison, the micro-positron emission tomography imaging study shows that [18F]FDG preferentially localize to the U87MG tumor, with resulting tumor/muscle ratios ranging from 3.89 to 4.08 after 30 min to 2 h post-administration of the probe. In conclusion, the NIR fluorescent glucose analog, Cy5.5-2DG and Cy5.5-NHS both demonstrate tumor targeting abilities in cell culture and in living mice. More studies are warranted to further explore their application for optical tumor imaging. In order to develop NIR glucose analog with ability to targeting GLUTs/hexokinase, it is highly important to select NIR dyes with reasonable molecular size. PMID:16704203

Curcumin Encapsulated into Methoxy Poly(Ethylene Glycol) Poly(ε-Caprolactone) Nanoparticles Increases Cellular Uptake and Neuroprotective Effect in Glioma Cells.

PubMed

Marslin, Gregory; Sarmento, Bruno Filipe Carmelino Cardoso; Franklin, Gregory; Martins, José Alberto Ribeiro; Silva, Carlos Jorge Ribeiro; Gomes, Andreia Ferreira Castro; Sárria, Marisa Passos; Coutinho, Olga Maria Fernandes Pereira; Dias, Alberto Carlos Pires

2017-03-01

Curcumin is a natural polyphenolic compound isolated from turmeric ( Curcuma longa ) with well-demonstrated neuroprotective and anticancer activities. Although curcumin is safe even at high doses in humans, it exhibits poor bioavailability, mainly due to poor absorption, fast metabolism, and rapid systemic elimination. To overcome these issues, several approaches, such as nanoparticle-mediated targeted delivery, have been undertaken with different degrees of success. The present study was conducted to compare the neuroprotective effect of curcumin encapsulated in poly( ε -caprolactone) and methoxy poly(ethylene glycol) poly( ε -caprolactone) nanoparticles in U251 glioblastoma cells. Prepared nanoparticles were physically characterized by laser doppler anemometry, transmission electron microscopy, and X-ray diffraction. The results from laser doppler anemometry confirmed that the size of poly( ε -caprolactone) and poly(ethylene glycol) poly( ε -caprolactone) nanoparticles ranged between 200-240 nm for poly( ε -caprolactone) nanoparticles and 30-70 nm for poly(ethylene glycol) poly( ε -caprolactone) nanoparticles, and transmission electron microscopy images revealed their spherical shape. Treatment of U251 glioma cells and zebrafish embryos with poly( ε -caprolactone) and poly(ethylene glycol) poly( ε -caprolactone) nanoparticles loaded with curcumin revealed efficient cellular uptake. The cellular uptake of poly(ethylene glycol) poly( ε -caprolactone) nanoparticles was higher in comparison to poly( ε -caprolactone) nanoparticles. Moreover, poly(ethylene glycol) poly( ε -caprolactone) di-block copolymer-loaded curcumin nanoparticles were able to protect the glioma cells against tBHP induced-oxidative damage better than free curcumin. Together, our results show that curcumin-loaded poly(ethylene glycol) poly( ε -caprolactone) di-block copolymer nanoparticles possess significantly stronger neuroprotective effect in U251 human glioma cells compared to free curcumin and curcumin-loaded poly( ε -caprolactone) nanoparticles. Georg Thieme Verlag KG Stuttgart · New York.

Quantification of Tumor Vascular Permeability and Blood Volume by Positron Emission Tomography

PubMed Central

Chen, Haojun; Tong, Xiao; Lang, Lixin; Jacobson, Orit; Yung, Bryant C.; Yang, Xiangyu; Bai, Ruiliang; Kiesewetter, Dale O.; Ma, Ying; Wu, Hua; Niu, Gang; Chen, Xiaoyuan

2017-01-01

Purpose: Evans Blue (EB) is an azo dye that binds quantitatively with serum albumin. With an albumin binding, NOTA conjugated truncated Evan's blue (NEB) dye derived PET tracer, we aimed to establish a strategy for evaluating vascular permeability in malignant tumors via non-invasive PET. Experimental design: Sixty-minute dynamic PET using [18F]FAl-NEB was performed in three xenograft tumor models including INS-1 rat insulinoma, UM-SCC-22B human head and neck carcinoma and U-87 MG human glioblastoma. Tumor vascular permeability was quantified by the difference of the slopes between tumor and blood time-activity curve (TACs, expressed as Ps). The method was further substantiated by EB extraction and colorimetric assay and correlates with that calculated from dynamic contrast enhanced magnetic resonance imaging (DCE-MRI). The changes in tumor vasculature at different time points were assessed with NEB PET in U-87 MG and UM-SCC-22B tumor models after treatment with bevacizumab or doxorubicin. Result: The Ps values calculated from tumor and blood TACs from multiple time-point static images are consistent with those from dynamic images. Moreover, the Ps showed a positive and significant correlation with extracted EB concentration and KPS-MRI generated from DCE-MRI, which further confirmed the soundness of this methodology. The antiangiogenic effect of bevacizumab could be revealed by NEB PET in U-87 MG tumors as early as 8 hrs after therapy, demonstrated by a substantial decrease of Ps. On the contrary, there was no significant change of Ps in bevacizumab treated UM-SCC-22B tumors, compared with control group. However, the significant changes of Pswere overestimated in doxorubicin treated UM-SCC-22B tumors. Conclusions: We successfully developed a relatively convenient and novel strategy to evaluate vascular permeability and blood volume using NEB PET. This method will be advantageous in evaluating vascular permeability, promoting drug delivery, and monitoring tumor response to therapeutics that affect tumor angiogenesis. PMID:28744320

Quantitative correlational study of microbubble-enhanced ultrasound imaging and magnetic resonance imaging of glioma and early response to radiotherapy in a rat model.

PubMed

Yang, Chen; Lee, Dong-Hoon; Mangraviti, Antonella; Su, Lin; Zhang, Kai; Zhang, Yin; Zhang, Bin; Li, Wenxiao; Tyler, Betty; Wong, John; Wang, Ken Kang-Hsin; Velarde, Esteban; Zhou, Jinyuan; Ding, Kai

2015-08-01

Radiotherapy remains a major treatment method for malignant tumors. Magnetic resonance imaging (MRI) is the standard modality for assessing glioma treatment response in the clinic. Compared to MRI, ultrasound imaging is low-cost and portable and can be used during intraoperative procedures. The purpose of this study was to quantitatively compare contrast-enhanced ultrasound (CEUS) imaging and MRI of irradiated gliomas in rats and to determine which quantitative ultrasound imaging parameters can be used for the assessment of early response to radiation in glioma. Thirteen nude rats with U87 glioma were used. A small thinned skull window preparation was performed to facilitate ultrasound imaging and mimic intraoperative procedures. Both CEUS and MRI with structural, functional, and molecular imaging parameters were performed at preradiation and at 1 day and 4 days postradiation. Statistical analysis was performed to determine the correlations between MRI and CEUS parameters and the changes between pre- and postradiation imaging. Area under the curve (AUC) in CEUS showed significant difference between preradiation and 4 days postradiation, along with four MRI parameters, T2, apparent diffusion coefficient, cerebral blood flow, and amide proton transfer-weighted (APTw) (all p < 0.05). The APTw signal was correlated with three CEUS parameters, rise time (r = - 0.527, p < 0.05), time to peak (r = - 0.501, p < 0.05), and perfusion index (r = 458, p < 0.05). Cerebral blood flow was correlated with rise time (r = - 0.589, p < 0.01) and time to peak (r = - 0.543, p < 0.05). MRI can be used for the assessment of radiotherapy treatment response and CEUS with AUC as a new technique and can also be one of the assessment methods for early response to radiation in glioma.

Quantitative metabolome analysis profiles activation of glutaminolysis in glioma with IDH1 mutation.

PubMed

Ohka, Fumiharu; Ito, Maki; Ranjit, Melissa; Senga, Takeshi; Motomura, Ayako; Motomura, Kazuya; Saito, Kaori; Kato, Keiko; Kato, Yukinari; Wakabayashi, Toshihiko; Soga, Tomoyoshi; Natsume, Atsushi

2014-06-01

Isocitrate dehydrogenase 1 (IDH1), which localizes to the cytosol and peroxisomes, catalyzes the oxidative decarboxylation of isocitrate to α-ketoglutarate (α-KG) and in parallel converts NADP(+) to NADPH. IDH1 mutations are frequently detected in grades 2-4 gliomas and in acute myeloid leukemias (AML). Mutations of IDH1 have been identified at codon 132, with arginine being replaced with histidine in most cases. Mutant IDH1 gains novel enzyme activity converting α-KG to D-2-hydroxyglutarate (2-HG) which acts as a competitive inhibitor of α-KG. As a result, the activity of α-KG-dependent enzyme is reduced. Based on these findings, 2-HG has been proposed to be an oncometabolite. In this study, we established HEK293 and U87 cells that stably expressed IDH1-WT and IDH1-R132H and investigated the effect of glutaminase inhibition on cell proliferation with 6-diazo-5-oxo-L-norleucine (DON). We found that cell proliferation was suppressed in IDH1-R132H cells. The addition of α-KG restored cell proliferation. The metabolic features of 33 gliomas with wild type IDH1 (IDH1-WT) and with IDH1-R132H mutation were examined by global metabolome analysis using capillary electrophoresis time-of-flight mass spectrometry (CE-TOFMS). We showed that the 2-HG levels were highly elevated in gliomas with IDH1-R132H mutation. Intriguingly, in gliomas with IDH1-R132H, glutamine and glutamate levels were significantly reduced which implies replenishment of α-KG by glutaminolysis. Based on these results, we concluded that glutaminolysis is activated in gliomas with IDH1-R132H mutation and that development of novel therapeutic approaches targeting activated glutaminolysis is warranted.

Lipid nanocapsule functionalization by lipopeptides derived from human papillomavirus type-16 capsid for nucleic acid delivery into cancer cells.

PubMed

Weyland, M; Griveau, A; Bejaud, J; Benoit, J-P; Coursaget, P; Garcion, E

2013-10-01

Plasmid DNA (pDNA) and small interfering RNAs (siRNAs) are very useful tools for the treatment of cancer. However, pDNA and siRNAs efficacy is restricted by their negative charge and susceptibility to degradation by endonucleases that prevent them penetrating tissue and cellular barriers such as the plasma and endolysosomal membranes. Viral vectors have some advantages but their use is largely limited by their immunogenicity. On the other hand, synthetic nanoparticles have advantage of being relatively non-immunogenic but their ability to deliver nucleic acids remains less efficient than their viral counterparts. The present study is focussed on the development and evaluation of biomimetic lipid nanocapsules (LNCs) functionalized with a L1 papillomavirus type-16 capsid-derived lipopeptide on their surface, for transfection of U87MG glioma cells and Caco-2 colorectal adenocarcinoma cells with pDNA or siRNAs. Since the L1-peptide has been described as a nuclear localization signal able to complex with nucleic acids and bind to heparan sulfate on the cell surface, the structure and function of L1-peptide bound to LNCs (L1-LNCs) were investigated. Although L1-LNCs were shown to complex with both pDNA and siRNAs, the pDNA-L1-LNC complexes showed only weak transfection efficiency. In contrast, siRNA-L1-LNC complexes appeared as effective repressors of targeted messengers. Copyright © 2013 Elsevier B.V. All rights reserved.

Tumor-Associated Macrophages Are Predominant Carriers of Cyclodextrin-Based Nanoparticles into Gliomas

PubMed Central

Alizadeh, Darya; Zhang, Leying; Hwang, Jungyeon; Schluep, Thomas; Badie, Behnam

2009-01-01

The goal of this study was to evaluate the mechanism of cyclodextrin-based nanoparticle (CDP-NP) uptake into a murine glioma model. Using mixed in vitro culture systems, we demonstrated that CDP-NP was preferentially taken up by BV2 and N9 microglia (MG) cells as compared to GL261 glioma cells. Fluorescent microscopy and flow cytometry analysis of intracranial (i.c.) GL261 gliomas confirmed these findings and demonstrated a predominant CDP-NP uptake by macrophages (MP) and MG within and around the tumor site. Interestingly, in mice bearing bilateral i.c. tumor, MG and MP carrying CDP-NP were able to migrate to the contralateral tumors. In conclusion, these studies better characterize the cellular distribution of CDP-NP in i.c. tumors and demonstrate that MP and MG could potentially be used as nanoparticle drug carriers into malignant brain tumors. PMID:19836468

Regulation of IL-10 expression by upstream stimulating factor (USF-1) in glioma-associated microglia.

PubMed

Zhang, Leying; Handel, Michelle Van; Schartner, Jill M; Hagar, Aaron; Allen, Grant; Curet, Marjorie; Badie, Behnam

2007-03-01

Understanding the local CNS immune response to neoplasms is essential in the development of immune-based treatments for malignant brain tumors. Using rodent glioma models, we have recently found tumor-associated microglia/macrophages (MG/MP) to be less responsive to known MG/MP activators such as CpG, LPS and IFN-gamma. To understand the mechanism of MG/MP suppression, nuclear extracts from rodent intracranial C6 gliomas, C6 glioma-associated MG/MP, normal brain, and normal MG/MP were obtained and studied using Electrophoretic Mobility Shift Assay (EMSA). Among the nuclear factors studied (AP-1, IRF, USF-1 and Stat-1) only USF-1, which is constitutively expressed in most cells, was down-regulated in tumor-associated MG/MP, but not normal MG/MP. Because tumor-associated MG/MP had higher expression of IL-10 (but not TNF-alpha or TGF-beta), we evaluated the role of USF-1 on IL-10 expression. siRNA mediated inhibition of USF-1 expression in primary MG/MP cultures resulted in up-regulation of IL-10 mRNA but not TNF-alpha or TGF-beta. These findings suggest that USF-1 may play a role in IL-10 regulation in MG/MP in brain tumors.

Nrf2 suppresses the function of dendritic cells to facilitate the immune escape of glioma cells.

PubMed

Wang, Jialiang; Liu, Peng; Xin, Shaoyan; Wang, Zongbao; Li, Jun

2017-11-15

Nrf2 is presented in dendritic cells (DCs) and contributes to the maintenance of redox homeostasis. However, the expression pattern and function of Nrf2 in the maturation of DCs in the glioma-infiltrated microenvironment remain unrevealed. Our study aims to investigate the roles of Nrf2 in glioma cell-tamed DCs and their impact on the downstream T cell proliferation and cytotoxicity to glioma cells. It was showed that the inducible maturation of DCs was significantly suppressed after stimulation with tumor-conditioned medium (TCM) prepared from glioma cells (LN-18 and U118MG), as suggested by the decreased CD80, CD86 and IL-12 p70 expression and higher levels of IL-10 than the normal astrocyte medium treated DCs. Moreover, the TCM-exposed DCs had significantly increased expression and transcriptional activity of Nrf2 compared to the negative control. Nrf2 inhibition in DC cells substantially antagonized the inhibitory effects of TCM on the maturation and activation of DC cells, reflected by the elevated maturation markers and IL-12 p70. We further confirmed that Nrf2 inhibition in TCM-exposed DC cells promoted the proliferation of T cells as evaluated by the CFSE-labeled assay and Th1 response shown by the elevated production of IFN-γ. The cytotoxic T lymphocyte assay revealed that Nrf2 genetic suppression in DC cells greatly enhanced the capacity of T cells in the cytotoxicity to glioma cells dependent on the E:T ratio. Collectively, our study demonstrated that Nrf2 inhibition in DCs in glioma-exposed microenvironment could enhance the maturation of DCs and the subsequent activation of T cells and their cytotoxicity on glioma cells. Copyright © 2017. Published by Elsevier Inc.

MicroRNA-223 Enhances Radiation Sensitivity of U87MG Cells In Vitro and In Vivo by Targeting Ataxia Telangiectasia Mutated

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liang, Liping; Zhu, Ji; Zaorsky, Nicholas G.

2014-03-15

Purpose: Ataxia telangiectasia mutated (ATM) protein is important in the DNA damage response because it repairs radiation-induced damage in cancers. We examined the effect of microRNA-223 (miR-223), a regulator of ATM expression, on radiation sensitivity of cancer cells. Methods and Materials: Human embryonic kidney 293 T (293T) cells were infected with pLL3.7-miR-223 plasmid to generate the pLL3.7-miR-223 and -empty virus (EV) lentivirus (miR-223 and EV). A dual luciferase assay in which the reporter contained wild-type 3′ untranslated region (UTR) of ATM was performed. U87MG cells were infected with miR-223 or EV to establish the overexpressed stable cell lines (U87-223 or U87-EV, respectively).more » Cells were irradiated in vitro, and dose enhancement ratios at 2 Gy (DER{sub 2}) were calculated. Hind legs of BALB/c athymic mice were injected with U87-223 or U87-EV cells; after 2 weeks, half of the tumors were irradiated. Tumor volumes were tracked for a total of 5 weeks. Results: The dual luciferase reporter assay showed a significant reduction in luciferase activity of 293T cells cotransfected with miR-223 and the ATM 3′UTR compared to that in EV control. Overexpression of miR-223 in U87MG cells showed that ATM expression was significantly downregulated in the U87-223 cells compared to that in U87-EV (ATM/β-actin mRNA 1.0 vs 1.5, P<.05). U87-223 cells were hypersensitive to radiation compared to U87-EV cells in vitro (DER{sub 2} = 1.32, P<.01). Mice injected with miR-223-expressing tumors had almost the same tumors after 3 weeks (1.5 cm{sup 3} vs 1.7 cm{sup 3}). However, irradiation significantly decreased tumor size in miR-223-expressing tumors compared to those in controls (0.033 cm{sup 3} vs 0.829 cm{sup 3}). Conclusions: miR-223 overexpression downregulates ATM expression and sensitizes U87 cells to radiation in vitro and in vivo. MicroRNA-223 may be a novel cancer-targeting therapy, although its cancer- and patient-specific roles are currently undefined.« less

Study of interaction of GNR with glioblastoma cells

NASA Astrophysics Data System (ADS)

Hole, Arti; Cardoso-Avila, P. E.; Sridharan, Sangita; Sahu, Aditi; Nair, Jyothi; Dongre, Harsh; Goda, Jayant S.; Sawant, Sharada; Dutt, Shilpee; Pichardo-Molina, J. L.; Murali Krishna, C.

2018-01-01

Radiation resistance is one of the major causes of recurrence and failure of radiotherapy. Different methods have been used to increase the efficacy of radiation therapy and at the same time restrict the radiation resistivity. From last few years nanoparticles have played a key role in the enhancement of radiosensitization. The densely packed nanoparticles can selectively scatter or absorb the high radiations, which allow better targeting of cellular components within the tumor hence resulting in increased radiation damage to the cancer cells. Glioblastoma multiforme (GBM) is one of the highly radioresistant brain cancer. Current treatment methods are surgical resection followed by concurrent chemo and radiation therapy. In this study we have used in-house engineered gold nano rodes (GNR) and analyzed their effect on U-87MG cell lines. MTT assay was employed to determine the cytotoxic concentration of the nanoparticles. Raman spectroscopy was used to analyze the effect of gold nanoparticles on glioma cells, which was followed by transmission electron microscopic examinations to visualize their cellular penetration. Our data shows that GNR were able to penetrate the cells and induce cytotoxicity at the concentration of 198 μM as determined by MTT assay at 24 post GNP treatment. Additionally, we show that Raman spectroscopy, could classify spectra between untreated and cells treated with nanoparticles. Taken together, this study shows GNR penetration and cytotoxicity in glioma cells thereby providing a rationale to use them in cancer therapeutics. Future studies will be carried out to study the biological activity of the formulation as a radiosensitizer in GBM.

PPARγ agonists regulate the expression of stemness and differentiation genes in brain tumour stem cells

PubMed Central

Pestereva, E; Kanakasabai, S; Bright, J J

2012-01-01

Background: Brain tumour stem cells (BTSCs) are a small population of cancer cells that exhibit self-renewal, multi-drug resistance, and recurrence properties. We have shown earlier that peroxisome proliferator-activated receptor gamma (PPARγ) agonists inhibit the expansion of BTSCs in T98G and U87MG glioma. In this study, we analysed the influence of PPARγ agonists on the expression of stemness and differentiation genes in BTSCs. Methods: The BTSCs were isolated from T98G and DB29 glioma cells, and cultured in neurobasal medium with epidermal growth factor+basic fibroblast growth factor. Proliferation was measured by WST-1 (4-[3-(4-iodophenyl)-2-(4-nitrophenyl)-2 H-5-tetrazolio]-1,3-benzene disulphonate) and 3H thymidine uptake assays, and gene expression was analysed by quantitative reverse--transcription PCR and Taqman array. The expression of CD133, SRY box 2, and nanog homeobox (Nanog) was also evaluated by western blotting, immunostaining, and flow cytometry. Results: We found that PPARγ agonists, ciglitazone and 15-deoxy-Δ12,14-ProstaglandinJ2, inhibited cell viability and proliferation of T98G- and DB29-BTSCs. The PPARγ agonists reduced the expansion of CD133+ BTSCs and altered the expression of stemness and differentiation genes. They also inhibited Sox2 while enhancing Nanog expression in BTSCs. Conclusion: These findings highlight that PPARγ agonists inhibit BTSC proliferation in association with altered expression of Sox2, Nanog, and other stemness genes. Therefore, targeting stemness genes in BTSCs could be a novel strategy in the treatment of glioblastoma. PMID:22531638

Astrocyte-specific expression of human T-cell lymphotropic virus type 1 (HTLV-1) Tax: induction of tumor necrosis factor alpha and susceptibility to lysis by CD8+ HTLV-1-specific cytotoxic T cells.

PubMed

Méndez, E; Kawanishi, T; Clemens, K; Siomi, H; Soldan, S S; Calabresi, P; Brady, J; Jacobson, S

1997-12-01

Human T-cell lymphotropic virus type 1 (HTLV-1) is associated with a chronic neurological disease termed HTLV-1-associated myelopathy/tropical spastic paraperesis (HAM/TSP). Although the pathogenesis of this disease remains to be elucidated, the evidence suggests that immunopathological mechanisms are involved. Since HTLV-1 tax mRNA was colocalized with glial acidic fibrillary protein, a marker for astrocytes, we developed an in vitro model to assess whether HTLV-1 infection activates astrocytes to secrete cytokines or present viral immunodominant epitopes to virus-specific T cells. Two human astrocytic glioma cell lines, U251 and U373, were transfected with the 3' portion of the HTLV-1 genome and with the HTLV-1 tax gene under astrocyte-specific promoter control. In this study, we report that Tax-expressing astrocytic glioma transfectants activate the expression of tumor necrosis factor alpha mRNA in vitro. Furthermore, these Tax-expressing glioma transfectants can serve as immunological targets for HTLV-1-specific cytotoxic T lymphocytes (CTL). We propose that these events could contribute to the neuropathology of HAM/TSP, since infected astrocytes can become a source for inflammatory cytokines upon HTLV-1 infection and serve as targets for HTLV-1-specific CTL, resulting in parenchymal damage by direct lysis and/or cytokine release.

