Blockage of glioma cell survival by truncated TEAD-binding domain of YAP.

BACKGROUND: Gliomas are highly aggressive and lack of efficient targeted therapy. YAP, as a Hippo pathway downstream effector, plays a key role in promoting tumor development through the interaction with transcription factor TEAD on the NH3-terminal proline-rich domain. Therefore, targeting TEAD-interacting domain of YAP may provide a novel approach for the treatment of gliomas. MATERIALS AND METHODS: We generated a truncated YAP protein which includes the TEAD-binding domain (YAPBD), and supposed YAPBD can interact with endogenous TEAD but lost the function to activate YAP target gene expressions. The association of YAP expression with the malignant characters of glioma tissues were determined by immunohistochemistry. TEAD-binding capacity of YAPBD was determined by co-immunoprecipitation. The cell proliferation and migration were determined by MTT assay, xenograft assay, wound healing assay and transwell assay, respectively. YAP target genes were detected by Western blot. RESULTS: YAP was highly expressed in glioma tissues and associated with tumor malignancy. YAPBD could block the TEAD-YAP complex formation by competing with YAP binding to TEAD. YAPBD could inhibit glioma cell growth both in vitro and in vivo, through the induction of cell cycle arrest and apoptosis. The cell cycle-related gene cyclin D1 and c-myc, and anti-apoptotic gene Bcl-2, Bcl-xL and survivin were inhibited after YAPBD overexpression. Furthermore, YAPBD also decreased cell migration and invasion, and repressed epithelial-mesenchymal transition. CONCLUSION: YAPBD can block glioma cell survival and repress YAP-dependent gene expressions, indicating gene therapy which targets TEAD-YAP complex would be a potential and significant novel approach for human malignant gliomas.

Truncated TEAD-binding protein of TAZ inhibits glioma survival through the induction of apoptosis and repression of epithelial-mesenchymal transition.

Transcriptional coactivator with PDZ-binding motif (TAZ), a Hippo pathway downstream effector, promotes tumor progression by serving as a transcriptional coactivator with TEAD. Here, we introduced a new construct which can express the TEAD-binding domain of TAZ protein (TAZBD), and determined its antitumor effect in malignant glioma both in vitro and in vivo. We first observed that TAZ was upregulated in glioma tissues and related to malignant clinicopathologic characteristic, indicating the crucial role of TAZ during glioma progression. In U87 and U251 cells, TAZBD expression increased the proportion of apoptotic cells, and suppressed the colony formation and tumorigenicity. Further, TAZBD also decreased cell metastasis through the repression of epithelial-mesenchymal transition. The mechanistic study showed that TAZBD suppression of glioma cells was predominantly through blocking the TAZ-TEAD complex formation by competing with endogenous TAZ. Thus, the gene therapy of malignant glioma through blocking TAZ-TEAD complex by TAZBD may provide a new way for the targeted therapy of glioma.

Malignant Intracranial High Grade Glioma and Current Treatment Strategy.

Malignant high-grade glioma (HGG) is the most common and extremely fatal type of primary intracranial tumor. These tumors recurred within 2 to 3 cm of the primary region of tumor resection in the majority of cases. Furthermore, the blood-brain barrier significantly limited the access of many systemically administered chemotherapeutics to the tumor, pointing towards a stringent need for new therapeutic patterns. Therefore, targeting therapy using local drug delivery for HGG becomes a priority for the development of novel therapeutic strategies. The main objectives to the effective use of chemotherapy for HGG include the drug delivery to the tumor region and the infusion of chemotherapeutic agents into the vascular supply of a tumor directly, which could improve the pharmacokinetic profile by enhancing drug delivery to the neoplasm tissue. Herein, we reviewed clinical and molecular features, different methods of chemotherapy application in HGGs, especially the existing and promising targeting therapies using local drug delivery for HGG which could effectively inhibit tumor invasion, proliferation and recurrence of HGG to combat the deadly disease. Undoubtedly, novel chemical medicines targeting these HGG may represent one of the most important directions in the Neuro-oncology.

