cyclin-d1 and wogonin

Wogonin has been reported to exhibit various biological activities such as anti-inflammation, anti-microbial, and anti-tumor. Previous studies have demonstrated that wogonin could down-regulate Cyclin D1 activity on multiple cancers. However, the related mechanisms have not been fully elucidated so far.. The aim of the current study was to explore whether wogonin can suppress hepatocellular carcinoma (HCC) progression and the mechanism of wogonin in inhibiting Cyclin D1 expression.. Herein, we assessed the anti-tumor activity of wogonin against hepatocellular carcinoma (HCC) by MTT assay, clonogenic assay, cell cycle analysis and orthotopic xenograft mouse models. Western blot, immunofluoscence assay, co-immunoprecipitation assay, docking program, surface plasmon resonance, site-directed mutagenesis assay and immunohistochemical assay were performed for exploring the underlying mechanisms of wogonin-induced growth inhibition in HCC.. Our results showed that non-toxic dosage of wogonin (10, 20 µM) could inhibit cells proliferation and suppress cells cycle progression in MHCC97L and HepG2 cell. Moreover, the findings from the western blot and immunofluoscence assay confirmed the inhibition action of wogonin (10, 20 µM) on Cyclin D1 expression in MHCC97L cells, and wogonin (10, 20 µM) pre-treatment was capable of promoting Cyclin D1 ubiquitination and degradation in MHCC97L cell. In addition, wogonin promoted phosphorylation of Cyclin D1 on threonine-286 site, the mutation of threonine-286 to alanine-286A blocked Cyclin D1 proteolysis induced by wogonin. Wogonin-promoted Cyclin D1 phosphorylation and subsequent proteolysis may associate with the activation of GSK3beta in cancer cells. The phosphorylated form of GSK3beta (active form) expression was significantly increased after wogonin (20 µM) exposure. Molecular docking study and Biacore SPR analysis of GSK3beta mutant further validated the high-affinity wogonin binding site on GSK3beta. Moreover, in vivo studies further confirmed that phospho-GSK3beta Tyr216 was over-expressed in HCC specimens after wogonin treatment while the amount of Cyclin D1 was significantly decreased.. In summary, our data reveal a novel molecular mechanism by which wogonin induces HCC cells cycle arrest and suppresses tumor proliferation.

Topics: Animals; Antineoplastic Agents; Carcinoma, Hepatocellular; Cell Cycle Checkpoints; Cell Proliferation; Cyclin D1; Down-Regulation; Enzyme Activation; Flavanones; Glycogen Synthase Kinase 3 beta; Hep G2 Cells; Humans; Liver Neoplasms; Male; Mice, Inbred BALB C; Molecular Docking Simulation; Phosphorylation; Xenograft Model Antitumor Assays

To explore the effects of wogonin on the apoptosis,invasion,migration and Wnt / β-catenin signaling pathway of gastric cancer cells SGC7901.. Three common gastric cancer cell lines( SGC7901,BGC-823 and MKN-45) were conventionally cultured to logarithmic growth. The cell proliferation was detected by MTT assay after treatment with different concentrations of wogonin( 0,20,50,100 and 200 μmol / L) for 24,48,72 and 96 h. The apoptosis rate, migration and invasion ability of SGC7901 cells were detected by Annexin V-FITC / PI annexin double staining flow cytometry, scratch test and transwell cell invasion assay, respectively. After treament with different concentrations of wogonin for 48 h,the protein levels of β-catenin, C-myc and Cyclin D1 were detected by Western blotting.. Wogonin within the range of 20 ~ 200 mol / L could inhibit the proliferation of SGC7901,BGC-823 and MKN-45 cells in a dose-and time-dependent manner. Compared with 0 μmol / L wogonin group,After treament with different concentrations of wogonin for 24 h and 48 h,SGC7901 cells had elevated apoptosis rate and decreased migration distance and number of penetrating membrane cell in the rest concentrations( P < 0. 05). The protein levels of β-catenin,C-myc and Cyclin D1 were lower in 20 ~ 200μmol /L wogonin groups than 0 μmol /L wogonin group except for the concentration of 20 μmol /L wogonin groups( P < 0. 05),and the effects in a dose-dependent manner.. Wogonin has the ability to inhibit the proliferation of gastric cancer cells,and can induce apoptosis and inhibit cell migration and invasion,which may be related to the inhibition of Wnt / β-catenin signaling pathway activation.

