Theacrine attenuates epithelial mesenchymal transition in human breast cancer MDA-MB-231 cells.

Theacrine, a purine alkaloid structurally similar to caffeine, has recently become of interest as a potential therapeutic compound. Here, we investigated the antimetastatic potential of theacrine on human breast cancer MDA-MB-231 cells. We observed that theacrine can reverse epithelial-to-mesenchymal transition (EMT), which resulted in a decrease in the levels of mesenchymal markers (Fibronectin, Vimentin, N-cadherin, Twist, and Snail) and an increase in the levels of epithelial markers (Occludin and E-cadherin) in the cells. Additionally, theacrine attenuates TGF-ß-induced EMT, cell adhesion, migration, and invasion in MDA-MB-231 cells. Overall, our results suggest that theacrine may inhibit the breast cancer cell metastasis by reversing the EMT process.

Pharmacological Utilization of Bergamottin, Derived from Grapefruits, in Cancer Prevention and Therapy.

Cancer still remains one of the leading causes of death worldwide. In spite of significant advances in treatment options and the advent of novel targeted therapies, there still remains an unmet need for the identification of novel pharmacological agents for cancer therapy. This has led to several studies evaluating the possible application of natural agents found in vegetables, fruits, or plant-derived products that may be useful for cancer treatment. Bergamottin is a furanocoumarin derived from grapefruits and is also a well-known cytochrome P450 inhibitor. Recent studies have demonstrated potent anti-oxidative, anti-inflammatory, and anti-cancer properties of grapefruit furanocoumarin both in vitro and in vivo. The present review focuses on the potential anti-neoplastic effects of bergamottin in different tumor models and briefly describes the molecular targets affected by this agent.

Bergamottin Inhibits Adipogenesis in 3T3-L1 Cells and Weight Regulation in Diet-Induced Obese Mice.

Obesity is a serious and increasing health problem worldwide, and the inhibition of adipogenesis is considered to be a potential therapeutic target for it. Bergamottin (BGM), a component of grapefruit juice, has been reported to regulate lipolysis. However, the physiological role of BGM in obesity has not been evaluated so far. In the present study, we investigated the effects of BGM on obesity in 3T3-L1 cells and in mice fed a high-fat diet (HFD). BGM inhibited adipogenic differentiation of 3T3-L1 cells along with a significant decrease in the lipid content by downregulating the expression of two critical adipogenic factors, CCAAT enhancer-binding protein-alpha (C/EBP[Formula: see text]) and peroxisome proliferator activated receptor-gamma (PPAR[Formula: see text]). The expressions of target proteins such as adipocyte fatty acid-binding protein (aP2), adiponectin, and resistin were also decreased by BGM. It activated AMP-activated protein kinase (AMPK) by increasing phosphorylation of AMPK and the downstream target acetyl-CoA carboxylase (ACC), indicating that BGM exerted its antiadipogenic effect through AMPK activation. In the HFD-induced obese mouse model, BGM administration significantly reduced the weight and sizes of white adipose tissue as well as the weight gain of mice fed HFD. Moreover, UCP1 and PGC1[Formula: see text] expressions, well-known as brown adipocyte marker genes, were higher in the BGM-treated HFD mice than that in the HFD-induced obese mice. This study suggests that BGM suppress adipogenesis by AMPK activation in vitro and reduces body weight in vivo.

2,5-Dihydroxyacetophenone Induces Apoptosis of Multiple Myeloma Cells by Regulating the MAPK Activation Pathway.