Nectin-like molecule 1 inhibits the migration and invasion of U251 glioma cells by regulating the expression of an extracellular matrix protein osteopontin.

PubMed

Yin, Bin; Li, Ke-han; An, Tai; Chen, Tao; Peng, Xiao-zhong

2010-06-01

To investigate the molecular mechanism of nectin-like molecule 1 (NECL1) inhibiting the migration and invasion of U251 glioma cells. We infected U251 glioma cells with adeno-nectin-like molecule 1 (Ad-NECL1) or empty adenovirus (Ad). Transwell and wound healing assays were performed to observe the migration of U251 cells incubated with the cell supernatant from Ad-NECL1 or Ad infected U251 cells. DNA microarray was applied to screen the gene expression profile after the restoration of NECL1 in U251 glioma cell lines. The differential expression of osteopontin (OPN), a gene related to migration and invasion, was further analyzed with semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR), Western blot, and immunohistochemistry. The restoration of NECL1 inhibited migration of U251 cells significantly (P<0.05). Altogether 195 genes were found differentially expressed by microarray, in which 175 were up-regulated and 20 down-regulated, including 9 extracellular matrix proteins involved in the migration of cells. Both mRNA and protein expressions of OPN, the most markedly reduced extracellular matrix protein, were found decreased in U251 cells after restoration of NECL1. Immunohistochemical assay also detected an increase of OPN in glioma tissues, related with the progressing of malignant grade. A link might exist between NECL1 and the extracellular matrix protein OPN in inhibiting the migration and invasion of U251 glioma cells.

First-in-human study of PET and optical dual-modality image-guided surgery in glioblastoma using 68Ga-IRDye800CW-BBN.

PubMed

Li, Deling; Zhang, Jingjing; Chi, Chongwei; Xiao, Xiong; Wang, Junmei; Lang, Lixin; Ali, Iqbal; Niu, Gang; Zhang, Liwei; Tian, Jie; Ji, Nan; Zhu, Zhaohui; Chen, Xiaoyuan

2018-01-01

Purpose : Despite the use of fluorescence-guided surgery (FGS), maximum safe resection of glioblastoma multiforme (GBM) remains a major challenge. It has restricted surgeons between preoperative diagnosis and intraoperative treatment. Currently, an integrated approach combining preoperative assessment with intraoperative guidance would be a significant step in this direction. Experimental design : We developed a novel 68 Ga-IRDye800CW-BBN PET/near-infrared fluorescence (NIRF) dual-modality imaging probe targeting gastrin-releasing peptide receptor (GRPR) in GBM. The preclinical in vivo tumor imaging and FGS were first evaluated using an orthotopic U87MG glioma xenograft model. Subsequently, the first-in-human prospective cohort study (NCT 02910804) of GBM patients were conducted with preoperative PET assessment and intraoperative FGS. Results : The orthotopic tumors in mice could be precisely resected using the near-infrared intraoperative system. Translational cohort research in 14 GBM patients demonstrated an excellent correlation between preoperative positive PET uptake and intraoperative NIRF signal. The tumor fluorescence signals were significantly higher than those from adjacent brain tissue in vivo and ex vivo (p < 0.0001). Compared with pathology, the sensitivity and specificity of fluorescence using 42 loci of fluorescence-guided sampling were 93.9% (95% CI 79.8%-99.3%) and 100% (95% CI 66.4%-100%), respectively. The tracer was safe and the extent of resection was satisfactory without newly developed neurologic deficits. Progression-free survival (PFS) at 6 months was 80% and two newly diagnosed patients achieved long PFS. Conclusions: This initial study has demonstrated that the novel dual-modality imaging technique is feasible for integrated pre- and intraoperative targeted imaging via the same molecular receptor and improved intraoperative GBM visualization and maximum safe resection.

Identification of ATP Citrate Lyase as a Positive Regulator of Glycolytic Function in Glioblastomas

PubMed Central

Beckner, Marie E.; Fellows-Mayle, Wendy; Zhang, Zhe; Agostino, Naomi R.; Kant, Jeffrey A.; Day, Billy W.; Pollack, Ian F.

2009-01-01

Glioblastomas, the most malignant type of glioma, are more glycolytic than normal brain tissue. Robust migration of glioblastoma cells has been previously demonstrated under glycolytic conditions and their pseudopodia contain increased glycolytic and decreased mitochondrial enzymes. Glycolysis is suppressed by metabolic acids, including citric acid which is excluded from mitochondria during hypoxia. We postulated that glioma cells maintain glycolysis by regulating metabolic acids, especially in their pseudopodia. The enzyme that breaks down cytosolic citric acid is ATP citrate lyase (ACLY). Our identification of increased ACLY in pseudopodia of U87 glioblastoma cells on 1D gels and immunoblots prompted investigation of ACLY gene expression in gliomas for survival data and correlation with expression of ENO1, that encodes enolase 1. Queries of the NIH’s REMBRANDT brain tumor database based on Affymetrix data indicated that decreased survival correlated with increased gene expression of ACLY in gliomas. Queries of gliomas and glioblastomas found an association of upregulated ACLY and ENO1 expression by chi square for all probe sets (reporters) combined and correlation for numbers of probe sets indicating shared upregulation of these genes. Real-time quantitative PCR confirmed correlation between ACLY and ENO1 in 21 glioblastomas (p < 0.001). Inhibition of ACLY with hydroxycitrate suppressed (p < 0.05) in vitro glioblastoma cell migration, clonogenicity and brain invasion under glycolytic conditions and enhanced the suppressive effects of a Met inhibitor on cell migration. In summary, gene expression data, proteomics and functional assays support ACLY as a positive regulator of glycolysis in glioblastomas. PMID:19795461

Metabolic Remodeling Precedes Mitochondrial Outer Membrane Permeabilization in Human Glioma Xenograft Cells

PubMed Central

Ponnala, Shivani; Chetty, Chandramu; Veeravalli, Krishna Kumar; Dinh, Dzung H.; Klopfenstein, Jeffrey D.; Rao, Jasti S.

2011-01-01

Glioma cancer cells adapt to changing microenvironment and shift from mitochondrial oxidative phosphorylation to aerobic glycolysis for their metabolic needs irrespective of oxygen availability. In the present study, we show that silencing MMP-9 in combination with uPAR/cathepsin B switch glioma cells glycolytic metabolism to oxidative phosphorylation (OXPHOS) and generate reactive oxygen species (ROS) to predispose glioma cells to mitochondrial outer membrane permeabilization. shRNA for MMP-9 and uPAR (pMU) as well as shRNA for MMP-9 and cathepsin B (pMC) activated complexes of mitochondria involved in OXPHOS and inhibited glycolytic hexokinase expression. The decreased interaction of hexokinase 2 with mitochondria in the treated cells indicated the inhibition of glycolysis activation. Overexpression of Akt reversed the pMU- and pMC-mediated glycolysis to OXPHOS switch. OXPHOS un-coupler oligomycin A altered the expression levels of the Bcl-2 family of proteins; treatment with pMU or pMC reversed this effect and induced mitochondrial outer membrane permeabilization. In addition, our results show changes in mitochondrial pore transition to release cytochrome c due to change in the VDAC-Bcl-XL and BAX-BAK interaction with pMU and pMC treatments. Taken together, our results suggest that pMU and pMC treatments switch glioma cells from glycolytic to OXPHOS pathway through an inhibitory effect on Akt, ROS induction, and an increase of cytosolic cytochrome c accumulation. These results demonstrate the potential of pMU and pMC as therapeutic candidates for treatment of glioma. PMID:22076676

Metabolic remodeling precedes mitochondrial outer membrane permeabilization in human glioma xenograft cells.

PubMed

Ponnala, Shivani; Chetty, Chandramu; Veeravalli, Krishna Kumar; Dinh, Dzung H; Klopfenstein, Jeffrey D; Rao, Jasti S

2012-02-01

Glioma cancer cells adapt to changing microenvironment and shift from mitochondrial oxidative phosphorylation to aerobic glycolysis for their metabolic needs irrespective of oxygen availability. In the present study, we show that silencing MMP-9 in combination with uPAR/cathepsin B switch the glycolytic metabolism of glioma cells to oxidative phosphorylation (OXPHOS) and generate reactive oxygen species (ROS) to predispose glioma cells to mitochondrial outer membrane permeabilization. shRNA for MMP-9 and uPAR (pMU) as well as shRNA for MMP-9 and cathepsin B (pMC) activated complexes of mitochondria involved in OXPHOS and inhibited glycolytic hexokinase expression. The decreased interaction of hexokinase 2 with mitochondria in the treated cells indicated the inhibition of glycolysis activation. Overexpression of Akt reversed the pMU- and pMC-mediated OXPHOS to glycolysis switch. The OXPHOS un-coupler oligomycin A altered the expression levels of the Bcl-2 family of proteins; treatment with pMU or pMC reversed this effect and induced mitochondrial outer membrane permeabilization. In addition, our results show changes in mitochondrial pore transition to release cytochrome c due to changes in the VDAC-Bcl-XL and BAX-BAK interaction with pMU and pMC treatments. Taken together, our results suggest that pMU and pMC treatments switch glioma cells from the glycolytic to the OXPHOS pathway through an inhibitory effect on Akt, ROS induction and an increase of cytosolic cytochrome c accumulation. These results demonstrate the potential of pMU and pMC as therapeutic candidates for the treatment of glioma.

Evaluation of 111In-Labeled Cyclic RGD Peptides: Effects of Peptide and Linker Multiplicity on Their Tumor Uptake, Excretion Kinetics and Metabolic Stability

PubMed Central

Shi, Jiyun; Zhou, Yang; Chakraborty, Sudipta; Kim, Young-Seung; Jia, Bing; Wang, Fan; Liu, Shuang

2011-01-01

Purpose: The purpose of this study was to demonstrate the valence of cyclic RGD peptides, P-RGD (PEG4-c(RGDfK): PEG4 = 15-amino-4,710,13-tetraoxapentadecanoic acid), P-RGD2 (PEG4-E[c(RGDfK)]2, 2P-RGD4 (E{PEG4-E[c(RGDfK)]2}2, 2P4G-RGD4 (E{PEG4-E[G3-c(RGDfK)]2}2: G3 = Gly-Gly-Gly) and 6P-RGD4 (E{PEG4-E[PEG4-c(RGDfK)]2}2) in binding to integrin αvβ3, and to assess the impact of peptide and linker multiplicity on biodistribution properties, excretion kinetics and metabolic stability of their corresponding 111In radiotracers. Methods: Five new RGD peptide conjugates (DOTA-P-RGD (DOTA =1,4,7,10-tetraazacyclododecane-1,4,7,10-tetracetic acid), DOTA-P-RGD2, DOTA-2P-RGD4, DOTA-2P4G-RGD4, DOTA-6P-RGD4), and their 111In complexes were prepared. The integrin αvβ3 binding affinity of cyclic RGD conjugates were determined by a competitive displacement assay against 125I-c(RGDyK) bound to U87MG human glioma cells. Biodistribution, planar imaging and metabolism studies were performed in athymic nude mice bearing U87MG human glioma xenografts. Results: The integrin αvβ3 binding affinity of RGD conjugates follows the order of: DOTA-6P-RGD4 (IC50 = 0.3 ± 0.1 nM) ~ DOTA-2P4G-RGD4 (IC50 = 0.2 ± 0.1 nM) ~ DOTA-2P-RGD4 (IC50 = 0.5 ± 0.1 nM) > DOTA-3P-RGD2 (DOTA-PEG4-E[PEG4-c(RGDfK)]2: IC50 = 1.5 ± 0.2 nM) > DOTA-P-RGD2 (IC50 = 5.0 ± 1.0 nM) >> DOTA-P-RGD (IC50 = 44.3 ± 3.5 nM) ~ c(RGDfK) (IC50 = 49.9 ± 5.5 nM) >> DOTA-6P-RGK4 (IC50 = 437 ± 35 nM). The fact that DOTA-6P-RGK4 had much lower integrin αvβ3 binding affinity than DOTA-6P-RGD4 suggests that the binding of DOTA-6P-RGD4 to integrin αvβ3 is RGD-specific. This conclusion is consistent with the lower tumor uptake for 111In(DOTA-6P-RGK4) than that for 111In(DOTA-6P-RGD4). It was also found that the G3 and PEG4 linkers between RGD motifs have a significant impact on the integrin αvβ3-targeting capability, biodistribution characteristics, excretion kinetics and metabolic stability of 111In-labeled cyclic RGD peptides. Conclusion: On the basis of their integrin αvβ3 binding affinity and tumor uptake of their corresponding 111In radiotracers, it was conclude that 2P-RGD4, 2P4G-RGD4 and 6P-RGD4 are most likely bivalent in binding to integrin αvβ3, and extra RGD motifs might contribute to the long tumor retention times of 111In(DOTA-2P-RGD4), 111In(DOTA-2P4G-RGD4) and 111In(DOTA-6P-RGD4) than that of 111In(DOTA-3P-RGD3) at 72 h p.i. Among the 111In-labeled cyclic RGD tetramers evaluated in the glioma model, 111In(DOTA-2P4G-RGD4) has very high tumor uptake with the best tumor/kidney and tumor/liver ratios, suggesting that 90Y(DOTA-2P4G-RGD4) and 177Lu(DOTA-2P4G-RGD4) might have the potential for targeted radiotherapy of integrin αvβ3-positive tumors. PMID:21850213

Antiproliferative activity of synthetic fatty acid amides from renewable resources.

PubMed

dos Santos, Daiane S; Piovesan, Luciana A; D'Oca, Caroline R Montes; Hack, Carolina R Lopes; Treptow, Tamara G M; Rodrigues, Marieli O; Vendramini-Costa, Débora B; Ruiz, Ana Lucia T G; de Carvalho, João Ernesto; D'Oca, Marcelo G Montes

2015-01-15

In the work, the in vitro antiproliferative activity of a series of synthetic fatty acid amides were investigated in seven cancer cell lines. The study revealed that most of the compounds showed antiproliferative activity against tested tumor cell lines, mainly on human glioma cells (U251) and human ovarian cancer cells with a multiple drug-resistant phenotype (NCI-ADR/RES). In addition, the fatty methyl benzylamide derived from ricinoleic acid (with the fatty acid obtained from castor oil, a renewable resource) showed a high selectivity with potent growth inhibition and cell death for the glioma cell line-the most aggressive CNS cancer. Copyright © 2014 Elsevier Ltd. All rights reserved.

STAT3 Activation Promotes Oncolytic HSV1 Replication in Glioma Cells

PubMed Central

Okemoto, Kazuo; Wagner, Benjamin; Meisen, Hans; Haseley, Amy; Kaur, Balveen; Chiocca, Ennio Antonio

2013-01-01

Recent studies report that STAT3 signaling is a master regulator of mesenchymal transformation of gliomas and that STAT3 modulated genes are highly expressed in the mesenchymal transcriptome of gliomas. A currently studied experimental treatment for gliomas consists of intratumoral injection of oncolytic viruses (OV), such as oncolytic herpes simplex virus type 1 (oHSV). We have described one particular oHSV (rQNestin34.5) that exhibits potent anti-glioma activity in animal models. Here, we hypothesized that alterations in STAT3 signaling in glioma cells may affect the replicative ability of rQNestin34.5. In fact, human U251 glioma cells engineered to either over-express STAT3 or with genetic down-regulation of STAT3 supported oHSV replication to a significantly higher or lesser degree, respectively, when compared to controls. Administration of pharmacologic agents that increase STAT3 phosphorylation/activation (Valproic Acid) or increase STAT3 levels (Interleukin 6) also significantly enhanced oHSV replication. Instead, administration of inhibitors of STAT3 phosphorylation/activation (LLL12) significantly reduced oHSV replication. STAT3 led to a reduction in interferon signaling in oHSV infected cells and inhibition of interferon signaling abolished the effect of STAT3 on oHSV replication. These data thus indicate that STAT3 signaling in malignant gliomas enhances oHSV replication, likely by inhibiting the interferon response in infected glioma cells, thus suggesting avenues for possible potentiation of oncolytic virotherapy. PMID:23936533

Bioactive oleanane-type saponins from the rhizomes of Anemone taipaiensis.

PubMed

Wang, Xiao-Yang; Gao, Hui; Zhang, Wei; Li, Yuan; Cheng, Guang; Sun, Xiao-Li; Tang, Hai-Feng

2013-10-15

Investigation of the n-BuOH extract of the rhizomes of Anemone taipaiensis led to the isolation of five new oleanane-type triterpenoid saponins (1-5), together with seven known saponins (6-12). Their structures were determined by the extensive use of (1)D and (2)D NMR experiments along with ESIMS analyses and acid hydrolysis. The aglycone of 1, 2 and 4 was determined as siaresinolic acid, which was reported in this genus for the first time. The cytotoxicities of the saponins 1-12, prosapogenins 4a, 5a, 10a-12a and sapogenins siaresinolic acid (SA), oleanolic acid (OA), hederagenin (HE) were evaluated against five human cancer cell lines, including HepG2, HL-60, A549, HeLa and U87MG. The monodesmosidic saponins 6-8, 5a, 10a-12a and sapogenins SA, OA, HE exhibited cytotoxic activity toward all cancer cell lines, with IC50 values ranging from 2.25 to 57.28 μM. Remarkably, the bisdesmosidic saponins 1-4 and 9 showed selective cytotoxicity against the U87MG cells. Copyright © 2013 Elsevier Ltd. All rights reserved.

A glioma-derived analog to platelet-derived growth factor: demonstration of receptor competing activity and immunological crossreactivity.

PubMed Central

Nistér, M; Heldin, C H; Wasteson, A; Westermark, B

1984-01-01

A human clonal glioma cell line, U-343 MGa Cl 2, cultured under serum-free conditions, was found to release a factor that competed with 125I-labeled platelet-derived growth factor (125I-PDGF) for binding to human foreskin fibroblasts. The concentration of competing activity in conditioned medium was equal to 20-30 ng of PDGF per ml. The PDGF receptor competing activity had an elution position on Sephadex G-200 close to that of tracer PDGF. The same fractions in the chromatogram also contained growth-promoting activity and material active in a PDGF radioimmunoassay. Incubation of partially purified, 125I-labeled glioma factor with fibroblasts, or rabbit anti-PDGF serum, led to the selective binding of a component with an estimated Mr of 31,000, as shown by NaDodSO4/gel electrophoresis under nonreducing conditions. After reduction this component migrated as a Mr 18,000 protein. Thus, the behavior in NaDodSO4/gel electrophoresis was similar to that of PDGF. Furthermore, incubation of partially purified glioma factor with immobilized PDGF antibodies markedly decreased the amount of PDGF receptor competing activity remaining in the supernatant. These results suggest that the factor produced by glioma cells has structural, immunological, and functional resemblance to PDGF. We previously reported that a human osteosarcoma cell line produces a PDGF-like molecule with growth-promoting activity. Taken together with the recent finding that PDGF is homologous to the transforming gene product of simian sarcoma virus, our present data give additional support for the idea that an autocrine activation of the PDGF receptor may be operational in the growth of human tumors of mesenchymal or glial origin. Images PMID:6322178

The brain-penetrating CXCR4 antagonist, PRX177561, increases the antitumor effects of bevacizumab and sunitinib in preclinical models of human glioblastoma.

PubMed

Gravina, Giovanni Luca; Mancini, Andrea; Marampon, Francesco; Colapietro, Alessandro; Delle Monache, Simona; Sferra, Roberta; Vitale, Flora; Richardson, Peter J; Patient, Lee; Burbidge, Stephen; Festuccia, Claudio

2017-01-05

Glioblastoma recurrence after treatment with the anti-vascular endothelial growth factor (VEGF) antibody bevacizumab is characterized by a highly infiltrative and malignant behavior that renders surgical excision and chemotherapy ineffective. It has been demonstrated that anti-VEGF/VEGFR therapies control the invasive phenotype and that relapse occurs through the increased activity of CXCR4. We therefore hypothesized that combining bevacizumab or sunitinib with the novel CXCR4 antagonist, PRX177561, would have superior antitumor activity. The effects of bevacizumab, sunitinib, and PRX177561 were tested alone or in combination in subcutaneous xenografts of U87MG, U251, and T98G cells as well as on intracranial xenografts of luciferase tagged U87MG cells injected in CD1-nu/nu mice. Animals were randomized to receive vehicle, bevacizumab (4 mg/kg iv every 4 days), sunitinib (40 mg/kg po qd), or PRX177561 (50 mg/kg po qd). The in vivo experiments demonstrated that bevacizumab and sunitinib increase the in vivo expression of CXCR4, SDF-1α, and TGFβ1. In addition, we demonstrate that the co-administration of the novel brain-penetrating CXCR4 antagonist, PRX177561, with bevacizumab or sunitinib inhibited tumor growth and reduced the inflammation. The combination of PRX177561 with bevacizumab resulted in a synergistic reduction of tumor growth with an increase of disease-free survival (DSF) and overall survival (OS), whereas the combination of PRX177561 with sunitinib showed a mild additive effect. The CXC4 antagonist PRX177561 may be a valid therapeutic complement to anti-angiogenic therapy, particularly when used in combination with VEGF/VEGFR inhibitors. Therefore, this compound deserves to be considered for future clinical evaluation.

Zero-valent Fe confined mesoporous silica nanocarriers (Fe(0) @ MCM-41) for targeting experimental orthotopic glioma in rats

PubMed Central

Shevtsov, M. A.; Parr, M. A.; Ryzhov, V. A.; Zemtsova, E. G.; Arbenin, A. Yu; Ponomareva, A. N.; Smirnov, V. M.; Multhoff, G.

2016-01-01

Mesoporous silica nanoparticles (MSNs) impregnated with zero-valent Fe (Fe(0) @ MCM-41) represent an attractive nanocarrier system for drug delivery into tumor cells. The major goal of this work was to assess whether MSNs can penetrate the blood-brain barrier in a glioblastoma rat model. Synthesized MSNs nanomaterials were characterized by energy dispersive X-ray spectroscopy, measurements of X-ray diffraction, scanning electron microscopy and Mössbauer spectroscopy. For the detection of the MSNs by MR and for biodistribution studies MSNs were labeled with zero-valent Fe. Subsequent magnetometry and nonlinear-longitudinal-response-M2 (NLR-M2) measurements confirmed the MR negative contrast enhancement properties of the nanoparticles. After incubation of different tumor (C6 glioma, U87 glioma, K562 erythroleukemia, HeLa cervix carcinoma) and normal cells such as fibroblasts and peripheral blood mononuclear cells (PBMCs) MSNs rapidly get internalized into the cytosol. Intracellular residing MSNs result in an enhanced cytotoxicity as Fe(0) @ MCM-41 promote the reactive oxygen species production. MRI and histological studies indicated an accumulation of intravenously injected Fe(0) @ MCM-41 MSNs in orthotopic C6 glioma model. Biodistribution studies with measurements of second harmonic of magnetization demonstrated an increased and dose-dependent retention of MSNs in tumor tissues. Taken together, this study demonstrates that MSNs can enter the blood-brain barrier and accumulate in tumorous tissues. PMID:27386761

Flavonoids Activated Caspases for Apoptosis in Human Glioblastoma T98G and U87MG Cells But Not in Human Normal Astrocytes

PubMed Central

Das, Arabinda; Banik, Naren L.; Ray, Swapan K.

