Browsing by Subject "Tetrandrine"
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Item TETRANDRINE INDUCED INHIBITION OF SIGNAL TRANSDUCER AND ACTIVATOR OF TRANSCRIPTION (STAT) 3 CAUSES THE REDUCTION OF CELL SURVIVAL, PROLIFERATION, AND ANGIOGENESIS IN TNBC(2014-03) Gibbs, Lee D.; Chaudhary, Pankaj; Vishwanatha, JamboorNatural agents may be promising to combat aggressive behavior of the triple-negative breast cancer (TNBC). STAT3 is a protein that is highly expressed in breast cancer tissues compared to non-malignant breast tissues. Our objective for the present study is to analyze the anti-tumorogenic effects of a Chinese herbal drug, tetrandrine, in TNBC progression through inhibition of STAT3 phosphorylation. Purpose (a): The most successful therapies for breast cancer target the estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (Her-2). Hormonal therapies are not useful in combating triple negative breast cancer (TNBC), which lacks these targeted hormonal receptors. In fact, some of these patients that undergo hormone deprivation and/or Herceptin therapy acquire resistance. The triple negative breast cancer (TNBC) phenotype, which lacks the presence of Her-2, ER, and PR are even more aggressive and resistant. Therefore, there is an urgent clinical need to identify novel agents that can kill tumor cells with no additional toxicity to normal cells and this would have great impact on treatment of such patients. Tetrandrine, a bis-benzylisoquinoline alkaloid isolated from the root of Stephania tetrandra, is a calcium channel blocker used in Chinese medicine for the treatment of silicosis and arthritis. Studies have shown that tetrandrine also has anti-tumor and anti-growth activities. Our objective is to study the effects of tetrandrine on STAT3 signaling that plays an important role in cell proliferation, survival, chemoresistance and angiogenesis. STAT3 protein is highly expressed in breast cancer tissues compared to non-malignant breast tissues. We hypothesize that tetrandrine treatment inhibits the phosphorylation of STAT3 and its associated downstream signaling lead to the reduction of cell survival, proliferation, and angiogenesis in TNBC cells. Methods (b): TNBC cell lines, MDA-MB-231 and HCC70, and non-tumorigenic epithelial cell line MCF-10A were cultured in ATCC recommended medium. MTT assays were carried out to determine the effect of tetrandrine on cell viability. Additionally, cells were subjected to various concentrations of tetrandrine and Western blotting was performed for analysis of protein expression and phosphorylation. Results (c): Our data indicate that tetrandrine selectively inhibits the growth of MDA-MB-231 and HCC70 cells compared to non-tumorigenic MCF-10A cells. In the MTT assay, Tetrandrine concentrations ranging from 0 to 40μM gradually decreased MDA-MB-231, HCC70 and MCF-10A cell viability, corresponding to IC50 values of 25, 20 and 75 μM (n = 8), respectively, after 48 hours of treatment. Our results show that tetrandrine inhibited the phosphorylation of STAT3 in a concentration dependent manner. Furthermore, the inhibition of STAT3 activation by tetrandrine led to the suppression of proteins involved in proliferation (cyclin D1), survival (Bcl-2, Bcl-xL, and Mcl-1), and angiogenesis (VEGF). This effect correlated with the inhibition of proliferation and apoptosis in TNBC cells. Conclusions (d): Our preliminary results suggest that tetrandrine inhibits the proliferation of TNBC cells through inhibition of constitutive STAT3 phosphorylation and it’s associated down stream signaling and has therapeutic potential in the treatment of TNBC.Item TETRANDRINE INDUCES ROS-DRIVEN CASPASE-DEPENDENT APOPTOSIS OF PROSTATE CANCER CELLS VIA MITOCHONDRIAL AND CELL DEATH RECEPTOR PATHWAY(2014-03) Chaudhary, Pankaj; Vishwanatha, Jamboor K.Androgen deprivation is still the standard systemic therapy for prostate cancer, but patients invariably relapse with a more aggressive form of prostate cancer termed hormone refractory, androgen independent, or castration resistant prostate cancer. Once prostate cancer becomes castration-resistant, metastasis is a significant problem and treatment options are limited. Therefore, identification of novel agents that can selectively kill tumor cells with no additional toxicity to normal tissue would have significant impact on prostate cancer therapy. Purpose (a): Tetrandrine, a bisbenzylisoquinoline alkaloid, isolated from the root of Stephania tetrandra is used in traditional Chinese medicine as an anti-rheumatic, anti-inflammatory, and