Inhibition of eIF4F complex loading inhibits the survival of malignant glioma.

The eukaryotic initiation factor (eIF)4E­binding proteins (4E­BPs) regulate cap­dependent protein translation and control the assembly of the eIF4F complex. In the present study, a phosphorylation­deficient truncated 4E­BP2 (eIF4FD) was constructed into the eukaryotic expression vector pSecTag2, and the in vitro and in vivo effects on malignant glioma survival were determined through inhibiting eIF4F complex assembly. Cell cycle distribution analysis and TUNEL staining show that overexpression of eIF4FD suppressed cell proliferation and induced apoptosis in U251 cells. Western blotting showed that the cell cycle­related genes cyclin D1 and C­myc, and anti­apoptotic genes B­cell lymphoma 2 (Bcl­2), Bcl­extra large and survivin were reduced following the overexpression of eIF4FD. Furthermore, eIF4FD suppressed glioma vascularization via reductions in the expression of ß­catenin and vascular endothelial growth factor. In the orthotopic xenograft model, the stable expression of eIF4FD in U251 cells attenuated cell growth and increased the rate of apoptosis. Accordingly, pSecTag2­PTD­eIF4FD injection via the tail vein of mice also lead to cell growth inhibition and the induction of apoptosis. Therefore, the study showed that phosphorylation­deficient truncated 4E­BP2 efficiently inhibited eIF4E and prevented the formation of the eIF4F complex, which further contributed to the inhibition of cell proliferation and vascularization, and the induction of apoptosis. Therefore, the 4E­BP2­based phosphorylation­deficient truncation designed in the present study may represent a novel approach for the targeted therapy of human malignant glioma though inhibition of the translation initiation complex.

Does Early Postsurgical Temozolomide Plus Concomitant Radiochemotherapy Regimen Have Any Benefit in Newly-diagnosed Glioblastoma Patients? A Multi-center, Randomized, Parallel, Open-label, Phase II Clinical Trial.

BACKGROUND: The radiochemotherapy regimen concomitantly employing temozolomide (TMZ) chemotherapy and radiotherapy (RT) 4 weeks after surgery, followed by 6 cycles of TMZ is a common treatment for glioblastoma (GBM). However, its median overall survival (OS) is only 14.6 months. This study was to explore the effectiveness and safety of early TMZ chemotherapy between surgery and chemoradiotherapy plus the standard concomitant radiochemotherapy regimen. METHODS: A randomized, parallel group, open-label study of 99 newly diagnosed GBM patients was conducted at 10 independent Chinese neurosurgical departments from June 2008 to June 2012. Patients were treated with concomitant radiochemotherapy regimen plus early postsurgical temozolomide (early TMZ group) or standard concomitant radiochemotherapy regimen (control group). Overall response was assessed based on objective tumor assessments, administration of corticosteroid and neurological status test. Hematological, biochemical, laboratory, adverse event (AE), and neurological condition were measured for 24 months of follow-up. The primary efficacy endpoint of this study was overall survival (OS). The secondary endpoint was progression free survival (PFS). RESULTS: The median OS time in the early TMZ group was 17.6 months, compared with 13.2 months in the control group (log-rank test P = 0.021). In addition, the OS rate in the early TMZ group was higher at 6, 12, and 18 months than in the control group, respectively (P < 0.05). The median PFS time was 8.7 months in the early TMZ group and 10.4 months in the control group (log-rank test P = 0.695). AEs occurred in 29 (55.8%) and 31(73.8%) patients respectively in early and control groups, including nausea (15.4% vs. 33.3%), vomiting (7.7% vs. 28.6%), fever (7.7% vs. 11.9%), and headache (3.8% vs. 23.8%). Only 30.8% and 33.3% were drug-related, respectively. CONCLUSIONS: Addition of TMZ chemotherapy in the early break of the standard concomitant radiochemotherapy regimen was well tolerated and significantly improved the OS of the GBM patients, compared with standard concomitant radiochemotherapy regimen. However, a larger randomized trial is warranted to verify these results.