2,5-Dihydroxyacetophenone (DHAP) is an active compound obtained from Radix rehmanniae preparata, which is widely used as a herbal medicine in many Asian countries. DHAP has been found to possess anti-inflammatory, anti-anxiety, and neuroprotective qualities. For the present study, we evaluated the anti-cancer effects of DHAP on multiple myeloma cells. It was discovered that DHAP downregulated the expression of oncogenic gene products like Bcl-xl, Bcl-2, Mcl-1, Survivin, Cyclin D1, IAP-1, Cyclin E, COX-2, and MMP-9, and upregulated the expression of Bax and p21 proteins, consistent with the induction of G2/M phase cell cycle arrest and apoptosis in U266 cells. DHAP inhibited cell proliferation and induced apoptosis, as characterized by the cleavage of PARP and the activation of caspase-3, caspase-8, and caspase-9. Mitogen-activated protein kinase (MAPK) pathways have been linked to the modulation of the angiogenesis, proliferation, metastasis, and invasion of tumors. We therefore attempted to determine the effect of DHAP on MAPK signaling pathways, and discovered that DHAP treatment induced a sustained activation of JNK, ERK1/2, and p38 MAPKs. DHAP also potentiated the pro-apoptotic and anti-proliferative effects of bortezomib in U266 cells. Our results suggest that DHAP can be an effective therapeutic agent to target multiple myeloma.

Korean Red Ginseng Extract Enhances the Anticancer Effects of Sorafenib through Abrogation of CREB and c-Jun Activation in Renal Cell Carcinoma.

Although application of sorafenib in the treatment of human renal cell carcinoma (RCC) remains one of the best examples of successful targeted therapy, majority of RCC patients suffer from its side effects as well as develop resistance to this targeted therapy. Thus, there is a need to promote novel alternative therapies for the treatment of RCC. In this study, we investigated whether Korean red ginseng extract (KRGE) could inhibit the proliferation and induce chemosensitization in human renal cancer cells. Also, we used a human phospho-antibody array containing 46 antibodies against signaling molecules to examine a subset of phosphorylation events after KRGE and sorafenib combination treatment. Korean red ginseng extract suppressed the proliferation of two RCC cell lines; activated caspase-3; caused poly(ADP-ribose) polymerase cleavage; abrogated the expression of B-cell lymphoma 2, B-cell lymphoma extra large, survivin, inhibitors of apoptosis proteins-1/2, cyclooxygenase-2, cyclin D1, matrix metallopeptidase-9, and vascular endothelial growth factor; and upregulated pro-apoptotic gene products. Interestingly, KRGE enhanced the cytotoxic and apoptotic effects of sorafenib in RCC cells. The combination treatment of KRGE and sorafenib more clearly suppressed cyclic adenosine monophosphate response element-binding protein and c-Jun phosphorylation and induced phosphorylation of p53 than did the individual treatment regimen. Our results clearly demonstrate that KRGE can enhance the anticancer activity of sorafenib and may have a substantial potential in the treatment of RCC. Copyright © 2017 John Wiley & Sons, Ltd.

Ginkgolic Acid C 17:1, Derived from Ginkgo biloba Leaves, Suppresses Constitutive and Inducible STAT3 Activation through Induction of PTEN and SHP-1 Tyrosine Phosphatase.

Ginkgolic acid C 17:1 (GAC 17:1) extracted from Ginkgo biloba leaves, has been previously reported to exhibit diverse antitumor effect(s) through modulation of several molecular targets in tumor cells, however the detailed mechanism(s) of its actions still remains to be elucidated. Signal transducer and activator of transcription 3 (STAT3) is an oncogenic transcription factor that regulates various critical functions involved in progression of diverse hematological malignancies, including multiple myeloma, therefore attenuating STAT3 activation may have a potential in cancer therapy. We determined the anti-tumor mechanism of GAC 17:1 with respect to its effect on STAT3 signaling pathway in multiple myeloma cell lines. We found that GAC 17:1 can inhibit constitutive activation of STAT3 through the abrogation of upstream JAK2, Src but not of JAK1 kinases in U266 cells and also found that GAC can suppress IL-6-induced STAT3 phosphorylation in MM.1S cells. Treatment of protein tyrosine phosphatase (PTP) inhibitor blocked suppression of STAT3 phosphorylation by GAC 17:1, thereby indicating a critical role for a PTP. We also demonstrate that GAC 17:1 can induce the substantial expression of PTEN and SHP-1 at both protein and mRNA level. Further, deletion of PTEN and SHP-1 genes by siRNA can repress the induction of PTEN and SHP-1, as well as abolished the inhibitory effect of drug on STAT3 phosphorylation. GAC 17:1 down-regulated the expression of STAT3 regulated gene products and induced apoptosis of tumor cells. Overall, GAC 17:1 was found to abrogate STAT3 signaling pathway and thus exert its anticancer effects against multiple myeloma cells.