2011-01-01

BACKGROUND Human glioblastoma is a deadly brain cancer that continues to defy all current therapeutic strategies. We induced apoptosis in human glioblastoma T98G and U87MG cells following treatment with apigenin (APG), (−)-epigallocatechin (EGC), (−)-epigallocatechin-3-gallate (EGCG), and genistein (GST) that did not induce apoptosis in human normal astrocytes (HNA). METHODS Induction of apoptosis was examined using Wright staining and ApopTag assay. Production of reactive oxygen species (ROS) and increase in intracellular free [Ca2+] were measured by fluoresent probes. Analysis of mRNA and Western blotting indicated increases in expression and activities of the stress kinases and cysteine proteases for apoptosis. JC-1 showed changes in mitochondrial membrane potential (ΔΨm) and use of specific inhibitors confirmed activation of kinases and proteases in apoptosis. RESULTS Treatment of glioblastoma cells with APG, EGC, EGCG, or GST triggered ROS production that induced apoptosis with phosphorylation of p38 MAPK and activation of the redox-sensitive JNK1 pathway. Pretreatment of cells with ascorbic acid attenuated ROS production and p38 MAPK phosphorylation. Increases in intracellular free [Ca2+] and activation of caspase-4 indicated involvement of endoplasmic reticulum stress in apoptosis. Other events in apoptosis included overexpression of Bax, loss of ΔΨm, mitochondrial release of cytochrome c and Smac into the cytosol, down regulation of baculoviral inhibitor-of-apoptosis repeat containing proteins, and activation of calpain, caspase-9, and caspase-3. EGC and EGCG also induced caspase-8 activity. APG, EGC, EGCG, or GST did not induce apoptosis in HNA. CONCLUSION Results strongly suggest that flavonoids are potential therapeutic agents for induction of apoptosis in human glioblastoma cells. PMID:19894226

Overexpression of SASH1 related to the decreased invasion ability of human glioma U251 cells.

PubMed

Yang, Liu; Liu, Mei; Gu, Zhikai; Chen, Jianguo; Yan, Yaohua; Li, Jian

2012-12-01

The purpose of this study was to investigate the impact of SAM- and SH3-domain containing 1 (SASH1) on the biological behavior of glioma cells, including its effects on cellular growth, proliferation, apoptosis, invasion, and metastasis, and thereby to provide an experimental basis for future therapeutic treatments. A pcDNA3.1-SASH1 eukaryotic expression vector was constructed and transfected into the U251 human glioma cell line. Using the tetrazolium-based colorimetric (MTT) assay, flow cytometry analyses, transwell invasion chamber experiments, and other methods, we examined the impact of SASH1 on the biological behaviors of U251 cells, including effects on viability, cell cycle, apoptosis, and invasion. Furthermore, the effect of SASH1 on the expression of cyclin D1, caspase-3, matrix metalloproteinase (MMP)-2, MMP-9, and other proteins was observed. Compared to the empty vector and blank control groups, the pcDNA3.1-SASH1 group of U251 cells exhibited significantly reduced cell viability, proliferation, and invasion (p < 0.05), although there was no difference between the empty vector and blank control groups. The pcDNA3.1-SASH1 group demonstrated a significantly higher apoptotic index than did the empty vector and blank control groups (p < 0.05), and the percentage of apoptotic cells was similar between the empty vector and blank control groups. In addition, the pcDNA3.1-SASH1 group expressed significantly lower protein levels of cyclin D1 and MMP-2/9 compared to the control and empty vector groups (p < 0.05) and significantly higher protein levels of caspase-3 than the other two groups (p < 0.05). Cyclin D1, caspase-3, and MMP-2/9 expression was unchanged between the empty vector and blank control groups. SASH1 gene expression might be related to the inhibition of the growth, proliferation, and invasion of U251 cells and the promotion of U251 cells apoptosis.

Phase I trial of capecitabine rapidly disintegrating tablets and concomitant radiation therapy in children with newly diagnosed brainstem gliomas and high-grade gliomas

PubMed Central

Kilburn, Lindsay B.; Kocak, Mehmet; Schaedeli Stark, Franziska; Meneses-Lorente, Georgina; Brownstein, Carrie; Hussain, Sazzad; Chintagumpala, Murali; Thompson, Patrick A.; Gururangan, Sri; Banerjee, Anuradha; Paulino, Arnold C.; Kun, Larry; Boyett, James M.; Blaney, Susan M.

2013-01-01

Background We conducted a phase I study to estimate the maximum tolerated dose and describe the dose-limiting toxicities and pharmacokinetics of oral capecitabine rapidly disintegrating tablets given concurrently with radiation therapy to children with newly diagnosed brainstem or high-grade gliomas. Methods Children 3–21 y with newly diagnosed intrinsic brainstem or high-grade gliomas were eligible for enrollment. The starting dose was 500 mg/m2, given twice daily, with subsequent cohorts enrolled at 650 mg/m2 and 850 mg/m2 using a 3 + 3 phase I design. Children received capecitabine at the assigned dose daily for 9 wks starting from the first day of radiation therapy (RT). Following a 2-wk break, patients received 3 courses of capecitabine 1250 mg/m2 twice daily for 14 days followed by a 7-day rest. Pharmacokinetic sampling was performed in consenting patients. Six additional patients with intrinsic brainstem gliomas were enrolled at the maximum tolerated dose to further characterize the pharmacokinetic and toxicity profiles. Results Twenty-four patients were enrolled. Twenty were fully assessable for toxicity. Dose-limiting toxicities were palmar plantar erythroderma (grades 2 and 3) and elevation of alanine aminotransferase (grades 2 and 3). Systemic exposure to capecitabine and metabolites was similar to or slightly lower than predicted based on adult data. Conclusions Capecitabine with concurrent RT was generally well tolerated. The recommended phase II capecitabine dose when given with concurrent RT is 650 mg/m2, administered twice daily. A phase II study to evaluate the efficacy of this regimen in children with intrinsic brainstem gliomas is in progress (PBTC-030). PMID:23592571

Nicotinamide augments the survival and incidence of apoptosis in glioma cells following photodynamic therapy in vitro

NASA Astrophysics Data System (ADS)

Bisland, Stuart K.; Modi, Nayan; Wilson, Brian C.

2004-10-01

The ability to customize photodynamic therapy (PDT) parameters with regards to timing and dosing of administered drug and light can be beneficial in determining target specificity and mode of cell death. Sustained, low level PDT or metronomic PDT (mPDT) may afford enhanced apoptotic cell death. This is of particular importance when considering PDT for the treatment of brain tumors as unlike apoptosis, necrotic cell death often leads to inflammation with increased intracranial pressure. The ability, therefore, to 'fine tune' PDT in favour of apoptosis is paramount. We have studied both acute (one time treatment) PDT (aPDT) and mPDT delivery strategies in combination with nicotinamide (NA) in an attempt to maximize the number of tumor cells dieing by apoptosis. Using several different glioma cell lines (9L, U87-MG and CNS-1) we now confirm that NA provides a dose-dependent (0.1-0.5 mM) increase in apoptotic cells following d-aminolevulinic acid-mediated aPDT or mPDT. Furthermore, using the 9L cell line stably transfected with the luciferase gene, NA was shown to delay the depletion of bioluminscence signal in aPDT and mPDT treated cells, inferring that adenosine triphosphate levels are maintained for longer following NA treatment. NA has previously been reported as promoting neuronal and vascular cell survival in normal brain following a number of neurological insults in which reactive oxygen species are implicated including, stroke, Alzheimer's disease and toxin-induced lesions. It is likely that the effects of NA reflect its capacity as an antioxidant as well as its ability to inhibit poly (adenosine diphosphate-ribose) polymerase-mediated depletion of ATP. Our results indicate that NA may prove therapeutically advantageous when used in combination with PDT treatment of brain tumors.

MiR-18a increased the permeability of BTB via RUNX1 mediated down-regulation of ZO-1, occludin and claudin-5.

PubMed

Miao, Yin-Sha; Zhao, Ying-Yu; Zhao, Li-Ni; Wang, Ping; Liu, Yun-Hui; Ma, Jun; Xue, Yi-Xue

2015-01-01

The purposes of this study were to investigate the possible molecular mechanisms of miR-18a regulating the permeability of blood-tumor barrier (BTB) via down-regulated expression and distribution of runt-related transcription factor 1 (RUNX1). An in vitro BTB model was established with hCMEC/D3 cells and U87MG cells to obtain glioma vascular endothelial cells (GECs). The endogenous expressions of miR-18a and RUNX1 were converse in GECs. The overexpression of miR-18a significantly impaired the integrity and increased the permeability of BTB, which respectively were detected by TEER and HRP flux assays, accompanied by down-regulated mRNA and protein expressions and distributions of ZO-1, occludin and claudin-5 in GECs. Dual-luciferase reporter assay was carried out and revealed RUNX1 is a target gene of miR-18a. Meanwhile, mRNA and protein expressions and distribution of RUNX1 were downregulated by miR-18a. Most important, miR-18a and RUNX1 could reversely regulate the permeability of BTB as well as the expressions and distributions of ZO-1, occludin and claudin-5. Finally, chromatin immunoprecipitation verified that RUNX1 interacted with "TGGGGT" DNA sequence in promoter region of ZO-1, occludin and claudin-5 respectively. Taken together, our present study indicated that miR-18a increased the permeability of BTB via RUNX1 mediated down-regulation of tight junction related proteins ZO-1, occludin and claudin-5, which would attract more attention to miR-18a and RUNX1 as potential targets of drug delivery across BTB and provide novel strategies for glioma treatment. Copyright © 2014 Elsevier Inc. All rights reserved.

Local delivery of cancer-cell glycolytic inhibitors in high-grade glioma

PubMed Central

Wicks, Robert T.; Azadi, Javad; Mangraviti, Antonella; Zhang, Irma; Hwang, Lee; Joshi, Avadhut; Bow, Hansen; Hutt-Cabezas, Marianne; Martin, Kristin L.; Rudek, Michelle A.; Zhao, Ming; Brem, Henry; Tyler, Betty M.

2015-01-01

Background 3-bromopyruvate (3-BrPA) and dichloroacetate (DCA) are inhibitors of cancer-cell specific aerobic glycolysis. Their application in glioma is limited by 3-BrPA's inability to cross the blood-brain-barrier and DCA's dose-limiting toxicity. The safety and efficacy of intracranial delivery of these compounds were assessed. Methods Cytotoxicity of 3-BrPA and DCA were analyzed in U87, 9L, and F98 glioma cell lines. 3-BrPA and DCA were incorporated into biodegradable pCPP:SA wafers, and the maximally tolerated dose was determined in F344 rats. Efficacies of the intracranial 3-BrPA wafer and DCA wafer were assessed in a rodent allograft model of high-grade glioma, both as a monotherapy and in combination with temozolomide (TMZ) and radiation therapy (XRT). Results 3-BrPA and DCA were found to have similar IC50 values across the 3 glioma cell lines. 5% 3-BrPA wafer-treated animals had significantly increased survival compared with controls (P = .0027). The median survival of rats with the 50% DCA wafer increased significantly compared with both the oral DCA group (P = .050) and the controls (P = .02). Rats implanted on day 0 with a 5% 3-BrPA wafer in combination with TMZ had significantly increased survival over either therapy alone. No statistical difference in survival was noted when the wafers were added to the combination therapy of TMZ and XRT, but the 5% 3-BrPA wafer given on day 0 in combination with TMZ and XRT resulted in long-term survivorship of 30%. Conclusion Intracranial delivery of 3-BrPA and DCA polymer was safe and significantly increased survival in an animal model of glioma, a potential novel therapeutic approach. The combination of intracranial 3-BrPA and TMZ provided a synergistic effect. PMID:25053853

Quantitative correlational study of microbubble-enhanced ultrasound imaging and magnetic resonance imaging of glioma and early response to radiotherapy in a rat model

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Chen; Lee, Dong-Hoon; Zhang, Kai

Purpose: Radiotherapy remains a major treatment method for malignant tumors. Magnetic resonance imaging (MRI) is the standard modality for assessing glioma treatment response in the clinic. Compared to MRI, ultrasound imaging is low-cost and portable and can be used during intraoperative procedures. The purpose of this study was to quantitatively compare contrast-enhanced ultrasound (CEUS) imaging and MRI of irradiated gliomas in rats and to determine which quantitative ultrasound imaging parameters can be used for the assessment of early response to radiation in glioma. Methods: Thirteen nude rats with U87 glioma were used. A small thinned skull window preparation was performedmore » to facilitate ultrasound imaging and mimic intraoperative procedures. Both CEUS and MRI with structural, functional, and molecular imaging parameters were performed at preradiation and at 1 day and 4 days postradiation. Statistical analysis was performed to determine the correlations between MRI and CEUS parameters and the changes between pre- and postradiation imaging. Results: Area under the curve (AUC) in CEUS showed significant difference between preradiation and 4 days postradiation, along with four MRI parameters, T{sub 2}, apparent diffusion coefficient, cerebral blood flow, and amide proton transfer-weighted (APTw) (all p < 0.05). The APTw signal was correlated with three CEUS parameters, rise time (r = − 0.527, p < 0.05), time to peak (r = − 0.501, p < 0.05), and perfusion index (r = 458, p < 0.05). Cerebral blood flow was correlated with rise time (r = − 0.589, p < 0.01) and time to peak (r = − 0.543, p < 0.05). Conclusions: MRI can be used for the assessment of radiotherapy treatment response and CEUS with AUC as a new technique and can also be one of the assessment methods for early response to radiation in glioma.« less

Phase I Pharmacokinetic Study of the VEGFR Tyrosine Kinase Inhibitor Vatalanib (PTK787) plus Imatinib and Hydroxyurea for Malignant Glioma

PubMed Central

Reardon, David A.; Egorin, Merrill J.; Desjardins, Annick; Vredenburgh, James J.; Beumer, Jan H.; Lagattuta, Theodore F.; Gururangan, Sridharan; Herndon, James E.; Salvado, August J.; Friedman, Henry S.

2009-01-01

Background We determined the maximum tolerated dose (MTD) and dose-limiting toxicities (DLT) of the oral vascular endothelial growth factor receptor (VEGFR) inhibitor, vatalanib, when administered with imatinib and hydroxyurea on a continuous daily schedule among recurrent malignant glioma patients. Methods All patients received 500 mg of hydroxyurea twice daily. Imatinib was dosed at 400 mg per day for patients not taking enzyme-inducing antiepileptic drugs (EIAEDs; stratum A) and at 500 mg twice-a-day for patients taking EIAEDs (stratum B). Vatalanib was escalated from 500 mg to 1250 mg twice daily in successive cohorts, independently for each stratum. Pharmacokinetics of each drug were assessed. Results Thirty-seven recurrent patients, including 34 (92%) with glioblastoma and 3 (8%) with grade 3 malignant glioma, were enrolled. Nineteen patients (51%) were taking EIAEDs. The MTD of vatalanib for all patients was 1000 mg twice-a-day. DLTs were hematologic, gastrointestinal, renal and hepatic. No patients developed intracranial hemorrhage. Concurrent administration of imatinib and hydroxyurea did not affect vatalanib exposure, but EIAEDs decreased vatalanib and imatinib plasma exposures. Conclusion Vatalanib doses up to 1000 mg twice-a-day combined with imatinib and hydroxyurea are well tolerated. Strategies to target tumor blood vessel endothelial cells and pericytes by inhibiting VEGFR and PDGFR, respectively, are safe among recurrent malignant glioma patients and may enhance anti-angiogenesis activity. PMID:19248046

The therapeutic potential of polymersomes loaded with 225Ac evaluated in 2D and 3D in vitro glioma models.

PubMed

de Kruijff, R M; van der Meer, A J G M; Windmeijer, C A A; Kouwenberg, J J M; Morgenstern, A; Bruchertseifer, F; Sminia, P; Denkova, A G

2018-06-01

Alpha emitters have great potential in targeted tumour therapy, especially in destroying micrometastases, due to their high linear energy transfer (LET). To prevent toxicity caused by recoiled daughter atoms in healthy tissue, alpha emitters like 225 Ac can be encapsulated in polymeric nanocarriers (polymersomes), which are capable of retaining the daughter atoms to a large degree. In the translation to a (pre-)clinical setting, it is essential to evaluate their therapeutic potential. As multicellular tumour spheroids mimic a tumour microenvironment more closely than a two-dimensional cellular monolayer, this study has focussed on the interaction of the polymersomes with U87 human glioma spheroids. We have found that polymersomes distribute themselves throughout the spheroid after 4 days which, considering the long half-life of 225 Ac (9.9 d) (Vaidyanathan and Zalutsky, 1996), allows for irradiation of the entire spheroid. A decrease in spheroidal growth has been observed upon the addition of only 0.1 kBq 225 Ac, an effect which was more pronounced for the 225 Ac in polymersomes than when only coupled to DTPA. At higher activities (5 kBq), the spheroids have been found to be destroyed completely after two days. We have thus demonstrated that 225 Ac containing polymersomes effectively inhibit tumour spheroid growth, making them very promising candidates for future in vivo testing. Copyright © 2018 Elsevier B.V. All rights reserved.

Preparation and in vitro characterization of SN-38-loaded, self-forming polymeric depots as an injectable drug delivery system.

PubMed

Manaspon, Chawan; Hongeng, Suradej; Boongird, Atthaporn; Nasongkla, Norased

2012-10-01

This work describes the preparation and characterization of anticancer-loaded injectable polymeric depots that consisted of D,L-lactide (LA), ε-caprolactone (CL), and poly(ethylene glycol) (PEG) or [poly(ε-caprolactone)-random-poly(D,L-lactide)]-block-poly(ethylene glycol)-block-[poly(ε-caprolactone)-random-poly(D,L-lactide)] (PLEC) copolymers for malignant gliomas treatment. PLECs were polymerized with different percentages of LA to deliver 7-ethyl-10-hydroxycamptothecin (SN-38), a highly potent anticancer drug. SN-38-loaded depots could form directly in phosphate buffer saline with more than 98% encapsulation efficiency. The release rate of SN-38 from depots was found to depend on the amount of LA in PLECs, loading content of SN-38 in the depots, and depot weight. Encapsulation of SN-38 inside depots could enhance the stability of SN-38 where all of SN-38 released after 60 days was in an active form. Depots without SN-38 were evaluated as noncytotoxic against U-87MG, whereas SN-38-loaded depots showed cytotoxic effect as a function of concentration. Copyright © 2012 Wiley Periodicals, Inc.

Quantitative imaging of magnesium distribution at single-cell resolution in brain tumors and infiltrating tumor cells with secondary ion mass spectrometry (SIMS)

PubMed Central

Chandra, Subhash; Parker, Dylan J.; Barth, Rolf F.; Pannullo, Susan C.

2016-01-01

Glioblastoma multiforme (GBM) is one of the deadliest forms of human brain tumors. The infiltrative pattern of growth of these tumors includes the spread of individual and/or clusters of tumor cells at some distance from the main tumor mass in parts of the brain protected by an intact blood-brain-barrier. Pathophysiological studies of GBM could be greatly enhanced by analytical techniques capable of in situ single-cell resolution measurements of infiltrating tumor cells. Magnesium homeostasis is an area of active investigation in high grade gliomas. In the present study, we have used the F98 rat glioma as a model of human GBM and an elemental/isotopic imaging technique of secondary ion mass spectrometry (SIMS), a CAMECA IMS-3f ion microscope, for studying Mg distributions with single-cell resolution in freeze-dried brain tissue cryosections. Quantitative observations were made on tumor cells in the main tumor mass, contiguous brain tissue, and infiltrating tumor cells in adjacent normal brain. The brain tissue contained a significantly lower total Mg concentration of 4.70 ± 0.93 mmol/Kg wet weight (mean ± SD) in comparison to 11.64 ± 1.96 mmol/Kg wet weight in tumor cells of the main tumor mass and 10.72 ± 1.76 mmol/Kg wet weight in infiltrating tumor cells (p<0.05). The nucleus of individual tumor cells contained elevated levels of bound Mg. These observations demonstrate enhanced Mg-influx and increased binding of Mg in tumor cells and provide strong support for further investigation of GBMs for altered Mg homeostasis and activation of Mg-transporting channels as possible therapeutic targets. PMID:26703785

Memantine-derived drugs as potential antitumor agents for the treatment of glioblastoma.

PubMed

Cacciatore, Ivana; Fornasari, Erika; Marinelli, Lisa; Eusepi, Piera; Ciulla, Michele; Ozdemir, Ozlem; Tatar, Abdulgani; Turkez, Hasan; Di Stefano, Antonio

2017-11-15

Glioblastoma is one of the most aggressive malignant primary brain cancer in adults. To date, surgery, radiotherapy and current pharmacological treatments are not sufficient to manage this pathology that has a high mortality rate (median survival 12-15months). Recently, anticancer multi-targeted compounds have attracted much attention with the aim to obtain new drugs able to hit different biological targets that are involved in the onset and progression of the disease. Here, we report the synthesis of novel memantine-derived drugs (MP1-10) and their potential antitumor activities in human U87MG glioblastoma cell line. MP1-10 were synthetized joining memantine, which is a NMDA antagonist, to different histone deacetylase inhibitors to obtain one molecule with improved therapeutic efficacy. Biological results indicated that MP1 and MP2 possessed more potent anti-proliferative effects on U87MG cells than MP3-10 in a dose-dependent manner. MP1 and MP2 induced significant cell death by apoptosis characterized by apoptotic morphological changes in Hoechst staining. Both drugs also exhibited non-genotoxic and only mild cytotoxic effects in human whole blood cells. However, only MP1, showing good chemico-physical properties (solubility, LogP) and enzymatic stabilities in gastric and intestinal fluids, can be considered a suitable candidate for in vivo pharmacokinetic studies. Copyright © 2017 Elsevier B.V. All rights reserved.

Genetically Engineered Mouse Model of Diffuse Intrinsic Pontine Glioma as a Preclinical Tool

DTIC Science & Technology

2014-11-01

LIMITATION OF ABSTRACT 18 . NUMBER OF PAGES 19a. NAME OF RESPONSIBLE PERSON USAMRMC a. REPORT U b. ABSTRACT U c. THIS PAGE U UU 39 19b. TELEPHONE...for this mouse cohort was included in Figure 1. Out of 23 GFAP tv- a; p53 floxed mice infected with RCAS-PDGF-B and RCAS-Cre, 18 developed...being used as part of the DIPG preclinical consortium (together with several laboratories that are using human DIPG xenografts ) to help prioritize

Induction of antigen-specific cytotoxic T-cell response by dendritic cells generated from ecto-mesenchymal stem cells infected with an adenovirus containing the MAGE-D4a gene.