The cap-translation inhibitor 4EGI-1 induces mitochondrial dysfunction via regulation of mitochondrial dynamic proteins in human glioma U251 cells.

Translation initiation factors (eIFs) are over-activated in many human cancers and may contribute to their progression. The small molecule 4EGI-1, a potent inhibitor of translation initiation through disrupting eIF4E/eIF4G interaction, has been shown to exert anti-cancer effects in human cancer cells. The goal of the present study was to evaluate the anti-cancer effects of 4EGI-1 in human glioma U251 cells. We found that 4EGI-1 impaired the assembly of the eIF4F complex, and inhibited proliferation of U251 cells via inducing apoptosis. 4EGI-1 treatment induced collapse of mitochondrial membrane potential (MMP) and production of intracellular reactive oxygen species (ROS), which were prevented by the ROS scavenger N-acetyl-cysteine (NAC). In addition, 4EGI-1 inhibited mitochondrial ATP synthesis via suppressing complex I activity, but had no effects on mitochondrial biogenesis. The results of fluorescence staining showed that 4EGI-1 indeed fragmented the mitochondrial network of U251 cells. We found a significant decrease in optic atrophy type 1 (Opa-1) and mitofusin 1 (Mfn-1) related to fusion proteins as well as an increase in fission protein dynamin-related protein 1 (Drp-1). Furthermore, the anti-cancer effects of 4GI-1 were partially nullified by knock down of Drp-1 using siRNA. These data indicate that the use of inhibitors that directly target the translation initiation complex eIF4F could represent a potential novel approach for human glioma therapy.

The expression of hepatoma-derived growth factor in primary central nervous system lymphoma and its correlation with angiogenesis, proliferation and clinical outcome.

Hepatoma-derived growth factor (HDGF), a potential predictive and prognostic marker in several human cancers, is the firstly reported member of the HDGF family of proteins containing a well-conserved N-terminal amino acid sequence. HDGF is implicated in tumorigenesis by direct angiogenic activity, and its expression is correlated with aggressive biological ability of cancer cells including proliferation and angiogenesis. So, we propose that HDGF may be a valuable factor in progression and prognosis for primary central nervous system lymphoma (PCNSL) through its angiogenic and proliferative activity. So, HDGF, CD31 and Ki67 expression in the specimens of 60 patients suffering from PCNSL was investigated by immunohistochemistry in this study. Their correlations with clinicopathologic features and prognosis were evaluated to determine whether HDGF, CD31 and Ki67 expression levels correlate with the prognosis of the 60 patients suffering from PCNSL. We found that all PCNSL specimens showed HDGF, CD31 and Ki67 expression with different expression levels. Statistical analysis showed that HDGF had a positive correlation with CD31, but not with Ki67. Patients with higher HDGF and CD31 expression level had poorer overall survival rates than those with lower expression levels of HDGF and CD31, while Ki67 expression level did not correlate with overall survival. Multivariate analysis revealed that postoperative adjuvant chemotherapy and high expression of HDGF was independent prognostic indicator of patient survival.

Neuritin expression and its relation with proliferation, apoptosis, and angiogenesis in human astrocytoma.

Neuritin, a new member of the neurotrophic factor family, plays an important role in promoting neuronal survival, differentiation, function, and repair. However, whether neuritin is expressed in human astrocytoma and involved in their proliferation, apoptosis, and angiogenesis remains unclear. The expression of neuritin messenger RNA, protein and the relationship with proliferation, apoptosis, and angiogenesis were examined in human astrocytoma samples and three glioma cell lines by immunohistochemistry, Western blot, and quantitative real-time RT-PCR and so on. And neuritin immunoreactivity score (IRS), proliferative index (PI), apoptotic index (AI), overall daily growth (ODG), and microvessel density (MVD) in brain astrocytoma were measured. The results showed that neuritin was overexpressed in human astrocytoma samples, and the overexpression correlated positively with the malignancy of astrocytomas as reflected by changes in proliferation, apoptosis, and angiogenesis markers. In our study, we found neuritin is overexpressed in astrocytoma, which may be an important factor in tumorigenesis and progression of astrocytoma, and can be used as a target for biological therapy.