Resveratrol inhibits STAT3 signaling pathway through the induction of SOCS-1: Role in apoptosis induction and radiosensitization in head and neck tumor cells.

BACKGROUND: Signal transducer and activator of transcription 3 (STAT3) is persistently activated in squamous cell carcinoma of the head and neck (SCCHN) and can cause uncontrolled cellular proliferation and division. HYPOTHESIS: Thus, its targeted abrogation could be an effective strategy to reduce the risk of SCCHN. Resveratrol is known for its anti-cancer efficacy in a variety of cancer models. STUDY DESIGN: The effect resveratrol on STAT3 activation, associated protein kinases, phosphatases, cellular proliferation and apoptosis was investigated. METHODS: We evaluated the effect of resveratrol on STAT3 signaling cascade and its regulated functional responses in SCCHN cells. RESULTS: We found that HN3 and FaDu cells expressed strongly phosphorylated STAT3 on both tyrosine 705 and serine 727 residues as compared to other SCCHN cells. The phosphorylation was completely suppressed by resveratrol in FaDu cells, but not substantially in HN3 cells. STAT3 suppression was mediated through the inhibition of activation of upstream JAK2, but not of JAK1 and Src kinases. Treatment with the protein tyrosine phosphatase (PTP) inhibitor pervanadate reversed the resveratrol-induced down-regulation of STAT3, thereby indicating a critical role for a PTP. We also found that resveratrol induced the expression of the SOCS-1 protein and mRNA. Further, deletion of SOCS-1 gene by siRNA suppressed the induction of SOCS-1, and reversed the inhibition of STAT3 activation. Resveratrol down-regulated various STAT3-regulated gene products, inhibited proliferation, invasion, as well as induced the cell accumulation in the sub-G1 phase and caused apoptosis. Beside, this phytoalexin also exhibited the enhancement of apoptosis when combined with ionizing radiation treatment. CONCLUSION: Our results suggest that resveratrol blocks STAT3 signaling pathway through induction of SOCS-1, thus attenuating STAT3 phosphorylation and proliferation in SCCHN cells.

Embelin Inhibits Invasion and Migration of MDA-MB-231 Breast Cancer Cells by Suppression of CXC Chemokine Receptor 4, Matrix Metalloproteinases-9/2, and Epithelial-Mesenchymal Transition.

Embelin (EB) is a benzoquinone derivative isolated from Embelia ribes Burm plant. Recent scientific evidence shows that EB induces apoptosis and inhibits migration and invasion in highly metastatic human breast cancer cells. However, the exact mechanisms of EB in tumor metastasis and invasion have not been fully elucidated. Here, we investigated the underlying mechanisms of antimetastatic activities of EB in breast cancer cells. The EB downregulated the chemokine receptor 4 (CXCR4) as well as matrix metalloproteinase (MMP)-9/2 expression and upregulated the tissue inhibitor of metalloproteinase 1 expression in MDA-MB-231 cells under noncytotoxic concentrations but not in MCF-7 cells. Additionally, EB inhibited the CXC motif chemokine ligand 12 induced invasion and migration activities of MDA-MB-231 cells. A detailed study of underlying mechanisms revealed that the regulation of the downregulation of CXCR4 was at the transcriptional level, as indicated by the downregulation of mRNA expression and suppression of nuclear factor-kappa B (NF-κB) activation. It further reduced the binding of NF-κB to the CXCR4 promoter. Besides, EB downregulated mesenchymal marker proteins (neural cadherin and vimentin) and concurrently upregulated epithelial markers (epithelial cadherin and occludin). Overall, these findings suggest that EB can abrogate breast cancer cell invasion and metastasis by suppression of CXCR4, MMP-9/2 expressions, and inhibition of epithelial-mesenchymal transition and thus may have a great potential to suppress metastasis of breast cancer. Copyright © 2016 John Wiley & Sons, Ltd.

