phenanthrenes and Glioma

Isocitrate dehydrogenase (IDH) mutation is a common genetic abnormality in human malignancies characterized by remarkable metabolic reprogramming. Our present study demonstrated that IDH1-mutated cells showed elevated levels of reactive oxygen species and higher demands on Nrf2-guided glutathione de novo synthesis. Our findings showed that triptolide, a diterpenoid epoxide from

Topics: Amino Acid Transport System y+; Animals; Biosynthetic Pathways; Cell Line, Tumor; Diterpenes; Epoxy Compounds; Gene Expression Regulation, Neoplastic; Glioma; Glutamate-Cysteine Ligase; Glutathione; Humans; Isocitrate Dehydrogenase; Mice; Mutation; NF-E2-Related Factor 2; Phenanthrenes; Reactive Oxygen Species; Synthetic Lethal Mutations; Xenograft Model Antitumor Assays

Immunosuppression is one of hallmark features in many cancers including glioma. Triptolide, a natural compound purified from the Chinese herb Tripterygium wilfordii, has been reported to inhibit PD-L1 otherwise known as the B7 homolog 1 (B7-H1) expression in breast cancer. The purpose of this paper is to test the effects of Triptolide on T cell inhibition in glioma cells.. We labeled T cells and cocultured with Interferon-γ (IFN-γ) and Triptolide treated glioma cells. The effect on inhibition of T cells as well as subtypes of T cells was measured by Flow Cytometry. We also tested the expression of PD-L1 in six glioma cell lines.. We found that Triptolide could reverse T cell inhibition especially CD4+ T cell and induced IFN-γ secretion. In addition, Triptolide could also induce interleukin-2 secretion and overcome interleukin-10 inhibition caused by glioma cells under IFN-γ treated condition. Triptolide could also down-regulate IFN-γ induced PD-L1 surface expression in glioma cells.. These results suggest that Triptolide may be used to reverse CD4+ T cell inhibition caused by glioma cells and is an alternative candidate for targeting PD-L1, one of the checkpoint inhibitors for the treatment of glioma.

Topics: Antineoplastic Agents, Alkylating; B7-H1 Antigen; CD4-Positive T-Lymphocytes; Coculture Techniques; Diterpenes; Down-Regulation; Epoxy Compounds; Gene Expression Regulation, Neoplastic; Glioma; Humans; Interferon-gamma; Phenanthrenes; T-Lymphocytes, Helper-Inducer; Tumor Cells, Cultured

Malignant gliomas (MGs) are one of the most common primary brain cancers in adults with a high mortality rate and relapse rate. Thus, finding better effective approaches to treat MGs has become very urgent. Here, we studied the effects of cryptotanshinone (CTS) on MGs in vitro and in vivo, and explored the underlying mechanisms. Effects of CTS in vitro on cell proliferation, cycle, migration and invasion were evaluated. The activation of JAK/STATs signaling was detected by western blot and immunofluorescenc staining. SHP-2 inhibitor or SiRNA were used to determine the involvement of SHP-2. The in vivo anti-MGs activity of CTS was studied with nude mice bearing intracerebral U87 xenografts. Our results revealed that CTS significantly inhibited the proliferation of MGs in vitro via inhibiting STAT3 signal pathway. The cell cycle was arrested at G0/G1 phase. Although CTS did not change the expression of total SHP-2 protein, the tyrosine phosphatase activity of SHP-2 protein was increased by CTS treatment in a dose-dependent manner in vivo and in vitro. SHP-2 inhibitor or SiRNA could reverse the inhibitory effect of CTS on phosphorylation of STAT3 Tyr705. In vivo study also showed that CTS inhibited the intracranial tumor growth and extended survival of nude mice bearing intracerebral U87 xenografts, confirming an inhibitory effect of CTS on MGs. Our results indicated CTS may be a potential therapeutic agent for MGs. The inhibitory action of CTS is largely attributed to the inhibition of STAT3 Tyr705 phosphorylation with a novel mechanism of upregulating the tyrosine phosphatase activity of SHP-2 protein.