Hemorrhagic pituitary macroadenoma: characteristics, endoscopic endonasal transsphenoidal surgery, and outcomes.

PURPOSE: This study aims to assess the effect of endoscopic endonasal transsphenoidal surgery (EETSS) of hemorrhagic pituitary macroadenoma (HPMA). PATIENTS AND METHODS: We retrospectively reviewed 52 cases with HPMA collected from the Xijing Hospital from April 1995 to April 2009. There were 39 males and 13 females, ranging in age from 18 to 79 years (average 51.6 years). Patients presented with headache or acute ophthalmological symptoms after adenoma hemorrhage. Computed tomography (CT) scan and magnetic resonance imaging (MRI) showed pituitary macroadenoma with hemorrhage in all cases. Twenty-eight adenomas showed marked suprasellar extension, 19 showed moderate extension, and another 5 showed slight extension. All patients were promptly treated by emergency EETSS, usually within 24 h after hospitalization. RESULTS: Total removal of tumor was achieved in 46 cases (88.5%), and subtotal removal in 6 cases (11.5%). Postoperative radiotherapy and reoperation of the tumor were required in five patients with either residual or relapsed tumors. Follow-up ranged from 8 to 93 months (mean 41.6 months) for 43 cases. Visual acuity and visual field recovery and improvement was recorded in 92.1% and 94.3% of patients who had preoperative visual symptoms, respectively. CONCLUSIONS: The majority of macroadenomas are hemorrhagic, and they often occur in middle-aged, male subjects. Detection by imaging in the setting of pituitary apoplexy accurately predicts the nature of the apoplectic process and helps to guide the type and timing of surgery. Early EETSS is the most effective therapy and significantly improves visual outcomes and systemic conditions.

Chordoid glioma: a case report and literature review.

BACKGROUND: chordoid glioma is a rare tumor (World Health Organization grade II) with both glial and chordoid features, often located in the suprasellar region and anterior third ventricle. It was first described by Brat in 1998. Because there is no detailed information available from the clinical perspective, we reviewed the literature. METHODS: a literature search through PUBMED and CNKI revealed 64 cases of chordoid glioma. Information on the clinical course was very limited. We reviewed the literature and studied the pathologic and imaging features, postoperative mortality and morbidity in relation to surgical extension and approaches, and the importance of adjuvant treatment. CONCLUSIONS: mortality in the immediate postoperative period is 28%, and postoperative morbidity is 60%, which are statistically higher after gross total resection as compared with subtotal resection. Translamina terminalis approach is considered to be the best approach. The current study cannot document that patients have longer survival and higher quality of life after gross total resection than subtotal resection. The role of postoperative radiotherapy is uncertain and there is no report on the use of chemotherapy. More information about the optimal treatment strategy is needed.

Up-regulation of EphA2 and down-regulation of EphrinA1 are associated with the aggressive phenotype and poor prognosis of malignant glioma.

