stilbenes and Fibrosarcoma

Several components isolated from rhubarb, the root of Rheum undulatum L., including emodin, rhein, rhaponticin, and piceatannol, have been reported to induce cell death and inhibit metastasis in various types of cancer. Recently, piceatannol-3-O-β-D-glucopyranoside (PG) isolated from rhubarb was demonstrated to improve vascular dysfunction by inhibiting arginase activity.. In this study, we examined the anti-cancer activities of PG, including effects on the proliferation, metastasis, and angiogenesis of endothelial and malignant cancer cells.. We found that PG did not affect the proliferation of human fibrosarcoma (HT1080) and human umbilical vein endothelial cells (HUVECs) at treatments up to 100  μM. However, PG efficiently suppressed the metastatic ability of HT1080 cells, as determined by scratch wound migration, transwell migration/invasion assay, and three-dimensional (3D) spheroid invasion assay. PG significantly suppressed the phorbol 12-myristate 13-acetate (PMA)-induced increase of matrix metalloproteinase (MMP)-9 expression as well as gelatinolytic MMP-9 activity, which are essential for cancer metastasis. In addition, PG treatment reduced the production of proangiogenic factors in HT1080 cells under normoxic and hypoxic conditions and suppressed hypoxia-induced activation of the hypoxia-inducible factor (HIF)-1α pathway. We also found that HUVEC angiogenic activity, including migration and tubular structure formation, were significantly reduced by PG treatment. Moreover, in an in ovo chick chorioallantoic membrane assay, spontaneous and vascular endothelial growth factor (VEGF)-induced vessel formation were significantly inhibited by PG treatment.. These results collectively indicate that PG has potent anti-metastatic and anti-angiogenic activities with no cytotoxicity. Thus, PG may be useful to limit the hyperplasia of malignant tumors and the spread of cancer to distant secondary organs.

Topics: Adult; Angiogenesis Inhibitors; Animals; Cell Line, Tumor; Cell Movement; Chick Embryo; Chorioallantoic Membrane; Fibrosarcoma; Glucosides; Human Umbilical Vein Endothelial Cells; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Matrix Metalloproteinase 9; Neovascularization, Pathologic; Stilbenes; Tetradecanoylphorbol Acetate; Vascular Endothelial Growth Factor A

Metastatic fibrosarcomas still represent a therapeutic dilemma. Commonly used chemotherapeutic agents such as doxorubicin have been proven effective in fewer than 30% of all cases disseminated of fibrosarcoma. Elderly patients with cardiac disease are not suitable for systemic chemotherapy with doxorubicin. We therefore tested the apoptotic effects of the natural and well-tolerated compound resveratrol on human fibrosarcoma cells (HT1080).. Vital, apoptotic and necrotic cells were quantified using flow cytometric analysis. Gene expression was analyzed by RNA microarrays.. Application of resveratrol induced apoptotic cell death and significantly reduced proliferation of HT1080 cells. Correspondingly, expression of apoptosis-associated genes was altered in microarray analysis.. This in vitro study demonstrates the anticancer activity of resveratrol against human fibrosarcoma cells. These results provide experimental support for in vivo trials assessing the effect of the natural polyphenol resveratrol.

Topics: Apoptosis; Cell Line, Tumor; Fibrosarcoma; Flow Cytometry; Gene Expression; Humans; Oligonucleotide Array Sequence Analysis; Resveratrol; Stilbenes

Environmental conditions or chemical agents can interfere with the function of the endoplasmic reticulum, and the resulting endoplasmic reticulum (ER) stress can be toxic to the cell if it is not relieved. The classical compensatory response to ER stress is the unfolded protein response (UPR) that reduces protein load in the ER. However, autophagy may also compensate by removing large insoluble protein aggregates. Agents that stress the ER can have anti-cancer activity, and novel applications of ER stress inducing agents are being investigated. Plant stilbenes are a class of stress responsive molecules that includes resveratrol, which are being investigated as potential therapeutics in humans for conditions such as aging or cancer.. We performed a screen of 1726 small, drug like molecules to identify those that could activate an ER-stress responsive luciferase gene. After secondary screening, we determined that the plant stilbenes pterostilbene and piceatannol were the most potent inducers of ER stress from this group. ER stress can be particularly toxic to cells with high ER load, so we examined their effect on cells expressing the Wnt family of secreted glycoprotein growth factors. Molecular analysis determined that these ER stress-inducing stilbenes could block Wnt processing and also induce autophagy in acute lymphoblastic leukemia cells expressing Wnt16. Combining pterostilbene (to induce ER stress) with chloroquine (to inhibit autophagy) lead to significant cellular toxicity in cells from aggressive acute lymphoblastic leukemia.. Plant stilbenes are potent inducers of ER stress. However, their toxicity is more pronounced in cancer cells expressing Wnt growth factors. The toxicity of stilbenes in these ALL cells can be potentiated by the addition of autophagy inhibitors, suggesting a possible therapeutic application.