Topics: Animals; Cell Line, Tumor; Cell Proliferation; Female; Glioma; Humans; Mice; Mice, Inbred BALB C; Mice, Nude; Neoplasm Proteins; Phenanthrenes; Protein Tyrosine Phosphatase, Non-Receptor Type 11; Signal Transduction; STAT3 Transcription Factor; Xenograft Model Antitumor Assays

Among primary brain cancers, gliomas are the most deadly and most refractory to current treatment modalities. Previous reports overwhelmingly support the role of the RNA-binding protein Hu antigen R (HuR) as a positive regulator of glioma disease progression. HuR expression is consistently elevated in tumor tissues, and a cytoplasmic localization appears essential for HuR-dependent oncogenic transformation. Here, we report HuR aggregation (multimerization) in glioma and the analysis of this tumor-specific HuR protein multimerization in clinical brain tumor samples. Using a split luciferase assay, a bioluminescence resonance energy transfer technique, and site-directed mutagenesis, we examined the domains involved in HuR multimerization. Results obtained with the combination of the split HuR luciferase assay with the bioluminescence resonance energy transfer technique suggested that multiple (at least three) HuR molecules come together during HuR multimerization in glioma cells. Using these data, we developed a model of HuR multimerization in glioma cells. We also demonstrate that exposing glioma cells to the HuR inhibitor tanshinone group compound 15,16-dihydrotanshinone-I or to the newly identified compound 5 disrupts HuR multimerization modules and reduces tumor cell survival and proliferation. In summary, our findings provide new insights into HuR multimerization in glioma and highlight possible pharmacological approaches for targeting HuR domains involved in cancer cell-specific multimerization.

Topics: Brain Neoplasms; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cell Transformation, Neoplastic; ELAV-Like Protein 1; Furans; Glioma; Humans; Neoplasm Proteins; Phenanthrenes; Protein Aggregation, Pathological; Protein Domains; Quinones

Inhibitors of PARP-1(Poly(ADP-ribose) polymerase-1) act by competing with NAD(+), the enzyme physiological substrate, which play a protective role in many pathological conditions characterized by PARP-1 overactivation. It has been shown that PARP-1 also promotes tumor growth and progression through its DNA repair activity. Since angiogenesis is an essential requirement for these activities, we sought to determine whether PARP inhibition might affect rat brain microvascular endothelial cells (GP8.3) migration, stimulated by C6-glioma conditioned medium (CM). Through wound-healing experiments and MTT analysis, we demonstrated that PARP-1 inhibitor PJ-34 [N-(6-Oxo-5,6-dihydrophenanthridin-2-yl)-N,N-dimethylacetamide] abolishes the migratory response of GP8.3 cells and reduces their viability. PARP-1 also acts in a DNA independent way within the Extracellular-Regulated-Kinase (ERK) signaling cascade, which regulates cell proliferation and differentiation. By western analysis and confocal laser scanning microscopy (LSM), we analyzed the effects of PJ-34 on PARP-1 expression, phospho-ERK and phospho-Elk-1 activation. The effect of MEK (mitogen-activated-protein-kinase-kinase) inhibitor PD98059 (2-(2-Amino-3-methoxyphenyl)-4 H-1-benzopyran-4-one) on PARP-1 expression in unstimulated and in CM-stimulated GP8.3 cells was analyzed by RT-PCR. PARP-1 expression and phospho-ERK activation were significantly reduced by treatment of GP8.3 cells with PJ-34 or PD98059. By LSM, we further demonstrated that PARP-1 and phospho-ERK are coexpressed and share the same subcellular localization in GP8.3 cells, in the cytoplasm as well as in nucleoplasm. Based on these data, we propose that PARP-1 and phospho-ERK interact in the cytosol and then translocate to the nucleus, where they trigger a proliferative response. We also propose that PARP-1 inhibition blocks CM-induced endothelial migration by interfering with ERK signal-transduction pathway.

Topics: Active Transport, Cell Nucleus; Animals; Brain; Brain Neoplasms; Cell Line, Tumor; Cell Movement; Culture Media, Conditioned; Endothelial Cells; ets-Domain Protein Elk-1; Extracellular Signal-Regulated MAP Kinases; Glioma; Microvessels; Paracrine Communication; Phenanthrenes; Phosphorylation; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerase Inhibitors; Poly(ADP-ribose) Polymerases; Protein Binding; Protein Kinase Inhibitors; Rats; Signal Transduction