Topics: Apoptosis; beta Catenin; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Flavanones; Humans; Stomach Neoplasms; Wnt Signaling Pathway

Wogonin, a naturally occurring flavonoid, has been shown to have tumor therapeutic potential both in vitro and in vivo. To better understand its anticancer mechanism, we examined the effect of wogonin on human cervical carcinoma HeLa cells. In this study, we observed that G1 phase arrest was involved in wogonin-induced growth inhibition in HeLa cells. Over a 24 h exposure of HeLa cells to 90 micromol x L(-1) wogonin, the promoters of G1-S transition, including cyclin D1/Cdk4 and pRb, decreased within 12 h and E2F-1 depleted in the nucleus at the same time. As the G1 phase arrest developed, p53 and the Cdk inhibitor p21Cip1 elevated both at protein and mRNA levels. Furthermore, the up-regulation of p21Cip1 induced by wogonin was dramatically inhibited by siRNA-mediated p53 gene silencing. Collectively, our data suggested that wogonin induced G1 phase arrest in HeLa cells by modulating several key G1 regulatory proteins, such as Cdk4 and cyclin D1, as well as up-regulation of a p53-mediated p21Cip1 expression. This mechanism of wogonin may play an important role in the killing of cancerous cells and offer a potential mechanism for its anticancer action in vivo.

Topics: Apoptosis; Cyclin D1; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinase Inhibitor p21; Flavanones; G1 Phase; HeLa Cells; Humans; RNA, Small Interfering; Tumor Suppressor Protein p53

Wogonin is a plant monoflavonoid which has been reported to inhibit cell growth and/or induce apoptosis in various tumors. Herein, we investigated the in vitro and in vivo anticancer effects and associated mechanisms of wogonin in human breast cancer. Effects of wogonin were examined in estrogen receptor (ER)-positive and -negative human breast cancer cells in culture for proliferation, cell cycle progression, and apoptosis. The in vivo effect of oral wogonin was examined on tumor xenograft growth in athymic nude mice. The molecular changes associated with the biological effects of wogonin were analyzed by immunoblotting. Cell growth was attenuated by wogonin (50-200 microM), independently of its ER status, in a time- and concentration-dependent manner. Apoptosis was enhanced and accompanied by upregulation of PARP and Caspase 3 cleavages as well as proapoptotic Bax protein. Akt activity was suppressed and reduced phosphorylation of its substrates, GSK-3beta and p27, was observed. Suppression of Cyclin D1 expression suggested the downregulation of the Akt-mediated canonical Wnt signaling pathway. ER expression was downregulated in ER-positive cells, while c-ErbB2 expression and its activity were suppressed in ER-negative SK-BR-3 cells. Wogonin feeding to mice showed inhibition of tumor growth of T47D and MDA-MB-231 xenografts by up to 88% without any toxicity after 4 weeks of treatment. As wogonin was effective both in vitro and in vivo, our novel findings open the possibility of wogonin as an effective therapeutic and/or chemopreventive agent against both ER-positive and -negative breast cancers, particularly against the more aggressive and hormonal therapy-resistant ER-negative types.

Topics: Animals; Apoptosis; Breast Neoplasms; Caspases; Cell Proliferation; Cyclin D1; Drugs, Chinese Herbal; Estrogen Receptor alpha; Female; Flavanones; Humans; Mice; Mice, Inbred BALB C; Mice, Nude; Receptor, ErbB-2; Signal Transduction; Tumor Cells, Cultured; Xenograft Model Antitumor Assays