Malignant gliomas display over-expression of the receptor tyrosine kinase EphA2. However, expression levels of the EphA2 ligand, EphrinA1, have not been fully elucidated. Seventy-eight patients with primary gliomas were included in this study who underwent surgical resection, radiation, and chemotherapy. The expression of EphA2 and EphrinA1 in tumors was assessed by immunohistochemistry and was statistically analyzed in combination with the follow-up data of patients. EphA2 was highly expressed in most malignant gliomas, but EphrinA1 was expressed at low levels in these tumors. The increased EphA2 expression is associated with higher-grade histology and poor patient prognosis. Contrary to this, the increased EphrinA1 expression is associated with lower-grade histology, but not associated with poor patient prognosis. Moreover, patients with tumors positive for EphA2 and negative for EphrinA1 had significantly shorter overall and progression-free survival than patients with tumors positive for both EphA2 and EphrinA1, negative for both EphA2 and EphrinA1, or negative for EphA2 and positive for EphrinA1. RNAi-mediated suppression of endogenous EphA2 in human glioblastoma multiforme cells resulted in increased EphrinA1 levels, as well as decreased cell viability, anchorage independence and in vitro invasion, and increased apoptosis. Furthermore, suppression of EphA2 resulted in delayed tumor growth in mice xenografts. Together, these data indicate that up-regulation of EphA2 and down-regulation of Ephrina1 may correlate with poor prognosis for patients with high-grade glioma. EphA2 suppression partially reversed the aggressive phenotypes of malignant gliomas, possibly through up-regulating EphrinA1 expression, which may help explain how EphA2 modulates the malignant progression of gliomas.

Prognostic value of pretreatment 18F-FDG PET in patients with primary central nervous system lymphoma: SUV-based assessment.

The purpose of this retrospective study was to evaluate the prognostic value of 18F-fluorodeoxyglucose (FDG) positron emission tomography (PET) in immunocompetent patients with primary central nervous system lymphoma (PCNSL). We also investigated whether FDG uptake was related to angiogenesis in the tumors. Seventeen patients with newly diagnosed and histologically confirmed PCNSL were investigated with FDG-PET before the treatment. FDG uptake was assessed by standardized uptake value of the tumor showing the maximum uptake (SUVmax). The Kaplan­Meier method was used to estimate the overall and the progression-free survival times. The difference of the survival curves between the low to moderate FDG uptake (SUVmax < 12) and high FDG uptake (SUVmax ≥ 12) groups was statistically analyzed. The relationship between FDG SUVmax and microvessel density (MVD) in the tumor specimens determined by CD34 immunohistochemical staining was examined. Finally, the difference of the overall survival curve between the low MVD (<20) and high MVD (≥ 20) groups was statistically analyzed. After the completion of initial treatment, 7 of the 8 low to moderate FDG uptake patients showed complete response (CR) and 1 showed partial response (PR). On the other hand, 5 of the 9 high FDG uptake patients showed CR and 4 showed PR. However, the difference of treatment response (CR vs PR) between the low to moderate and high FDG uptake groups was not statistically significant (P = 0.36). Median survival time was more than 26 months in low to moderate FDG uptake patients and 12 months in high FDG uptake patients. The overall survival time of patients with low to moderate FDG uptake was significantly longer than that of patients with high FDG uptake (P < 0.05). The progression free survival (PFS) was also significantly longer in patients with low to moderate FDG uptake compared to the patients with high FDG uptake (P < 0.05). There was no significant relationship between FDG SUVmax and MVD in the tumors. The survival time tended to be longer in patients with high MVD than in low MVD, but the difference was not statistically significant (P = 0.15). Pretreatment FDG uptake may have a prognostic value in newly diagnosed PCNSL.

Correlation of L-methyl-11C-methionine (MET) uptake with L-type amino acid transporter 1 in human gliomas.

L-type amino acid transporter 1 (LAT1) is a neutral amino acid transport system and is a major route for the transport of large neutral amino acids, including methionine, through the plasma membrane. LAT1 requires the heavy chain of 4F2 cell surface antigen (4F2hc) for its functional expression. Positron emission tomography (PET) with L-[methyl-(11)C] methionine (MET) provides information about amino acid metabolism in brain tumors. We conducted a clinicopathologic study to elucidate the correlation of LAT1 and 4F2hc expression with MET uptake in patients with newly diagnosed human gliomas. Thirty-three newly diagnosed glioma patients were enrolled in this study. Uptake of MET in the tumor was evaluated with the maximum standardized uptake value (SUVmax). Expression of the LAT1, 4F2hc, and CD34, and Ki-67 labeling index of the tumor were analyzed by immunohistochemical staining, and the correlation with the SUVmax in the tumors was examined. Expression of LAT1 and 4F2hc was higher in high-grade gliomas than in low-grade gliomas. The grade of LAT1 immunostaining increased with glioma grade. LAT1 was mainly expressed in the tumor cytoplasm and vascular endothelium and 4F2hc was mainly expressed in the tumor cytoplasm and plasma membrane. Expression of LAT1 but not 4F2hc was significantly correlated with MET SUVmax. Expression of LAT1 in the tumor vascular endothelium is significantly correlated with CD34 positive microvessel density. In conclusion, MET SUVmax correlates with LAT1 expression in the tumor in newly diagnosed gliomas. MET transport may be increased by an increased number of microvessels combined with a higher density or activity of LAT1 in the tumor endothelial cells in high-grade gliomas. Use of MET-PET as a molecular target combined with anti-angiogenesis in glioma therapy should be addressed in future studies.

