One-pot (2 + 4) cycloaddition of propargyl alcohols with o-hydroxyphenyl substituted secondary phosphine oxides.

The present study was undertaken to determine the roles of insulin in the growth of transplanted breast cancer in nude mice, and the proliferation and migration of MCF-7 human breast cancer cells and assess its influence on downstream signaling pathways. In a xenograft mouse model with injection of MCF-7 human breast cancer cells, tumor size was measured every other day. The insulin level and insulin receptor (IR) were increased in the breast cancer patient tissues. Insulin injected subcutaneously around the tumor site in mice caused increase in the size and weight of tumor masses, and promoted proliferation and migration of MCF-7 cells. The effects of insulin on the increase in the proliferation and migration of MCF-7 human breast cancer cells were abolished by pretreatment with the extracellular regulated kinase (ERK) inhibitor PD98059. Insulin increased the phosphorylation of ERK in the MCF-7 cells. These results indicate that insulin promotes the growth of breast cancer in nude mice, and increases the proliferation and migration of MCF-7 human breast cancer cells via the ERK pathway.

Obesity has been associated with an increased risk of postmenopausal breast cancer, which may be due to the expression of leptin. The aim of this study was to determine the role of leptin in the growth of breast cancer cells in nude mice, the proliferation and migration of MCF-7 human breast cancer cells and its downstream signaling pathway. The xenograft mouse model was elicited by injecting MCF-7 human breast cancer cells into the left back axilla and the tumor size was measured every other day. Leptin injected subcutaneously around the tumor site led to an increase in the size and weight of the tumor, whereas the leptin antagonist (LA) significantly inhibited the size and weight of the tumor. Leptin promoted the proliferation and migration of MCF-7 cells and LA inhibited it. The effects of leptin on increasing the size and weight of the tumor in the nude mice and the proliferation and migration of MCF-7 human breast cancer cells were eradicated by pretreatment with LA, the extracellular-signal regulated kinase (ERK) inhibitor PD98059. In the xenograft mouse model the leptin level was increased and leptin increased the phosphorylation of ERK in the MCF-7 cells, whereas LA significantly reduced the phosphorylation of ERK. These results indicated that leptin promotes the growth of breast cancer in the nude mice and increases the proliferation and migration of MCF-7 human breast cancer cells via the ERK pathway.

Activator of protein 1 (AP-1) is a heterodimeric transcription factor composed of various members of the Jun and Fos families and binds to DNA at specific AP-1 binding sites. AP-1 transcriptional activity is increased by phosphorylation at serine residues in the c‑Jun component of AP-1. In the present study, the proliferation of MCF-7 breast cancer cells was found to be suppressed by tamoxifen (TAM)-activated c-Jun through the protein kinase C (PKC) pathway. The molecular mechanism by which c‑Jun activation induces antiproliferative signals in estrogen receptor (ER)-positive MCF-7 human breast cancer cells remains unknown. TAM inhibited the proliferation of ER-positive MCF-7 human breast cancer cells and ER-negative MDA-MB-435 human breast cancer cells and 48 h incubation with 10 µM TAM led to inhibition of 80% of proliferation. In addition, no significant difference in c-Jun mRNA and protein levels was detected in MCF-7 and MDA-MB-435 cells stimulated by TAM for 48 h. TAM treatment of MCF-7 cells activated the transcriptional activity of AP-1, which responds specifically to phorbol ester. To determine the role of c-Jun in the antiproliferation of MCF-7 cells stimulated by TAM, the inhibition rates of MCF‑7 cells were correlated with c‑Jun expression and stimulation of TAM. Results showed that the inhibition rate of TAM-stimulated MCF-7 cells was positively regulated by overexpression of c-Jun and negatively regulated by underexpression of c-Jun. Overall, these results indicate that the TAM-stimulated antiproliferation of MCF-7 cells is positively regulated by c-Jun through activation of the PKC pathway.

High salt diet may promote progression of breast tumor through eliciting immune response