PubMed

Hu, Shijie; Li, Bing; Shen, Xuefeng; Zhang, Rui; Gao, Dakuan; Guo, Qingdong; Jin, Yan; Fei, Zhou

2016-04-01

The present study aimed to investigate the feasibility of using ecto-mesenchymal stem cell (EMSC)-derived dendritic cells (DCs) for glioma immunotherapy following infection by a recombinant adenovirus containing the melanoma-associated antigen D4a (MAGE-D4a) gene. The ex vivo cultured EMSCs were infected by the adenoviral plasmid containing MAGE-D4a (pAd/MAGE-D4a). Efficiency of transfection was evaluated through the detection of green fluorescent protein-marked MAGE-D4a. The MAGE-EMSCs were induced to differentiate into DCs, termed as MAGE-EMSCs-DCs. The morphology was subsequently analyzed under a microscope, and methyl thiazolyl tetrazolium (MTT) and interferon-γ (IFN-γ) assays were performed to analyze the cytotoxicity of the MAGE-EMSC-DCs on the human glioma U251 cell line. Following purification by magnetic-activated cell sorting, the EMSCs grew into swirls, with a long spindle shape and were fibroblast-like. The gene transfected with recombinant adenovirus vectors maintained high and stable expression levels of MAGE-D4a, and its efficiency was increased in a multiplicity of infection-dependent manner. The results of the MTT assay indicated that the T cells, primed by the recombinant MAGE-D4a-infected EMSC-DCs in vitro , recognized MAGE-D4a-expressing tumor cell lines in a human leukocyte antigen class I-restricted manner, and evoked a higher cytotoxic T cell (CTL) response. The CTL response induced by the MAGE-EMSC-DCs, co-cultured with the U251 cells for 24 h, produced 765.0 pg/ml IFN-γ, which was significantly greater when compared to the control wells. T lymphocytes stimulated by MAGE-EMSC-DCs evoke a higher CTL response to human glioma cell lines, and may serve as a promising therapeutic modality for the treatment of MAGE-D4a-expressing glioma.

Automatic lesion detection and segmentation of 18F-FET PET in gliomas: A full 3D U-Net convolutional neural network study.

PubMed

Blanc-Durand, Paul; Van Der Gucht, Axel; Schaefer, Niklaus; Itti, Emmanuel; Prior, John O

2018-01-01

Amino-acids positron emission tomography (PET) is increasingly used in the diagnostic workup of patients with gliomas, including differential diagnosis, evaluation of tumor extension, treatment planning and follow-up. Recently, progresses of computer vision and machine learning have been translated for medical imaging. Aim was to demonstrate the feasibility of an automated 18F-fluoro-ethyl-tyrosine (18F-FET) PET lesion detection and segmentation relying on a full 3D U-Net Convolutional Neural Network (CNN). All dynamic 18F-FET PET brain image volumes were temporally realigned to the first dynamic acquisition, coregistered and spatially normalized onto the Montreal Neurological Institute template. Ground truth segmentations were obtained using manual delineation and thresholding (1.3 x background). The volumetric CNN was implemented based on a modified Keras implementation of a U-Net library with 3 layers for the encoding and decoding paths. Dice similarity coefficient (DSC) was used as an accuracy measure of segmentation. Thirty-seven patients were included (26 [70%] in the training set and 11 [30%] in the validation set). All 11 lesions were accurately detected with no false positive, resulting in a sensitivity and a specificity for the detection at the tumor level of 100%. After 150 epochs, DSC reached 0.7924 in the training set and 0.7911 in the validation set. After morphological dilatation and fixed thresholding of the predicted U-Net mask a substantial improvement of the DSC to 0.8231 (+ 4.1%) was noted. At the voxel level, this segmentation led to a 0.88 sensitivity [95% CI, 87.1 to, 88.2%] a 0.99 specificity [99.9 to 99.9%], a 0.78 positive predictive value: [76.9 to 78.3%], and a 0.99 negative predictive value [99.9 to 99.9%]. With relatively high performance, it was proposed the first full 3D automated procedure for segmentation of 18F-FET PET brain images of patients with different gliomas using a U-Net CNN architecture.

Downregulation of Pygopus 2 inhibits vascular mimicry in glioma U251 cells by suppressing the canonical Wnt signaling pathway

PubMed Central

WANG, HAIDONG; FU, JIANHUA; XU, DIANSHUANG; XU, WEIWEI; WANG, SHIYONG; ZHANG, LIU; XIANG, YONGSHENG

2016-01-01

Gliomas are the most common type of malignant primary brain tumor, and the Wnt signaling pathway is associated with glioma malignancy. Pygopus protein plays an important role in developmental brain patterning, and has been identified to be a component of the Wnt signaling pathway. In the present study, the Pygopus 2 (Pygo2) protein was examined in 80 glioma tissue samples. Short hairpin (sh)RNA-Pygo2 was transfected into glioma U251 cells, and the cell proliferation, colony formation and bromodeoxyuridine (BrdU) incorporation were analyzed. Western blot analysis and reverse transcription-polymerase chain reaction were used to detect the expression of Pygo2. A vascular mimicry assay was performed to examine the vascular mimicry of U251 cells. A luciferase reporter assay was used to detect the β-catenin/Wnt system. The cyclin D1 protein was also detected using western blot analysis. The results demonstrated that inhibition of the expression of Pygo2 significantly triggered the decrease of cell proliferation, colony formation and BrdU incorporation compared with the cells treated with scramble control shRNA (shRNA-Scr). shRNA-Pygo2 transfection was found to inhibit vascular-mimicry and block the Wnt signaling pathway compared to the cells transfected with shRNA-Scr. The transfection of shRNA-Pygo2 also decreased the expression of the Wnt target gene cyclin D1. In conclusion, shRNA-Pygo2 suppressed glioma cell proliferation effectively and inhibited vascular mimicry by inhibiting the expression of cyclin D1 in the canonical Wnt/β-catenin pathway in brain glioma cells. PMID:26870266

Characterization of peripheral blood human immunodeficiency virus isolates from Hispanic women with cognitive impairment

PubMed Central

Toro Nieves, Dianedis M; Plaud, Marinés; Wojna, Valerie; Skolasky, Richard; Meléndez, Loyda M

2009-01-01

Human immunodeficiency virus type 1 (HIV-1) tropism plays an important role in HIV-associated dementia. In this study, aimed at determining if the tropism and coreceptor usage of circulating viruses correlates with cognitive function, the authors isolated and characterized HIV from the peripheral blood of 21 Hispanic women using antiretroviral therapy. Macrophage tropism was determined by inoculation of HIV isolates onto monocyte-derived macrophages and lymphocyte cultures. To define coreceptor usage, the HIV isolates were inoculated onto the U87.CD4 glioma cell lines with specific CCR5 and CXCR4 coreceptors. HIV isolates from cognitively impaired patients showed higher levels of replication in mitogen-stimulated peripheral blood mononuclear cells than did isolates from patients with normal cognition (P < .05). The viral growth of HIV primary isolates in macrophages and lymphocytes did not differ between patients with and those without cognitive impairment. However, isolates from the cognitively impaired women preferentially used the X4 coreceptor (P < .05). These phenotypic studies suggest that cognitively impaired HIV-infected women receiving treatment may have a more highly replicating and more pathogenic X4 virus in the circulation that could contribute to their neuropathogenesis. PMID:17849315

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cervia, Davide, E-mail: d.cervia@unitus.it; Department of Biomedical and Clinical Sciences, “Luigi Sacco” University Hospital, University of Milan, Milano; Catalani, Elisabetta

Water-soluble protein signals (pheromones) of the ciliate Euplotes have been supposed to be functional precursors of growth factors and cytokines that regulate cell–cell interaction in multi-cellular eukaryotes. This work provides evidence that native preparations of the Euplotes raikovi pheromone Er-1 (a helical protein of 40 amino acids) specifically increases viability, DNA synthesis, proliferation, and the production of interferon-γ, tumor necrosis factor-α, interleukin (IL)-1β, IL-2, and IL-13 in human Jurkat T-cells. Also, Er-1 significantly decreases the mRNA levels of the β and γ subunits of IL-2 receptor (IL-2R), while the mRNA levels of the α subunit appeared to be not affected.more » Jurkat T-cell treatments with Er-1 induced the down-regulation of the IL-2Rα subunit by a reversible and time-dependent endocytosis, and increased the levels of phosphorylation of the extracellular signal-regulated kinases (ERK). The cell-type specificity of these effects was supported by the finding that Er-1, although unable to directly influence the growth of human glioma U-373 cells, induced Jurkat cells to synthesize and release factors that, in turn, inhibited the U-373 cell proliferation. Overall, these findings imply that Er-1 coupling to IL-2R and ERK immuno-enhances T-cell activity, and that this effect likely translates to an inhibition of glioma cell growth. -- Highlights: ► Euplotes pheromone Er-1 increases the growth of human Jurkat T-cells. ► Er-1 increases the T-cell production of specific cytokines. ► Er-1 activates interleukin-2 receptor and extracellular signal-regulated kinases. ► The immuno-enhancing effect of Er-1 on Jurkat cells translates to an inhibition of human glioma cell growth.« less

Curcumin inhibits vasculogenic mimicry through the downregulation of erythropoietin-producing hepatocellular carcinoma-A2, phosphoinositide 3-kinase and matrix metalloproteinase-2

PubMed Central

LIANG, YIMING; HUANG, MIN; LI, JIANWEN; SUN, XINLIN; JIANG, XIAODAN; LI, LIANGPING; KE, YIQUAN

2014-01-01

Glioblastomas (GBMs) are the most common and aggressive malignant primary brain tumors found in humans. In high-grade gliomas, vasculogenic mimicry (VM) is often detected. VM is the formation of de novo vascular networks by highly invasive tumor cells, instead of endothelial cells. An understanding of the mechanisms of VM formation will contribute to the targeted therapy of GBMs. In the present study, the efficacy of curcumin (CCM) on VM formation and its mechanisms were investigated. It was found that CCM inhibits the VM formation, proliferation, migration and invasion of human glioma U251 cells in a dose-dependent manner. Furthermore, CCM downregulated the protein and mRNA expression of erythropoietin-producing hepatocellular carcinoma-A2, phosphoinositide 3-kinase and matrix metalloproteinase-2, indicating that CCM may function through these factors for the inhibition of VM formation. These data provide novel insights into the use of CCM to antagonize VM, and may contribute to the angiogenesis-targeted therapy of malignant glioma. PMID:25202424

Improved Intratumoral Oxygenation Through Vascular Normalization Increases Glioma Sensitivity to Ionizing Radiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

McGee, Mackenzie C.; Hamner, J. Blair; Williams, Regan F.

2010-04-15

Purpose: Ionizing radiation, an important component of glioma therapy, is critically dependent on tumor oxygenation. However, gliomas are notable for areas of necrosis and hypoxia, which foster radioresistance. We hypothesized that pharmacologic manipulation of the typically dysfunctional tumor vasculature would improve intratumoral oxygenation and, thus, the antiglioma efficacy of ionizing radiation. Methods and Materials: Orthotopic U87 xenografts were treated with either continuous interferon-beta (IFN-beta) or bevacizumab, alone, or combined with cranial irradiation (RT). Tumor growth was assessed by quantitative bioluminescence imaging; the tumor vasculature using immunohistochemical staining, and tumor oxygenation using hypoxyprobe staining. Results: Both IFN-beta and bevaziumab profoundly affectedmore » the tumor vasculature, albeit with different cellular phenotypes. IFN-beta caused a doubling in the percentage of area of perivascular cell staining, and bevacizumab caused a rapid decrease in the percentage of area of endothelial cell staining. However, both agents increased intratumoral oxygenation, although with bevacizumab, the effect was transient, being lost by 5 days. Administration of IFN-beta or bevacizumab before RT was significantly more effective than any of the three modalities as monotherapy or when RT was administered concomitantly with IFN-beta or bevacizumab or 5 days after bevacizumab. Conclusion: Bevacizumab and continuous delivery of IFN-beta each induced significant changes in glioma vascular physiology, improving intratumoral oxygenation and enhancing the antitumor activity of ionizing radiation. Additional investigation into the use and timing of these and other agents that modify the vascular phenotype, combined with RT, is warranted to optimize cytotoxic activity.« less

Silencing Nrf2 impairs glioma cell proliferation via AMPK-activated mTOR inhibition

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jia, Yue; Wang, Handong, E-mail: njhdwang@hotmail.com; Wang, Qiang

Gliomas are the leading cause of death among adults with primary brain malignancies. Treatment for malignant gliomas remains limited, and targeted therapies have been incompletely explored. Nuclear factor erythroid 2-related factor 2 (Nrf2), a key transcription regulator for antioxidant and detoxification enzymes, is abundantly expressed in cancer cells. In this study, the role and mechanism of Nrf2 in cancer cell proliferation was investigated in multiple glioma cell lines. We first evaluated the expression patterns of Nrf2 in four glioma cell lines and found all four cell lines expressed Nrf2, but the highest level was observed in U251 cells. We further evaluatedmore » the biological functions of Nrf2 in U251 glioma cell proliferation by specific inhibition of Nrf2 using short hairpin RNA (shRNA). We found that Nrf2 depletion inhibited glioma cell proliferation. Nrf2 depletion also decreased colony formation in U251 cells stably expressing Nrf2 shRNA compared to scrambled control shRNA. Moreover, suppression of Nrf2 expression could lead to ATP depletion (with concomitant rise in AMP/ATP ratio) and consequently to AMPK-activated mTOR inhibition. Finally, activation of adenosine monophosphate–activated protein kinase (AMPK) by treated with phenformin, an AMPK agonist, can mimic the inhibitory effect of Nrf2 knockdown in U251 cells. In conclusion, our findings will shed light to the role and mechanism of Nrf2 in regulating glioma proliferation via ATP-depletion-induced AMPK activation and consequent mTOR inhibition, a novel insight into our understanding the role and mechanism of Nrf2 in glioma pathoetiology. To our knowledge, this is also the first report to provide a rationale for the implication of cross-linking between Nrf2 and mTOR signaling.« less

Characterization and evaluation of DOTA-conjugated Bombesin/RGD-antagonists for prostate cancer tumor imaging and therapy.

PubMed

Stott Reynolds, Tamila J; Schehr, Rebecca; Liu, Dijie; Xu, Jingli; Miao, Yubin; Hoffman, Timothy J; Rold, Tammy L; Lewis, Michael R; Smith, Charles J

2015-02-01

Here we present the metallation, characterization, in vivo and in vitro evaluations of dual-targeting, peptide-based radiopharmaceuticals with utility for imaging and potentially treating prostate tumors by virtue of their ability to target the αVβ3 integrin or the gastrin releasing peptide receptor (GRPr). [RGD-Glu-6Ahx-RM2] (RGD: Arg-Gly-Asp; Glu: glutamic acid; 6-Ahx: 6-amino hexanoic acid; RM2: (D-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2)) was conjugated to a DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) bifunctional chelator (BFCA) purified via reversed-phase high-performance liquid chromatography (RP-HPLC), characterized by electrospray ionization-mass spectrometry (ESI-MS), and radiolabeled with (111)In or (177)Lu. Natural-metallated compounds were assessed for binding affinity for the αVβ3 integrin or GRPr in human glioblastoma U87-MG and prostate PC-3 cell lines and stability prior to in vivo evaluation in normal CF-1 mice and SCID mice xenografted with PC-3 cells. Competitive displacement binding assays with PC-3 and U87-MG cells revealed high to moderate binding affinity for the GRPr or the αVβ3 integrin (IC50 range of 5.39±1.37 nM to 9.26±0.00 nM in PC-3 cells, and a range of 255±47 nM to 321±85 nM in U87-MG cells). Biodistribution studies indicated high tumor uptake in PC-3 tumor-bearing mice (average of 7.40±0.53% ID/g at 1h post-intravenous injection) and prolonged retention of tracer (mean of 4.41±0.91% ID/g at 24h post-intravenous injection). Blocking assays corroborated the specificity of radioconjugates for each target. Micro-single photon emission computed tomography (microSPECT) confirmed favorable radiouptake profiles in xenografted mice at 20h post-injection. [RGD-Glu-[(111)In-DO3A]-6-Ahx-RM2] and [RGD-Glu-[(177)Lu- DO3A]-6-Ahx-RM2] show favorable pharmacokinetic and radiouptake profiles, meriting continued evaluation for molecular imaging in murine U87-MG/PC-3 xenograft models and radiotherapy studies with (177)Lu and (90)Y conjugates. These heterovalent, peptide-targeting ligands perform comparably with many mono- and multivalent conjugates with the potential benefit of increased sensitivity for detecting cancer cells exhibiting differential expression of target receptors. Published by Elsevier Inc.

H2-P, a honokiol derivative, exerts anti-angiogenesis effects via c-MYC signaling pathway in glioblastoma.

PubMed

Wang, Ting; Chen, Wei; Wu, Jialin

2018-04-01

H2-P, a derivative of honokiol, was first synthesized in our laboratory. Compared with honokiol, H2-P has even high anti-tumor activity. In the present study, we evaluated the ability of H2-P to inhibit the survival rate in four gliomas cell lines. The result showed that H2-P could significantly inhibit proliferation of gliomas cells in a dose-dependent manner (IC50 U251  = 9.03, IC50 SHG-44  = 10.74, IC50 U78  = 19.87, and IC50 c6  = 22.56 nM). Furthermore, to determine the mechanism underlying the anti-gliomas effects of H2-P, six kinase activities was detected by Z'-LYTE™ system. The high-throughput screening shown that effect targets of H2-P were MEK and VEGFR2. We also studied the inhibition of H2-P vascular endothelial cells (EA.HY926). The data shown that H2-P could increase endothelial cells apoptosis rate, while inhibiting endothelial cell proliferation (IC50 EA.hy926  = 16.11 nM) and migration. Besides, we investigated anti-angiogenesis of H2-P in the rat thoracic aorta rings, chicken chorioallantoic membrane (CAM), and capillary tube formation models. H2-P showed strong inhibition of angiogenesis. Moreover, we found that H2-P also could reduce tumor volume in mice significantly (P < 0.01), and downregulate gene expression level of VEGFR2, MEK, and c-MYC in tumor. These data suggest that H2-P have an excellent anti-tumor activity by exerting anti-angiogenesis effects via c-MYC signaling pathway in glioblastoma (GBM). © 2017 Wiley Periodicals, Inc.

1800MHz Microwave Induces p53 and p53-Mediated Caspase-3 Activation Leading to Cell Apoptosis In Vitro

PubMed Central

Xing, Fuqiang; Zhan, Qiuqiang; He, Yiduo; Cui, Jiesheng; He, Sailing; Wang, Guanyu

2016-01-01

Recent studies have reported that exposure of mammalian cells to microwave radiation may have adverse effects such as induction of cell apoptosis. However, the molecular mechanisms underlying microwave induced mammalian cell apoptosis are not fully understood. Here, we report a novel mechanism: exposure to 1800MHz microwave radiation induces p53-dependent cell apoptosis through cytochrome c-mediated caspase-3 activation pathway. We first measured intensity of microwave radiation from several electronic devices with an irradiation detector. Mouse NIH/3T3 and human U-87 MG cells were then used as receivers of 1800MHz electromagnetic radiation (EMR) at a power density of 1209 mW/m2. Following EMR exposure, cells were analyzed for viability, intracellular reactive oxygen species (ROS) generation, DNA damage, p53 expression, and caspase-3 activity. Our analysis revealed that EMR exposure significantly decreased viability of NIH/3T3 and U-87 MG cells, and increased caspase-3 activity. ROS burst was observed at 6 h and 48 h in NIH/3T3 cells, while at 3 h in U-87 MG cells. Hoechst 33258 staining and in situ TUNEL assay detected that EMR exposure increased DNA damage, which was significantly restrained in the presence of N-acetyl-L-cysteine (NAC, an antioxidant). Moreover, EMR exposure increased the levels of p53 protein and p53 target gene expression, promoted cytochrome c release from mitochondrion, and increased caspase-3 activity. These events were inhibited by pretreatment with NAC, pifithrin-α (a p53 inhibitor) and caspase inhibitor. Collectively, our findings demonstrate, for the first time, that 1800MHz EMR induces apoptosis-related events such as ROS burst and more oxidative DNA damage, which in turn promote p53-dependent caspase-3 activation through release of cytochrome c from mitochondrion. These findings thus provide new insights into physiological mechanisms underlying microwave-induced cell apoptosis. PMID:27689798

Studies of the antitumor mechanism of action of dermaseptin B2, a multifunctional cationic antimicrobial peptide, reveal a partial implication of cell surface glycosaminoglycans.

PubMed

Dos Santos, Célia; Hamadat, Sabah; Le Saux, Karen; Newton, Clara; Mazouni, Meriem; Zargarian, Loussiné; Miro-Padovani, Mickael; Zadigue, Patricia; Delbé, Jean; Hamma-Kourbali, Yamina; Amiche, Mohamed

2017-01-01

Dermaseptin-B2 (DRS-B2) is a multifunctional cationic antimicrobial peptide (CAP) isolated from frog skin secretion. We previously reported that DRS-B2 possesses anticancer and antiangiogenic activities in vitro and in vivo. In the present study, we evaluated the antiproliferative activity of DRS-B2 on numerous tumor cell lines, its cell internalization and studies of its molecular partners as well as their influences on its structure. Confocal microscopy using ([Alexa594]-(Cys0)-DRS-B2) shows that in sensitive human tumor cells (PC3), DRS-B2 seems to accumulate rapidly at the cytoplasmic membranes and enters the cytoplasm and the nucleus, while in less sensitive tumor cells (U87MG), DRS-B2 is found packed in vesicles at the cell membrane. Furthermore FACS analysis shows that PC3 cells viability decreases after DRS-B2 treatment while U87 MG seems to be unaffected. However, "pull down" experiments performed with total protein pools from PC3 or U87MG cells and the comparison between the antiproliferative effect of DRS-B2 and its synthetic analog containing all D-amino acids suggest the absence of a stereo-selective protein receptor. Pretreatment of PC3 cells with sodium chlorate, decreases the antiproliferative activity of DRS-B2. This activity is partially restored after addition of exogenous chondroitin sulfate C (CS-C). Moreover, we demonstrate that at nanomolar concentrations CS-C potentiates the antiproliferative effect of DRS-B2. These results highlight the partial implication of glycosaminoglycans in the mechanism of antiproliferative action of DRS-B2. Structural analysis of DRS-B2 by circular dichroism in the presence of increasing concentration of CS-C shows that DRS-B2 adopts an α-helical structure. Finally, structure-activity-relationship studies suggest a key role of the W residue in position 3 of the DRS-B2 sequence for its antiproliferative activity.

Gallium Maltolate Disrupts Tumor Iron Metabolism and Retards the Growth of Glioblastoma by Inhibiting Mitochondrial Function and Ribonucleotide Reductase.

