naphthoquinones and Glioblastoma

Glioblastoma is a type of intracranial malignancy. Shikonin, a Chinese traditional medicine, has been shown to have anti-tumor efficacy toward human glioblastoma cells in vitro. However, shikonin cannot easily cross the blood-brain barrier. To address this issue, we evaluated the anti-tumor effects of direct intracranial infusion of shikonin in in vivo orthotopic syngeneic murine glioblastoma models using C57BL/6 mice.. The cytotoxic effects of shikonin against murine glioblastoma cells, SB28 and CT-2A, were reported resistance to temozolomide, were evaluated using an allophycocyanin-conjugated annexin V and propidium iodide assay with flow cytometry. Impedance-based real-time cell analysis (RTCA) was used to analyze the inhibitory effects of shikonin on growth and proliferation. To evaluate the anti-tumor activity of shikonin in vivo, we used orthotopic syngeneic murine glioblastoma models with SB28 and CT-2A cells.. In flow cytometry-based cytotoxic assays, shikonin induced apoptosis. RTCA indicated that shikonin decreased the cell index of murine glioblastoma cells, SB28 and CT-2A, in a dose-dependent manner (p < 0.0001 for both cell lines), while temozolomide did not (p = 0.91 and 0.82, respectively). In murine glioblastoma models, SB28 and CT-2A, direct intracranial infusion of shikonin, as a local chemotherapy, improved the overall survival of mice in a dose-dependent manner compared with control groups (p < 0.0001 and p = 0.02, respectively). While temozolomide did not (p = 0.48 and 0.52, respectively).. The direct intracranial infusion of shikonin has potential as a local therapy for patients with glioblastoma.

Topics: Animals; Antineoplastic Agents; Brain Neoplasms; Cell Line, Tumor; Glioblastoma; Humans; Mice; Mice, Inbred C57BL; Naphthoquinones; Temozolomide

Glioblastoma multiforme is a malignant neoplasia with a median survival of less than two years and without satisfactory therapeutic options. The so-called glioblastoma stem cells escape the established radio- and chemotherapies and lead to tumor recurrence in most cases. The alkaloid Shikonin with its various anti stem cell properties and the interstitial photodynamic therapy with 5-aminolevulinic acid seem to be promising new options in the therapy of glioblastoma. In this study, in vitro investigations were performed to observe the influence of Shikonin on viability, proliferation, induction of apoptosis and the capability of forming tumor spheres in U-87 MG and the primary glioblastoma cell line GB14. The combined effect with the chemotherapeutic temozolomide and photodynamic treatment on the mRNA expression of glioma specific stem cell markers and further examined intracellular protoporphyrin IX accumulation under Shikonin treatment was analyzed. Shikonin effectively inhibited the capability of forming tumor spheres and enhanced temozolomide effectiveness in the reduction of proliferation and in the induction of apoptosis. Additionally, Shikonin increased the mRNA expression of the tumor suppressing Neurofibromatosis type 1 (NF1) gene and showed modulating effects on intracellular protoporphyrin IX.

Topics: Aminolevulinic Acid; Brain Neoplasms; Cell Line, Tumor; Glioblastoma; Humans; Naphthoquinones; Neoplasm Recurrence, Local; Photochemotherapy; RNA, Messenger; Temozolomide

Glioblastoma (GBM) is the most frequent malignant brain tumor. It represents the most aggressive astrocytoma with an overall survival of 14 months. Despite improvements in surgery techniques, radio‑ and chemotherapy, most patients present treatment resistance, recurrence and disease progression. Therefore, development of effective alternative therapies is essential to overcome treatment failure. The purpose of the study was to evaluate the antitumoral activity of the synthetic compound LQB‑118, in vitro. Monolayer and three‑dimensional (3D) cell culture systems of human‑derived GBM cell lines were used to evaluate the effect of LQB‑118 on cell viability, cell death and migration. LQB‑118 reduced cell viability as determined by MTT and trypan blue exclusion assays and promoted apoptosis in monolayer cell lines with an intrinsic temozolomide (TMZ)‑resistance profile. In 3D culture models, LQB‑118 reduced cell viability as evaluated by APH assay and inhibited cell migration while the TMZ resistance profile was maintained. Moreover, LQB‑118 reduced p38 and AKT expression and phosphorylation, whereas it reduced only the phosphorylated ERK1/2 form. LQB‑118 reduced p38 and NRF2 expression, an axis that is associated with TMZ resistance, revealing a mechanism to overcome resistance. LQB‑118 also demonstrated an additional effect when combined with ionizing radiation and cisplatin. In conclusion, the present data demonstrated that LQB‑118 maintained its effectiveness in a 3D cell conformation, which shares more similarities with the tumor mass. LQB‑118 is a promising agent for GBM treatment as monotherapy and associated with radiotherapy or cisplatin. Its effect is associated with inhibition of GBM‑related survival signaling pathways.

