Antiproliferative activity of aucubin is through cell cycle arrest and apoptosis in human non-small cell lung cancer A549 cells

Aucubin, an iridoid glycoside isolated from the leaves of Aucuba japonica, inhibits human non-small cell lung cancer A549 cells by blocking cell cycle progression in the G(0)/G(1) phase and inducing apoptosis. An ELISA showed that the G(0)/G(1) phase arrest is due to p53-mediated induction of p21. Enhancement of Fas and its two ligands, membrane-bound and soluble Fas ligand, may be responsible for the apoptotic effect induced by aucubin. The present study shows, for the first time, that the induction of p53 and activity of the Fas/Fas ligand apoptotic system may participate in the antiproliferative activity of aucubin in A549 cells.     

Induction of cell cycle arrest and apoptosis in human non-small cell lung cancer A549 cells by casuarinin from the bark of Terminalia arjuna Linn

Casuarinin, a hydrolyzable tannin isolated from the bark of Terminalia arjuna Linn. (Combretaceae), inhibits human non-small cell lung cancer A549 cells by blocking cell cycle progression in the G0/G1 phase and inducing apoptosis. Enzyme-linked immunosorbent assay showed that the G0/G1 phase arrest is due to p53-dependent induction of p21/WAF1. An enhancement in Fas/APO-1 and the two forms of Fas ligand (FasL), membrane-bound FasL and soluble FasL, might be responsible for the apoptotic effect induced by casuarinin. Our study reports here for the first time that the induction of p53 and the activity of the Fas/FasL apoptotic system may participate in the antiproliferative activity of casuarinin in A549 cells.     

The antiproliferative activity of prodelphinidin B-2 3'-O-gallate from green tea leaf is through cell cycle arrest and Fas-mediated apoptotic pathway in A549 cells.

Prodelphinidin B-2 3'-O-gallate, a proanthocyanidin gallate isolated from green tea leaf, was investigated for its anti-proliferative activity in human non-small cell lung cancer A549 cells. The results showed that prodelphinidin B-2 3'-O-gallate inhibited the proliferation of A549 cells with no detectable toxic effects on normal WI-38 cells as measured by the XTT assay. Flow cytometric analysis showed that prodelphinidin B-2 3'-O-gallate blocked cell cycle progression in the G0/G1 phase. In addition, prodelphinidin B-2 3'-O-gallate effectively induced A549 cell apoptosis as determined by assessing the nucleosome level in cytoplasm. Enzyme-linked immunosorbent assay showed that the G0/G1 phase arrest is due to p53-independent induction of p21/WAF1. An enhancement in Fas/APO-1 and its two form ligands, membrane-bound Fas ligand (mFasL) and soluble Fas ligand (sFasL), might be responsible for the apoptotic effect induced by prodelphinidin B-2 3'-O-gallate. We suggested that prodelphinidin B-2 3'-O-gallate's activities might be potentially contribute to its overall chemopreventive effects against lung cancer, and can possibly be considered for future therapeutic application.     

Antiproliferative activity of paeoniflorin is through cell cycle arrest and the Fas/Fas ligand-mediated apoptotic pathway in human non-small cell lung cancer A549 cells

1. Paeoniflorin (PF), isolated from the paeony root, is reported to have immunoregulatory, neuromuscular blocking, anticonvulsant, antihyperglycaemic and antihypotensive effects. 2. The present study investigated the antiproliferative activity of PF. The results showed that PF inhibited the proliferation of A549 by blocking cell cycle progression in the G(0)/G(1) phase and inducing apoptosis. 3. An ELISA showed that G(0)/G(1) phase arrest may be due to p53-independent induction of p21/wild-type p53-activated fragment 1 (WAF1). Increased protein expression of Fas/apoptosis-1 (APO-1) and its two ligands, membrane-bound Fas ligand and soluble Fas ligand, may be responsible for the PF-induced apoptosis. 4. This is the first study to show that the induction of p21/WAF1 and the activity of the Fas/Fas ligand apoptotic system may participate in the antiproliferative activity of PF in A549 cells.     