PubMed

Chitambar, Christopher R; Al-Gizawiy, Mona M; Alhajala, Hisham S; Pechman, Kimberly R; Wereley, Janine P; Wujek, Robert; Clark, Paul A; Kuo, John S; Antholine, William E; Schmainda, Kathleen M

2018-06-01

Gallium, a metal with antineoplastic activity, binds transferrin (Tf) and enters tumor cells via Tf receptor1 (TfR1); it disrupts iron homeostasis leading to cell death. We hypothesized that TfR1 on brain microvascular endothelial cells (BMEC) would facilitate Tf-Ga transport into the brain enabling it to target TfR-bearing glioblastoma. We show that U-87 MG and D54 glioblastoma cell lines and multiple glioblastoma stem cell (GSC) lines express TfRs, and that their growth is inhibited by gallium maltolate (GaM) in vitro After 24 hours of incubation with GaM, cells displayed a loss of mitochondrial reserve capacity followed by a dose-dependent decrease in oxygen consumption and a decrease in the activity of the iron-dependent M2 subunit of ribonucleotide reductase (RRM2). IHC staining of rat and human tumor-bearing brains showed that glioblastoma, but not normal glial cells, expressed TfR1 and RRM2, and that glioblastoma expressed greater levels of H- and L-ferritin than normal brain. In an orthotopic U-87 MG glioblastoma xenograft rat model, GaM retarded the growth of brain tumors relative to untreated control ( P = 0.0159) and reduced tumor mitotic figures ( P = 0.045). Tumors in GaM-treated animals displayed an upregulation of TfR1 expression relative to control animals, thus indicating that gallium produced tumor iron deprivation. GaM also inhibited iron uptake and upregulated TfR1 expression in U-87 MG and D54 cells in vitro We conclude that GaM enters the brain via TfR1 on BMECs and targets iron metabolism in glioblastoma in vivo, thus inhibiting tumor growth. Further development of novel gallium compounds for brain tumor treatment is warranted. Mol Cancer Ther; 17(6); 1240-50. ©2018 AACR . ©2018 American Association for Cancer Research.

Studies of the antitumor mechanism of action of dermaseptin B2, a multifunctional cationic antimicrobial peptide, reveal a partial implication of cell surface glycosaminoglycans

PubMed Central

Dos Santos, Célia; Hamadat, Sabah; Le Saux, Karen; Newton, Clara; Mazouni, Meriem; Zargarian, Loussiné; Miro-Padovani, Mickael; Zadigue, Patricia; Delbé, Jean; Hamma-Kourbali, Yamina

2017-01-01

Dermaseptin-B2 (DRS-B2) is a multifunctional cationic antimicrobial peptide (CAP) isolated from frog skin secretion. We previously reported that DRS-B2 possesses anticancer and antiangiogenic activities in vitro and in vivo. In the present study, we evaluated the antiproliferative activity of DRS-B2 on numerous tumor cell lines, its cell internalization and studies of its molecular partners as well as their influences on its structure. Confocal microscopy using ([Alexa594]-(Cys0)-DRS-B2) shows that in sensitive human tumor cells (PC3), DRS-B2 seems to accumulate rapidly at the cytoplasmic membranes and enters the cytoplasm and the nucleus, while in less sensitive tumor cells (U87MG), DRS-B2 is found packed in vesicles at the cell membrane. Furthermore FACS analysis shows that PC3 cells viability decreases after DRS-B2 treatment while U87 MG seems to be unaffected. However, "pull down" experiments performed with total protein pools from PC3 or U87MG cells and the comparison between the antiproliferative effect of DRS-B2 and its synthetic analog containing all D-amino acids suggest the absence of a stereo-selective protein receptor. Pretreatment of PC3 cells with sodium chlorate, decreases the antiproliferative activity of DRS-B2. This activity is partially restored after addition of exogenous chondroitin sulfate C (CS-C). Moreover, we demonstrate that at nanomolar concentrations CS-C potentiates the antiproliferative effect of DRS-B2. These results highlight the partial implication of glycosaminoglycans in the mechanism of antiproliferative action of DRS-B2. Structural analysis of DRS-B2 by circular dichroism in the presence of increasing concentration of CS-C shows that DRS-B2 adopts an α-helical structure. Finally, structure-activity-relationship studies suggest a key role of the W residue in position 3 of the DRS-B2 sequence for its antiproliferative activity. PMID:28797092

Bevacizumab Plus Irinotecan in Recurrent WHO Grade 3 Malignant Gliomas

PubMed Central

Desjardins, Annick; Reardon, David A.; Herndon, James E.; Marcello, Jennifer; Quinn, Jennifer A.; Rich, Jeremy N.; Sathornsumetee, Sith; Gururangan, Sridharan; Sampson, John; Bailey, Leighann; Bigner, Darell D.; Friedman, Allan H.; Friedman, Henry S.; Vredenburgh, James J.

2013-01-01

Purpose Although patients with newly diagnosed WHO grade 3 malignant glioma have a more favorable prognosis than those with WHO grade 4 malignant glioma, salvage therapies following recurrence offer essentially palliative benefit. We did a phase II trial of bevacizumab, a monoclonal antibody to vascular endothelial growth factor, in combination with irinotecan for patients with recurrent grade 3 malignant glioma. Experimental Design Upon documentation of adequate safety among an initial cohort of nine patients treated with bevacizumab (10 mg/kg) and irinotecan every 14 days, a second cohort (n = 24) was treated with bevacizumab (15 mg/kg) every 3 weeks with irinotecan on days 1, 8, 22, and 29 of each 42-day cycle. For both cohorts, the dose of irinotecan was 340 mg/m2 for patients on enzyme-inducing antiepileptic drugs (EIAED) and 125 mg/m2 for patients not on EIAEDs. After each 6-week cycle, patients were evaluated with a physical examination and magnetic resonance imaging. Results The 6-month progression-free survival was 55% (95% confidence interval, 36–70%). The 6-month overall survival was 79% (95% confidence interval, 61–89%). Twenty patients (61%) had at least a partial response. Outcome did not differ between the two treatment cohorts. Significant adverse events were infrequent and included a central nervous system hemorrhage in one patient, and one patient who developed thrombotic thrombocytopenic purpura. Conclusion Bevacizumab and irinotecan is an active regimen with acceptable toxicity for patients with recurrent WHO grade 3 malignant glioma. PMID:18981004

Downregulation of mitochondrial UQCRB inhibits cancer stem cell-like properties in glioblastoma.

PubMed

Jung, Narae; Kwon, Ho Jeong; Jung, Hye Jin

2018-01-01

Glioblastoma stem cell targeted therapies have become a powerful strategy for the treatment of this deadliest brain tumor. We demonstrate for the first time that downregulation of mitochondrial ubiquinol-cytochrome c reductase binding protein (UQCRB) inhibits the cancer stem cell-like properties in human glioblastoma cells. The synthetic small molecules targeting UQCRB significantly suppressed not only the self-renewal capacity such as growth and neurosphere formation, but also the metastatic potential such as migration and invasion of glioblastoma stem‑like cells (GSCs) derived from U87MG and U373MG at subtoxic concentrations. Notably, the UQCRB inhibitors repressed c‑Met-mediated downstream signal transduction and hypoxia‑inducible factor‑1α (HIF‑1α) activation, thereby reducing the expression levels of GSC markers including CD133, Nanog, Oct4 and Sox2 in the GSCs. Furthermore, the UQCRB inhibitors decreased mitochondrial ROS generation and mitochondrial membrane potential in the GSCs, indicating that they regulate the mitochondrial function in GSCs. Indeed, the knockdown of UQCRB gene by UQCRB siRNA significantly inhibited the cancer stem cell-like phenotypes as well as the expression of stemness markers by blocking mitochondrial ROS/HIF‑1α/c‑Met pathway in U87MG GSCs. These findings suggest that UQCRB and its inhibitors could be a new therapeutic target and lead compounds for eliminating cancer stem cells in glioblastoma.

Modulation of substance P signaling by dipeptidyl peptidase-IV enzymatic activity in human glioma cell lines.

PubMed

Busek, P; Stremenová, J; Krepela, E; Sedo, A

2008-01-01

Dipeptidyl peptidase-IV (DPP-IV, CD26) is a serine protease almost ubiquitously expressed on cell surface and present in body fluids. DPP-IV has been suggested to proteolytically modify a number of biologically active peptides including substance P (SP) and the chemokine stromal cell derived factor-1alpha (SDF-1alpha, CXCL12). SP and SDF-1alpha have been implicated in the regulation of multiple biological processes and also induce responses that may be relevant for glioma progression. Both SP and SDF-1alpha are signaling through cell surface receptors and use intracellular calcium as a second messenger. The effect of DPP-IV on intracellular calcium mobilization mediated by SP and SDF-1alpha was monitored in suspension of wild type U373 and DPP-IV transfected U373DPPIV glioma cells using indicator FURA-2. Nanomolar concentrations of SP triggered a transient dose dependent increase in intracellular calcium rendering the cells refractory to repeated stimulation, while SDF-1 had no measurable effect. SP signaling in DPP-IV overexpressing U373DPPIV cells was not substantially different from that in wild type cells. However, preincubation of SP with the DPP-IV overexpressing cells lead to the loss of its signaling potential, which could be prevented with DPP-IV inhibitors. Taken together, DPP-IV may proteolytically inactivate local mediators involved in gliomagenesis.

CCNG2 Overexpression Mediated by AKT Inhibits Tumor Cell Proliferation in Human Astrocytoma Cells.

PubMed

Zhang, Danfeng; Wang, Chunhui; Li, Zhenxing; Li, Yiming; Dai, Dawei; Han, Kaiwei; Lv, Liquan; Lu, Yicheng; Hou, Lijun; Wang, Junyu

2018-01-01

The cyclin family protein CCNG2 has an important inhibitory role in cancer initiation and progression, but the exact mechanism is still unknown. In this study, we examined the relationship between CCNG2 and the malignancy of astrocytomas and whether the AKT pathway, which is upregulated in astrocytomas, may inhibit CCNG2 expression. CCNG2 expression was found to be negatively associated with the pathological grade and proliferative activity of astrocytomas, as the highest expression was found in control brain tissue ( N  = 31), whereas the lowest expression was in high-grade glioma tissue ( N  = 31). Additionally, CCNG2 overexpression in glioma cell lines, T98G and U251 inhibited proliferation and arrested cells in the G0/G1 phase. Moreover, CCNG2 overexpression could increase glioma cells apoptosis. In contrast, AKT activity increased in glioma cells that had low CCNG2 expression. Expression of CCNG2 was higher in cells treated with the AKT kinase inhibitor MK-2206 indicating that the presence of phosphorylated AKT may inhibit the expression of CCNG2. Inhibition of AKT also led to decreased colony formation in T98G and U251 cells and knocked down of CCNG2 reversed the result. Finally, overexpression of CCNG2 in glioma cells reduced tumor volume in a murine model. To conclude, low expression of CCNG2 correlated with the severity astrocytoma and CCNG2 overexpression could induce apoptosis and inhibit proliferation. Inhibition of AKT activity increased the expression of CCNG2. The present study highlights the regulatory consequences of CCNG2 expression and AKT activity in astrocytoma tumorigenesis and the potential use of CCNG2 in anticancer treatment.

Gene therapy for human glioblastoma using neurotropic JC virus-like particles as a gene delivery vector.

PubMed

Chao, Chun-Nun; Yang, Yu-Hsuan; Wu, Mu-Sheng; Chou, Ming-Chieh; Fang, Chiung-Yao; Lin, Mien-Chun; Tai, Chien-Kuo; Shen, Cheng-Huang; Chen, Pei-Lain; Chang, Deching; Wang, Meilin

2018-02-02

Glioblastoma multiforme (GBM), the most common malignant brain tumor, has a short period of survival even with recent multimodality treatment. The neurotropic JC polyomavirus (JCPyV) infects glial cells and oligodendrocytes and causes fatal progressive multifocal leukoencephalopathy in patients with AIDS. In this study, a possible gene therapy strategy for GBM using JCPyV virus-like particles (VLPs) as a gene delivery vector was investigated. We found that JCPyV VLPs were able to deliver the GFP reporter gene into tumor cells (U87-MG) for expression. In an orthotopic xenograft model, nude mice implanted with U87 cells expressing the near-infrared fluorescent protein and then treated by intratumoral injection of JCPyV VLPs carrying the thymidine kinase suicide gene, combined with ganciclovir administration, exhibited significantly prolonged survival and less tumor fluorescence during the experiment compared with controls. Furthermore, JCPyV VLPs were able to protect and deliver a suicide gene to distal subcutaneously implanted U87 cells in nude mice via blood circulation and inhibit tumor growth. These findings show that metastatic brain tumors can be targeted by JCPyV VLPs carrying a therapeutic gene, thus demonstrating the potential of JCPyV VLPs to serve as a gene therapy vector for the far highly treatment-refractory GBM.

Canine (Pet Dog) Tumor Microsurgery and Intratumoral Concentration and Safety of Metronomic Chlorambucil for Spontaneous Glioma: A Phase I Clinical Trial.

PubMed

Bentley, R Timothy; Thomovsky, Stephanie A; Miller, Margaret A; Knapp, Deborah W; Cohen-Gadol, Aaron A

2018-06-04

Metronomic (daily low-dose) chlorambucil requires further study before use in human patients with glioma. The aim of this study was to investigate distribution and safety of metronomic chlorambucil in naturally occurring canine glioma. Eight client-owned (pet) dogs with newly diagnosed spontaneous glioma were prospectively enrolled. Chlorambucil was administered preoperatively at 4 mg/m 2 every 24 hours for ≥3 days and continued postoperatively until death or dose-limiting adverse events. Chlorambucil concentrations in the surgical glioma specimen, cerebrospinal fluid, and serum were analyzed. Dogs additionally received lomustine postoperatively. Dogs were monitored for seizures, myoclonus, cytopenias, and tumor recurrence. Complete microsurgical resection was achieved in 7 oligodendrogliomas and 1 astrocytoma (6 high grade, 2 low grade). Median surgical glioma specimen chlorambucil concentration was 0.52 ng/g (range, 0-2.62 ng/g), or 37% (range, 0%-178%) of serum concentration. Median cerebrospinal fluid concentration was 0.1 ng/mL (range, 0-0.3 ng/mL). Chlorambucil was not associated with increase in seizure activity. Six dogs displayed prolonged seizure-free intervals. There was no myoclonus. Three dogs developed asymptomatic thrombocytopenia after 8-12 months of chlorambucil. Median progression-free survival was 253 days (range, 63-860 days). Median overall survival was 257 days (range, 64-860 days). The presence of intratumoral chlorambucil indicated an altered blood-brain barrier that varied from case to case. Despite sporadic previous reports of neurotoxicity, prolonged seizure-free intervals supported a high safety margin at this dose in this species. Metronomic chlorambucil was well tolerated. Spontaneous canine glioma offers a robust preclinical model. Copyright © 2018 Elsevier Inc. All rights reserved.

Apigenin Inhibits Cancer Stem Cell-Like Phenotypes in Human Glioblastoma Cells via Suppression of c-Met Signaling.

PubMed

Kim, Boram; Jung, Narae; Lee, Sanghun; Sohng, Jae Kyung; Jung, Hye Jin

2016-11-01

Glioblastoma (GBM) is a highly malignant human brain tumor with limited treatment choices. The extremely aggressive characteristics of GBM result from GBM stem cells (GSCs), a subpopulation in tumor having self-renewal potential and resistance to chemotherapy and radiotherapy. Therefore, eliminating GSCs is an effective strategy to treat this fatal disease. In this study, we investigated the therapeutic effects of dietary flavonoids, including apigenin, quercetin, and naringenin, against cancer stem cell-like phenotypes of human GBM cell lines U87MG and U373MG. Among flavonoids studied, apigenin and quercetin significantly suppressed not only the self-renewal capacity such as cell growth and clonogenicity, but also the invasiveness of GBM stem-like cells. Notably, apigenin blocked the phosphorylation of c-Met and its downstream effectors, transducer and activator of transcription 3, AKT (Protein kinase B), and mitogen-activated protein kinase in the GSCs, thereby reducing the expression levels of GSC markers such as CD133, Nanog, and Sox2. These results suggest that the GSC inhibition effect of apigenin may be caused by downregulation of c-Met signaling pathway. Copyright © 2016 John Wiley & Sons, Ltd.

The histone deacetylase SIRT6 suppresses the expression of the RNA-binding protein PCBP2 in glioma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Xin; Hao, Bin; Liu, Ying

Highlights: • PCBP2 expression is over-expressed in human glioma tissues and cell lines. • SIRT6 is decreased in glioma and correlated with PCBP2. • SIRT6 inhibits PCBP2 expression by deacetylating H3K9. • SIRT6 inhibits glioma growth in vitro and in vivo. - Abstract: More than 80% of tumors that occur in the brain are malignant gliomas. The prognosis of glioma patients is still poor, which makes glioma an urgent subject of cancer research. Previous evidence and our present data show that PCBP2 is over-expressed in human glioma tissues and predicts poor outcome. However, the mechanism by which PCBP2 is regulatedmore » in glioma remains elusive. We find that SIRT6, one of the NAD{sup +}-dependent class III deacetylase SIRTUINs, is down-regulated in human glioma tissues and that the level of SIRT6 is negatively correlated with PCBP2 level while H3K9ac enrichment on the promoter of PCBP2 is positively correlated with PCBP2 expression. Furthermore, we identify PCBP2 as a target of SIRT6. We demonstrate that PCBP2 expression is inhibited by SIRT6, which depends upon deacetylating H3K9ac. Finally, our results reveal that SIRT6 inhibits glioma cell proliferation and colony formation in vitro and glioma cell growth in vivo in a PCBP2 dependent manner. In summary, our findings implicate that SIRT6 inhibits PCBP2 expression through deacetylating H3K9ac and SIRT6 acts as a tumor suppressor in human glioma.« less

Naringin inhibits the invasion and migration of human glioblastoma cell via downregulation of MMP-2 and MMP-9 expression and inactivation of p38 signaling pathway.

PubMed

Aroui, Sonia; Najlaoui, Feten; Chtourou, Yassine; Meunier, Annie-Claire; Laajimi, Amel; Kenani, Abderraouf; Fetoui, Hamadi

2016-03-01

Gliomas are the most common and malignant primary brain tumors. They are associated with a poor prognosis despite the availability of multiple therapeutic options. Naringin, a common dietary flavonoid abundantly present in fruits and vegetables, is believed to possess strong anti-proliferative and anti-cancer properties. However, there are no reports describing its effects on the invasion and migration of glioblastoma cell lines. Our results showed that the treatment of U251 glioma cell lines with different concentrations of naringin inhibited the invasion and migration of these cells. In addition, we revealed a decrease in the levels of matrix metalloproteinases (MMP-2) and (MMP-9) expression as well as proteinase activity in U251 glioma cells. In contrast, the expression of tissue inhibitor of metalloproteinases (TIMP-1) and (TIMP-2) was increased. Furthermore, naringin treatment decreased significantly the phosphorylated level of p38. Combined treatment with a p38 inhibitor (SB203580) resulted in the synergistic reduction of MMP-2 and MMP-9 expressions correlated with an increase of TIMP-1 and TIMP-2 expressions and the anti-invasive properties. However, p38 chemical activator (anisomycin) could block these effects produced by naringin, suggesting a direct downregulation of the p38 signaling pathway. These data suggest that naringin may have therapeutic potential for controlling invasiveness of malignant gliomas by inhibiting of p38 signal transduction pathways.

A novel vaccine containing EphA2 epitope and LIGHT plasmid induces robust cellular immunity against glioma U251 cells.

PubMed

Chen, Hongjie; Yuan, Bangqing; Zheng, Zhaocong; Liu, Zheng; Wang, Shousen; Liu, Yong

2011-01-01

EphA2 is a receptor tyrosine kinase and can be acted as an attractive antigen for glioma vaccines. In addition, LIGHT plays an important role on enhancing T cell proliferation and cytokine production. To improve the CTL mediated immune response against glioma cells, we prepared the novel vaccine containing EphA2(883-891) peptide (TLADFDPRV) and LIGHT plasmid and utilized it to immunize the HLA-A2 transgenic HHD mice. In addition, trimera mice were immunized with the novel vaccine to elicit the antitumor immune response. The results demonstrated that the novel vaccine could induce robust cellular immunity against glioma U251 cells without lysing autologous lymphocytes. Moreover, the novel vaccine could significantly inhibit the tumor growth and prolong the life span of tumor bearing mice. These findings suggested that the novel vaccine containing EphA2 epitope and LIGHT plasmid could induce anti-tumor immunity against U251 cells expressing EphA2, and provided a promising strategy for glioma immunotherapy. Copyright © 2011 Elsevier Inc. All rights reserved.

Cell cycle phase dependent emergence of thymidylate synthase studied by monoclonal antibody (M-TS-4).

PubMed

Shibui, S; Hoshino, T; Iwasaki, K; Nomura, K; Jastreboff, M M

1989-05-01

A method of identifying thymidylate synthase (TS) at the cellular level was developed using anti-TS monoclonal antibody (M-TS-4), a monoclonal antibody created against purified TS from a HeLa cell line. In HeLa cells and four human glioma cell lines (U-251, U-87, 343-MGA, and SF-188), TS was identified primarily in the cytoplasm. Autoradiographic and flow cytometric studies showed that TS appeared mainly in the G1 phase and subsided early in the S phase; thus, the G1 phase can be divided into TS-positive and -negative fractions. Nuclear TS was not demonstrated unequivocally with M-TS-4, and the relationship between nuclear TS and DNA synthesis could not be determined. Although the percentage of TS-positive cells was larger than the S-phase fraction measured by autoradiography after a pulse of tritiated thymidine or by the immunoperoxidase method using BUdR, the ratios were within a similar range (1.2-1.4) in all cell lines studied. Therefore, the S-phase fraction can be estimated indirectly from the percentage of TS-positive cells measured by M-TS-4. Because the emergence of TS detected by our method is cell cycle dependent, M-TS-4 may be useful for biochemical studies of TS and for cytokinetic analysis.

A syngeneic glioma model to assess the impact of neural progenitor target cell age on tumor malignancy

PubMed Central

Mikheev, Andrei M; Stoll, Elizabeth A; Mikheeva, Svetlana A; Maxwell, John-Patrick; Jankowski, Pawel P; Ray, Sutapa; Uo, Takuma; Morrison, Richard S; Horner, Philip J; Rostomily, Robert C

2010-01-01

Summary Human glioma incidence, malignancy and treatment resistance are directly proportional to patient age. Cell intrinsic factors are reported to contribute to human age-dependent glioma malignancy but suitable animal models to examine the role of aging are lacking. Here we developed an orthotopic syngeneic glioma model to test the hypothesis that the age of neural progenitor cells (NPCs), presumed cells of glioma origin, influences glioma malignancy. Gliomas generated from transformed donor 3-, 12-, and 18-month-old NPCs in same-aged adult hosts all formed highly invasive glial tumors that phenocopied the human disease. Survival analysis indicated increased malignancy of gliomas generated from older 12- and 18-month-old transformed NPCs compared with their 3-month counterparts (median survival of 38.5 and 42.5 vs. 77 days, respectively). This study showed for the first time that age of target cells at the time of transformation can affect malignancy and demonstrated the feasibility of a syngeneic model using transformed NPCs for future examination of the relative impacts of age-related cell intrinsic and cell-extrinsic factors in glioma malignancy. PMID:19489742

Diagnostic accuracy of magnetic resonance imaging techniques for treatment response evaluation in patients with high-grade glioma, a systematic review and meta-analysis.

