Quercetin inhibits HGF/c-Met signaling and HGF-stimulated melanoma cell migration and invasion.

BACKGROUND: Melanoma is notorious for its propensity to metastasize, which makes treatment extremely difficult. Receptor tyrosine kinase c-Met is activated in human melanoma and is involved in melanoma progression and metastasis. Hepatocyte growth factor (HGF)-mediated activation of c-Met signaling has been suggested as a therapeutic target for melanoma metastasis. Quercetin is a dietary flavonoid that exerts anti-metastatic effect in various types of cancer including melanoma. In a previous report, we demonstrated that quercetin inhibited melanoma cell migration and invasion in vitro, and prevented melanoma cell lung metastasis in vivo. In this study, we sought to determine the involvement of HGF/c-Met signaling in the anti-metastatic action of quercetin in melanoma. METHODS: Transwell chamber assay was conducted to determine the cell migratory and invasive abilities. Western blotting was performed to determine the expression levels and activities of c-Met and its downstream molecules. And immunoblotting was performed in BS(3) cross-linked cells to examine the homo-dimerization of c-Met. Quantitative real-time PCR analysis was carried out to evaluate the mRNA expression level of HGF. Transient transfection was used to overexpress PAK or FAK in cell models. Student's t-test was used in analyzing differences between two groups. RESULTS: Quercetin dose-dependently suppressed HGF-stimulated melanoma cell migration and invasion. Further study indicated that quercetin inhibited c-Met phosphorylation, reduced c-Met homo-dimerization and decreased c-Met protein expression. The effect of quercetin on c-Met expression was associated with a reduced expression of fatty acid synthase. In addition, quercetin suppressed the phosphorylation of c-Met downstream molecules including Gab1 (GRB2-associated-binding protein 1), FAK (Focal Adhesion Kinase) and PAK (p21-activated kinases). More importantly, overexpression of FAK or PAK significantly reduced the inhibitory effect of quercetin on the migration of the melanoma cells. CONCLUSIONS: Our findings suggest that suppression of the HGF/c-Met signaling pathway contributes to the anti-metastatic action of quercetin in melanoma.

The herbal compound cryptotanshinone restores sensitivity in cancer cells that are resistant to the tumor necrosis factor-related apoptosis-inducing ligand.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) selectively induces apoptosis and kills cancer cells but not normal cells. However, TRAIL resistance due to low level of TRAIL receptor expression is widely found in cancer cells and hampers its development for cancer treatment. Thus, the agents that can sensitize the tumor cells to TRAIL-mediated apoptosis are urgently needed. We investigated whether tanshinones, the major bioactive compounds of Salvia miltiorrhiza (danshen), can up-regulate TRAIL receptor expression. Among the major tanshinones being tested, cryptotanshinone (CT) showed the best ability to induce TRAIL receptor 2 (DR5) expression. We further showed that CT was capable of promoting TRAIL-induced cell death and apoptosis in A375 melanoma cells. CT-induced DR5 induction was not cell type-specific, as DR5 induction was observed in other cancer cell types. DR5 knockdown abolished the enhancing effect of CT on TRAIL responses. Mechanistically, induction of the DR5 by CT was found to be p53-independent but dependent on the induction of CCAAT/enhancer-binding protein-homologous protein (CHOP). Knockdown of CHOP abolished CT-induced DR5 expression and the associated potentiation of TRAIL-mediated cell death. In addition, CT-induced ROS production preceded up-regulation of CHOP and DR5 and consequent sensitization of cells to TRAIL. Interestingly, CT also converted TRAIL-resistant lung A549 cancer cells into TRAIL-sensitive cells. Taken together, our results indicate that CT can potentiate TRAIL-induced apoptosis through up-regulation of DR5.

Lipidomics identification of metabolic biomarkers in chemically induced hypertriglyceridemic mice.