Topics: Brain Neoplasms; Cell Culture Techniques; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cell Survival; Drug Resistance, Neoplasm; Drug Synergism; Glioblastoma; Humans; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Naphthoquinones; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Protein Kinases; Proto-Oncogene Proteins c-akt; Pterocarpans; Temozolomide

Glioblastoma is the most common and malignant brain tumor, and current therapies confer only modest survival benefits. A major obstacle is our ability to monitor treatment effect on tumors. Current imaging modalities are ambiguous, and repeated biopsies are not encouraged. To scout for markers of treatment response, we used NMR spectroscopy to study the effects of a survivin inhibitor on the metabolome of primary glioblastoma cancer stem cells. Applying high resolution NMR spectroscopy (

Topics: Antineoplastic Agents; Biomarkers, Pharmacological; Brain Neoplasms; Cell Survival; Citric Acid; Citric Acid Cycle; Glioblastoma; Humans; Imidazoles; Lactic Acid; Magnetic Resonance Spectroscopy; Metabolome; Molecular Targeted Therapy; Naphthoquinones; Neoplastic Stem Cells; Primary Cell Culture; Principal Component Analysis; Survivin

Effective treatments that extend survival of malignant brain tumor glioblastoma (GBM) have not changed in more than a decade; however, there exists a minority patient group (<5%) whose survival is longer than 3 yr. We herein present a case report of a long-term surviving 51-yr-old female diagnosed with a

Topics: Brain Neoplasms; DNA Mismatch Repair; Drug Screening Assays, Antitumor; Female; Gene Regulatory Networks; Genotype; Germ-Line Mutation; Glioblastoma; Humans; Imidazoles; Middle Aged; Mutation; Naphthoquinones; Neoplasm Recurrence, Local; Phenotype; Whole Genome Sequencing

1,4-naphthoquinones, especially juglone, are known for their anticancer activity. However, plumbagin, lawsone, and menadione have been less investigated for these properties. Therefore, we aimed to determine the effects of plumbagin, lawsone, and menadione on C6 glioblastoma cell viability, ROS production, and mitochondrial function.. Cell viability was assessed spectrophotometrically using metabolic activity method, and by fluorescent Hoechst/propidium iodide nuclear staining. ROS generation was measured fluorometrically using DCFH-DA. Oxygen uptake rates were recorded by the high-resolution respirometer Oxygraph-2k.. Plumbagin and menadione displayed highly cytotoxic activity on C6 cells (IC. Plumbagin and menadione exhibit strong prooxidant, mitochondrial oxidative phosphorylation uncoupling and cytotoxic activity. In contrast, lawsone demonstrates a moderate effect on C6 cell viability and mitochondrial functions, and possesses strong antioxidant properties.

Topics: Animals; Antineoplastic Agents; Antioxidants; Apoptosis; Cell Line, Tumor; Cell Survival; Glioblastoma; Mitochondria; Naphthoquinones; Oxidants; Oxidation-Reduction; Oxidative Stress; Phosphorylation; Phytotherapy; Plant Extracts; Rats; Reactive Oxygen Species; Uncoupling Agents; Vitamin K 3