Isoliquiritigenin inhibits the proliferation and induces the apoptosis of human non-small cell lung cancer a549 cells

1. Isoliquiritigenin (ISL) is a natural pigment with the simple chalcone structure 4,2',4'-trihydroxychalcone. In the present study, we report, for the first time, ISL-induced inhibition of the proliferation of the human non-small cell lung cancer A549 cell line. 2. The results showed that ISL not only inhibited A549 cell proliferation, but also induced apoptosis and blocked cell cycle progression in the G1 phase. An ELISA assay demonstrated that ISL significantly increased the expression of p53 and p21/WAF1 protein, which caused cell cycle arrest. 3. An enhancement in Fas and its two ligands, namely membrane-bound Fas ligand (mFasL) and soluble Fas ligand (sFasL), may be responsible for the apoptotic effect induced by ISL. 4. Taken together, the results indicate that the p53 and Fas/FasL apoptotic system may participate in the antiproliferative activity of ISL in A549 cells.     

Upregulation of Fas/Fas ligand-mediated apoptosis by gossypol in an immortalized human alveolar lung cancer cell line

Gossypol has wide antineoplastic effects in vitro, but its effects on human lung cancer have not been explored. To evaluate the activity of gossypol against alveolar cell lung cancer and to provide information on the mechanism of action, we examined the effects of gossypol on the proliferation of A549 cells indirectly using an XTT assay and on the distribution of cells within the phases of the cell cycle using flow cytometry. We also examined several factors that may affect apoptosis, including p53, p21/WAF1, Fas receptor, Fas ligand (FasL) and caspase 8 activity. The results showed that gossypol inhibited proliferation of A549 cells at a concentration of 0.5 micromol/L after 12 h treatment. The effect was both dose- and time-dependent by the induction of apoptosis without the effect of p53 and p21/WAF1. Upregulation of Fas/FasL, in association with the activation of downstream caspase 8 activity, was observed following treatment with gossypol. The Fas/FasL pathway accounted for 75% of gossypol-mediated apoptosis. We suggest that the Fas/FasL apoptotic system is the major pathway for gossypol-mediated apoptosis of A549 cells. Gossypol had no effect on the distribution of A549 cells within the phases of the cell cycle. In conclusion, gossypol inhibited A549 cells mainly by induction of the Fas/FasL apoptotic pathway, but not the p53 and p21/WAF1 pathway.     

Acacetin inhibits the proliferation of Hep G2 by blocking cell cycle progression and inducing apoptosis

Flavonoids are a broadly distributed class of plant pigments, universally present in vascular plants and responsible for much of the coloring in nature. They are strong antioxidants that occur naturally in foods and can inhibit carcinogenesis in rodents. In this study, we examined acacetin (5,7-dihydroxy-4'-methoxyflavone), a flavonoid compound, for its effect on proliferation in a human liver cancer cell line, Hep G2. The results showed that acacetin inhibited the proliferation of Hep G2 by inducing apoptosis and blocking cell cycle progression in the G1 phase. Enzyme-linked immunosorbent assay showed that acacetin significantly increased the expression of p53 and p21/WAF1 protein, contributing to cell cycle arrest. An enhancement in Fas/APO-1 and its two form ligands, membrane-bound Fas ligand and soluble Fas ligand, as well as Bax protein, was responsible for the apoptotic effect induced by acacetin. Taken together, our study suggests that the induction of p53 and activity of the Fas/Fas ligand apoptotic system may participate in the antiproliferative activity of acacetin in Hep G2 cells.     

Casuarinin from the bark of Terminalia arjuna induces apoptosis and cell cycle arrest in human breast adenocarcinoma MCF-7 cells

Casuarinin, a hydrolyzable tannin isolated from the bark of Terminalia arjuna L. (Combretaceae), was investigated for its antiproliferative activity in human breast adenocarcinoma MCF-7 cells. The results showed that casuarinin inhibited the proliferation of MCF-7 by blocking cell cycle progression in the G0/G1 phase and inducing apoptosis. An enzyme-linked immunosorbent assay showed that casuarinin increased the expression of p21/WAF1 concomitantly as the MCF-7 cells underwent G0/G1 arrest. An enhancement in Fas/APO-1 and its two forms of ligands, membrane-bound Fas ligand (mFasL) and soluble Fas ligand (sFasL), might be responsible for the apoptotic effect induced by casuarinin. Our study reports here for the first time that the induction of p21/WAF1 and the activity of Fas/Fas ligand apoptotic system may participate in the antiproliferative activity of casuarinin in MCF-7 cells.     