anti-hypertensive agent for the past several years. During recent years, increasing number of studies have focused on the potential of tetrandrine in cancer therapy. Despite its great potential as an anti-cancer agent, the effect of tetrandrine in prostate cancer has not been studied. Therefore, in the present study, we demonstrate the cytotoxic efficacy of tetrandrine in human androgen-independent prostate cancer cells, PC3 and DU145, and delineate the mechanism of this effect. Methods (b): Prostate cancer cell lines, PC3 and DU145, and normal prostate PWR-1E cells were cultured in ATCC recommended medium. The toxicity of tetrandrine was analyzed by MTT assay and Vybrant Apoptosis Assay Kit. Western blotting was used to detect the expression of proteins involved in apoptosis. Results (c): Our results indicate that tetrandrine selectively inhibits the growth of PC3 and DU145 cancer cells compared to normal prostate PWR-1E cells. Treatment of cancer cells with tetrandrine caused the upregulation of Fas and Bax, downregulation of Bcl-2, cleavage of Bid, and release of cytochrome c, which were accompanied by activation of caspases-9, -3 and -8 and subsequently poly(ADP-ribose) polymerase cleavage. Pre-incubation with caspase-8 inhibitor significantly blocked the tetrandrine-induced Bid cleavage, reduction in mitochondrial membrane potential, and activation of caspase 3, and cell death. Together, these results suggest that the mitochondrial pathway is primarily involved in tetrandrine-induced apoptosis. Additionally, our results demonstrated that tetrandrine-induced apoptosis was caused by the generation of reactive oxygen species (ROS) and most of the signaling effects were attenuated with the preincubation of cells with N-acetylcysteine, thereby further confirming the involvement of ROS in these events. Conclusions (d): Our results demonstrated that treatment of prostate cancer cells with tetrandrine induces caspase-dependent apoptosis via Fas-mediated Bid cleavage and cytochrome c release.Item TETRANDRINE: A NOVEL CHEMOPREVENTIVE AGENT(2013-04-12) Gibbs, LeePurpose: The development and study of chemopreventive agents may be promising in combating aggressive behavior of triple-negative breast cancer. Tetrandrine, a bis-benzylisoquinoline alkaloid isolated from the root of Stephania tetrandra, is a calcium channel blocker used in Chinese medicine for the treatment of silicosis and arthritis. Studies have shown that tetrandrine also has anti-tumor and anti-growth activities. Our objective is to study the effects of tetrandrine on the localization of Annexin A2 (36 kDa calcium-dependent phospholipid binding protein) and determine the implication of this in the overall cell proliferation and cancer metastasis processes. We hypothesize that inhibition of calcium trafficking by tetrandrine will inhibit the migration, invasion and proliferation via attenuation of AnxA2 localization to the plasma membrane Methods: We have used Her-2 positive, triple-negative, and non-cancerous breast cell lines (MDA-MB-231, HCC70, BT474, SKBR3, and MCF-10A) to study the effects of tetrandrine. MTT assays were carried out to determine the effect of tetrandrine on cell viability. Additionally, cells were subjected to the Versene wash to analyze the effect of calcium depletion on the AnnXA2 localization in cells. Effects of tetrandrine on the expression and localization of Annexin A2 were also analyzed by the immunofluorescence and the Western blot analyses of the sub cellular fractions of the control and treated cells. Results: The viability of the cells at various concentrations of tetrandrine after incubation of 48 h measured by MTT assay showed IC50 values of 20uM, 25uM, 30uM, 40uM, and 75uM for HCC70, MDA MB231, SKBR3, BT474, and MCF10A respectively. Annexin A2 translocation to the membrane was analyzed by using a) versene (calcium ion chelating agent) washed cells, b) Western blot analyses of membrane and cytosolic extracts and c) Immunofluorescence of control and tetrandrine treated cells. Results of these studies indicated that treatment of breast cancer cells with tetrandrine did not show any significant effect on the Annx.A2 accumulation in plasma membrane of these cells when compared with the control cells. Conclusions: Our results suggest that tetrandrine inhibits the proliferation of breast cancer cells through mechanisms independent of Annexin A2. Significantly high IC50 value for MCF10A compared to the breast cancer cells indicates that this phyto-chemical may be used as a cancer chemopreventive agent.