In this study, we aim to identify the potential biomarkers in hTG pathogenesis in schisandrin B-induced hTG mouse model. To investigate whether these identified biomarkers are only specific to schisandrin B-induced hTG mouse model, we also measured these biomarkers in a high fat diet (HFD)-induced hTG mouse model. We employed a LC/MS/MS-based lipidomic approach for the study. Mouse liver and serum metabolites were separated by reversed phase liquid chromatography. Metabolite candidates were identified by matching with marker retention times, isotope distribution patterns, and high-resolution MS/MS fragmentation patterns. Subsequently, target candidates were quantified by quantitative MS. In the schisandrin B-induced hTG mice, we found that the plasma fatty acids, diglyceroids, and phospholipids were significantly increased. Palmitic acid and stearic acid were increased in the plasma; oleic acid, linoleic acid, linolenic acid, arachidonic acid, and docosahexaenoic acid were increased in both the plasma and the liver. Acetyl-CoA, malonyl-CoA, and succinyl-CoA were increased only in the liver. The changes in levels of these identified markers were also observed in HFD-induced hTG mouse model. The consistent results obtained from both hTG models not only suggest novel biomarkers in hTG pathogenesis, but they also provide insight into the underlying mechanism of the schisandrin B-induced hTG.

Inhibition of the STAT3 signaling pathway contributes to apigenin-mediated anti-metastatic effect in melanoma.

Signal transducer and activator of transcription 3 (STAT3) signaling is constantly activated in human melanoma, and promotes melanoma metastasis. The dietary flavonoid apigenin is a bioactive compound that possesses low toxicity and exerts anti-metastatic activity in melanoma. However, the anti-metastasis mechanism of apigenin has not been fully elucidated. In the present study, we showed that apigenin suppressed murine melanoma B16F10 cell lung metastasis in mice, and inhibited cell migration and invasion in human and murine melanoma cells. Further study indicated that apigenin effectively suppressed STAT3 phosphorylation, decreased STAT3 nuclear localization and inhibited STAT3 transcriptional activity. Apigenin also down-regulated STAT3 target genes MMP-2, MMP-9, VEGF and Twist1, which are involved in cell migration and invasion. More importantly, overexpression of STAT3 or Twist1 partially reversed apigenin-impaired cell migration and invasion. Our data not only reveal a novel anti-metastasis mechanism of apigenin but also support the notion that STAT3 is an attractive and promising target for melanoma treatment.

Quercetin exerts anti-melanoma activities and inhibits STAT3 signaling.

Melanoma is highly resistant to chemotherapy, and the mortality rate is increasing rapidly worldwide. STAT3 signaling has been implicated in the pathogenesis of melanoma and constitutive activated STAT3 has been validated can as a target for melanoma therapy. Quercetin, a noncarcinogenic dietary flavonoid with low toxicity, has been shown to exert anti-melanoma activity. However, the anti-melanoma mechanisms of quercetin are not fully understood. In this study, we sought to test the involvement of STAT3 signaling in the inhibitory effects of quercetin on melanoma cell growth, migration and invasion. Our results showed that exposure to quercetin resulted in inhibition of proliferation of melanoma cells, induction of cell apoptosis, and suppression of migratory and invasive properties. Mechanistic study indicated that quercetin inhibited the activation of STAT3 signaling by interfering with STAT3 phosphorylation, and reducing STAT3 nuclear localization. This inhibited STAT3 transcription activity and down-regulated STAT3 targeted genes Mcl-1, MMP-2, MMP-9 and VEGF, which are involved in cell growth, migration and invasion. Importantly, overexpression of constitutively active STAT3 partially rescued the growth inhibiting effects induced by quercetin. Furthermore, quercetin suppressed A375 tumor growth and STAT3 activities in xenografted mice model, and inhibited murine B16F10 cells lung metastasis in an animal model. Overall, these results indicate that the antitumor activity of quercetin is at least partially due to inhibition of STAT3 signaling in melanoma cells. Our findings provided new insight into the action of quercetin potently inhibits the STAT3 signaling pathway, suggesting it has a potential role in the prevention and treatment of melanoma.