Genes encoding growth-inhibitory proteins are postulated to be candidate tumor suppressors. The identification of such proteins may benefit the early diagnosis and therapy of tumors. Here we report the cloning and functional characterization of a novel human bone marrow stromal cell (BMSC)-derived growth inhibitor (BDGI) by large scale random sequencing of a human BMSC cDNA library. Human BDGI cDNA encodes a 477-amino acid residue protein that shares high homology with rat and mouse pregnancy-induced growth inhibitors. The C-terminal of BDGI is identical to a novel human pregnancy-induced growth inhibitor, OKL38. BDGI is also closely related to many other eukaryotic proteins, which together form a novel and highly conserved family of BDGI-like proteins. BDGI overexpression inhibits the proliferation, decreases anchorage-dependent growth, and reduces migration of MCF-7 human breast cancer cells, whereas down-regulation of BDGI expression promotes the proliferation of MCF-7 and HeLa cervix epitheloid carcinoma cells. Interestingly, the inhibitory effect of BDGI on MCF-7 cells is more potent than that of OKL38. We demonstrate that BDGI induces cell cycle arrest in S phase and subsequent apoptosis of MCF-7 cells, which is likely to account for the antiproliferative effects of BDGI. This process may involve up-regulation of p27Kip1 and down-regulation of cyclin A, Bcl-2, and Bcl-xL. The inhibitory effect of BDGI on cell proliferation and the induction of apoptosis were also observed in A549 lung cancer cells but not HeLa cells. These results indicate that BDGI might be a growth inhibitor for human tumor cells, especially breast cancer cells, possibly contributing to the development of new therapeutic strategies for breast cancer.

All-trans-retinoic acid (ATRA) is well known to inhibit the proliferation of human breast cancer cells. Much less is known about the antiproliferative activity of the naturally occurring metabolites and isomers of ATRA. In the present study, we investigated the antiproliferative activity of ATRA, its physiological catabolites 4-oxo-ATRA and 5,6-epoxy-ATRA and isomers 9-cis-RA and 13-cis-RA in MCF-7 human breast cancer cells by bromodeoxyuridine incorporation. MCF-7 cells were grown in steroid- and retinoid-free medium supplemented with growth factors. Under these culture conditions, ATRA and its naturally occurring catabolites and isomers showed significant antiproliferative activity in MCF-7 cells in a concentration-dependent manner (10[-11] M to 10[-6] M). The antiproliferative activity of ATRA catabolites and isomers was equal to that of the parent compound ATRA at concentrations of 10(-8) M and 10(-7) M. Only at 10(-6) M were the catabolites and the stereoisomer 13-cis-RA less potent. The stereoisomer 9-cis-RA was as potent as ATRA at all concentrations tested (10[-11] M to 10[-6] M). In addition, we show that the catabolites and isomers were formed from ATRA to only a limited extent. Together, our findings suggest that in spite of their high antiproliferative activity the catabolites and isomers of ATRA cannot be responsible for the observed growth inhibition induced by ATRA.

Objective: FHL2 was previously identified to be a novel interacting factor of Id family proteins. The aim of this study was to investigate, the effects of FHL2 on Id1-mediated transcriptional regulation activity and its oncogenic activity in human breast cancer cells. Methods: Cell transfection was performed by Superfect reagent. Id1 stably overexpressed MCF-7 cells was cloned by G418 screening. The protein level of Id1 was detected by western blot analysis. Dual relative luciferase assays were used to measure the effect of E47-mediated transcriptional activity in MCF-7 human breast cancer cells. MTT assay was used to measure cell proliferation. Transwell assay was used to measure the invasive capacity of MCF-7 cancer cells. Results: The basic helix-loop-helix (bHLH) factor E47-mediated transcription activity was markedly repressed by Id1 in MCF-7 cells. This Id1-mediated repression was effectively antagonized by FHL2 transduction. Overexpression of Id1 markedly promoted the proliferation rate and invasive capacity of MCF-7 cells; however, these effects induced by Id1 were significantly suppressed by overexpression of FHL2 in cells. Conclusion: FHL2 can inhibit the proliferation and invasiveness of human breast cancer cells by repressing the functional activity of Id1. These findings provide the basis for further investigating the functional roles of FHL2-Id1 signaling in the carcinogenesis and development of human breast cancer.

Complement system activation contributes to various immune and inflammatory diseases, as well as cancers.However, the role of complement activation in the proliferation of cancer cells is not clear. In the present study, we investigated the consequences of complement activation on the proliferation of breast cancer cells and its possible mechanisms. We focused our study on the potential roles of the anaphylatoxins C3a and C5a in the proliferation of human breast cancer, as two important immune mediators generated after complement activation. Our study revealed that C5a stimulation, but not C3a, enhanced the proliferation of human breast cancer cells in vitro. Moreover, the expression of response gene to complement 32 (RGC-32) was pronounced in breast cancer cells in response to C5a stimulation. Notably, blockade of the C5a receptor markedly reduced the expression of RGC-32 and the proliferation of breast cancer cells stimulated by C5a. Meanwhile, silencing of RGC-32 expression reduced the proliferation of breast cancer cells induced by C5a treatment. Further investigation revealed that Akt activation was involved in C5a-induced RGC-32 expression and breast cancer cell proliferation. In conclusion, the present study indicates that C5a may promote the proliferation of breast cancer cells through Akt1 activation of the RGC-32 gene.