Putranjivain A from Euphorbia jolkini inhibits proliferation of human breast adenocarcinoma MCF-7 cells via blocking cell cycle progression and inducing apoptosis

Putranjivain A, isolated from the whole plant of Euphorbia jolkini Bioss (Euphorbiaceae), was investigated for its antiproliferative activity in human breast adenocarcinoma MCF-7 cells. The results showed that putranjivain A inhibited the proliferation of MCF-7 by blocking cell cycle progression in the G0/G1 phase and inducing apoptosis. Enzyme-linked immunosorbent assay showed that putranjivain A increased the expression of p21/WAF1 concomitantly as MCF-7 cell underwent G0/G1 arrest. An enhancement in Fas/APO-1 and its two forms of ligands, membrane-bound Fas ligand (mFasL) and soluble Fas ligand (sFasL), might be responsible for the apoptotic effect induced by putranjivain A. Our study reports here for the first time that the induction of p21/WAF1 and the activity of Fas/Fas ligand apoptotic system may participate in the antiproliferative activity of putranjivain A in MCF-7 cells.     

Central role of mitochondria and p53 in PUVA-induced apoptosis in human keratinocytes cell line NCTC-2544

Despite strong evidence concerning the high efficiency of PUVA therapy (psoralen plus UVA light), its mechanism of action has not yet been fully elucidated. In this study, we have evaluated in a cell line of human keratinocytes (NCTC-2544) the effects of two linear psoralen derivatives, 8-methoxypsoralen (8-MOP) and 5-methoxypsoralen (5-MOP), that are widely used in PUVA therapy and two angular derivatives, Angelicin (ANG) and 4,6,4'-trymetyl angelicin (TMA). All derivatives photoinduce cellular death, TMA being the most active compound. The cell cycle analysis showed that the four derivatives induce, 24 h after irradiation, a cell cycle arrest in G1 phase later followed by massive apoptosis. The G1 arrest is correlated to an increase in the expression of p21(Waf1/Cip1), a protein associated with the cell cycle block and apoptosis. Furthermore, treatment of NCTC-2544 resulted in p53 activation by 5-MOP, 8-MOP, and ANG but not TMA and its phosphorylation at serine-15. The levels of p21(Waf1/Cip1) paralleled p53 protein staining pattern suggesting that p53 activation correlated with p21(Waf1/Cip1) induction. Simultaneous to p53 activation, psoralens induced mitochondrial depolarization, cytochrome c release, mitochondrial production of reactive oxygen species, as well as caspase-3 and -9 activation. Thus these results strongly indicate the necessity of p53 activation and the induction of the apoptotic machinery downstream of mitochondria.     

B1, a novel topoisomerase II inhibitor, induces apoptosis and cell cycle G1 arrest in lung adenocarcinoma A549 cells

In our previous studies, we demonstrated that 2,6-bis-(2-chloroacetamido) anthraquinone (B1) showed a highly significant cytotoxic effect. However, its influence in the cell cycle and apoptotic induction effects has not been investigated yet. Here we report the antiproliferative effect of B1, for which IC50 values were 0.57 μmol/l for lung cancer A549 cells, 0.63 μmol/l for colon cancer HT-29 cells, and 0.53 μmol/l for breast cancer MCF-7 cells. DNA topoisomerase II (Topo II), an essential enzyme in DNA synthesis and meiotic division, is highly expressed in cancer cells. Some currently used clinical anticancer drugs (doxorubicin and mitoxantrone) targeting Topo II are very effective antineoplastic agents. B1, sharing the basic structure of known Topo II inhibitors, demonstrated a significant inhibitory effect on Topo II bioactivity. In A549 cells, B1 increased apoptotic cell population with induction of Fas, Bax, and cleaved poly(ADP-ribose) polymerase and by reduction of Bcl-2 expression. Moreover, cell cycle analysis indicated that B1 induced G1 phase arrest through modulation of G1 cell cycle regulatory proteins, such as the downregulation of cyclin D1 and upregulation of Cip/p21, Kip1/p27, and p53. Thus, our study suggests that B1, with the ability to inhibit Topo II activity and cause cell cycle G1 arrest and apoptosis, has potential as a novel anticancer agent.     

Tetrazolium Violet Induced Apoptosis and Cell Cycle Arrest in Human Lung Cancer A549 Cells

Tetrazolium violet is a tetrazolium salt and has been proposed as an antitumor agent. In this study, we reported for the first time that tetrazolium violet not only inhibited human lung cancer A549 cell proliferation but also induced apoptosis and blocked cell cycle progression in the G1 phase. The results showed that tetrazolium violet significantly decreased the viability of A549 cells at 5-15 μM. Tetrazolium violet -induced apoptosis in A549 cells was confirmed by {"type":"entrez-nucleotide","attrs":{"text":"H33258","term_id":"978675","term_text":"H33258"}}H33258 staining assay. In A549, tetrazolium violet blocked the progression of the cell cycle at G1 phase by inducing p53 expression and further up-regulating p21/WAF1 expression. In addition, an enhancement in Fas/APO-1 and its two forms of ligands, membrane-bound Fas ligand (mFasL) and soluble Fas ligand (sFasL), as well as caspase, were responsible for the apoptotic effect induced by tetrazolium violet. The conclusion of this study is that tetrazolium violet induced p53 expression which caused cell cycle arrest and apoptosis. These findings suggest that tetrazolium violet has strong potential for development as an agent for treatment lung cancer.     