Indomethacin sensitizes TRAIL-resistant melanoma cells to TRAIL-induced apoptosis through ROS-mediated upregulation of death receptor 5 and downregulation of survivin.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has attracted considerable attention owing to its selective killing of tumor cells but not normal cells. Melanoma shows weak response to TRAIL because of its low level of TRAIL death receptors. Here, we investigated whether indomethacin, a nonsteroidal anti-inflammatory drug, can potentiate TRAIL-induced apoptosis in melanoma cells. We showed that indomethacin was capable of promoting TRAIL-induced cell death and apoptosis in A375 melanoma cells. Mechanistically, indomethacin induced cell surface expression of death receptor 5 (DR5) in melanoma cells and also in various types of cancer cells. DR5 knockdown abolished the enhancing effect of indomethacin on TRAIL responses. Induction of the DR5 by indomethacin was found to be p53 independent but dependent on the induction of CCAAT/enhancer-binding protein homologous protein (CHOP). Knockdown of CHOP abolished indomethacin-induced DR5 expression and the associated potentiation of TRAIL-mediated cell death. In addition, indomethacin-induced reactive oxygen species (ROS) production preceded upregulation of CHOP and DR5, and consequent sensitization of cells to TRAIL. We also found that indomethacin treatment downregulated survivin via ROS and the NF-κB-mediated signaling pathways. Interestingly, indomethacin also converted TRAIL-resistant melanoma MeWo and SK-MEL-5 cells into TRAIL-sensitive cells. Taken together, our results indicate that indomethacin can potentiate TRAIL-induced apoptosis through upregulation of death receptors and downregulation of survivin.

Comparisons of the chemical profiles, cytotoxicities and anti-inflammatory effects of raw and rice wine-processed Herba Siegesbeckiae.

ETHNOPHARMACOLOGICAL RELEVANCE: Although slightly toxic, the Chinese medicinal herb Herba Siegesbeckiae (HS) has long been used as a remedy for traditional Chinese medicine symptoms that resemble inflammatory joint disorders, because it can eliminate the wind-dampness and soothe painful joints. Proper processing can reduce the toxicity and/or enhance the efficacy of raw herbs. In this study, we aim to examine if processing with rice wine reduces the cytotoxicities and/or enhances the anti-inflammatory effects of HS, and to explore the chemical basis behind the potential changes of medicinal properties caused by the processing. MATERIALS AND METHODS: We used cell models to examine the cytotoxicities and anti-inflammatory effects of HS and rice wine-processed HS (WHS). The chemical profiles of HS and WHS were compared using the ultra-performance liquid chromatography/quadrupole-time-of-flight mass spectrometry (UPLC/Q-TOF-MS) analysis. RESULTS: We found that WHS was less toxic than HS in cultured cells as shown in the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Both HS and WHS had anti-inflammatory effects as demonstrated by their abilities to reduce nitric oxide (NO) production as well as protein and mRNA expressions of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. Interestingly, the anti-inflammatory effects of WHS were more potent than that of HS at the concentration of 100 µg/mL. By comparing the chemical profiles, we found that 19 peaks were lower, while 2 other peaks were higher in WHS than in HS. Four compounds including neo-darutoside, darutoside, stigmasterol and 16-O-acetyldarutoside corresponding to 4 individual changed peaks were tentatively identified by matching with empirical molecular formulae and mass fragments. CONCLUSION: Our study showed that processing with rice wine significantly reduced the cytotoxicities and enhanced the anti-inflammatory effects of HS as demonstrated in cell models. We also developed a UPLC/Q-TOF-MS method to clearly differentiate HS from WHS by their different chemical profiles. Further study is warranted to establish the relationship between the alteration of chemical profiles and the changes of medicinal properties caused by processing with rice wine.