Ginkgetin Blocks Constitutive STAT3 Activation and Induces Apoptosis through Induction of SHP-1 and PTEN Tyrosine Phosphatases.

Ginkgetin, a biflavone from Ginkgo biloba leaves, is known to exhibit antiinflammatory, antifungal, neuroprotective, and antitumor activities, but its precise mechanism of action has not been fully elucidated. Because the aberrant activation of STAT3 has been linked with regulation of inflammation, proliferation, invasion, and metastasis of tumors, we hypothesized that ginkgetin modulates the activation of STAT3 in tumor cells. We found that ginkgetin clearly suppressed constitutive phosphorylation of STAT3 through inhibition of the activation of upstream JAK1 and c-Src kinases and nuclear translocation of STAT3 on both A549 and FaDu cells. Treatment with sodium pervanadate reversed the ginkgetin-induced down-modulation of STAT3, thereby indicating a critical role for a PTP. We also found that ginkgetin strongly induced the expression of the SHP-1 and PTEN proteins and its mRNAs. Further, deletion of SHP-1 and PTEN genes by siRNA suppressed the induction of SHP-1 and PTEN, and reversed the inhibition of STAT3 activation. Ginkgetin induced apoptosis as characterized by an increased accumulation of cells in subG1 phase, positive Annexin V binding, loss of mitochondrial membrane potential, down-regulation of STAT3-regulated gene products, and cleavage of PARP. Overall, ginkgetin abrogates STAT3 signaling pathway through induction of SHP-1 and PTEN proteins, thus attenuating STAT3 phosphorylation and tumorigenesis.

Hypolipidemic effect and antioxidant activity of glycoprotein isolated from Ulmus davidiana Nakai in Triton WR-1339-treated mouse.

The glycoprotein isolated from Ulmus davidiana Nakai (UDN) (UDN glycoprotein) has a molecular weight of 116 kDa and consists of 78.65% carbohydrate content and 21.35% protein content. In the present study, we investigated the hypolipidemic effect of UDN glycoprotein on Triton WR-1339-induced mice. With pretreatment with UDN glycoprotein, the triacylglycerol (TAG), total cholesterol and low density lipoprotein-cholesterol (LDL-C) concentrations were significantly reduced, whereas high density lipoprotein-cholesterol (HDL-C) concentration was increased in the plasma of Triton WR-1339-induced mice. With respect to antioxidative activity, UDN glycoprotein significantly decreased the level of thiobarbituric acid reactive substances (TBARS) and improved activities of catalase and glutathione peroxidase (GPx), without an apparent change of superoxide dismutase (SOD) activity. Also UDN glycoprotein significantly increased nitric oxide (NO) production in Triton WR-1339-induced mice. These results indicate that UDN glycoprotein has a hypolipidemic effect, possesses antioxidant activity and has an ability to stimulate NO production. Thus, we speculate that UDN glycoprotein is an example of natural compound that lowers plasma lipid level together with having an antioxidant function in Triton WR-1339-induced mice.

Protective effect of glycoprotein isolated from Ulmus davidiana Nakai on carbon tetrachloride-induced mouse liver injury.