Topics: Antimalarials; Antineoplastic Agents; Antioxidants; Autophagy; Blotting, Western; Cell Proliferation; Chloroquine; Drug Synergism; Drug Therapy, Combination; Endoplasmic Reticulum Stress; Fibrosarcoma; High-Throughput Screening Assays; Humans; Plants; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Resveratrol; Small Molecule Libraries; Stilbenes; Tumor Cells, Cultured

Tumour vasculature is notoriously sinusoidal and leaky, and is hence susceptible to vascular disruption. Microtubule destabilising drugs such as the combretastatins form the largest group of tumour vascular disrupting agents and cause selective shutdown of tumour blood flow within minutes to hours, leading to secondary tumour cell death. Targeting the tumour vasculature is a proven anticancer strategy but early treatment response biomarkers are required for personalising treatment planning. Protein induction following treatment with combretastatin A4-phosphate was examined in a mouse fibrosarcoma model (fs188), where tumour cells express only the matrix-bound isoform of vascular endothelial growth factor A (VEGF188). These tumours are relatively resistant to vascular disruption by combretastatin A4-phosphate and hence a study of protein induction following treatment could yield insights into resistance mechanisms. The distribution of a number of proteins induced following treatment were visualised by MALDI-mass spectrometry imaging. Responses identified were validated by LC-ESI-MS/MS and immunohistochemical staining. Significant changes in proteins connected with necrosis, cell structure, cell survival and stress-induced molecular chaperones were identified. Protein-protein interactions were identified using STRING 9.0 proteomic network software. These relationship pathways provided an insight into the activity of the active tumour milieu and a means of linking the identified proteins to their functional partners.

Topics: Animals; Fibrosarcoma; Gene Expression Regulation, Neoplastic; Humans; Mice; Neovascularization, Pathologic; Protein Interaction Maps; Proteomics; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Stilbenes; Vascular Endothelial Growth Factor A

Based on the soil-to-seeds principle, we explored the small-molecular sequential dual-targeting theranostic strategy (SMSDTTS) for prolonged survival and imaging detectability in a xenograft tumor model.. Thirty severe combined immunodeficiency (SCID) mice bearing bilateral radiation-induced fibrosarcoma-1 (RIF-1) subcutaneously were divided into group A of SMSDTTS with sequential intravenous injections of combretastatin A4 phosphate (CA4P) and (131)I-iodohypericin ((131)I-Hyp) at a 24 h interval; group B of single targeting control with CA4P and vehicle of (131)I-Hyp; and group C of vehicle control (10 mice per group). Tumoricidal events were monitored by in vivo magnetic resonance imaging (MRI) and planar gamma scintiscan, and validated by ex vivo autoradiography and histopathology. Besides, 9 mice received sequential intravenous injections of CA4P and (131)I-Hyp were subjected to biodistribution analysis at 24, 72 and 120 h.. Gamma counting revealed fast clearance of (131)I-Hyp from normal organs but intense accumulation in necrotic tumor over 120 h. After only one treatment, significantly prolonged survival (p<0.001) was found in group A compared to group B and C with median survival of 33, 22, and 21 days respectively. Tumor volume on day 15 was 2.0 ± 0.89, 5.66 ± 1.66, and 5.02 ± 1.0 cm(3) with tumor doubling time 7.8 ± 2.8, 4.4 ± 0.67, and 4.5 ± 0.5 days respectively. SMSDTTS treated tumors were visualized as hot spots on gamma scintiscans, and necrosis over tumor ratio remained consistently high on MRI, autoradiography and histology.. The synergistic antitumor effects, multifocal targetability, simultaneous theranostic property, and good tolerance of the SMSDTTS were evident in this experiment, which warrants further development for preclinical and clinical applications.