Cell survival, cell death and cell cycle pathways are interconnected: implications for cancer therapy.

The partial cross-utilization of molecules and pathways involved in opposing processes like cell survival, proliferation and cell death, assures that mutations within one signaling cascade will also affect the other opposite process at least to some extent, thus contributing to homeostatic regulatory circuits. This review highlights some of the connections between opposite-acting pathways. Thus, we discuss the role of cyclins in the apoptotic process, and in the regulation of cell proliferation. CDKs and their inhibitors like the INK4-family (p16(Ink4a), p15(Ink4b), p18(Ink4c), p19(Ink4d)), and the Cip1/Waf1/Kip1-2-family (p21(Cip1/Waf1), p27(Kip1), p57(Kip2)) are shown both in the context of proliferation regulators and as contributors to the apoptotic machinery. Bcl2-family members (i.e. Bcl2, Bcl-X(L) Mcl-1(L); Bax, Bok/Mtd, Bak, and Bcl-X(S); Bad, Bid, Bim(EL), Bmf, Mcl-1(S)) are highlighted both for their apoptosis-regulating capacity and also for their effect on the cell cycle progression. The PI3-K/Akt cell survival pathway is shown as regulator of cell metabolism and cell survival, but examples are also provided where aberrant activity of the pathway may contribute to the induction of apoptosis. Myc/Mad/Max proteins are shown both as a powerful S-phase driving complex and as apoptosis-sensitizers. We also discuss multifunctional proteins like p53 and Rb (RBL1/p107, RBL2/p130) both in the context of G1-S transition and as apoptotic triggers. Finally, we reflect on novel therapeutic approaches that would involve redirecting over-active survival and proliferation pathways towards induction of apoptosis in cancer cells.     

Norsolorinic acid inhibits proliferation of T24 human bladder cancer cells by arresting the cell cycle at the G0/G1 phase and inducing a Fas/membrane-bound Fas ligand-mediated apoptotic pathway

1. Norsolorinic acid, isolated from Aspergillus nidulans, has been shown to have antiproliferative activity in T24 human bladder cancer cells by arresting the cell cycle at the G(0)/G(1) phase and inducing apoptosis. The aim of the present study was to investigate the antiproliferative activity of norsolorinic acid in T24 human bladder cancer cells. 2. The effects of norsolorinic acid (1, 5, 10 and 20 micromol/L) on the proliferation of T24 cells and on the distribution of cells within different phases of the cell cycle were investigated indirectly using an XTT assay and a flow cytometer, respectively. Factors affecting the cell cycle and apoptosis, including p53, p21, Fas receptor, Fas ligand (FasL) and caspase 8 activity, were examined using ELISA. 3. The results showed that norsolorinic acid inhibited proliferation of T24 cells in a dose-dependent manner, with an IC(50) of 10.5 micromol/L. The effect involved the induction of cell cycle arrest at the G(0)/G(1) phase and apoptosis. 4. These results demonstrate that G(0)/G(1) phase arrest is due to increased expression of p21 in cells treated with norsolorinic acid (10 and 20 micromol/L) for 24 h. Moreover, enhanced Fas and membrane-bound Fas ligand (mFasL) may be responsible for the apoptotic effect of norsolorinic acid. Thus, the present study reports, for the first time, that induction of p21 and the Fas/mFas ligand apoptotic system may participate in the antiproliferative action of norsolorinic acid in T24 human bladder cancer cells.     

Mechanisms of induction of cell cycle arrest and cell death by cryptolepine in human lung adenocarcinoma a549 cells.