Glioblastoma (GBM) cells with stem cell-like properties are called glioma stem cells (GSCs). GSCs display highly treatment resistance and are responsible for tumor recurrence. Napabucasin (BBI608), a novel small molecule inhibitor of STAT3, has been identified to eliminate stemness-like tumor cells in some cancers. However, the influence of Napabucasin on GBM cells, especially on GSCs, is currently unclear. In this study, we explored the influence and underlying mechanisms of Napabucasin on GBM cells.. STAT3 expression and its correlation with the glioma grade and patient survival were analyzed using CGGA and TCGA glioma databases. The influence of Napabucasin on proliferation, stemness, the cell cycle, apoptosis, and invasion of human GBM cell lines U87MG and LN229 was tested by CCK8, EdU incorporation, colony formation, Transwell invasion, and three-dimensional spheroid assays as well as flow cytometry, qPCR, and western blot analysis. The ability of Napabucasin to inhibit cell proliferation of U87MG tumor xenografts in mice was assessed using a live animal bioluminescence imaging system and immunohistochemistry.. Napabucasin suppressed the proliferation, colony formation, and invasion of U87MG and LN229 cells. Furthermore, Napabucasin induced cell cycle arrest and apoptosis. More importantly, Napabucasin treatment obviously inhibited expression of stemness-associated genes including STAT3 and suppressed the spheroid formation of glioma cells in vitro. Napabucasin also disrupted the NF-κB signaling pathway via downregulation of RelA (p65). Finally, glioma growth was effectively impaired by Napabucasin in nude mice bearing intracranial glioma xenografts.. Napabucasin treatment may be a novel approach for the treatment of GBM, particularly GSCs.

Topics: Animals; Benzofurans; Cell Proliferation; Female; Glioblastoma; Humans; Mice; Mice, Nude; Naphthoquinones; STAT3 Transcription Factor; Stem Cells

Glioblastoma multiform (GBM) is the most common and devastating type of primary brain tumor, being considered the deadliest of human cancers. In this context, extensive efforts have been undertaken to develop new drugs that exhibit both antiproliferation and antimetastasis effects on GBM. 1,4-Naphthoquinone (1,4-NQ) scaffold has been found in compounds able to inhibit important biological targets associated with cancer, which includes DNA topoisomerase, Hsp90 and monoamine oxidase. Among potential antineoplastic 1,4-NQs is the plant-derived lapachol (2-hydroxy-3-prenyl-1,4-naphthoquinone) that was found to be active against the Walker-256 carcinoma and Yoshida sarcoma. In the present study, we examined the effect of polyamine (PA)-conjugated derivatives of lapachol, nor-lapachol and lawsone on the growth and invasion of the human GBM cells. The conjugation with PA (a spermidine analog) resulted in dose-dependent and time-dependent increase of cytotoxicity of the 1,4-NQs. In addition, in-vitro inhibition of GBM cell invasion by lapachol was increased upon PA conjugation. Previous biochemical experiments indicated that these PA-1,4-NQs are capable of inhibiting DNA human topoisomerase II-α (topo2α), a major enzyme involved in maintaining DNA topology. Herein, we applied molecular docking to investigate the binding of PA-1,4-NQs to the ATPase site of topo2α. The most active molecules preferentially bind at the ATP-binding site of topo2α, which is energetically favored by the conjugation with PA. Taken together, these findings suggested that the PA-1,4-NQ conjugates might represent potential molecules in the development of new drugs in chemotherapy for malignant brain tumors.

Topics: Adenosine Triphosphate; Animals; Antineoplastic Agents; Antineoplastic Agents, Phytogenic; Astrocytes; Binding Sites; Brain Neoplasms; Cell Line, Tumor; Cerebral Cortex; DNA Topoisomerases, Type II; Glioblastoma; Humans; Mice; Naphthoquinones; Polyamines; Primary Cell Culture

Radiotherapy constitutes a standard arm of therapy in the multimodal treatment of patients with glioblastoma (GBM). Ironically, studies have recently revealed that radiation can augment malignant progression, by promoting migration and invasion, which make the disease especially difficult to cure. Here, we investigated the anticancer effects of YM155, a purported radiosensitizer, in GBM cell lines.. GBM cell lines U251 and U87 were treated with YM155 to assess cytotoxicity and activity of the molecule in vitro. Nude mice were implanted with cells to generate orthotopic xenografts for in vivo studies. Response of cells to treatment was examined using cell viability, immunofluorescence, wound healing, and the Transwell invasion assay. Molecules potentially mediating response were examined through western blot analysis, phospho-kinase arrays, and qPCR. Cells were transfected with siRNA knockdown and gene expression constructs to identify molecular mediators of response.. YM155 reduced viability of U251 and U87 cells and enhanced radiosensitivity through inhibition of homologous recombination. Besides, YM155 decreased invasion caused by radiation and led to expression changes in molecular markers associated with EMT. STAT3 was one of 10 molecules identified on a phosphokinase array exhibiting significant change in phosphorylation under YM155 treatment. Transfection with STAT3 siRNAs or expression constructs demonstrated that EMT changes were achieved by inhibiting the phosphorylation of STAT3 and were survivin-independent. Finally, combining YM155 and radiation in orthotopic xenografts reduced growth and prolonged overall survival of animals.. YM155 decreased radiation-induced invasion in GBM cell lines in vitro and in vivo through inhibition of STAT3.