Glioblastoma multiforme (GBM) is a lethal solid cancer in adults. Temozolomide (TMZ) is a first-line chemotherapeutic agent but the efficacy is limited by intrinsic and acquired resistance in GBM. Triptolide (TPL), a derivative from traditional Chinese medicine, demonstrated anti-tumor activity. In this study, we explored the interaction of TPL and TMZ in glioma-initiating cells (GICs) and the potential mechanism. A GIC line (GIC-1) was successfully established. Cell viability of GIC-1 after treatment was measured using a CCK-8 assay. The interaction between TPL and TMZ was calculated from Chou-Talalay equations and isobologram. Self-renewal was evaluated with tumor sphere formation assay. Apoptosis was assessed with flow cytometry and western blot. Luciferase assay was employed to measure NF-κB transcriptional activity. The expression of NF-κB downstream genes, NF-κB nuclear translocalization and phoshorylation of IκBα and p65 were evaluated using western blot. We found that GIC-1 cells were resistant to TMZ, with the expected IC50 of 705.7 μmol/L. Co-treatment with TPL yielded a more than three-fold dose reduction of TMZ. TPL significantly increased the percentage of apoptotic cells and suppressed the tumor sphere formation when combined with TMZ. Phosphorylation of IκBα and p65 coupled with NF-κB nuclear translocalization were notably inhibited after a combined treatment. Co-incubation synergistically repressed NF-κB transcriptional activity and downstream gene expression. TPL sensitizes GICs to TMZ by synergistically enhancing apoptosis, which is likely resulting from the augmented repression of NF-κB signaling. TPL is therefore a potential chemosensitizer in the treatment of GBM.

Topics: Animals; Antineoplastic Agents, Alkylating; Apoptosis; Cell Line, Tumor; Cell Transformation, Neoplastic; Dacarbazine; Disease Models, Animal; Diterpenes; Drug Synergism; Epoxy Compounds; Glioma; Humans; Mice; Neoplasm Transplantation; NF-kappa B; Phenanthrenes; Proto-Oncogene Proteins c-bcl-2; Signal Transduction; Stimulation, Chemical; Temozolomide; Transcription, Genetic; X-Linked Inhibitor of Apoptosis Protein

Triptolide (Trip), a diterpene triepoxide isolated from medicinal vine Trypterygium wilfordii Hook. F. possessed multiple biological activities including antineoplastic actions. However, no report concerning its effects on ion currents has been published. In this study, we attempted to determine whether this compound has any effects on ion currents in malignant glioma cells. The mRNA expression of KCNJ10 (Kir4.1) was detected in U373 glioma cells. The inwardly rectifying K(+) currents (IK(IR)) in U373 cells were almost fully blocked by BaCl2 (1mM). Trip (30 nM-10 μM) effectively decreased the amplitude of IK(IR) in a concentration-dependent manner with an IC50 value of 0.72 μM. In chlorotoxin-treated U373 cells, Trip-mediated block of IK(IR) remained effective. Addition of Trip (3 μM) slightly inhibited the amplitude of Ca(2+)-activated K(+) current and sustained K(+) outward current in U373 cells. In cell-attached configuration, when Trip was added to the bath, the activity of inwardly rectifying K(+) (Kir) channels diminished with no change in single-channel conductance. Its suppression of Kir channels was accompanied by a reduction in the slow component of mean open time. Under current-clamp conditions, addition of Trip depolarized the membrane along with changes in frequency histogram of resting potential. Block by this component of Kir4.1 channels may be an important mechanism underlying its actions on the functional activity of glioma cells. Targeting at Kir4.1 channels may be clinically useful as an adjunctive regimen to anti-cancer drugs.

Topics: Cell Line, Tumor; Diterpenes; Electrophysiological Phenomena; Epoxy Compounds; Gene Expression Regulation; Glioma; Humans; Kinetics; Membrane Potentials; Phenanthrenes; Potassium; Potassium Channels, Inwardly Rectifying; RNA, Messenger; Scorpion Venoms

A series of B-ring modified analogues of triptolide were synthesized and tested for their cytotoxicity against two human tumor cell lines (U251 and PC-3). From the current investigation, the structure-cytotoxic activity relationships of these analogues suggested that the introduction of hydroxyl, epoxide, halogen or olefinic groups on C5 and/or C6 could still retain the cytotoxicity, albeit a little less potency, and the C7,C8-β-epoxide group of triptolide was essential to its potent cytotoxic activity.

Topics: Antineoplastic Agents; Cell Proliferation; Diterpenes; Drug Screening Assays, Antitumor; Epoxy Compounds; Glioma; Humans; Male; Models, Molecular; Molecular Structure; Organic Chemicals; Phenanthrenes; Prostatic Neoplasms; Structure-Activity Relationship; Tumor Cells, Cultured