Topics: Administration, Intravenous; Animals; Anthracenes; Antineoplastic Agents; Disease Models, Animal; Fibrosarcoma; Histocytochemistry; Humans; Iodine Radioisotopes; Magnetic Resonance Imaging; Male; Mice; Mice, SCID; Perylene; Radiography; Radionuclide Imaging; Stilbenes; Survival Analysis; Transplantation, Heterologous; Treatment Outcome

Trans-3,4',5-trihydroxystilbene (resveratrol) is a grape polyphenol present in various plants, food products, red wine and grapes. Resveratrol has anti-inflammatory, anticarcinogenic, anti-oxidant and anti-aging properties. Matrix metalloproteinases (MMPs) are key enzymes involved in the degradation of the extracellular matrix, and their expression may be regulated in cancer metastasis. In the present study, we aimed to evaluate the effect of resveratrol on MMPs and cell migration, and to understand the mechanism of action in HT1080 human fibrosarcoma cells. We found that resveratrol inhibited HT1080 cell viability at various concentrations as detected by the MTT assay and FACS analysis. However, resveratrol dramatically increased the activation and expression of MMP-9 in a dose- and time-dependent manner, as determined by gelatin zymography assay and western blot analysis. We also discovered that resveratrol enhanced the migratory ability of HT1080 cells, as determined by the wound healing assay, and decreased the phosphorylation of p38 kinase. Moreover, the Akt kinase was inhibited by resveratrol in the HT1080 cells. The inhibition of p38 and Akt kinases with SB203580 and LY294002 further increased resveratrol-induced MMP-9 as well as cell migration in the HT1080 cells. Our results suggest that resveratrol regulates MMP-9 and migratory abilities through the p38 kinase and PI-3K pathways in HT1080 human fibrosarcoma cells.

Topics: Antineoplastic Agents, Phytogenic; Cell Line, Tumor; Cell Movement; Cell Survival; Dose-Response Relationship, Drug; Fibrosarcoma; Humans; MAP Kinase Signaling System; Matrix Metalloproteinase 9; p38 Mitogen-Activated Protein Kinases; Phosphatidylinositol 3-Kinases; Phosphorylation; Proto-Oncogene Proteins c-akt; Resveratrol; Stilbenes

Characterising the protein signatures in tumours following vascular-targeted therapy will help determine both treatment response and resistance mechanisms. Here, mass spectrometry imaging and MS/MS with and without ion mobility separation have been used for this purpose in a mouse fibrosarcoma model following treatment with the tubulin-binding tumour vascular disrupting agent, combretastatin A-4-phosphate (CA-4-P). Characterisation of peptides after in situ tissue tryptic digestion was carried out using Matrix-Assisted Laser Desorption/Ionisation-Mass Spectrometry (MALDI-MS) and Matrix-Assisted Laser Desorption/Ionisation-Ion Mobility Separation-Mass Spectrometry Imaging (MALDI IMS-MSI) to observe the spatial distribution of peptides. Matrix-Assisted Laser Desorption/Ionisation-Ion Mobility Separation-Tandem Mass Spectrometry (MALDI-IMS-MS/MS) of peaks was performed to elucidate any pharmacological responses and potential biomarkers. By taking tumour samples at a number of time points after treatment gross changes in the tissue were indicated by changes in the signal levels of certain peptides. These were identified as arising from haemoglobin and indicated the disruption of the tumour vasculature. It was hoped that the use of PCA-DA would reveal more subtle changes taking place in the tumour samples however these are masked by the dominance of the changes in the haemoglobin signals.

Topics: Animals; Antineoplastic Agents, Phytogenic; Biomarkers, Tumor; Fibrosarcoma; Mice; Peptide Mapping; Proteins; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Stilbenes

Resveratrol, a grape polyphenol, is thought to have anti-inflammatory, cardioprotective, and cancer preventive properties. However, the mechanisms by which resveratrol might produce these effects are not clearly defined. A study was performed on whether resveratrol could prevent tumor cells from adhering to endothelial cells, which is an essential step during tumor metastasis. Phorbol 12-myristate 13-acetate (PMA) induced human fibrosarcoma HT1080 cells to adhere to endothelial ECV304 cells. Resveratrol inhibited PMA-induced HT1080 cells adhesion in a dose-dependent manner. To further study the mechanisms of this resveratrol-mediated blockade of tumor cell adhesion, the expression of the cell adhesion molecules intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and E-selectin were examined. PMA induced ICAM-1 expression in HT1080 cells. In contrast, the expression of VCAM-1 and E-selectin were not altered by PMA treatment. The increase in tumor cell adhesion to endothelial cells following PMA treatment was partially inhibited by ICAM-1 siRNA or neutralizing antibodies. Resveratrol reduced the PMA-induced ICAM-1 expression in HT1080 cells as determined by RT-PCR, flow cytometry and ELISA. As the induction of ICAM-1 requires activation of the transcription factor NF-kappaB, the effects of resveratrol on the activation of this factor in HT1080 cells was also investigated. Resveratrol inhibited the PMA-induced NF-kappaB activation and NF-kappaB-dependent luciferase activity. These results suggest that resveratrol may exert an antimetastatic effect by inhibiting NF-kappaB activation and ICAM-1 expression, leading to suppression of tumor cell adhesion to endothelial cells.