Topics: Animals; Cell Line, Tumor; Cell Proliferation; Cell Survival; Epithelial-Mesenchymal Transition; Glioblastoma; Homologous Recombination; Humans; Imidazoles; Mice, Nude; Naphthoquinones; Neoplasm Invasiveness; STAT3 Transcription Factor; Survivin

Juglone is a natural pigment, which has cytotoxic effect against various human tumor cells. However, its cytotoxicity to glioma cells, especially to tumor stem-like cells (TSCs) has not been demonstrated.. TSCs of glioma were enriched from U87 and two primary cells (SHG62, and SHG66) using serum-free medium supplemented with growth factors, including bFGF, EGF and B27. After treatment of juglone with gradient concentrations (0, 10, 20, and 40 μM), the viability and apoptosis of TSCs were evaluated by WST-8 assay and flow cytometry. Reactive oxygen species (ROS) was labeled by the cell-permeable fluorescent probe and detected with flow cytometry. ROS scavenger (NAC) and p38-MAPK inhibitor (SB203580) were applied to resist the cytotoxic effect. Caspase 9 cleavage and p38 phosphorylation (P-p38) were quantified by western blot. Juglone as well as temozolomide (TMZ) were administrated in intracranial xenografts and MR scan was performed every week to evaluate the anti-tumor effect in vivo.. Juglone could obviously inhibit the proliferation of TSCs in glioma by decreasing cell viability (P < 0.01) and inducing apoptosis (P < 0.01), which was accompanied by increased caspase 9 cleavage in a dose-dependent manner (P < 0.01). In the meantime, juglone could generate ROS significantly and increase p38 phosphorylation (P < 0.01). In addition, pretreatment with ROS scavenger or p38-MAPK inhibitor could reverse juglone-induced cytotoxicity (P < 0.01). More importantly, juglone could also suppress tumor growth in vivo and improve the survival of U87-bearing mice compared with control (P < 0.05), although TMZ seemed to have better effect.. Juglone could inhibit the growth of TSCs in gliomas through the activation of ROS-p38-MAPK pathway in vitro, and the anti-glioma effect was validated in vivo, which offers a potential therapeutic agent to gliomas.

Topics: Animals; Antineoplastic Agents; Apoptosis; Brain Neoplasms; Cell Line, Tumor; Cell Survival; Glioblastoma; Humans; Mice; Naphthoquinones; Neoplastic Stem Cells; p38 Mitogen-Activated Protein Kinases; Reactive Oxygen Species; Xenograft Model Antitumor Assays

Glioblastomas (GBM) comprise 17% of all primary brain tumors. These tumors are extremely aggressive due to their infiltrative capacity and chemoresistance, with glial-to-mesenchymal transition (GMT) proteins playing a prominent role in tumor invasion. One compound that has recently been used to reduce the expression of these proteins is shikonin (SHK), a naphthoquinone with anti-tumor properties. Temozolomide (TMZ), the most commonly used chemotherapeutic agent in GBM treatment, has so far not been studied in combination with SHK. Here, we investigated the combined effects of these two drugs on the proliferation and motility of GBM-derived cells.. The cytotoxic and proliferative effects of SHK and TMZ on human GBM-derived cells were tested using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), Ki67 staining and BrdU incorporation assays. The migration capacities of these cells were evaluated using a scratch wound assay. The expression levels of β3 integrin, metalloproteinases (MMPs) and GMT-associated proteins were determined by Western blotting and immunocytochemistry.. We found that GBM-derived cells treated with a combination of SHK and TMZ showed decreases in their proliferation and migration capacities. These decreases were followed by the suppression of GMT through a reduction of β3 integrin, MMP-2, MMP-9, Slug and vimentin expression via inactivation of PI3K/AKT signaling.. From our results we conclude that dual treatment with SHK and TMZ may constitute a powerful new tool for GBM treatment by reducing therapy resistance and tumor recurrence.