Brain Tumor Stem-Like Cells Identified by Neural Stem Cell Marker CD15.

In recent years, a small number of cells that have stem cell properties were identified in human gliomas called brain tumor stem cells (BTSCs), which were thought to mainly contribute to the initiation and development of gliomas and could be identified by the surface marker CD133. However, recent studies indicated that the expression of CD133 might be regulated by environmental conditions such as hypoxia and that there might be CD133(-) BTSCs. Genetic mouse models demonstrated that some gliomas originated from transformed neural stem cells (NSCs). Therefore, we investigated the expression of CD15, a surface marker for NSCs, in tumor spheres derived from astrocytoma and ependymoma. CD15(+) cells isolated from these tumor spheres had properties of BTSCs including self-renewal, multidifferentiation, and the ability to recapitulate the phenocopy of primary tumors. CD15 exhibited stable expression in long-term cultured tumor spheres, which sustained BTSCs properties, whereas CD133 expression decreased significantly in late passages. Furthermore, CD15(+)CD133(-) cells isolated from early or late passages of tumor spheres showed similar characteristics of BTSCs. Examination of glioma samples by immunohistochemistry showed that CD15 was expressed in a subset of human brain tumors. Therefore, CD15 can be used as a marker of stem-like cells derived from brain tumors that might contain CD133(-) BTSCs.

Gross-total hematoma removal of hypertensive basal ganglia hemorrhages: a long-term follow-up.

BACKGROUND AND PURPOSE: Hypertensive basal ganglia hemorrhage (HBGH) accounts for 35%-44% of cases of hypertensive intracranial hemorrhage (ICH), which is one of the most devastating forms of cerebrovascular disease. In this study, intracerebral hematoma was evacuated with a burr hole craniectomy. The relationships of residue hematoma volume to brain edema, inflammation factors and the long-term prognosis of HBGH patients were studied. METHODS: One hundred and seventy-six patients with HBGH were randomly divided into gross-total removal of hematoma (GTRH) and sub-total removal of hematoma (STRH) groups. The pre-operative and post-operative data of the patients in the two groups were compared. The pre-operative data included age, sex, hematoma volume, time from the ictus to the operation, Glasgow Coma Scale (GCS) scores, and the European Stroke Scale (ESS) scores. The post-operative information included edema grade, level of thromboxane B2 (TXB2), 6-keto-prostaglandin F1a (6-K-PGF1a), tumor necrosis factor-a (TNF-a) and endothelin (ET) in hematoma drainage or cerebral spinal fluid (CSF), ESS and Barthel Index (BI). RESULTS: There was no statistical difference between the two groups (P>0.05) in the pre-operative data. The levels of TXB2, 6-K-PGF1a, TNF-a and ET in the GTRH group were significantly lower than those in the STRH group at different post-operative times. The ESS in the GTRH group increased rapidly after the operation and was higher than that in the STRH group. There was a significant difference between the two groups (P<0.05). The post-operative CT scan at different times showed that the brain edema grades were better in the GTRH group than in the STRH group. The BI was higher in the GTRH group than in the STRH group (P<0.05). CONCLUSIONS: GTRH is an effective method to decrease ICH-induced injury to brain tissue. Such effect is related to decreased perihematomal edema formation and secondary injury by coagulation end products activated inflammatory cascade.