This study was carried out to evaluate the hepatoprotective activity of glycoprotein isolated from the stems of Ulmus davidiana Nakai (UDN), which has been used as an anti-inflammatory agent in folk medicine. We evaluated lipid peroxidation in glucose/glucose oxidase (G/GO)-induced BNL CL.2 cells and measured thiobarbituric acid reactive substances (TBARS), lactate dehydrogenase (LDH), nitric oxide (NO), antioxidant enzyme (superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx)), activity of cytotoxic-related signals (hepatic cytochrome c, nuclear factor-kappa B (NF-kappaB) and activator protein-1 (AP-1)) and levels of plasma lipids (triglyceride (TG) and total cholesterol (TC)) in carbon tetrachloride (CCl(4,) 1.0 mL kg(-1))-induced A/J mouse. The results in G/GO-induced BNL CL.2 cells showed that UDN glycoprotein had a dose-dependent inhibitory effect on lipid peroxidation. The results in carbon tetrachloride (CCl(4,) 1.0 mL kg(-1))-induced A/J mouse indicated that treatment with UDN glycoprotein (40 mg kg -1) lowered LDH activity and TBARS formation, and increased NO production and antioxidant enzymes activity, compared with control. Also, our finding from CCl(4)-treated mice after pretreatment with UDN glycoprotein demonstrated that the activity of cytotoxic-related signals decreased but the levels of plasma lipids increased, compared with CCl(4) treatment alone. Here, we speculate that UDN glycoprotein has a protective character to CCl(4)-induced mouse liver injury.

Glycoprotein isolated from Gardenia jasminoides Ellis has a scavenging activity against oxygen radicals and inhibits the oxygen radical-induced protein kinase C alpha and nuclear factor-kappa B in NIH/3T3 cells.

This study was earned out to investigate the antioxidative and anti-apoptotic effects of glycoprotein isolated from Gardenia jasminoides Ellis fruit (GJE glycoprotein), which has been used to heal hepatic and inflammatory diseases in folk medicine. GJE glycoprotein showed a single band with a molecular weight of 27kDa on the 15% sodium dodecyl sulfate polyacrylamide gel. It consists of a carbohydrate component (57.65%) and a protein component (42.35%). GJE glycoprotein has dose-dependent scavenging activities for DPPH, lipid peroxyl, superoxide anion and hydroxyl radicals in cell-free systems. We also evaluated the protective and anti-apoptotic activities of GJE glycoprotein on the glucose/glucose oxidase (G/GO)-induced or hypoxanthine/xanthine oxidase (HX/XO)-induced cytotoxicity and apoptosis systems in NIH/3T3 cells, using 3-(4,5-diinettiylthiazol-2-yl)-2,5 diphenyl tetrazolium bromide (MTT), DNA fragmentation and H33342/ethidium bromide staining assays, respectively. Results in this experiment showed that GJE glycoprotein has dose-dependent blocking activities against G/GO- or HX/XO-induced cytotoxicity and apoptosis. In addition, we investigated whether GJE glycoprotein blocks the activation of redox-sensitive signal mediators, protein kinase C alpha (PKCα) and nuclear factor-kappa B (NF-κB) in G/GO or HX/XO-induced apoptotic NIH/3T3 cells, using a Western blot analysis and an electrophoretic mobility shift assay (EMSA). We found that 100µg/ml GJE glycoprotein has an inhibitory effect on PKCα translocation and the DNA binding activity of (NF-κB). Here, we speculate that GJE glycoprotein is a natural antioxidant and one of the modulators of apoptotic signal pathways in NIH/3T3 cells.

Glycine- and proline-rich glycoprotein isolated from Solanum nigrum Linne activates caspase-3 through cytochrome c in HT-29 cells.

This study was carried out to investigate apoptotic effects of the glycoprotein (SNL glycoprotein, 150 kDa) isolated from Solanum nigrum Linne, which has been used as an anti-pyretic and anti-cancer agent in folk medicine. We found that the SNL glycoprotein consists of carbohydrate (69.74%) and protein content (30.26%), which has >50% hydrophobic amino acids containing glycine and proline. LDH assay indicated that the SNL glycoprotein has obvious cytotoxic and apoptotic effects (>50% cell death) at 40 microg/ml SNL glycoprotein for 2 h in HT-29 cells. The results showed that the SNL glycoprotein has a stimulatory effect on the release of mitochondrial cytochrome c, cleavages of pro-caspase-9, pro-caspase-3, and poly(ADP-ribose) polymerase (PARP) proteins in HT-29 cells. However, the SNL glycoprotein did not significantly stimulate or change the levels of intracellular reactive oxygen species (ROS). The results of this experiment suggest that the SNL glycoprotein activates caspase-3 in HT-29 cells, independent of ROS.