Topics: Antineoplastic Combined Chemotherapy Protocols; Brain Neoplasms; Cell Line, Tumor; Cell Movement; Cell Proliferation; Dacarbazine; Drug Resistance, Neoplasm; Epithelial-Mesenchymal Transition; Glioblastoma; Humans; Naphthoquinones; Temozolomide

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.

Topics: Animals; Antineoplastic Agents; Brain Neoplasms; Cell Growth Processes; Cell Line, Tumor; Cell Movement; Chick Embryo; Chorioallantoic Membrane; Glioblastoma; Human Umbilical Vein Endothelial Cells; Humans; Naphthoquinones; Neovascularization, Pathologic; NIMA-Interacting Peptidylprolyl Isomerase; Signal Transduction

Glioma has become a significant global health problem with substantial morbidity and mortality, underscoring the importance of elucidating its underlying molecular mechanisms. Recent studies have identified mir-218 as an anti-oncogene; however, the specific functions of mir-218-1 and mir-218-2 remain unknown, especially the latter. The objective of this study was to further investigate the role of mir-218-2 in glioma. Our results indicated that mir-218-2 is highly overexpressed in glioma. Furthermore, we showed that mir-218-2 is positively correlated with the growth, invasion, migration, and drug susceptibility (to β-lapachone) of glioma cells. In vitro, the overexpression of mir-218-2 promoted glioma cell proliferation, invasion, and migration. In addition, the overexpression of mir-218-2 in vivo was found to increase glioma tumor growth. Accordingly, the inhibition of mir-218-2 resulted in the opposite effects. Cell division cycle 27 (CDC27), the downstream target of mir-218-2, is involved in the regulation of glioma cells. Our results indicate that the overexpression of CDC27 counteracted the function of mir-218-2 in glioma cells. These novel findings provide new insight in the application of mir-218-2 as a potential glioma treatment.

Topics: Animals; Antineoplastic Agents; Apc3 Subunit, Anaphase-Promoting Complex-Cyclosome; Brain Neoplasms; Cell Line, Tumor; Cell Movement; Cell Proliferation; Drug Resistance, Neoplasm; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Glioblastoma; Humans; Male; Mice, Inbred BALB C; Mice, Nude; MicroRNAs; Naphthoquinones; Neoplasm Invasiveness; Signal Transduction; Time Factors; Transfection; Tumor Burden; Xenograft Model Antitumor Assays

In this study we evaluated the impact of juglone on rat glioma C6 cell culture viability, proliferation and invasiveness in vitro. Juglone induced C6 cell death with EC50 concentrations equal to 10.4 ± 1.6 µM after 24h incubation. At relatively low concentrations juglone significantly decreased cell proliferation, reduced spheroid invasiveness and suppressed "wound" healing. In addition, generation of intracellular reactive oxygen and nitrogen species (RS) was detected in cells treated with juglone. Noteworthy, juglone was relatively stable in cell culture medium and levels of H2O2 generated from juglone due to its probable reaction with medium components were not sufficient to affect the viability of glioma cells. Moreover, addition of catalase to the cell medium did not reduce the cytotoxicity of juglone. Therefore, we propose that cell death may be induced through the action of RS other than H2O2, However the direct effect of juglone on the cellular targets could not be excluded either. In conclusion, juglone exerted cytotoxic, anti-proliferative and anti-invasive effects on C6 rat glioma cells in vitro.