Inhibitory effects of 5-heptadecylresorcinol on the proliferation of human MCF-7 breast cancer cells through modulating PI3K/Akt/mTOR pathway

The discovery of the amphibian skin tetradecapeptide bombesin led to the identification of several mammalian bombesin-like neuroendocrine peptides such as gastrinreleasing peptides (GRP'-27 and GRP8-27) and neuromedin B. GRP and neuromedin B are products of two distinct genes (Krane et al., 1988). Bombesin and its homologues stimulate gastric acid secretion and cause release of various peptide hormones (Dockray, 1987). Bombesin is also mitogenic, causing gastrin cell hyperplasia in rats (Lezoche et al., 1981) and stimulating proliferation of 3T3 murine fibroblasts (Rozengurt & Sinnett-Smith, 1980) and normal human bronchial epithelial cells (Willey et al., 1984) in vitro. Bombesin-like peptides may act as autocrine mitogens for small cell carcinoma of the lung (SCCL); SCCL lines secrete bombesinlike immunoreactivity (BLI) and are stimulated to proliferate by exogenous bombesin (Cuttita et al., 1985). Some SCCL lines are, however, insensitive to exogenous bombesin and do not express detectable GRP receptors (Kado-Fong & Malfroy, 1989). BLI has been found in rat mammary tumours (Gaudino et al., 1984) and in a small proportion of human breast cancer and breast carcinoid specimens (Foster & Tan, 1984; McKillop et al., 1988; Nesland et al., 1985) but is undetectable in normal breast tissue (Bostwick & Bensch, 1985). Recently, it was shown that both bombesin and GRP stimulate inositol phospholipid hydrolysis and Ca2 efflux in MCF-7 and T47D human breast cancer cells suggesting a role in mitogenic signalling (Patel & Schrey, 1990). BLI has been detected in MCF-7 and BT-20 breast cancer cell pellets (Weber et al., 1989). This suggests that a BLI-autocrine stimulatory loop may operate in some breast cancer cell lines such as is the case for SCCL cells. In the light of these observations we have investigated the effects of bombesin on the proliferation of breast cancer cells in culture. The cell lines examined were oestrogen-dependent (MCF-7), oestrogen-responsive (ZR-75-1, T47D) (Dickson & Lippman, 1986), and oestrogenindependent (MDA-MB-436) (Clarke et al., 1983). ZR-75-1, T47D and MDA-MB-436 human breast cancer cells were obtained from the European Collection of Animal Cell Cultures (Porton Down, UK), MCF-7 cells (originally from Dr M.E. Lippman, Georgetown, USA) were a gift from Dr C.R. Green, Liverpool University. All tissue culture media and foetal calf serum (FCS) were obtained from Flow Labs, Irvine, Scotland, UK. The same reserved batch of FCS was used throughout this investigation and all cells were routinely passaged in the presence of phenol red. ZR-75-1 cells were cultured in RPMI 1640 medium containing 5% FCS; MCF-7 in Eagle's minimal essential medium (MEM) with 5% FCS; T47D cells in DMEM with 10% FCS; and MDA-MB-436 cells in Liebowitz-15 medium with 10% FCS. Bombesin (Bachem, Bubendorf, Switzerland) stock solutions, made up in Earle's balanced salts solution containing 0.1% bovine serum albumin, were gassed with nitrogen to prevent oxidation and were stored frozen in liquid nitrogen. Heatand charcoal-treated foetal calf serum (DCC-FCS) was prepared by heating 100 ml of FCS, 10 g acid-washed NoritA activated charcoal and 1 g Dextran T40 at 53'C for 1 h (charcoal and dextran previously stirred at 4'C overnight). The suspension was centrifuged and then filtered through a 0.45 iLm membrane filter and finally filter-sterilised through a 0.22 pm membrane filter. To study effects of the peptides on proliferation, cells were inoculated into 24-well cluster plates (Costar, Northumbria Biologicals, UK) at 1 x 104 (MDA-MB-436) or 4 x 104 (ZR75-1, T47D, MCF-7) cells per well and incubated at 37'C in a humidified atmosphere containing 5% CO2. After 24 h, plating efficiency was determined in 8 replicate wells by electronic particle counting (Coulter counter model ZBI) of trypsinised cells. Medium was removed from the remaining wells and replaced either with medium containing 5% or 10% serum or with the same medium containing peptide, as indicated. Cells from replicate wells were detached by treatment with trypsin and counted at the times indicated; experimental and control media were either unchanged during the course of the experiment or were replaced on days 3 and 6 of incubation. In serum-free conditions, 0. 1% bovine serum albumin replaced the serum component. All growth studies were confirmed in at least three independent determinations. Bombesin did not stimulate cell proliferation of any of the lines in the presence of untreated FCS. The growth of ZR75-1 and T47D cells, however, was significantly and consistently stimulated above control in the presence of DCC-FCS (Figures 1 and 2). There were slight interexperimental variations in the effective dose-response ranges for each cell line but, under the culture conditions described, significant stimulation of cell division was always found with bombesin in the picomolar range. Bombesin stimulates proliferation of ZR-75-1 cells grown in the presence of 5% DCC-FCS, producing significant and increasing elevations above control from day 3 to day 9 of incubation (Figure la). Figure lb shows a typical dose-