116 kDa glycoprotein isolated from Ulmus davidiana Nakai (UDN) inhibits glucose/glucose oxidase (G/GO)-induced apoptosis in BNL CL.2 cells.

Ulmus davidiana Nakai (UDN) has been used in folk medicine for its anti-inflammatory activity. In the present study, we investigated the antiapoptotic effect of UDN glycoprotein in glucose/glucose oxidase (G/GO)-induced BNL CL.2 cells. To evaluate the antiapoptotic effect of UDN glycoprotein, experiments were carried out using Western blot analysis for nuclear factor-kappa B (NF-kappaB), caspase-3, and poly(ADP-ribose) polymerase (PARP). We also examined nitric oxide (NO) production and nuclear staining. When BNL CL.2 cells were treated with G/GO (50 mU/ml), viability of the cells was 54.1%. However, the number of living cells after the addition of UDN glycoprotein in the presence of G/GO increased. UDN glycoprotein protected from cell damage caused by G/GO. Interestingly, UDN glycoprotein decreased NF-kappaB activation and stimulated NO production in G/GO-induced BNL CL.2 cells. In apoptotic parameters, UDN glycoprotein inhibited activations of caspase-3 and PARP cleavage in G/GO-induced BNL CL.2 cells. The results of nuclear staining indicated that UDN glycoprotein (50 microg/ml) has a protective ability from apoptotic cell death caused G/GO (50 mU/ml). In conclusion, UDN glycoprotein has a protective effect on apoptosis induced by G/GO through the inhibition of NF-kappaB, caspase-3, and PARP activity, and the stimulation of NO production in BNL CL.2 cells.

150 kDa glycoprotein isolated from Solanum nigrum Linne enhances activities of detoxicant enzymes and lowers plasmic cholesterol in mouse.

This study was carried out to investigate the modulation of detoxicant enzyme activity and plasma lipidemic levels by 150 kDa glycoprotein isolated from Solanum nigrum Linne (SNL), which has been used as a hepatoprotective and anticancer agent in folk medicine. Our results in this study showed that SNL glycoprotein has a band with 150 kDa on the 10% sodium dodecyl sulfate polyacrylamide gel, and that it has a strong scavenging activity against lipid peroxyl radicals. We also evaluated the lipidemic levels of SNL glycoprotein, based on lipoproteins and activities of detoxicant enzymes in treatment with Triton WR-1339 or corn oil in vivo. When mice were treated with either Triton WR-1339 or corn oil in the absence of SNL glycoprotein, the number of plasma lipoproteins [triglyceride (TG), total cholesterol (TC) and low density lipoprotein (LDL)] increased. However, when the mice were treated with either Triton WR-1339 or corn oil in the presence of SNL glycoprotein, the plasma lipoprotein levels (TG, TC and LDL) were significantly reduced. Similar results of SNL glycoprotein treatment were also produced in the activities of detoxicant enzymes. Namely, the activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) were remarkably increased after treatment with SNL glycoprotein. In addition, the activity of hepatic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase was reduced by SNL glycoprotein in cholestyramine-treated mice. For example, we found that it inhibits the activity of cholestyramine-induced hepatic HMG-CoA reductase at 40 microg head body weight g(-1) SNL glycoprotein. Collectively, these results pointed out that SNL glycoprotein can enhance the activities of detoxicant enzymes and bring about the inhibition of HMG-CoA reductase activity in vivo. Therefore, we speculate that SNL glycoprotein can be used as a cholesterol-lowering agent even at low concentrations.