Topics: Animals; Antineoplastic Agents; Cell Death; Cell Line, Tumor; Cell Proliferation; Cell Survival; Glioblastoma; Humans; Hydrogen Peroxide; Naphthoquinones; Neoplasm Invasiveness; Rats; Reactive Oxygen Species

Overexpression and mutation of the epidermal growth factor receptor (EGFR) gene play a causal role in tumorigenesis and resistance to treatment of glioblastoma (GBM). EGFR inhibitors such as erlotinib are currently used for the treatment of GBM; however, their efficacy has been limited due to drug resistance. New treatment strategies are therefore urgently needed. Shikonin, a natural naphthoquinone, induces both apoptosis and necroptosis in human glioma cells, but the effectiveness of erlotinib-shikonin combination treatment as well as the underlying molecular mechanisms is unknown yet. In this study, we investigated erlotinib in combination with shikonin and 14 shikonin derivatives in parental U87MG and transfected U87MG.ΔEGFR GBM cells. Most of the shikonin derivatives revealed strong cytotoxicity. Shikonin together with five other derivatives, namely deoxyshikonin, isobutyrylshikonin, acetylshikonin, β,β-dimethylacrylshikonin and acetylalkannin showed synergistic cytotoxicity toward U87MG.ΔEGFR in combination with erlotinib. Moreover, the combined cytotoxic effect of shikonin and erlotinib was further confirmed with another three EGFR-expressing cell lines, BS153, A431 and DK-MG. Shikonin not only dose-dependently inhibited EGFR phosphorylation and decreased phosphorylation of EGFR downstream molecules, including AKT, P44/42MAPK and PLCγ1, but also together with erlotinib synergistically inhibited ΔEGFR phosphorylation in U87MG.ΔEGFR cells as determined by Loewe additivity and Bliss independence drug interaction models. These results suggest that the combination of erlotinib with shikonin or its derivatives might be a potential strategy to overcome drug resistance to erlotinib.

Topics: Anthraquinones; Antineoplastic Combined Chemotherapy Protocols; Cell Line, Tumor; Drug Synergism; ErbB Receptors; Erlotinib Hydrochloride; Glioblastoma; Humans; Mitogen-Activated Protein Kinases; Naphthoquinones; Phosphorylation; Proto-Oncogene Proteins c-akt; Quinazolines; Signal Transduction

2-Deoxy-d-glucose (2-DG), a synthetic glycolytic inhibitor, is currently under clinical evaluation as a promising anticancer agent. However, 2-DG treatment in cancer cells activates prosurvival Akt signaling that might limit its clinical efficacy. The NADPH oxidase 4 (Nox-4)/reactive oxygen species/Akt signaling is known to regulate survival, proliferation, infiltration, and invasion in glioblastomas (GBMs). The enhanced motility, invasiveness, and therapy resistance in GBMs are attributed to metabolic adaptation through increased aerobic glycolysis. Therefore, we hypothesized that inhibition of the Nox-4 might enhance 2-DG-induced suppression of glycolysis, migration, and invasion in GBM cells.. We identified the natural naphthoquinone compound shikonin as a potent inhibitor of the Nox-4/Akt signaling pathway. The combined treatment of shikonin+2-DG suppressed the glycolytic phenotype, migration, and invasion through modulation of the Akt/HIF1α/hexokinase-2 signaling axis in GBM cells. The combination also exhibited enhanced antiproliferative and antiangiogenic effects in vivo.. Our data for the first time demonstrate that inhibition of the Nox-4-associated prosurvival signaling pathway by shikonin enhances the antiproliferative and antiangiogenic potential of 2-DG in GBM cells.. In summary, the combined inhibition of Nox-4 and glycolysis may have therapeutic implications for the management of GBMs.

Topics: Antineoplastic Combined Chemotherapy Protocols; Cell Line, Tumor; Cell Movement; Deoxyglucose; Enzyme Inhibitors; Glioblastoma; Glycolysis; Humans; NADPH Oxidases; Naphthoquinones; Neoplasm Invasiveness; Proto-Oncogene Proteins c-akt; Signal Transduction

Though previous study demonstrated that shikonin could exert its antitumor activity by inducing apoptosis and necrosis, the pro-survival mechanisms involved in its antitumor process are still little to know. In the present study, for the first time, we found a protective mechanism was simultaneously activated which caused the reduced sensitivity of glioblastoma stem cells (GSCs) to the cytotoxicity of shikonin. Reduced active caspase-9 expression and enhanced mitochondrial membrane potential (MMP) were intriguingly observed within 24 h treatment by shikonin in GSCs. Further investigation identified that Endoplasmic Reticulum Stress (ERS) was involved in its antitumor process, which compromised the cytotoxicity of shikonin toward GSCs. Inhibiting ERS by 4-phenylbutyric acid (4-PBA) markedly enhanced the cytotoxicity of shikonin in GSCs. The consistent result was simultaneously observed in the GSCs-xenografted mice. Furthermore, our results identified that JNK/c-Jun pathway was involved in the antitumor process of shikonin, providing a mechanism by which ERS reduced the cytotoxicity of shikonin toward GSCs. Altogether, the novel observation in the present study identified that inhibiting ERS would be an attractive new approach to enhance the therapeutic potency of shikonin toward GSCs.