We investigated p53-dependent and -independent molecular events associated with cell cycle alteration and cell death in human lung adenocarcinoma A549 cells using cryptolepine, a DNA-damaging agent. After a 24-h treatment, cryptolepine caused an accumulation of p53 at concentrations of 1.25-10 microM and induction of p21(Cip1/WAF1) but only at concentrations up to 5muM. p21(Cip1/WAF1) was also strongly induced by cryptolepine (2.5-5 microM) in cells with p53 largely ablated via small interfering RNA-mediated gene silencing. Cryptolepine induced G1-phase block at 1.25-2.5 microM, S-phase and G2/M-phase block at 2.5-5 microM, and cell death at 10 microM. The dead cells displayed condensed and fragmented nuclei, features of apoptosis. Wortmannin, an inhibitor of ataxia telangiectasia-mutated and DNA-dependent protein kinase (DNA-PK), caused cell cycle arrest at G1 phase without inducing p53 and p21(Cip1/WAF1) expression and cell death. The addition of wortmannin partially prevented cryptolepine-induced expression of p53 and p21(Cip1/WAF1) together with the S-phase block and sensitized cells to induction of cell death. NU7026, a DNA-PK-specific inhibitor, showed neither induction of cell cycle arrest and apoptosis nor the expression of p53 and p21(Cip1/WAF1). The presence of NU7026 caused further reduction of cells in G1 phase induced by cryptolepine at 5 microM without affecting the induction of p53 and p21(Cip1/WAF1) and cell death. This study using the A549 cell as a model demonstrated that cryptolepine selects different molecular pathways to cell cycle checkpoint activation in a dose-specific manner and evokes a wortmannin-sensitive antiapoptosis response.     

Cell cycle- and protein kinase C-specific effects of resiniferatoxin and resiniferonol 9,13,14-ortho-phenylacetate in intestinal epithelial cells

We have previously reported that protein kinase C (PKC) signaling can trigger hallmark events of cell cycle withdrawal in intestinal epithelial cells, including downregulation of cyclin D1, induction of p21(Waf1/Cip1), and activation of the growth suppressor function of pocket proteins. In the current study, we compared the cell cycle- and PKC-specific effects of the vanilloid resiniferatoxin (RTX), its parent diterpene resiniferonol 9,13,14-ortho-phenylacetate (ROPA), and the PKC agonist PMA in the IEC-18 non-transformed intestinal crypt cell line. ROPA and PMA were found to produce strikingly similar alterations in cell cycle progression and PKC activity in IEC-18 cells, although PMA was approximately 1000-fold more potent in producing these effects. Both agents induced a transient PKC-dependent blockade in G1---> S progression associated with transient downregulation of cyclin D1 and induction of p21(Waf1/Cip1). In contrast, RTX produced a prolonged PKC-independent cell cycle arrest in G(0)/G(1) phase which was maintained for longer than 24h. This arrest was vanilloid receptor-independent and associated with prolonged downregulation of cyclin D1 mRNA and protein, with little effect on levels of p21(Waf1/Cip1). Combined exposure to RTX and ROPA produced a sustained and complete cell cycle blockade in IEC-18 cells, associated with depletion of cyclin D1 and sustained enhancement of p21(Waf1/Cip1) levels. PMA, ROPA, RTX and the RTX/ROPA combination were capable of activating ERK1/2 signaling in IEC-18 cells, albeit with different kinetics. In contrast, only PMA and ROPA activated JNK1/2 and p38 in this system. Notably, some preparations of commercially obtained RTX produced effects indistinguishable from those of the RTX/ROPA combination, suggesting that certain batches of the compound may contain significant amounts of ROPA (or another PKC agonist activity). Together, these data demonstrate that structurally related compounds can produce similar cell cycle-specific effects but through distinct mechanisms. In addition, they add to a growing body of evidence that vanilloids can have antiproliferative effects in a variety of cell types.     

Norsolorinic acid from Aspergillus nidulans inhibits the proliferation of human breast adenocarcinoma MCF-7 cells via Fas-mediated pathway

Norsolorinic acid, isolated from the Aspergillus nidulans, was investigated for its antiproliferative activity in human breast adenocarcinoma MCF-7 cells. To identity the anticancer mechanism of norsolorinic acid, we assayed its effect on apoptosis, cell cycle distribution, and levels of p53, p21/WAF1, Fas/APO-1 receptor and Fas ligand. The results showed that norsolorinic acid induced apoptosis of MCF-7 cells without mediation of p53 and p21/WAF1. We suggest that Fas/Fas ligand apoptotic system is the main pathway of norsolorinic acid-mediated apoptosis of MCF-7 cells. Our study reports here for the first time that the activity of the Fas/Fas ligand apoptotic system may participate in the antiproliferative activity of norsolorinic acid in MCF-7 cells.