Malignant gliomas (MGs) are among the most aggressive types of cancers in the human brain. Frequent tumor recurrence caused by a lack of effective therapeutic approaches results in a poor prognosis. Signal transducer and activator of transcription 3 (STAT3), an oncogenic protein, is constitutively activated in MGs and predicts a poor clinical outcome. STAT3 therefore is considered to be a promising target for the treatment of MGs. Cryptotanshinone (CTS), the main bioactive compound from the root of Salvia miltiorrhiza Bunge, has been reported to have various pharmacological effects. However, little is known about its function in MG cells. In this study, we evaluated the effect of CTS on the proliferation of human glioma cell lines (T98G and U87). Our results revealed that CTS significantly suppresses glioma cell proliferation. The phosphorylation of STAT3 Tyr705, but not Ser727, was inhibited by CTS, and STAT3 nuclear translocation was attenuated. Overexpression of constitutively active mutant STAT3C reversed the inhibitory effect of CTS, while knockdown STAT3 showed a similar inhibitory effect as CTS treatment. Following the downregulation of STAT3-regulated proteins cyclinD1 and survivin, cell cycle progression significantly arrested in G1/G0 phase. These results indicate that CTS may be a potential antiproliferation agent for the treatment of MGs and that its mechanism may be related to the inhibition of STAT3 signaling.

Topics: Animals; Brain Neoplasms; Cell Line, Tumor; Cell Proliferation; G1 Phase; Glioma; Humans; Mutant Proteins; Phenanthrenes; Phosphorylation; Phosphotyrosine; Rats; RNA, Small Interfering; S Phase; Signal Transduction; STAT3 Transcription Factor

Whole-body molecular imaging is able to directly map spatial distribution of molecules and monitor its biotransformation in intact biological tissue sections. Imaging mass spectrometry (IMS), a label-free molecular imaging method, can be used to image multiple molecules in a single measurement with high specificity. Herein, a novel easy-to-implement, whole-body IMS method was developed with air flow-assisted ionization in a desorption electrospray ionization mode. The developed IMS method can effectively image molecules in a large whole-body section in open air without sample pretreatment, such as chemical labeling, section division, or matrix deposition. Moreover, the signal levels were improved, and the spatial assignment errors were eliminated; thus, high-quality whole-body images were obtained. With this novel IMS method, in situ mapping analysis of molecules was performed in adult rat sections with picomolar sensitivity under ambient conditions, and the dynamic information of molecule distribution and its biotransformation was provided to uncover molecular events at the whole-animal level. A global view of the differential distribution of an anticancer agent and its metabolites was simultaneously acquired in whole-body rat and model mouse bearing neuroglioma along the administration time. The obtained drug distribution provided rich information for identifying the targeted organs and predicting possible tumor spectrum, pharmacological activity, and potential toxicity of drug candidates.

Topics: Air; Animals; Antineoplastic Agents; Glioma; Indolizines; Male; Mass Spectrometry; Mice; Molecular Imaging; Phenanthrenes; Phenanthrolines; Rats; Rats, Wistar; Whole Body Imaging

Malignant glioma is the most devastating and aggressive tumor in brain, characterized by rapid proliferation and diffuse invasion. Chemotherapy and radiotherapy are the pivotal strategies after surgery; however, high drug resistance of malignant glioma and the blood-brain barrier usually render chemotherapy drugs ineffective. Here, we find that triptolide, a small molecule with high lipid solubility, is capable of inhibiting proliferation and invasion of malignant glioma cells effectively. In both investigated malignant glioma cell lines, triptolide repressed cell proliferation via inducing cell cycle arrest in G0/G1 phase, associated with downregulation of G0/G1 cell cycle regulators cyclin D1, CDK4, and CDK6 followed by reduced phosphorylation of retinoblastoma protein (Rb). In addition, triptolide induced morphological change of C6 cells through downregulation of protein expression of MAP-2 and inhibition of activities of GTPases Cdc42 and Rac1/2/3, thus significantly suppressing migratory and invasive capacity. Moreover, in an in vivo tumor model, triptolide delayed growth of malignant glioma xenografts. These findings suggest an important inhibitory action of triptolide on proliferation and invasion of malignant glioma, and encourage triptolide as a candidate for glioma therapy.

Topics: Animals; Antineoplastic Agents, Alkylating; Apoptosis; Blotting, Western; Brain Neoplasms; cdc42 GTP-Binding Protein; Cell Cycle; Cell Movement; Cell Proliferation; Cyclin-Dependent Kinases; Diterpenes; Epoxy Compounds; Female; Flow Cytometry; Glioma; Mice; Mice, Nude; Neoplasm Invasiveness; Phenanthrenes; rac GTP-Binding Proteins; rac1 GTP-Binding Protein; RAC2 GTP-Binding Protein; Rats; Retinoblastoma Protein; Tumor Cells, Cultured