To investigate the effects of 50% ethyl alcohol (EtOH) extracts from Danzhi Xiaoyao Pill (, DXP) on the proliferation of MCF-7 human breast cancer cells and potential mechanisms. ATP-Lite assay was performed to test the proliferation of the MCF-7 breast cancer cell line; and antioxidant activity was measured by the oxygen radical absorbance capacity (ORAC). The effects of DXP on nitric oxide (NO) production were tested by lipopolysaccharide (LPS)-stimulated RAW 264.7 murine macrophages using the Griess reaction. The 50% EtOH DXP extracts displayed a cytotoxic response on MCF-7 cells at 0.10, 0.25 and 0.50 mg/mL dose-dependently with the proliferation inhibited by more than 85%. The ORAC value of the DXP was 820 micro moL Trolox equivalent/g, about 40% of the vitamin C value. DXP extracts had significant inhibitory effect on NO production at the concentration from 0.0625 mg/mL to 0.5 mg/mL (P<0.05, P<0.01). The extracts of DXP could significantly inhibit the proliferation of MCF-7 cells, with the effect possibly related to its antioxidant activity and the inhibition of NO production.

Visfatin stimulates proliferation of MCF-7 human breast cancer cells.

Polychlorinated biphenyls and their hydroxylated metabolites become more and more concerned for their endocrine disruption. At the presented study, experiment was designed to obtain the disruption effect of PCB101 and its hydroxylated metabolites . The proliferation of MCF-7 human breast cancer cells was detected at different exposed concentration and time period. MCF-7 cells were cultured with the concentration of PCB101 and twor of its hydroxylated metabolites at 5×10-95×10-85×10-75×10-6mol/L. And the concentrations of 17β-Estradiol (E2) were2×10-92×10-82×10-72×10-6mol/L respectively. And the proliferation rate was detected after 24h,48h, 72h, 96h exposure.Ethanol was used as solvent control.The results indicated that PCB101 and its hydroxylated metabolites had no effect on MCF-7 cell proliferation at designed concentrations and exposure time.

To observe the effects of arecoline on proliferation and apoptosis of MCF-7 human breast cancer cells and to ex-plore its possible mechanism. Human breast cancer MCF-7 cells were treated with arecoline at the concentrations of 0,10,30,50, 100,300,500μmol/L, the cell proliferation were detected by MTT assay, cell apoptosis were analyzed by Hoechst 33342 staining and flow cy-tometry, the protein expression of Bax,Bcl-2 and P53 were detected by Western blot. Low concentration(0,10,30, 50 μmol/L) arecoline had no effect on the proliferation and apoptosis of MCF-7. However, high concentration(100,300,500μmol/L) arecoline inhibited proliferation and induced apoptosis of MCF-7 cells in a concentration-dependent manner, arecoline also significantly increased P53 and Bax protein expression and decreased Bcl-2 protein expression. High concentration arecoline inhibited the proliferation and induced the apoptosis of MCF-7 cells, the mechanism was probably corrected with increasing P53 and Bax protein expression and decreasing Bcl-2 pro-tein expression.

Lithium, which is used to treat bipolar psychiatric disorders, can stimulate proliferation of a number of cells in tissue culture. Proliferation of MCF-7 human breast cancer cells, which also respond to EGF and estrogens, was stimulated by LiCl (1-5 mM) within the concentration range that is encountered during human therapy with lithium. Stimulation of growth was specific for lithium; rubidium, potassium, and sodium showed no such effect. In the presence of antiestrogen, lithium stimulated the growth of hormone-dependent breast cancer cells MCF-7, ZR-75-1, and T47D but not hormone-independent MDA-MB-231 cells or an estrogen-independent clone of MCF-7 cells. Lithium-stimulated proliferation was limited by cytotoxicity which could be moderated by added potassium chloride (5-20 mM) in the medium. Each of the mitogens lithium, 17 beta-estradiol, and EGF increased the rate of uptake of myo-inositol into MCF-7 cells. Whether normalized to inositol lipids, to protein, or to DNA, steady-state levels of inositol phosphates were elevated by each of the mitogens including lithium, which inhibits the breakdown of inositol phosphates in the phosphoinositide signaling pathway. These data indicate that therapeutic concentrations of lithium can stimulate the proliferation of human breast cancer cells by a mechanism that may involve the phosphoinositide pathway.