Topics: Animals; Antineoplastic Agents, Phytogenic; Cells, Cultured; Drugs, Chinese Herbal; Endoplasmic Reticulum Stress; Glioblastoma; Humans; JNK Mitogen-Activated Protein Kinases; Male; MAP Kinase Signaling System; Mice, Nude; Naphthoquinones; Neoplastic Stem Cells

Shikonin is an anthraquinone derivative extracted from the root of lithospermum. Shikonin is traditionally used in the treatment of inflammatory and infectious diseases such as hepatitis. Shikonin also inhibits proliferation and induces apoptosis in various tumors. However, the effect of shikonin on gliomas has not been fully elucidated. In the present study, we aimed to investigate the effects of shikonin on the migration and invasion of human glioblastoma cells as well as the underlying mechanisms. U87 and U251 human glioblastoma cells were treated with shikonin at 2.5, 5, and 7.5 μmol/L and cell viability, migration and invasiveness were assessed with CCK8, scratch wound healing, in vitro Transwell migration, and invasion assays. The expression and activity of matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) and the expression of phosphorylated β-catenin (p-β-catenin) and phosphorylated PI3K/Akt were also checked. Results showed that shikonin significantly inhibited the cell proliferation, migration, invasion, and expression of MMP-2 and MMP-9 in U87 and U251 cells. The expression of p-β-catenin showed contrary trends in two cell lines. It was significantly inhibited in U87 cells and promoted in U251 cells. Results in this work indicated that shikonin displayed an inhibitory effect on the migration and invasion of glioma cells by inhibiting the expression and activity of MMP-2 and -9. In addition, shikonin also inhibited the expression of p-PI3K and p-Akt to attenuate cell migration and invasion and MMP-2 and MMP-9 expression in both cell lines, which could be reversed by the PI3K/Akt pathway agonist, insulin-like growth factor-1 (IGF-1).

Topics: beta Catenin; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cell Survival; Drugs, Chinese Herbal; Glioblastoma; Humans; Insulin-Like Growth Factor I; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Medicine, Chinese Traditional; Naphthoquinones; Neoplasm Invasiveness; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Phosphorylation; Proto-Oncogene Proteins c-akt; RNA Interference; RNA, Small Interfering; Signal Transduction

Natural plant products may possess much potential in palliative therapy and supportive strategies of current cancer treatments with lesser cytotoxicity to normal cells compared to conventional chemotherapy. In the current study, anti-cancer properties of plumbagin, a plant-derived naphthoquinone, on brain cancer cells were determined. Plumbagin treatment resulted in the induction of DNA damage, cell cycle arrest and apoptosis, followed by suppression of the colony forming ability of the brain tumour cells. These effects were substantiated by upregulation of PTEN, TNFRSF1A and downregulation of E2F1 genes, along with a drop in MDM2, cyclin B1, survivin and BCL2 protein expression. Plumbagin induced elevated levels of caspase-3/7 activity as well. For the first time, we show here that plumbagin inhibits telomerase in brain tumour cells and results in telomere shortening following chronic long-term treatment. This observation implies considerable cytotoxicity of plumbagin towards cancer cells with higher telomerase activity. Collectively, our findings suggest plumbagin as a potential chemotherapeutic phytochemical in brain tumour treatment modalities.