Signal transducer and activator of transcription 3 (STAT3) is usually constitutively activated in a variety of malignancies. Thus, STAT3 may be a promising target for treatment of tumor cells. Recently, Tanshinone IIA (Tan IIA), a major active constituent from the root of Salvia miltiorrhiza Bunge, was reported to have apoptosis inducing effects on a large variety of cancer cells. In this study, we evaluate the anti-proliferation and apoptosis inducing effects of Tan IIA on C6 glioma cells. Cell growth and proliferation were measured by MTT assay, cell apoptosis was observed by flow cytometry and DNA-fragmentation analysis. Further more, we investigated inhibitory effects of Tan IIA on STAT3 activity and its downstream targets: Bcl-XL, cyclin D1. Alteration of STAT3 activity was examined by measuring their DNA binding activity and tyrosine phosphorylation. Changes in the expression levels of Bcl-XL and cyclin D1 were examined by Western blot analysis. We found that the cellular growth were inhibited and cell apoptosis were observed after the treatment with Tan IIA. The STAT3 activity was significantly reduced by Tan IIA parallel with a significant attenuation of expression of Bcl-XL and cyclin D1. These results suggest that Tan IIA may serve as an effective adjunctive reagent in the treatment of glioma for its targeting of constitutive STAT3 signaling.

Topics: Abietanes; Animals; Antineoplastic Agents, Phytogenic; Apoptosis; bcl-X Protein; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cyclin D1; DNA Fragmentation; Dose-Response Relationship, Drug; Glioma; Phenanthrenes; Rats; STAT3 Transcription Factor; Time Factors

Tanshinone IIA is a derivative of phenanthrene-quinone isolated from Danshen, a widely used Chinese herbal medicine. It has antioxidant properties, cytotoxic activities against multiple human cancer cells, inducing apoptosis and differentiation of some human cancer cells. The purpose of this study is to confirm its anticancer activity on human glioma cells, and to elucidate mechanism of its activity. Human glioma cells were tested in vitro for cytotoxicity, colony formation inhibition, BrdU incorporation after treatment with tanshinone IIA. Its effect of apoptosis induction was detected through EB/AO staining, cell cycle analysis and the expressions of ADPRTL1 and CYP1A1 genes, the differentiation induction effect was investigated through morphology, mRNA and protein expressions of GFAP and nestin genes by RT-PCR and immunocytochemistry. Tanshinone IIA demonstrated a dose- and time-dependent inhibitory effect on cell growth, IC(50) was 100 ng/ml, and it significantly inhibited colony formation and BrdU incorporation of human glioma cells. After treatment with 25-100 ng/ml of tanshinone IIA, the apoptotic cells increased significantly (P < 0.01), the cells in G(0)/G(1) phase increased (P < 0.01), and decreased in S phase, ADPRTL1 and CYP1A1 mRNA expression increased 1-2 folds. The cells treated with 100 ng/ml tanshinone IIA demonstrated astrocytes or neuron-like morphology, GFAP mRNA and protein expressions increased, nestin mRNA and protein expressions decreased significantly. The findings in this study suggested that tanshinone IIA exhibited strong effects on growth inhibition and induction of apoptosis and differentiation in human glioma cells. It might serve as a novel promising differentiation-inducing and/or therapeutic agent for human gliomas, and need to be investigated further.

Topics: Abietanes; Analysis of Variance; Antineoplastic Agents, Phytogenic; Apoptosis; Brain Neoplasms; Cell Differentiation; Dose-Response Relationship, Drug; Drugs, Chinese Herbal; Glial Fibrillary Acidic Protein; Glioma; Humans; Intermediate Filament Proteins; Nerve Tissue Proteins; Nestin; Phenanthrenes; RNA, Messenger; Statistics, Nonparametric; Tumor Cells, Cultured

Researches indicated that Tripterygium wilfordii possess antitumor activity. The current study was designed to investigate inhibitive effect of several monomes of Tripterygium wilfordii on the proliferation of glioma cells.. The effect of three monomes from Tripterygium wilfordii on the proliferation of glioma cell lines SHG44, C6, and U251 in vitro was examined by using MTT assay. Immunohistochemistry was used to examine the expression of Bax, Bcl-2 after treatment of triptolide and celastrol.. The proliferation of glioma cells was remarkably inhibited by triptolide and celastrol. They both increased expression of Bax and decreased expression of Bcl-2 in the SHG44 cells.. Triptolide and celastrol inhibit the proliferation of glioma cells in vitro, which was associated with promoting the expression of Bax and inhibiting the expression of Bcl-2 and accelerating cell apotosis.

Topics: Antineoplastic Agents; Cell Division; Diterpenes; Epoxy Compounds; Glioma; Humans; Inhibitory Concentration 50; Molecular Structure; Pentacyclic Triterpenes; Phenanthrenes; Plant Extracts; Tripterygium; Triterpenes; Tumor Cells, Cultured