PubMed

van Dijken, Bart R J; van Laar, Peter Jan; Holtman, Gea A; van der Hoorn, Anouk

2017-10-01

Treatment response assessment in high-grade gliomas uses contrast enhanced T1-weighted MRI, but is unreliable. Novel advanced MRI techniques have been studied, but the accuracy is not well known. Therefore, we performed a systematic meta-analysis to assess the diagnostic accuracy of anatomical and advanced MRI for treatment response in high-grade gliomas. Databases were searched systematically. Study selection and data extraction were done by two authors independently. Meta-analysis was performed using a bivariate random effects model when ≥5 studies were included. Anatomical MRI (five studies, 166 patients) showed a pooled sensitivity and specificity of 68% (95%CI 51-81) and 77% (45-93), respectively. Pooled apparent diffusion coefficients (seven studies, 204 patients) demonstrated a sensitivity of 71% (60-80) and specificity of 87% (77-93). DSC-perfusion (18 studies, 708 patients) sensitivity was 87% (82-91) with a specificity of 86% (77-91). DCE-perfusion (five studies, 207 patients) sensitivity was 92% (73-98) and specificity was 85% (76-92). The sensitivity of spectroscopy (nine studies, 203 patients) was 91% (79-97) and specificity was 95% (65-99). Advanced techniques showed higher diagnostic accuracy than anatomical MRI, the highest for spectroscopy, supporting the use in treatment response assessment in high-grade gliomas. • Treatment response assessment in high-grade gliomas with anatomical MRI is unreliable • Novel advanced MRI techniques have been studied, but diagnostic accuracy is unknown • Meta-analysis demonstrates that advanced MRI showed higher diagnostic accuracy than anatomical MRI • Highest diagnostic accuracy for spectroscopy and perfusion MRI • Supports the incorporation of advanced MRI in high-grade glioma treatment response assessment.

Dynamic-contrast-enhanced-MRI with extravasating contrast reagent: Rat cerebral glioma blood volume determination

NASA Astrophysics Data System (ADS)

Li, Xin; Rooney, William D.; Várallyay, Csanád G.; Gahramanov, Seymur; Muldoon, Leslie L.; Goodman, James A.; Tagge, Ian J.; Selzer, Audrey H.; Pike, Martin M.; Neuwelt, Edward A.; Springer, Charles S.

2010-10-01

The accurate mapping of the tumor blood volume (TBV) fraction ( vb) is a highly desired imaging biometric goal. It is commonly thought that achieving this is difficult, if not impossible, when small molecule contrast reagents (CRs) are used for the T1-weighted (Dynamic-Contrast-Enhanced) DCE-MRI technique. This is because angiogenic malignant tumor vessels allow facile CR extravasation. Here, a three-site equilibrium water exchange model is applied to DCE-MRI data from the cerebrally-implanted rat brain U87 glioma, a tumor exhibiting rapid CR extravasation. Analyses of segments of the (and the entire) DCE data time-course with this "shutter-speed" pharmacokinetic model, which admits finite water exchange kinetics, allow TBV estimation from the first-pass segment. Pairwise parameter determinances were tested with grid searches of 2D parametric error surfaces. Tumor blood volume ( vb), as well as ve (the extracellular, extravascular space volume fraction), and Ktrans (a CR extravasation rate measure) parametric maps are presented. The role of the Patlak Plot in DCE-MRI is also considered.

Distribution of the phosphatidylinositol 3-kinase inhibitors Pictilisib (GDC-0941) and GNE-317 in U87 and GS2 intracranial glioblastoma models-assessment by matrix-assisted laser desorption ionization imaging.

PubMed

Salphati, Laurent; Shahidi-Latham, Sheerin; Quiason, Cristine; Barck, Kai; Nishimura, Merry; Alicke, Bruno; Pang, Jodie; Carano, Richard A; Olivero, Alan G; Phillips, Heidi S

2014-07-01

Glioblastoma multiforme (GBM) is the most common primary brain tumor in adults, and the limited available treatment options have not meaningfully impacted patient survival in the past decades. Such poor outcomes can be at least partly attributed to the inability of most drugs tested to cross the blood-brain barrier and reach all areas of the glioma. The objectives of these studies were to visualize and compare by matrix-assisted laser desorption ionization (MALDI) imaging mass spectrometry the brain and tumor distribution of the phosphatidylinositol 3-kinase (PI3K) inhibitors pictilisib (GDC-0941, 2-(1H-indazol-4-yl)-6-(4-methanesulfonyl-piperazin-1-ylmethyl)-4-morpholin-4-yl-thieno[3,2-d]pyrimidine) and GNE-317 [5-(6-(3-methoxyoxetan-3-yl)-7-methyl-4-morpholinothieno[3,2-d]pyrimidin-2-yl)pyrimidin-2-amine] in U87 and GS2 orthotopic models of GBM, models that exhibit differing blood-brain barrier characteristics. Following administration to tumor-bearing mice, pictilisib was readily detected within tumors of the contrast-enhancing U87 model whereas it was not located in tumors of the nonenhancing GS2 model. In both GBM models, pictilisib was not detected in the healthy brain. In contrast, GNE-317 was uniformly distributed throughout the brain in the U87 and GS2 models. MALDI imaging revealed also that the pictilisib signal varied regionally by up to 6-fold within the U87 tumors whereas GNE-317 intratumor levels were more homogeneous. Liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) analyses of the nontumored half of the brain showed pictilisib had brain-to-plasma ratios lower than 0.03 whereas they were greater than 1 for GNE-317, in agreement with their brain penetration properties. These results in orthotopic models representing either the contrast-enhancing or invasive areas of GBM clearly demonstrate the need for whole-brain distribution to potentially achieve long-term efficacy in GBM. Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.

Knockdown of DIXDC1 Inhibits the Proliferation and Migration of Human Glioma Cells.

PubMed

Chen, Jianguo; Shen, Chaoyan; Shi, Jinlong; Shen, Jianhong; Chen, Wenjuan; Sun, Jie; Fan, Shaocheng; Bei, Yuanqi; Xu, Peng; Chang, Hao; Jiang, Rui; Hua, Lu; Ji, Bin; Huang, Qingfeng

2017-08-01

DIX domain containing 1 (DIXDC1), the human homolog of coiled-coil-DIX1 (Ccd1), is a positive regulator of Wnt signaling pathway. Recently, it was found to act as a candidate oncogene in colon cancer, non-small-cell lung cancer, and gastric cancer. In this study, we aimed to investigate the clinical significance of DIXDC1 expression in human glioma and its biological function in glioma cells. Western blot and immunohistochemistry analysis showed that DIXDC1 was overexpressed in glioma tissues and glioma cell lines. The expression level of DIXDC1 was evidently linked to glioma pathological grade and Ki-67 expression. Kaplan-Meier curve showed that high expression of DIXDC1 may lead to poor outcome of glioma patients. Serum starvation and refeeding assay indicated that the expression of DIXDC1 was associated with cell cycle. To determine whether DIXDC1 could regulate the proliferation and migration of glioma cells, we transfected glioma cells with interfering RNA-targeting DIXDC1; investigated cell proliferation with Cell Counting Kit (CCK)-8, flow cytometry assays, and colony formation analyses; and investigated cell migration with wound healing assays and transwell assays. According to our data, knockdown of DIXDC1 significantly inhibited proliferation and migration of glioma cells. These data implied that DIXDC1 might participate in the development of glioma, suggesting that DIXDC1 can become a potential therapeutic strategy for glioma.

Osthole enhances TRAIL-mediated apoptosis through downregulation of c-FLIP expression in renal carcinoma Caki cells.

PubMed

Min, Kyoung-Jin; Han, Min Ae; Kim, Shin; Park, Jong-Wook; Kwon, Taeg Kyu

2017-04-01

Osthole, an active constituent isolated from the fruit of Cnidium monnieri (L.) Cusson, has been shown to induce various beneficial biochemical effects such as anti-inflammatory and antitumor. In the present study, we examined whether osthole could sensitize TNF-related apoptosis-inducing ligand (TRAIL)-induced apoptosis in human renal carcinoma Caki cells. We found that osthole and TRAIL alone, had no effect on apoptosis, but combined treatment with osthole and TRAIL markedly induced apoptosis in Caki (renal carcinoma), U251MG (glioma) and MDA-MB-231 (breast carcinoma) cells. In contrast, combined treatment with osthole and TRAIL did not induce apoptosis in normal human skin fibroblast cells. Osthole induced downregulation of cellular FLICE-like inhibitory protein (c-FLIP) expression, and overexpression of c-FLIP markedly blocked apoptosis induced by the combined treatment with osthole and TRAIL. In addition, osthole markedly reduced mitochondrial membrane potential levels, and increased cytosolic cytochrome c release in combined treatment with osthole and TRAIL. Therefore, these data suggest that osthole may be an efficient TRAIL sensitizer.

L-Boronophenylalanine-Mediated Boron Neutron Capture Therapy for Malignant Glioma Progressing After External Beam Radiation Therapy: A Phase I Study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kankaanranta, Leena; Seppaelae, Tiina; Koivunoro, Hanna

Purpose: To investigate the safety of boronophenylalanine-mediated boron neutron capture therapy (BNCT) in the treatment of malignant gliomas that progress after surgery and conventional external beam radiation therapy. Methods and Materials: Adult patients who had histologically confirmed malignant glioma that had progressed after surgery and external beam radiotherapy were eligible for this Phase I study, provided that >6 months had elapsed from the last date of radiation therapy. The first 10 patients received a fixed dose, 290 mg/kg, of L-boronophenylalanine-fructose (L-BPA-F) as a 2-hour infusion before neutron irradiation, and the remaining patients were treated with escalating doses of L-BPA-F, eithermore » 350 mg/kg, 400 mg/kg, or 450 mg/kg, using 3 patients on each dose level. Adverse effects were assessed using National Cancer Institute Common Toxicity Criteria version 2.0. Results: Twenty-two patients entered the study. Twenty subjects had glioblastoma, and 2 patients had anaplastic astrocytoma, and the median cumulative dose of prior external beam radiotherapy was 59.4 Gy. The maximally tolerated L-BPA-F dose was reached at the 450 mg/kg level, where 4 of 6 patients treated had a grade 3 adverse event. Patients who were given >290 mg/kg of L-BPA-F received a higher estimated average planning target volume dose than those who received 290 mg/kg (median, 36 vs. 31 Gy [W, i.e., a weighted dose]; p = 0.018). The median survival time following BNCT was 7 months. Conclusions: BNCT administered with an L-BPA-F dose of up to 400 mg/kg as a 2-hour infusion is feasible in the treatment of malignant gliomas that recur after conventional radiation therapy.« less

Saw Palmetto Extract Inhibits Metastasis and Antiangiogenesis through STAT3 Signal Pathway in Glioma Cell.

PubMed

Ding, Hong; Shen, Jinglian; Yang, Yang; Che, Yuqin

2015-01-01

Signal transducer and activator of transcription factor 3 (STAT3) plays an important role in the proliferation and angiogenesis in human glioma. Previous research indicated that saw palmetto extract markedly inhibited the proliferation of human glioma cells through STAT3 signal pathway. But its effect on tumor metastasis and antiangiogenesis is not clear. This study is to further clear the impact of saw palmetto extract on glioma cell metastasis, antiangiogenesis, and its mechanism. TUNEL assay indicated that the apoptotic cells in the saw palmetto treated group are higher than that in the control group (p < 0.05). The apoptosis related protein is detected and the results revealed that saw palmetto extract inhibits the proliferation of human glioma. Meanwhile pSTAT3 is lower in the experimental group and CD34 is also inhibited in the saw palmetto treated group. This means that saw palmetto extract could inhibit the angiogenesis in glioma. We found that saw palmetto extract was an important phytotherapeutic drug against the human glioma through STAT3 signal pathway. Saw palmetto extract may be useful as an adjunctive therapeutic agent for treatment of individuals with glioma and other types of cancer in which STAT3 signaling is activated.

Engineering cell-fluorescent ion track hybrid detectors.

PubMed

Niklas, Martin; Greilich, Steffen; Melzig, Claudius; Akselrod, Mark S; Debus, Jürgen; Jäkel, Oliver; Abdollahi, Amir

2013-06-11

The lack of sensitive biocompatible particle track detectors has so far limited parallel detection of physical energy deposition and biological response. Fluorescent nuclear track detectors (FNTDs) based on Al₂O₃:C,Mg single crystals combined with confocal laser scanning microscopy (CLSM) provide 3D information on ion tracks with a resolution limited by light diffraction. Here we report the development of next generation cell-fluorescent ion track hybrid detectors (Cell-Fit-HD). The biocompatibility of FNTDs was tested using six different cell lines, i.e. human non-small cell lung carcinoma (A549), glioblastoma (U87), androgen independent prostate cancer (PC3), epidermoid cancer (A431) and murine (VmDk) glioma SMA-560. To evaluate cell adherence, viability and conformal coverage of the crystals different seeding densities and alternative coating with extracellular matrix (fibronectin) was tested. Carbon irradiation was performed in Bragg peak (initial 270.55 MeV u⁻¹). A series of cell compartment specific fluorescence stains including nuclear (HOECHST), membrane (Glut-1), cytoplasm (Calcein AM, CM-DiI) were tested on Cell-Fit-HDs and a single CLSM was employed to co-detect the physical (crystal) as well as the biological (cell layer) information. The FNTD provides a biocompatible surface. Among the cells tested, A549 cells formed the most uniform, viable, tightly packed epithelial like monolayer. The ion track information was not compromised in Cell-Fit-HD as compared to the FNTD alone. Neither cell coating and culturing, nor additional staining procedures affected the properties of the FNTD surface to detect ion tracks. Standard immunofluorescence and live staining procedures could be employed to co-register cell biology and ion track information. The Cell-Fit-Hybrid Detector system is a promising platform for a multitude of studies linking biological response to energy deposition at high level of optical microscopy resolution.

Downregulation of HIF-1a sensitizes U251 glioma cells to the temozolomide (TMZ) treatment

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tang, Jun-Hai; Ma, Zhi-Xiong; Huang, Guo-Hao

Purpose: The aim of this study was to investigate the effect of downregulation of HIF-1α gene on human U251 glioma cells and examine the consequent changes of TMZ induced effects and explore the molecular mechanisms. Methods: U251 cell line stably expressing HIF-1α shRNA was acquired via lentiviral vector transfection. The mRNA and protein expression alterations of genes involved in our study were determined respectively by qRT-PCR and Western blot. Cell proliferation was measured by MTT assay and colony formation assay, cell invasion/migration capacity was determined by transwell invasion assay/wound healing assay, and cell apoptosis was detected by flow cytometry. Results:more » We successfully established a U251 cell line with highly efficient HIF-1α knockdown. HIF-1a downregulation sensitized U251 cells to TMZ treatment and enhanced the proliferation-inhibiting, invasion/migration-suppressing, apoptosis-inducing and differentiation-promoting effects exerted by TMZ. The related molecular mechanisms demonstrated that expression of O{sup 6}-methylguanine DNA methyltransferase gene (MGMT) and genes of Notch1 pathway were significantly upregulated by TMZ treatment. However, this upregulation was abrogated by HIF-1α knockdown. We further confirmed important regulatory roles of HIF-1α in the expression of MGMT and activation of Notch1 pathways. Conclusion: HIF-1α downregulation sensitizes U251 glioma cells to the temozolomide treatment via inhibiting MGMT expression and Notch1 pathway activation. - Highlights: • TMZ caused more significant proliferation inhibition and apoptosis in U251 cells after downregulating HIF-1α. • Under TMZ treatment, HIF-1 downregulated U251 cells exhibited weaker mobility and more differentiated state. • TMZ caused MGMT over-expression and Notch1 pathway activation, which could be abrogated by HIF-1α downregulation.« less

The immunohistochemical expression of calcitonin receptor-like receptor (CRLR) in human gliomas

PubMed Central

Benes, L; Kappus, C; McGregor, G P; Bertalanffy, H; Mennel, H D; Hagner, S

2004-01-01

Background: Gliomas are the most common primary tumours of the central nervous system and exhibit rapid growth that is associated with neovascularisation. Adrenomedullin is an important tumour survival factor in human carcinogenesis. It has growth promoting effects on gliomas, and blockade of its actions has been experimentally shown to reduce the growth of glioma tissues and cell lines. There is some evidence that the calcitonin receptor-like receptor (CRLR) mediates the tumorigenic actions of adrenomedullin. Aim: To determine whether CRLR is expressed in human gliomas and the probable cellular targets of adrenomedullin. Methods: Biopsies from 95 human gliomas of varying grade were processed for immunohistochemical analysis using a previously developed and characterised antibody to CRLR. Results: All tumour specimens were positive for CRLR. As previously found in normal peripheral tissues, CRLR immunostaining was particularly intense in the endothelial cells. This was evident in all the various vascular conformations that were observed, and which are typical of gliomas. In addition, clear immunostaining of tumour cells with astrocyte morphology was observed. These were preferentially localised around vessels. Conclusions: This study has shown for the first time that the CRLR protein is present in human glioma tissue. The expression of the receptor in endothelial cells and in astrocytic tumour cells is consistent with the evidence that its endogenous ligand, adrenomedullin, may influence glioma growth by means of both direct mitogenic and indirect angiogenic effects. CRLR may be a valuable target for effective therapeutic intervention in these malignant tumours. PMID:14747444

The immunohistochemical expression of calcitonin receptor-like receptor (CRLR) in human gliomas.

PubMed

Benes, L; Kappus, C; McGregor, G P; Bertalanffy, H; Mennel, H D; Hagner, S

2004-02-01

Gliomas are the most common primary tumours of the central nervous system and exhibit rapid growth that is associated with neovascularisation. Adrenomedullin is an important tumour survival factor in human carcinogenesis. It has growth promoting effects on gliomas, and blockade of its actions has been experimentally shown to reduce the growth of glioma tissues and cell lines. There is some evidence that the calcitonin receptor-like receptor (CRLR) mediates the tumorigenic actions of adrenomedullin. To determine whether CRLR is expressed in human gliomas and the probable cellular targets of adrenomedullin. Biopsies from 95 human gliomas of varying grade were processed for immunohistochemical analysis using a previously developed and characterised antibody to CRLR. All tumour specimens were positive for CRLR. As previously found in normal peripheral tissues, CRLR immunostaining was particularly intense in the endothelial cells. This was evident in all the various vascular conformations that were observed, and which are typical of gliomas. In addition, clear immunostaining of tumour cells with astrocyte morphology was observed. These were preferentially localised around vessels. This study has shown for the first time that the CRLR protein is present in human glioma tissue. The expression of the receptor in endothelial cells and in astrocytic tumour cells is consistent with the evidence that its endogenous ligand, adrenomedullin, may influence glioma growth by means of both direct mitogenic and indirect angiogenic effects. CRLR may be a valuable target for effective therapeutic intervention in these malignant tumours.

Predominant antitumor effects by fully human anti-TRAIL-receptor2 (DR5) monoclonal antibodies in human glioma cells in vitro and in vivo

PubMed Central

Nagane, Motoo; Shimizu, Saki; Mori, Eiji; Kataoka, Shiro; Shiokawa, Yoshiaki

2010-01-01

Tumor necrosis factor–related apoptosis-inducing ligand (TRAIL/Apo2 L) preferentially induces apoptosis in human tumor cells through its cognate death receptors DR4 or DR5, thereby being investigated as a potential agent for cancer therapy. Here, we applied fully human anti-human TRAIL receptor monoclonal antibodies (mAbs) to specifically target one of death receptors for TRAIL in human glioma cells, which could also reduce potential TRAIL-induced toxicity in humans. Twelve human glioma cell lines treated with several fully human anti-human TRAIL receptor mAbs were sensitive to only anti-DR5 mAbs, whereas they were totally insensitive to anti-DR4 mAb. Treatment with anti-DR5 mAbs exerted rapid cytotoxicity and lead to apoptosis induction. The cellular sensitivity was closely associated with cell-surface expression of DR5. Expression of c-FLIPL, Akt, and Cyclin D1 significantly correlated with sensitivity to anti-DR5 mAbs. Primary cultures of glioma cells were also relatively resistant to anti-DR5 mAbs, exhibiting both lower DR5 and higher c-FLIPL expression. Downregulation of c-FLIPL expression resulted in the sensitization of human glioma cells to anti-DR5 mAbs, whereas overexpression of c-FLIPL conferred resistance to anti-DR5 mAb. Treatment of tumor-burden nude mice with the direct agonist anti-DR5 mAb KMTR2 significantly suppressed growth of subcutaneous glioma xenografts leading to complete regression. Similarly, treatment of nude mice bearing intracerebral glioma xenografts with KMTR2 significantly elongated lifespan without tumor recurrence. These results suggest that DR5 is the predominant TRAIL receptor mediating apoptotic signals in human glioma cells, and sensitivity to anti-DR5 mAbs was determined at least in part by the expression level of c-FLIPL and Akt. Specific targeting of death receptor pathway through DR5 using fully human mAbs might provide a novel therapeutic strategy for intractable malignant gliomas. PMID:20511188

uPAR and Cathepsin B Downregulation Induces Apoptosis by Targeting Calcineurin A to BAD via Bcl-2 in Glioma

PubMed Central

Malla, Rama Rao; Gopinath, Sreelatha; Gondi, Christopher S.; Alapati, Kiranmai; Dinh, Dzung H.; Tsung, Andrew J.; Rao, Jasti S.

2011-01-01

Cathepsin B and urokinase plasminogen activator receptor (uPAR) are postulated to play key roles in glioma invasion. Calcineurin is one of the key regulators of mitochondrial-dependent apoptosis, but its mechanism is poorly understood. Hence, we studied subcellular localization of calcineurin after transcriptional downregulation of uPAR and cathepsin B in glioma. In the present study, efficient downregulation of uPAR and cathepsin B increased the translocation of calcineurin A from the mitochondria to the cytosol, decreased pBAD (S136) expression and its interaction with 14-3-3ζ, and increased the interaction of BAD with Bcl-Xl. Co-depletion of uPAR and cathepsin B induced mitochondrial translocation of BAD and caspase 3 as well as PARP activation, cytochrome c and SMAC release. These effects were inhibited by FK506 (10 μM), a specific inhibitor of calcineurin. Calcineurin A was co-localized and also co-immunoprecipitated with Bcl-2. This interaction decreased with co-depletion of uPAR and cathepsin B and also with Bcl-2 inhibitor, HA 14-1 (20 μg/mL). Altered localization and interaction of calcineurin A with Bcl-2 was also observed in vivo when uPAR and cathepsin B were downregulated. In conclusion, downregulation of uPAR and cathepsin B induced apoptosis by targeting calcineurin A to BAD via Bcl-2 in glioma. PMID:21964739

Quantitative multiparametric MRI assessment of glioma response to radiotherapy in a rat model.