Topics: Cell Adhesion; Dose-Response Relationship, Drug; Endothelial Cells; Fibrosarcoma; Humans; Intercellular Adhesion Molecule-1; NF-kappa B; Resveratrol; RNA, Small Interfering; Stilbenes; Tetradecanoylphorbol Acetate; Transfection

Tubulin-binding vascular-disrupting agents (VDA) are currently in clinical trials for cancer therapy but the factors that influence tumor susceptibility to these agents are poorly understood. We evaluated the consequences of modifying tumor vascular morphology and function on vascular and therapeutic response to combretastatin-A4 3-O-phosphate (CA-4-P), which was chosen as a model VDA. Mouse fibrosarcoma cell lines that are capable of expressing all vascular endothelial growth factor (VEGF) isoforms (control) or only single isoforms of VEGF (VEGF120, VEGF164, or VEGF188) were developed under endogenous VEGF promoter control. Once tumors were established, VEGF isoform expression did not affect growth or blood flow rate. However, VEGF188 was uniquely associated with tumor vascular maturity, resistance to hemorrhage, and resistance to CA-4-P. Pericyte staining was much greater in VEGF188 and control tumors than in VEGF120 and VEGF164 tumors. Vascular volume was highest in VEGF120 and control tumors (CD31 staining) but total vascular length was highest in VEGF188 tumors, reflecting very narrow vessels forming complex vascular networks. I.v. administered 40 kDa FITC-dextran leaked slowly from the vasculature of VEGF188 tumors compared with VEGF120 tumors. Intravital microscopy measurements of vascular length and RBC velocity showed that CA-4-P produced significantly more vascular damage in VEGF120 and VEGF164 tumors than in VEGF188 and control tumors. Importantly, this translated into a similar differential in therapeutic response, as determined by tumor growth delay. Results imply differences in signaling pathways between VEGF isoforms and suggest that VEGF isoforms might be useful in vascular-disrupting cancer therapy to predict tumor susceptibility to VDAs.

Topics: Angiogenesis Inhibitors; Animals; Cell Line, Tumor; Enzyme-Linked Immunosorbent Assay; Female; Fibrosarcoma; Immunohistochemistry; Mice; Mice, SCID; Neovascularization, Pathologic; Protein Isoforms; RNA, Messenger; Stilbenes; Vascular Endothelial Growth Factor A

We carried out this investigation to examine the effects on angiogenesis-mediated processes and to define anti-angiogenesis mechanisms for flavonoids. We examined the effects and mechanisms of the flavonoid resveratrol on angiogenesis and tumor growth using the chick chorioallantoic membrane (CAM) model of angiogenesis, the CAM tumor growth model, and the effect on p53 in fibroblast growth factor-2 (FGF2) stimulated human endothelial cells using immunoassay. Resveratrol demonstrated potent inhibition (effective dose50=0.7+/-0.1 microM) of FGF2-induced angiogenesis and tumor growth. Furthermore, resveratrol significantly (P<0.01) inhibited platelet/fibrin clot-promoted human colon and fibrosarcoma tumor growth in the CAM tumor model. Resveratrol in a concentration-dependent (1-3 microM) manner significantly promoted apoptosis in FGF2-stimulated endothelial cells by increasing p53 protein production. These data indicated potent anti-angiogenesis efficacy, inhibition of tumor growth, and clot-mediated enhanced tumor growth. These data suggest potential anticancer benefits as a chemopreventive and chemotherapeutic for the flavonoid resveratrol.

Topics: Angiogenesis Inhibitors; Animals; Apoptosis; Cell Line; Chick Embryo; Chorioallantoic Membrane; Colonic Neoplasms; Endothelium, Vascular; Fibroblast Growth Factor 2; Fibrosarcoma; Flavonoids; Humans; Neovascularization, Pathologic; Platelet Aggregation Inhibitors; Resveratrol; Stilbenes; Tumor Suppressor Protein p53

Topics: Allantois; Angiogenesis Inhibitors; Animals; Cell Division; Chick Embryo; Chorion; Cornea; Endothelial Growth Factors; Endothelium, Vascular; Enzyme Activation; Fibroblast Growth Factor 2; Fibrosarcoma; Lymphokines; Mice; Mitogen-Activated Protein Kinases; Neoplasms; Neovascularization, Pathologic; Neovascularization, Physiologic; Resveratrol; Rosales; Stilbenes; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors; Wine; Wound Healing