Topics: Antineoplastic Agents, Phytogenic; Apoptosis; Brain Neoplasms; Cell Cycle; Cell Line, Tumor; Cell Proliferation; DNA Damage; Gene Expression Regulation, Neoplastic; Glioblastoma; Humans; Naphthoquinones; Telomerase; Telomere; Telomere Shortening

Pediatric glioblastoma is a malignant disease with an extremely poor clinical outcome. Patients usually suffer from resistance to radiation therapy, so targeted drug treatment may be a new possibility for glioblastoma therapy. Survivin is also overexpressed in glioblastoma. YM155, a novel small-molecule survivin inhibitor, has not been examined for its use in glioblastoma therapy.. The human glioblastoma cell line M059K, which expresses normal DNA-dependent protein kinase (DNA-PK) activity and is radiation-resistant, and M059J, which is deficient in DNA-PK activity and radiation-sensitive, were used in the study. Cell viability, DNA fragmentation, and the expression of survivin and securin following YM155 treatment were examined using MTT (methylthiazolyldiphenyl-tetrazolium) assay, ELISA assay, and Western blot analysis, respectively.. YM155 caused a concentration-dependent cytotoxic effect, inhibiting the cell viability of both M059K and M059J cells by 70% after 48 hours of treatment with 50 nM YM155. The half-maximal inhibitory concentration (IC50) was around 30-35 nM for both cell lines. Apoptosis was determined to have occurred in both cell lines because immunoreactive signals from the DNA fragments in the cytoplasm were increased 24 hours after treatment with 30 nM YM155. The expression of survivin and securin in the M059K cells was greater than that measured in the M059J cells. Treatment with 30 nM YM155, for both 24 and 48 hours, significantly suppressed the expression of survivin and securin in both cell lines.. The novel survivin inhibitor YM155 elicits potent cytotoxicity in glioblastoma cells in vitro via DNA-PK-independent mechanisms. YM155 could be used as a new therapeutic agent for the treatment of human glioblastomas.

Topics: Apoptosis; Brain Neoplasms; Cell Line, Tumor; Cell Survival; DNA-Activated Protein Kinase; Glioblastoma; Humans; Imidazoles; Inhibitor of Apoptosis Proteins; Naphthoquinones; Neoplasm Proteins; Securin; Survivin

KILLER/DR5 is a death-domain-containing proapoptotic receptor that binds to the cytotoxic ligand TRAIL. It was originally reported that induction of KILLER/DR5 mRNA following DNA damage was p53-dependent, but some drugs that induce apoptosis can upregulate KILLER/DR5 mRNA expression in cell lines with mutated p53. We further extend those findings by classifying the capability of various apoptosis-inducing drugs to increase the expression of KILLER/DR5 mRNA in a p53-independent manner. beta-Lapachone, a topoisomerase inhibitor, increased KILLER/DR5 mRNA in colon cancer cell lines with wild-type p53 but not with mutant p53. In contrast, betulinic acid, a novel chemotherapeutic compound, induced apoptosis and KILLER/DR5 mRNA in melanoma and glioblastoma cells through a p53-independent mechanism. The synthetic glucocorticoid dexamethasone elevated KILLER/DR5 mRNA in glioblastoma, ovarian cancer, and colon cancer cell lines with mutant p53 undergoing apoptosis, and this induction was inhibited by the transcriptional inhibitor actinomycin D. Although another glucocorticoid, prednisolone, also induced apoptosis, it did not increase KILLER/DR5 mRNA. Finally, the cytokine interferon-gamma (IFN-gamma) induced apoptosis and KILLER/DR5 in cell lines with mutant p53, and the induction of KILLER/DR5 mRNA by IFN-gamma was delayed in cells lacking wild-type STAT1, a transcription factor implicated in IFN-gamma signaling. Similarly, the induction of KILLER/DR5 mRNA by the cytokine TNF-alpha was also delayed in cell lines with mutated STAT1. These findings suggest that KILLER/DR5 may play a role in p53-independent apoptosis induced by specific drugs and warrants further investigation as a novel target for chemotherapy of tumors lacking wild-type p53.

Topics: Antibiotics, Antineoplastic; Apoptosis; Betulinic Acid; Colonic Neoplasms; Dactinomycin; Dexamethasone; DNA-Binding Proteins; Female; Glioblastoma; Glucocorticoids; Humans; Interferon-gamma; Melanoma; Mutation; Naphthoquinones; Ovarian Neoplasms; Pentacyclic Triterpenes; Prednisolone; Receptors, TNF-Related Apoptosis-Inducing Ligand; Receptors, Tumor Necrosis Factor; RNA, Messenger; STAT1 Transcription Factor; Trans-Activators; Triterpenes; Tumor Cells, Cultured; Tumor Suppressor Protein p53; Up-Regulation