PubMed

Hong, Xiaohua; Liu, Li; Wang, Meiyun; Ding, Kai; Fan, Ying; Ma, Bo; Lal, Bachchu; Tyler, Betty; Mangraviti, Antonella; Wang, Silun; Wong, John; Laterra, John; Zhou, Jinyuan

2014-06-01

The inability of structural MRI to accurately measure tumor response to therapy complicates care management for patients with gliomas. The purpose of this study was to assess the potential of several noninvasive functional and molecular MRI biomarkers for the assessment of glioma response to radiotherapy. Fourteen U87 tumor-bearing rats were irradiated using a small-animal radiation research platform (40 or 20 Gy), and 6 rats were used as controls. MRI was performed on a 4.7 T animal scanner, preradiation treatment, as well as at 3, 6, 9, and 14 days postradiation. Image features of the tumors, as well as tumor volumes and animal survival, were quantitatively compared. Structural MRI showed that all irradiated tumors still grew in size during the initial days postradiation. The apparent diffusion coefficient (ADC) values of tumors increased significantly postradiation (40 and 20 Gy), except at day 3 postradiation, compared with preradiation. The tumor blood flow decreased significantly postradiation (40 and 20 Gy), but the relative blood flow (tumor vs contralateral) did not show a significant change at most time points postradiation. The amide proton transfer weighted (APTw) signals of the tumor decreased significantly at all time points postradiation (40 Gy), and also at day 9 postradiation (20 Gy). The blood flow and APTw maps demonstrated tumor features that were similar to those seen on gadolinium-enhanced T1-weighted images. Tumor ADC, blood flow, and APTw were all useful imaging biomarkers by which to predict glioma response to radiotherapy. The APTw signal was most promising for early response assessment in this model. © The Author(s) 2013. Published by Oxford University Press on behalf of the Society for Neuro-Oncology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Significance of perivascular tumour cells defined by CD109 expression in progression of glioma.

PubMed

Shiraki, Yukihiro; Mii, Shinji; Enomoto, Atsushi; Momota, Hiroyuki; Han, Yi-Peng; Kato, Takuya; Ushida, Kaori; Kato, Akira; Asai, Naoya; Murakumo, Yoshiki; Aoki, Kosuke; Suzuki, Hiromichi; Ohka, Fumiharu; Wakabayashi, Toshihiko; Todo, Tomoki; Ogawa, Seishi; Natsume, Atsushi; Takahashi, Masahide

2017-12-01

In the progression of glioma, tumour cells often exploit the perivascular microenvironment to promote their survival and resistance to conventional therapies. Some of these cells are considered to be brain tumour stem cells (BTSCs); however, the molecular nature of perivascular tumour cells has not been specifically clarified because of the complexity of glioma. Here, we identified CD109, a glycosylphosphatidylinositol-anchored protein and regulator of multiple signalling pathways, as a critical regulator of the progression of lower-grade glioma (World Health Organization grade II/III) by clinicopathological and whole-genome sequencing analysis of tissues from human glioma. The importance of CD109-positive perivascular tumour cells was confirmed not only in human lower-grade glioma tissues but also in a mouse model that recapitulated human glioma. Intriguingly, BTSCs isolated from mouse glioma expressed high levels of CD109. CD109-positive BTSCs exerted a proliferative effect on differentiated glioma cells treated with temozolomide. These data reveal the significance of tumour cells that populate perivascular regions during glioma progression, and indicate that CD109 is a potential therapeutic target for the disease. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

A Novel Oncolytic Herpes Simplex Virus that Synergizes with Phosphatidylinositol 3-Kinase/Akt Pathway Inhibitors to Target Glioblastoma Stem Cells

PubMed Central

Kanai, Ryuichi; Wakimoto, Hiroaki; Martuza, Robert L.; Rabkin, Samuel D.

2011-01-01

Purpose To develop a new oncolytic herpes simplex virus (oHSV) for glioblastoma therapy that will be effective in glioblastoma stem cells (GSCs), an important and untargeted component of glioblastoma. One approach to enhance oHSV efficacy is by combination with other therapeutic modalities. Experimental design MG18L, containing a US3 deletion and an inactivating LacZ insertion in UL39, was constructed for the treatment of brain tumors. Safety was evaluated after intracerebral injection in HSV-susceptible mice. The efficacy of MG18L in human GSCs and glioma cell lines in vitro was compared to other oHSVs, alone or in combination with PI3K/Akt inhibitors (LY294002, triciribine, GDC-0941, BEZ235). Cytotoxic interactions between MG18L and PI3K/Akt inhibitors were determined using Chou-Talalay analysis. In vivo efficacy studies were performed using a clinically relevant mouse model of GSC-derived glioblastoma. Results MG18L was severely neuroattenuated in mice, replicated well in GSCs, and had anti-glioblastoma activity in vivo. PI3K/Akt inhibitors displayed significant but variable anti-proliferative activities in GSCs, while their combination with MG18L synergized in killing GSCs and glioma lines, but not human astrocytes, through enhanced induction of apoptosis. Importantly, synergy was independent of inhibitor sensitivity. In vivo, the combination of MG18L and LY294002 significantly prolonged survival of mice, as compared to either agent alone, achieving 50% long-term survival in glioblastoma-bearing mice. Conclusions This study establishes a novel therapeutic strategy: oHSV manipulation of critical oncogenic pathways to sensitize cancer cells to molecularly-targeted drugs. MG18L is a promising agent for the treatment of glioblastoma, being especially effective when combined with PI3K/Akt pathway-targeted agents. PMID:21505062

MicroRNA-139-5p acts as a tumor suppressor by targeting ELTD1 and regulating cell cycle in glioblastoma multiforme

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dai, Shouping; Wang, Xianjun; Li, Xiao

MicroRNA-139-5p was identified to be significantly down-regulated in glioblastoma multiform (GBM) by miRNA array. In this report we aimed to clarify its biological function, molecular mechanisms and direct target gene in GBM. Twelve patients with GBM were analyzed for the expression of miR-139-5p by quantitative RT-PCR. miR-139-5p overexpression was established by transfecting miR-139-5p-mimic into U87MG and T98G cells, and its effects on cell proliferation were studied using MTT assay and colony formation assays. We concluded that ectopic expression of miR-139-5p in GBM cell lines significantly suppressed cell proliferation and inducing apoptosis. Bioinformatics coupled with luciferase and western blot assays alsomore » revealed that miR-139-5p suppresses glioma cell proliferation by targeting ELTD1 and regulating cell cycle. - Highlights: • miR-139-5p is downregulated in GBM. • miR-139-5p regulates cell proliferation through inducing apoptosis. • miR-139-5p regulates glioblastoma tumorigenesis by targeting 3′UTR of ELTD1. • miR-139-5p is involved in cell cycle regulation.« less

Down-regulation of MDR1 by Ad-DKK3 via Akt/NFκB pathways augments the anti-tumor effect of temozolomide in glioblastoma cells and a murine xenograft model.

PubMed

Fujihara, Toshitaka; Mizobuchi, Yoshifumi; Nakajima, Kohei; Kageji, Teruyoshi; Matsuzaki, Kazuhito; Kitazato, Keiko T; Otsuka, Ryotaro; Hara, Keijiro; Mure, Hideo; Okazaki, Toshiyuki; Kuwayama, Kazuyuki; Nagahiro, Shinji; Takagi, Yasushi

2018-05-19

Glioblastoma multiforme (GBM) is the most malignant of brain tumors. Acquired drug resistance is a major obstacle for successful treatment. Earlier studies reported that expression of the multiple drug resistance gene (MDR1) is regulated by YB-1 or NFκB via the JNK/c-Jun or Akt pathway. Over-expression of the Dickkopf (DKK) family member DKK3 by an adenovirus vector carrying DKK3 (Ad-DKK3) exerted anti-tumor effects and led to the activation of the JNK/c-Jun pathway. We investigated whether Ad-DKK3 augments the anti-tumor effect of temozolomide (TMZ) via the regulation of MDR1. GBM cells (U87MG and U251MG), primary TGB105 cells, and mice xenografted with U87MG cells were treated with Ad-DKK3 or TMZ alone or in combination. Ad-DKK3 augmentation of the anti-tumor effects of TMZ was associated with reduced MDR1 expression in both in vivo and in vitro studies. The survival of Ad-DKK3-treated U87MG cells was inhibited and the expression of MDR1 was reduced. This was associated with the inhibition of Akt/NFκB but not of YB-1 via the JNK/c-Jun- or Akt pathway. Our results suggest that Ad-DKK3 regulates the expression of MDR1 via Akt/NFκB pathways and that it augments the anti-tumor effects of TMZ in GBM cells.

Assessment Effects of Resveratrol on Human Telomerase Reverse Transcriptase Messenger Ribonucleic Acid Transcript in Human Glioblastoma.

PubMed

Mirzazadeh, Azin; Kheirollahi, Majid; Farashahi, Ehsan; Sadeghian-Nodoushan, Fatemeh; Sheikhha, Mohammad Hasan; Aflatoonian, Behrouz

2017-01-01

Glioblastoma (GBM) is the most common and aggressive brain tumor, which has a poor prognosis despite the advent of different therapeutic strategies. There are numerous molecular biomarkers to contribute diagnosis, prognosis, and prediction of response to the current therapy in GBM. One of the most important markers that are potentially valuable is immortalization-specific or immortalization-associated marker named "hTERT messenger ribonucleic acid (mRNA)" the key subunit of telomerase enzyme, which is expressed in more than 85% of cancer cells, in spite of the majority of normal somatic cells. In this study, we investigated the effects of resveratrol (RSV) on this mRNA marker level, leading to cancer progression. U-87MG cell line was obtained from Pasteur Institute of Iran and treated with various concentrations of 0-160 μg/mL of RSV and at different time points (24, 48, and 72 h). To evaluate viability of U-87MG cells, standard 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was performed. Real-time polymerase chain reaction (RT-PCR) was used for comparative and quantitative assessment of human telomerase reverse transcriptase (hTERT) mRNA copy number versus control-untreated group. The results of our investigation suggested that RSV effectively inhibited cell growth and caused cell death in dose-dependent ( P < 0.05) and not in time-dependent manner ( P > 0.05), in vitro . Interestingly, quantitative RT-PCR analysis demonstrated that at half inhibition concentration, RSV dramatically decreased mRNA expression of hTERT, the catalytic subunit of telomerase enzyme, which leads to prevention of cell division and tumor progression. With regard to downregulation of this immortalization-associated marker, RSV may potentially be used as a therapeutic agent against GBM.

Downregulation of TES by hypermethylation in glioblastoma reduces cell apoptosis and predicts poor clinical outcome.

PubMed

Bai, Yu; Zhang, Quan-Geng; Wang, Xin-Hua

2014-12-11

Gliomas are the most common human brain tumors. Glioblastoma, also known as glioblastoma multiform (GBM), is the most aggressive, malignant, and lethal glioma. The investigation of prognostic and diagnostic molecular biomarkers in glioma patients to provide direction on clinical practice is urgent. Recent studies demonstrated that abnormal DNA methylation states play a key role in the pathogenesis of this kind of tumor. In this study, we want to identify a novel biomarker related to glioma initiation and find the role of the glioma-related gene. We performed a methylation-specific microarray on the promoter region to identify methylation gene(s) that may affect outcome of GBM patients. Normal and GBM tissues were collected from Tiantan Hospital. Genomic DNA was extracted from these tissues and analyzed with a DNA promoter methylation microarray. Testis derived transcript (TES) protein expression was analyzed by immunohistochemistry in paraffin-embedded patient tissues. Western blotting was used to detect TES protein expression in the GBM cell line U251 with or without 5-aza-dC treatment. Cell apoptosis was evaluated by flow cytometry analysis using Annexin V/PI staining. We found that the TES promoter was hypermethylated in GBM compared to normal brain tissues under DNA promoter methylation microarray analysis. The GBM patients with TES hypermethylation had a short overall survival (P <0.05, log-rank test). Among GBM samples, reduced TES protein level was detected in 33 (89.2%) of 37 tumor tissues by immunohistochemical staining. Down regulation of TES was also correlated with worse patient outcome (P <0.05, log-rank test). Treatment on the GBM cell line U251 with 5-aza-dC can greatly increase TES expression, confirming the hypermethylation of TES promoter in GBM. Up-regulation of TES prompts U251 apoptosis significantly. This study demonstrated that both TES promoter hypermethylation and down-regulated protein expression significantly correlated with worse patient outcome. Treatment on the GBM cell line (U251) with 5-aza-dC can highly release TES expression resulting in significant apoptosis in these cells. Our findings suggest that the TES gene is a novel tumor suppressor gene and might represent a valuable prognostic marker for glioblastoma, indicating a potential target for future GBM therapy.

Antioxidant and antiproliferative activities in different maturation stages of broccoli (Brassica oleracea Italica) biofortified with selenium.

PubMed

Bachiega, Patricia; Salgado, Jocelem Mastrodi; de Carvalho, João Ernesto; Ruiz, Ana Lúcia T G; Schwarz, Kélin; Tezotto, Tiago; Morzelle, Maressa Caldeira

2016-01-01

In this work, three different broccoli maturity stages subjected to biofortification with selenium were evaluated for antioxidant and antiproliferative activities. Antioxidant trials have shown that the maturation stages biofortified with selenium had significantly higher amounts of phenolic compounds and antioxidant activity, especially seedlings. Although non-polar extracts of all samples show antiproliferative activity, the extract of broccoli seedlings biofortified with selenium stood out, presenting cytocidal activity for a glioma line (U251, GI50 28.5 mg L(-1)). Copyright © 2015 Elsevier Ltd. All rights reserved.

Saw Palmetto Extract Inhibits Metastasis and Antiangiogenesis through STAT3 Signal Pathway in Glioma Cell

PubMed Central

Ding, Hong; Shen, Jinglian; Yang, Yang; Che, Yuqin

2015-01-01

Signal transducer and activator of transcription factor 3 (STAT3) plays an important role in the proliferation and angiogenesis in human glioma. Previous research indicated that saw palmetto extract markedly inhibited the proliferation of human glioma cells through STAT3 signal pathway. But its effect on tumor metastasis and antiangiogenesis is not clear. This study is to further clear the impact of saw palmetto extract on glioma cell metastasis, antiangiogenesis, and its mechanism. TUNEL assay indicated that the apoptotic cells in the saw palmetto treated group are higher than that in the control group (p < 0.05). The apoptosis related protein is detected and the results revealed that saw palmetto extract inhibits the proliferation of human glioma. Meanwhile pSTAT3 is lower in the experimental group and CD34 is also inhibited in the saw palmetto treated group. This means that saw palmetto extract could inhibit the angiogenesis in glioma. We found that saw palmetto extract was an important phytotherapeutic drug against the human glioma through STAT3 signal pathway. Saw palmetto extract may be useful as an adjunctive therapeutic agent for treatment of individuals with glioma and other types of cancer in which STAT3 signaling is activated. PMID:26788112

Cytotoxic effect of menadione and sodium orthovanadate in combination on human glioma cells.

PubMed

Delwar, Zahid M; Avramidis, Dimitrios; Follin, Elna; Hua, Yan; Siden, Åke; Cruz, Mabel; Paulsson, Kajsa M; Yakisich, Juan Sebastian

2012-08-01

Gliomas are the most common primary brain tumor, and their treatment is still a challenge. Here, we evaluated the antiproliferative effect of a novel combination of two potent oxidative stress enhancers: menadione (M) and sodium orthovanadate (SO). We observed both short-term and prolonged growth inhibitory effects of M or SO alone as well as in combination (M:SO) on DBTRG.05MG human glioma cells. A stronger antiproliferative effect was observed in the short-term proliferation assay with the M:SO combination compared to either investigated agent alone. In the long-term proliferation assay, a 10-day exposure to M:SO at concentrations of 10 μM:17.5 μM or 17.5 μM:10 μM was enough to kill 100% of the cells; no cell regrowth was observed after re-incubation in drug-free media. When used in combination, the single concentration of M and SO could be decreased by 2.5- to 5-fold of those used for each experimental drug alone and still obtain a similar antiproliferative effect. The underlying molecular mechanism was investigated by co-incubating M:SO with dithiothreitol (DTT) and genistein. Both substances partially neutralized the effects of the M:SO combination, showing additive effects. This observation suggests a role of oxidative stress and tyrosine kinase stimulation in the M:SO cytotoxic effect. Our results indicate that M:SO combination is an attractive alternative for glioma treatment that encourages further study. The neutralizing effects of genistein and DTT reveal a possibility for their use in the minimization of potential M:SO systemic toxicity.

Phase I Clinical Trial Assessing Temozolomide and Tamoxifen With Concomitant Radiotherapy for Treatment of High-Grade Glioma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Patel, Shilpen, E-mail: Shilpenp@uw.edu; DiBiase, Steven; Meisenberg, Barry

2012-02-01

Purpose: The new standard treatment of glioblastoma multiforme is concurrent radiotherapy (RT) and temozolomide. The proliferation of high-grade gliomas might be partly dependent on protein kinase C-mediated pathways. Tamoxifen has been shown in vitro to inhibit protein kinase C through estrogen receptor-independent antineoplastic effects. This Phase I trial was designed to determine the maximal tolerated dose (MTD) of tamoxifen when given with temozolomide and concurrent RT to patients with high-grade gliomas. Methods and Materials: A total of 17 consecutive patients in four cohorts with World Health Organization Grade 3 (n = 2) and 4 (n = 15) gliomas were givenmore » tamoxifen twice daily during 6 weeks of concurrent RT and temozolomide. Eligibility included histologic diagnosis, age >18 years old, Karnofsky performance status {>=}60, and no previous brain RT or chemotherapy. The starting dose was 50 mg/m{sup 2} divided twice daily. If no dose-limiting toxicities (DLTs) occurred in 3 patients, the dose was escalated in 25-mg/m{sup 2} increments until the MTD was reached. When {>=}2 patients within a cohort experienced a DLT, the MTD had been exceeded. Temozolomide was given with RT at 75 mg/m{sup 2}. A dose of 60 Gy in 2 Gy/d fractions to a partial brain field was delivered. Results: A total of 6 patients in Cohort 4 had received tamoxifen at 125 mg/m{sup 2}. One patient was excluded, and the fourth patient developed Grade 4 thrombocytopenia (DLT). Thus, 3 more patients needed to be enrolled. A deep venous thrombosis (DLT) occurred in the sixth patient. Thus, the MTD was 100 mg/m{sup 2}. Conclusions: The MTD of tamoxifen was 100 mg/m{sup 2} when given concurrently with temozolomide 75 mg/m{sup 2} and RT. Tamoxifen might have a role in the initial treatment of high-grade gliomas and should be studied in future Phase II trials building on the newly established platform of concurrent chemoradiotherapy.« less

Some phenolic compounds of extracts obtained from Origanum species growing in Turkey.

PubMed

Ozkan, Gülcan; Ozcan, Mehmet Musa

2014-08-01

Caffeic acid, rosmarinic acid, rutin, apigenin 7-O-glucoside, apigenin, and acesetin were the main phenolic compounds of Origanum onites extracts in all applications. While acesetin contents ranged from 133.59 mg/100 g (U1) to 437.25 mg/100 g (S3), rosmarinic acid changed between 215.94 mg/100 g (U4) and 1120.56 mg/100 g (S2) in Origanum vulgare L. subsp. hirtum (Link) Ietsw. Both rosmarinic acid and acesetin were not found in U5 application. Only caffeic acid (19.39 mg/100 g) was found in U5 application. Rosmarinic acid contents of O. onites extract changed between 158.62 mg/100 g (U5) and 799.87 mg/100 g (S2). Generally, dominant phenolic compound of Origanum extracts was rosmarinic acid compared with other extracts. In addition, methanol:water:acetic acid mixture (S2) (95:4.5:0.5) was found as the best application. Phenolic contents of extracts obtained with U series mixtures were found low.

Sphingosines Derived from Marine Sponge as Potential Multi-Target Drug Related to Disorders in Cancer Development

PubMed Central

Biegelmeyer, Renata; Schröder, Rafael; Rambo, Douglas F.; Dresch, Roger R.; Carraro, João L. F.; Mothes, Beatriz; Moreira, José Cláudio F.; da Frota Junior, Mário L. C.; Henriques, Amélia T.

2015-01-01

Haliclona tubifera, marine sponge species abundant in Brazilian coastline, presents only a few papers published in the literature. Recently, we have reported the isolation of two modified C18 sphingoid bases: (2R,3R,6R,7Z)-2-aminooctadec-7-ene-1,3,6-triol and and (2R,3R,6R)-2-aminooctadec-1,3,6-triol. In order to continue our research, in this work aimed at the biological investigation of fractions that led to the isolation of these compounds. We evaluated the cytotoxic effect of marine sponge H. tubifera fractions in glioma (U87) and neuroblastoma (SH-SY5Y) human cell lines. In addition, considering the link between cancer, imbalance of reactive oxygen species and coagulation disorders, we also investigated the in vitro effects on blood coagulation and their redox properties. We showed that the ethyl acetate (EtOAc) fraction, rich in sphingoid bases, had important cytotoxic effects in both cancer cell lines with an IC50 < 15 μg/mL and also can inhibit the production of peroxyl radicals. Interestingly, this fraction increased the recalcification time of human blood, showing anticoagulant properties. The present study indicates the sphingosines fraction as a promising source of chemical prototypes, especially multifunctional drugs in cancer therapy. PMID:26308014

89Zr-DFO-AMG102 Immuno-PET to Determine Local Hepatocyte Growth Factor Protein Levels in Tumors for Enhanced Patient Selection.

PubMed

Price, Eric W; Carnazza, Kathryn E; Carlin, Sean D; Cho, Andrew; Edwards, Kimberly J; Sevak, Kuntal K; Glaser, Jonathan M; de Stanchina, Elisa; Janjigian, Yelena Y; Lewis, Jason S

2017-09-01

The hepatocyte growth factor (HGF) binding antibody rilotumumab (AMG102) was modified for use as a 89 Zr-based immuno-PET imaging agent to noninvasively determine the local levels of HGF protein in tumors. Because recent clinical trials of HGF-targeting therapies have been largely unsuccessful in several different cancers (e.g., gastric, brain, lung), we have synthesized and validated 89 Zr-DFO-AMG102 as a companion diagnostic for improved identification and selection of patients having high local levels of HGF in tumors. To date, patient selection has not been performed using the local levels of HGF protein in tumors. Methods: The chelator p -SCN-Bn-DFO was conjugated to AMG102, radiolabeling with 89 Zr was performed in high radiochemical yields and purity (>99%), and binding affinity of the modified antibody was confirmed using an enzyme-linked immunosorbent assay (ELISA)-type binding assay. PET imaging, biodistribution, autoradiography and immunohistochemistry, and ex vivo HGF ELISA experiments were performed on murine xenografts of U87MG (HGF-positive, MET-positive) and MKN45 (HGF-negative, MET-positive) and 4 patient-derived xenografts (MET-positive, HGF unknown). Results: Tumor uptake of 89 Zr-DFO-AMG102 at 120 h after injection in U87MG xenografts (HGF-positive) was high (36.8 ± 7.8 percentage injected dose per gram [%ID/g]), whereas uptake in MKN45 xenografts (HGF-negative) was 5.0 ± 1.3 %ID/g and a control of nonspecific human IgG 89 Zr-DFO-IgG in U87MG tumors was 11.5 ± 3.3 %ID/g, demonstrating selective uptake in HGF-positive tumors. Similar experiments performed in 4 different gastric cancer patient-derived xenograft models showed low uptake of 89 Zr-DFO-AMG102 (∼4-7 %ID/g), which corresponded with low HGF levels in these tumors (ex vivo ELISA). Autoradiography, immunohistochemical staining, and HGF ELISA assays confirmed that elevated levels of HGF protein were present only in U87MG tumors and that 89 Zr-DFO-AMG102 uptake was closely correlated with HGF protein levels in tumors. Conclusion: The new immuno-PET imaging agent 89 Zr-DFO-AMG102 was successfully synthesized, radiolabeled, and validated in vitro and in vivo to selectively accumulate in tumors with high local levels of HGF protein. These results suggest that 89 Zr-DFO-AMG102 would be a valuable companion diagnostic tool for the noninvasive selection of patients with elevated local concentrations of HGF in tumors for planning any HGF-targeted therapy, with the potential to improve clinical outcomes. © 2017 by the Society of Nuclear Medicine and Molecular Imaging.