5,6-Dimethylxanthenone-4-acetic acid (DMXAA) and combretastatin A4 phosphate (CA-4-P) markedly inhibit tumor blood flow in mice and are both currently in clinical trial. One of the challenges in clinical evaluation of antivascular agents is the monitoring of tumor blood flow inhibition in individual patients. This study investigates, using mouse models, whether a new marker for tissue hypoxia, (99m)technetium-labeled 2,2'-(1,4-diaminobutane)bis(2-methyl-3-butanone) dioxime (99mTc-labeled HL-91; Prognox)] has potential for the scintigraphic monitoring of tumor response to antivascular agents. Determination of radioactivity in dissected tissues 3 h after DMXAA (80 micromol/kg) or CA-4-P (227 micromol/kg) was injected indicated that both drugs inhibited blood flow (86RbCl uptake; 84 and 87%, respectively) and increased 99mTc-labeled HL-91 levels (350 and 300%, respectively) selectively in murine RIF-1 tumors. Planar imaging of 99mTc-labeled HL-91 3 h after DMXAA injection showed a dose-dependent increase in tumor levels above a threshold of 50 micromol/kg; this same threshold was observed for the inhibition of tumor blood flow (determined using Hoechst 33342). DMXAA also inhibited blood flow--and increased 99mTc-labeled HL-91 uptake--in MDAH-MCa-4 mouse mammary carcinomas and in NZMN10 human melanoma xenografts. Whether 99mTc-labeled HL-91 might also be useful as a biomarker for tumor cell killing was investigated by clonogenic assay of surviving cells 15 h after imaging 99mTc-labeled HL-91 in RIF-1 tumors. Log cell kill in individual tumors showed a statistically significant linear correlation (P < 0.001) with 99mTc-labeled HL-91 uptake after 60 micromol/kg (r2 = 0.79) and 70 micromol/kg (r2 = 0.44) but not at 80 micromol/kg DMXAA. The lack of correlation at high doses presumably reflects the insensitivity of the tumor-averaged 99mTc-labeled HL-91 signal to small regions in which tumor blood flow is preserved (which will limit log cell kill). The results indicate the potential of 99mTc-labeled HL-91 for the noninvasive imaging of tumor blood flow inhibition by antivascular drugs in humans.

Topics: Angiogenesis Inhibitors; Animals; Antineoplastic Agents; Biomarkers, Tumor; Cell Hypoxia; Fibrosarcoma; Humans; Mammary Neoplasms, Experimental; Melanoma; Mice; Mice, Inbred C3H; Neoplasm Transplantation; Neoplasms, Experimental; Organotechnetium Compounds; Oximes; Radionuclide Imaging; Radiopharmaceuticals; Stilbenes; Xanthenes; Xanthones

The mechanisms of estrogen-induced cancer are still a matter of debate. Previous studies with stilbene estrogens and steroidal estrogens have shown that the in vitro transformation of primary Syrian hamster embryo (SHE) fibroblasts is a good experimental system for discriminating hormonal from nonhormonal events. We have now extended these studies to the triphenylethylene-type antiestrogens tamoxifen (TAM), 3-hydroxy-TAM, 4-hydroxy-TAM and 3,4-dihydroxy-TAM. Furthermore, a structural isomer of diethylstilbestrol (DES) with the hydroxy groups shifted into the meta-positions (3,3'-DES) was included. Our data clearly show that TAM and 4-hydroxy-TAM give rise to morphologic and neoplastic transformation of SHE cells in culture. In contrast, neither 3-hydroxy-TAM nor 3,3'-DES led to a significant transformation, in spite of the fact that both compounds are hormonally active. These results support the notion that nonhormonal events are essential for SHE cell transformation and suggest certain structural features to be necessary for the transforming capability. The putative carcinogenicity of TAM and 4-hydroxy-TAM but not of 3-hydroxy-TAM, as implied by these in vitro transformation studies, is corroborated by recent reports on ovarian and Leydig cell tumors in mice and hepatocellular carcinomas in rats.

Topics: Animals; Cell Transformation, Neoplastic; Cells, Cultured; Cricetinae; Estrogen Antagonists; Estrogens, Non-Steroidal; Fibrosarcoma; Mesocricetus; Mice; Mice, Nude; Stilbenes; Structure-Activity Relationship