Progress on potential strategies to target brain tumor stem cells.

The identification of brain tumor stem cells (BTSCs) leads to promising progress on brain tumor treatment. For some brain tumors, BTSCs are the driving force of tumor growth and the culprits that make tumor revive and resistant to radiotherapy and chemotherapy. Therefore, it is specifically significant to eliminate BTSCs for treatment of brain tumors. There are considerable similarities between BTSCs and normal neural stem cells (NSCs), and diverse aspects of BTSCs have been studied to find potential targets that can be manipulated to specifically eradicate BTSCs without damaging normal NSCs, including their surface makers, surrounding niche, and aberrant signaling pathways. Many strategies have been designed to kill BTSCs, and some of them have reached, or are approaching, effective therapeutic results. Here, we will focus on advantages in the issue of BTSCs and emphasize on potential therapeutic strategies targeting BTSCs.

Ardipusilloside I induces apoptosis in human glioblastoma cells through a caspase-8-independent FasL/Fas-signaling pathway.

Ardipusilloside I, a triterpenoid saponin isolated from Ardisia pusilla A. DC, suppresses the growth of a variety of cancer cells, and has certain immunomodulative properties. Herein, we investigated its effect on glioblastoma cell line U87MG cells and primary cultured human glioblastoma cells, and examined the underlying mechanism of action. Ardipusilloside I substantially decreased the number of viable cells of both cell lines in a time- and concentration-dependent manner, with a similar IC(50) of 4.05µM. Microscopy revealed apoptotic characteristics, including chromatin condensation and cell nucleus fragmentation, demonstrating that ardipusilloside I-induced apoptosis. Ardipusilloside I exposure also gradually increased the sub-G1 fraction (the apoptotic cell population) and an S phase-arrest of both glioblastoma cells. Furthermore, ardipusilloside I increased the expression of Fas and its ligand (FasL), and enhanced the activation of caspase-8 and caspase-3. Additionally, we observed a significant decreased apoptosis after the trigger effection of FasL was abolished by the neutralization antibody anti-FasL antibody and an unchanged apoptosis level when the activation of caspase-8 was interrupted by specific inhibitor z-IETD-fmk, which suggested that a casepase-8 independent FasL/Fas-signaling-mediated death receptor pathway is involved. These data suggested that ardipusilloside I could be developed as a chemotherapeutic agent for the management of gliomas.

Apoptosis induced by ardipusilloside III through BAD dephosphorylation and cleavage in human glioblastoma U251MG cells.

Ardipusilloside III is a saponin newly isolated from Ardisia pusilla A.DC. Since saponins have exhibited broad anti-cancer and pro-apoptotic activity, we investigated the ability of ardipusilloside III to induce apoptosis in human glioblastoma U251MG cells, as well as the involvement of apoptotic signaling pathways. Ardipusilloside III markedly suppressed proliferation of U251MG cells in a time- and dose-dependent manner (P < 0.05, IC50 = 8.2 microg/ml), but did not affect the growth of primary cultures of human astrocytes. Ardipusilloside III-treated U251MG cells underwent typical apoptotic changes. Exposure to a low dose of ardipusilloside III provoked G2/M-phase cell cycle arrest, which preceded apoptosis characterized by the appearance of cells with sub-G1 DNA content. However, a higher dose of ardipusilloside III induced apoptosis without first causing cell cycle arrest. In addition, ardipusilloside III exposure resulted in time-dependent BAD dephosphorylation and cleavage as well as activation of caspase-8 and caspase-3. Therefore, both the intrinsic pathway of apoptosis, mediated by BAD dephosphorylation and cleavage, and the extrinisic pathway of apoptosis, mediated by caspase-8 and caspase-3 activation, were involved in ardipusilloside III-induced apoptosis. These data suggest that ardipusilloside III is a reliable candidate for chemotherapeutic treatment of human glioblastomas, and should be investigated further.