Functional analysis of the DEPDC1 oncoantigen in malignant glioma and brain tumor initiating cells.

PubMed

Kikuchi, Ryogo; Sampetrean, Oltea; Saya, Hideyuki; Yoshida, Kazunari; Toda, Masahiro

2017-06-01

DEP domain containing 1 (DEPDC1) is a novel oncoantigen expressed in cancer cells, which presents oncogenic activity and high immunogenicity. Although DEPDC1 has been predicted to be a useful antigen for the development of a cancer vaccine, its pathophysiological roles in glioma have not been investigated. Here, we analyzed the expression and function of DEPDC1 in malignant glioma. DEPDC1 expression in glioma cell lines, glioma tissues, and brain tumor initiating cells (BTICs) was assessed by western blot and quantitative polymerase chain reaction (PCR). The effect of DEPDC1 downregulation on cell growth and nuclear factor kappa B (NFκB) signaling in glioma cells was investigated. Overall survival was assessed in mouse glioma models using human glioma cells and induced mouse brain tumor stem cells (imBTSCs) to determine the effect of DEPDC1 suppression in vivo. DEPDC1 expression was increased in glioma cell lines, tissues, and BTICs. Suppression of endogenous DEPDC1 expression by small interfering RNA (siRNA) inhibited glioma cell viability and induced apoptosis through NFκB signaling. In mouse glioma models using human glioma cells and imBTSCs, downregulation of DEPDC1 expression prolonged overall survival. These results suggest that DEPDC1 represents a target molecule for the treatment of glioma.

ATM inhibitor KU-55933 increases the TMZ responsiveness of only inherently TMZ sensitive GBM cells.

PubMed

Nadkarni, Aditi; Shrivastav, Meena; Mladek, Ann C; Schwingler, Paul M; Grogan, Patrick T; Chen, Junjie; Sarkaria, Jann N

2012-12-01

Ataxia telangiectasia mutated (ATM) kinase is critical in sensing and repairing DNA double-stranded breaks (DSBs) such as those induced by temozolomide (TMZ). ATM deficiency increases TMZ sensitivity, which suggests that ATM inhibitors may be effective TMZ sensitizing agents. In this study, the TMZ sensitizing effects of 2 ATM specific inhibitors were studied in established and xenograft-derived glioblastoma (GBM) lines that are inherently sensitive to TMZ and derivative TMZ-resistant lines. In parental U251 and U87 glioma lines, the addition of KU-55933 to TMZ significantly increased cell killing compared to TMZ alone [U251 survival: 0.004 ± 0.0015 vs. 0.08 ± 0.01 (p < 0.001), respectively, and U87 survival: 0.02 ± 0.005 vs. 0.04 ± 0.002 (p < 0.001), respectively] and also elevated the fraction of cells arrested in G2/M [U251 G2/M fraction: 61.8 ± 1.1 % vs. 35 ± 0.8 % (p < 0.001), respectively, and U87 G2/M fraction 25 ± 0.2 % vs.18.6 ± 0.4 % (p < 0.001), respectively]. In contrast, KU-55933 did not sensitize the resistant lines to TMZ, and neither TMZ alone or combined with KU-55933 induced a G2/M arrest. While KU-55933 did not enhance TMZ induced Chk1/Chk2 activation, it increased TMZ-induced residual γ-H2AX foci in the parental cells but not in the TMZ resistant cells. Similar sensitization was observed with either KU-55933 or CP-466722 combined with TMZ in GBM12 xenograft line but not in GBM12TMZ, which is resistant to TMZ due to MGMT overexpression. These findings are consistent with a model where ATM inhibition suppresses the repair of TMZ-induced DSBs in inherently TMZ-sensitive tumor lines, which suggests an ATM inhibitor potentially could be deployed with an improvement in the therapeutic window when combined with TMZ.

Temozolomide does not influence the transcription or activity of matrix metalloproteinases 9 and 2 in glioma cell lines.

PubMed

Suzuki, Yuta; Fujioka, Kouki; Ikeda, Keiichi; Murayama, Yuichi; Manome, Yoshinobu

2017-07-01

Glioblastoma multiforme (GBM) is a treatment-resistant malignancy with poor prognosis. Temozolomide (TMZ) is widely used as a first-line drug for GBM. Although this improves patient prognosis, it does not completely eradicate the tumour. Even after total surgical resection, GBM can exhibit uncontrollable invasiveness at the tumour margins owing to activation of matrix metalloproteinases (MMPs) such as MMP-2 and -9; these degrade collagen IV in the basement membrane, which normally prevents cancer invasion. TMZ induces DNA damage and activates transcription factors including c-jun, c-fos, nuclear factor-κβ, and early growth response protein-1, which have putative binding sites on the MMP-9 promoter. TMZ may therefore enhance tumour invasion by stimulating MMP-9 transcription and enzymatic activity. To test this hypothesis, we investigated MMP-2 and -9 mRNA transcription and activity in GBM cell lines treated with TMZ. Human A172 GBM cells were exposed to TMZ (25% and 50% inhibitory concentrations) for 24 or 48h; cell cycle distribution and mRNA levels of MMP-2 and -9 were evaluated using flow cytometry and semi-quantitative reverse transcription PCR, respectively. MMP-2 and -9 enzymatic activities were assessed using gelatin zymography in human A172 and U373 MG GBM cells exposed to TMZ under the same conditions. TMZ altered A172 cell cycle distribution, but not MMP-2 or -9 mRNA levels. TMZ did not affect MMP-2 or -9 enzymatic activities in A172 or U373 MG cells. These findings indicated that TMZ is therefore unlikely to promote GBM invasiveness. Copyright © 2017 Elsevier Ltd. All rights reserved.

Grading of Gliomas by Using Monoexponential, Biexponential, and Stretched Exponential Diffusion-weighted MR Imaging and Diffusion Kurtosis MR Imaging

PubMed Central

Bai, Yan; Lin, Yusong; Tian, Jie; Shi, Dapeng; Cheng, Jingliang; Haacke, E. Mark; Hong, Xiaohua; Ma, Bo; Zhou, Jinyuan

2016-01-01

Purpose To quantitatively compare the potential of various diffusion parameters obtained from monoexponential, biexponential, and stretched exponential diffusion-weighted imaging models and diffusion kurtosis imaging in the grading of gliomas. Materials and Methods This study was approved by the local ethics committee, and written informed consent was obtained from all subjects. Both diffusion-weighted imaging and diffusion kurtosis imaging were performed in 69 patients with pathologically proven gliomas by using a 3-T magnetic resonance (MR) imaging unit. An isotropic apparent diffusion coefficient (ADC), true ADC, pseudo-ADC, and perfusion fraction were calculated from diffusion-weighted images by using a biexponential model. A water molecular diffusion heterogeneity index and distributed diffusion coefficient were calculated from diffusion-weighted images by using a stretched exponential model. Mean diffusivity, fractional anisotropy, and mean kurtosis were calculated from diffusion kurtosis images. All values were compared between high-grade and low-grade gliomas by using a Mann-Whitney U test. Receiver operating characteristic and Spearman rank correlation analysis were used for statistical evaluations. Results ADC, true ADC, perfusion fraction, water molecular diffusion heterogeneity index, distributed diffusion coefficient, and mean diffusivity values were significantly lower in high-grade gliomas than in low-grade gliomas (U = 109, 56, 129, 6, 206, and 229, respectively; P < .05). Pseudo-ADC and mean kurtosis values were significantly higher in high-grade gliomas than in low-grade gliomas (U = 98 and 8, respectively; P < .05). Both water molecular diffusion heterogeneity index (area under the receiver operating characteristic curve [AUC] = 0.993) and mean kurtosis (AUC = 0.991) had significantly greater AUC values than ADC (AUC = 0.866), mean diffusivity (AUC = 0.722), and fractional anisotropy (AUC = 0.500) in the differentiation of low-grade and high-grade gliomas (P < .05). Conclusion Water molecular diffusion heterogeneity index and mean kurtosis values may provide additional information and improve the grading of gliomas compared with conventional diffusion parameters. © RSNA, 2015 Online supplemental material is available for this article. PMID:26230975

A pilot clinical study of apatinib plus irinotecan in patients with recurrent high-grade glioma: Clinical Trial/Experimental Study.

PubMed

Wang, Lei; Liang, Lijun; Yang, Tao; Qiao, Yun; Xia, Youyou; Liu, Liang; Li, Chao; Lu, Peizhi; Jiang, Xiaodong

2017-12-01

Malignant glioma is the most common primary malignant brain tumor that displays high vascularity, making vascular endothelial growth factor receptors become promising targets. This study was conducted to evaluate the efficacy and safety of apatinib, a new potent oral small-molecule tyrosine kinase inhibitor targeted vascular endothelial growth factor receptor 2, combined with irinotecan, in patients with recurrent malignant glioma. Ten patients with recurrent malignant glioma who were experiencing relapse after treatment of temozolomide were enrolled in this study. They received oral apatinib (500 mg qd) in conjunction with irinotecan (340 mg/m or 125 mg/m depending on use of enzyme-inducing antiepileptic drugs) for 6 cycles. After that the patients continued to take apatinib as maintenance. Dosage adjustment occurred in only 3 (30.0%) patients. Among the 10 patients, 9 were available for the efficacy evaluation. There were 5 with partial response, 2 with stable disease and 2 with progressive disease. The objective response rate and the disease control rate (DCR) were 55% (5/9) and 78% (7/9), respectively. The median progress free survival time was 8.3 months. As for safety analysis, the most 3 common adverse events were gastrointestinal reaction (31.8%), hypertension (22.7%), and myelosuppression (18.0%). Apatinib combined with irinotecan seems to be a promising therapeutic option for recurrent malignant glioma patients. Perspective clinical studies with adequate sample size are required to validate our results. NCT02848794 /Ahead-BG306.

Products of cells from gliomas: VIII. Multiple-well immunoperoxidase assay of immunoreactivity of primary hybridoma supernatants with human glioma and brain tissue and cultured glioma cells.

PubMed

McKeever, P E; Wahl, R L; Shakui, P; Jackson, G A; Letica, L H; Liebert, M; Taren, J A; Beierwaltes, W H; Hoff, J T

1990-06-01

To test the feasibility of primary screening of hybridoma supernatants against human glioma tissue, over 5000 combinations of hybridoma supernatants with glioma tissue, cultured glioma cells, and normal central neural tissue were screened with a new multiple-well (M-well) screening system. This is an immunoperoxidase assay system with visual endpoints for screening 20-30 hybridoma supernatants per single microscope slide. There were extensive differences between specificities to tissue and to cultured glioma cells when both were screened with M-wells and when cultured cells were screened with standard semi-automated fluorescence. Primary M-well screening with glioma tissue detected seven hybridoma supernatants that specifically identified parenchymal cells of glioma tissue and that were not detected with cultured cells. Immunoreactivities of individual supernatants for vascular components (nine supernatants), necrosis (five supernatants), and nuclei (three supernatants) were detected. Other supernatants bound multiple sites on glioma tissue and/or subpopulations of neurons and glia of normal tissue. The results show that primary screening with glioma tissue detects a number of different specificities of hybridoma supernatants to gliomas not detected by conventional screening with cultured cells. These are potentially applicable to diagnosis and therapy.

High expression of Bruton's tyrosine kinase (BTK) is required for EGFR-induced NF-κB activation and predicts poor prognosis in human glioma.

PubMed

Yue, Chenglong; Niu, Mingshan; Shan, Qian Qian; Zhou, Ting; Tu, Yiming; Xie, Peng; Hua, Lei; Yu, Rutong; Liu, Xuejiao

2017-09-25

Malignant glioma is the most common primary brain tumor in adults and has a poor prognosis. However, there are no effective targeted therapies for glioma patients. Thus, the development of novel targeted therapeutics for glioma is urgently needed. In this study, we examined the prognostic significance BTK expression in patients with glioma. Furthermore, we investigated the mechanism and therapeutic potential of ibrutinib in the treatment of human glioma in vitro and in vivo. Our data demonstrate that high expression of BTK is a novel prognostic marker for poor survival in patients with glioma. BTK-specific inhibitor ibrutinib effectively inhibits the proliferation, migration and invasion ability of glioma cells. Furthermore, ibrutinib can induce G1 cell-cycle arrest by regulating multiple cell cycle-associated proteins. More importantly, we found that BTK inhibition significantly blocks the degradation of IκBα and prevents the nuclear accumulation of NF-κB p65 subunit induced by EGF in glioma cells. Taken together, our study suggests that BTK is a novel prognostic marker and molecular therapeutic target for glioma. BTK is required for EGFR-induced NF-κB activation in glioma cells. These findings provide the basis for future clinical studies of ibrutinib for the treatment of glioma.

GLUT-1-independent infection of the glioblastoma/astroglioma U87 cells by the human T cell leukemia virus type 1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jin Qingwen; Agrawal, Lokesh; Walther Cancer Institute, Indianapolis, IN 46208

2006-09-15

The human glucose transporter protein 1 (GLUT-1) functions as a receptor for human T cell leukemia virus (HTLV). GLUT-1 is a twelve-transmembrane cell surface receptor with six extracellular (ECL) and seven intracellular domains. To analyze HTLV-1 cytotropism, we utilized polyclonal antibodies to a synthetic peptide corresponding to the large extracellular domain of GLUT-1. The antibodies caused significant blocking of envelope (Env)-mediated fusion and pseudotyped virus infection of HeLa cells but had no significant effect on infection of U87 cells. This differential effect correlated with the detection of high-level surface expression of GLUT-1 on HeLa cells and very weak staining ofmore » U87 cells. To investigate this in terms of viral cytotropism, we cloned GLUT-1 cDNA from U87 cells and isolated two different versions of cDNA clones: the wild-type sequence (encoding 492 residues) and a mutant cDNA with a 5-base pair deletion (GLUT-1{delta}5) between nucleotides 1329 and 1333. The deletion, also detected in genomic DNA, resulted in a frame-shift and premature termination producing a truncated protein of 463 residues. Transfection of the wild-type GLUT-1 but not GLUT-1{delta}5 cDNA into CHO cells resulted in efficient surface expression of the human GLUT-1. Co-expression of GLUT-1 with GLUT-1{delta}5 produces a trans-inhibition by GLUT-1{delta}5 of GLUT-1-mediated HTLV-1 envelope (Env)-mediated fusion. Co-immunoprecipitation experiments demonstrated physical interaction of the wild-type and mutant proteins. Northern blot and RT-PCR analyses demonstrated lower GLUT-1 RNA expression in U87 cells. We propose two mechanisms to account for the impaired cell surface expression of GLUT-1 on U87 cells: low GLUT-1 RNA expression and the formation of GLUT-1/GLUT-1{delta}5 heterodimers that are retained intracellularly. Significant RNAi-mediated reduction of endogenous GLUT-1 expression impaired HTLV-1 Env-mediated fusion with HeLa cells but not with U87 cells. We propose a GLUT-1-independent mechanism of HTLV-1 infection of U87 cells. The results may have important implications for HTLV-1 neurotropism and pathogenesis.« less

Combination of photodynamic therapy and temozolomide on glioma in a rat C6 glioma model.

PubMed

Zhang, Xiaoming; Guo, Mian; Shen, Lei; Hu, Shaoshan

2014-12-01

For glioma, temozolomide (TMZ) is a commonly used chemotherapy drug and photodynamic therapy (PDT) is an important adjuvant therapy. The aim of this study was to evaluate the effect of their combination for the treatment of glioma. A rat C6 glioma model using male Wistar rats (n=180) weighing 280-300 g was established. Glioma-bearing rats (n=100) were treated with mock, hematoporphyrin monomethyl ether (HMME), laser or PDT. The expression of P-glycoprotein (P-gp) in endothelial cells of the blood-tumor-barrier and in glioma tissues was detected using immunohistochemistry and western blot, respectively. Glioma-bearing rats (n=40) were treated with normal saline, TMZ (60 mg/m(2) for five consecutive days), PDT (630 nm for 10 min) or a combination of TMZ and PDT. TMZ concentration in glioma tissues was detected using liquid chromatography/mass spectrometry/mass spectrometry (LC/MS/MS) and cell death was observed using transmission microscopy. Concurrently, another batch of 40 glioma-bearing rats was subjected to the same treatment, and the survival of these rats was estimated using Kaplan-Meier analysis. PDT significantly decreased the expression of P-gp in endothelial cells comprising the blood-tumor-barrier and in glioma tissues. The combination of TMZ with PDT significantly increased TMZ concentration in glioma tissues, enhanced glioma cell apoptosis and prolonged the median survival of glioma-bearing rats. The combination of PDT with TMZ shows synergistic effect in rat C6 glioma model, indicating its potential clinical use in glioma treatment. Copyright © 2014 Elsevier B.V. All rights reserved.

A gene delivery system with a human artificial chromosome vector based on migration of mesenchymal stem cells towards human glioblastoma HTB14 cells.

PubMed

Kinoshita, Yusuke; Kamitani, Hideki; Mamun, Mahabub Hasan; Wasita, Brian; Kazuki, Yasuhiro; Hiratsuka, Masaharu; Oshimura, Mitsuo; Watanabe, Takashi

2010-05-01

Mesenchymal stem cells (MSCs) have been expected to become useful gene delivery vehicles against human malignant gliomas when coupled with an appropriate vector system, because they migrate towards the lesion. Human artificial chromosomes (HACs) are non-integrating vectors with several advantages for gene therapy, namely, no limitations on the size and number of genes that can be inserted. We investigated the migration of human immortalized MSCs bearing a HAC vector containing the herpes simplex virus thymidine kinase gene (HAC-tk-hiMSCs) towards malignant gliomas in vivo. Red fluorescence protein-labeled human glioblastoma HTB14 cells were implanted into a subcortical region in nude mice. Four days later, green fluorescence protein-labeled HAC-tk-hiMSCs were injected into a contralateral subcortical region (the HTB14/HAC-tk-hiMSC injection model). Tropism to the glioma mass and the route of migration were visualized by fluorescence microscopy and immunohistochemical staining. HAC-tk-hiMSCs began to migrate toward the HTB14 glioma area via the corpus callosum on day 4, and gathered around the HTB14 glioma mass on day 7. To test whether the delivered gene could effectively treat glioblastoma in vivo, HTB14/HAC-tk-hiMSC injected mice were treated with ganciclovir (GCV) or PBS. The HTB14 glioma mass was significantly reduced by GCV treatment in mice injected with HAC-tk-hiMSCs. It was confirmed that gene delivery by our HAC-hiMSC system was effective after migration of MSCs to the glioma mass in vivo. Therefore, MSCs containing HACs carrying an anticancer gene or genes may provide a new tool for the treatment of malignant gliomas and possibly of other tumor types.

Medical decision-making capacity in patients with malignant glioma.

PubMed

Triebel, Kristen L; Martin, Roy C; Nabors, Louis B; Marson, Daniel C

2009-12-15

Patients with malignant glioma (MG) must make ongoing medical treatment decisions concerning a progressive disease that erodes cognition. We prospectively assessed medical decision-making capacity (MDC) in patients with MG using a standardized psychometric instrument. Participants were 22 healthy controls and 26 patients with histologically verified MG. Group performance was compared on the Capacity to Consent to Treatment Instrument (CCTI), a psychometric measure of MDC incorporating 4 standards (choice, understanding, reasoning, and appreciation), and on neuropsychological and demographic variables. Capacity outcomes (capable, marginally capable, or incapable) on the CCTI standards were identified for the MG group. Within the MG group, scores on demographic, clinical, and neuropsychological variables were correlated with scores on each CCTI standard, and significant bivariate correlates were subsequently entered into exploratory stepwise regression analyses to identify multivariate cognitive predictors of the CCTI standards. Patients with MG performed significantly below controls on consent standards of understanding and reasoning, and showed a trend on appreciation. Relative to controls, more than 50% of the patients with MG demonstrated capacity compromise (marginally capable or incapable outcomes) in MDC. In the MG group, cognitive measures of verbal acquisition/recall and, to a lesser extent, semantic fluency predicted performance on the appreciation, reasoning, and understanding standards. Karnofsky score was also associated with CCTI performance. Soon after diagnosis, patients with malignant glioma (MG) have impaired capacity to make treatment decisions relative to controls. Medical decision-making capacity (MDC) impairment in MG seems to be primarily related to the effects of short-term verbal memory deficits. Ongoing assessment of MDC in patients with MG is strongly recommended.

Fisetin suppresses ADAM9 expression and inhibits invasion of glioma cancer cells through increased phosphorylation of ERK1/2.

PubMed

Chen, Chien-Min; Hsieh, Yi-Hsien; Hwang, Jin-Ming; Jan, Hsun-Jin; Hsieh, Shu-Ching; Lin, Shin-Huey; Lai, Chung-Yu

2015-05-01

Fisetin (3,3',4',7-tetrahydroxyflavone) is a naturally occurring flavonoid which is widely distributed in plants. It has been reported to possess some anticancer and anti-invasive capabilities. We set out to explore the effects of fisetin on antimetastatic and its mechanism of action in GBM8401 cells. The results indicated that fisetin exhibited effective inhibition of cell migration and inhibited the invasion of GBM8401 cells under non-cytotoxic concentrations. To identify the potential targets of fisetin, human proteinase antibody array analysis was performed, and the results indicated that the fisetin treatment inhibited the expression of ADAM9 protein and mRNA, which are known to contribute to the progression of glioma cancer. Our results showed that fisetin phosphorylated ERK1/2 in a sustained way that contributed to the inhibited ADAM9 protein and mRNA expression determined by Western blot and RT-PCR. Moreover, inhibition of ERK1/2 by U0126 or transfection with the siERK plasmid significantly abolished the fisetin-inhibited migration and invasion through activation of the ERK1/2 pathway. In summary, our results suggest that fisetin might be a potential therapeutic agent against human glioma cells based on its capacity to activate ERK1/2 and to inhibit ADAM9 expression.