Toxicity Evaluation of Geniposide on MCF-7 Cancer Cells

As one of the adjuvant treatments for cancer treatment, traditional Chinese medicine treatment has a wide range of cancer treatments, such as preventing metastasis and relapse, improving the efficacy of radiotherapy and chemotherapy, reducing the side effects of chemotherapy, improving body function, extending life and improving the life quality. Geniposide (GEN) is a bioactive substance extracted from the fruit of gardenia. In recent years, it has attracted attention due to its anti-tumor effect. In this study, we aimed to investigate whether GEN could inhibit the proliferation of human breast cancer cells (MCF-7) and promote its apoptosis. The half-inhibitory concentration (IC50) values of GEN were firstly determined as 16.06, 14.85 and 13.14 mg/mL by the CCK-8 experiment after cells treated for 24 h, 48 h, and 72 h, respectively. The inhibitory effect of GEN on MCF-7 cells was in concentration- and time-dependent manners from the results of CCK-8 experiment and Live/Dead staining. AO/EB staining result has shown that GEN has induced MCF-7 cell apoptosis.     

Migratory activity of human breast cancer cells is modulated by differential expression of xanthine oxidoreductase

Xanthine oxidoreductase (XOR) may exert an important, but poorly defined, role in the pathogenesis of breast cancer (BC). Loss of XOR expression was linked to aggressive BC, and recent clinical observations have suggested that decreasing XOR may be functionally linked to BC aggressiveness. The goal of the present investigation was to determine whether the decreased XOR observed in clinically aggressive BC was an intrinsic property of highly invasive mammary epithelial cells (MEC). Expression of XOR was investigated using HC11 mouse MEC, HB4a and MCF-10A normal human MEC, and several human mammary tumor cells including MCF-7 and MDA-MB-231. Consistent with clinical observations, data shown here revealed high levels of XOR in normal HC11 and MCF-10A cells that was markedly reduced in highly invasive mammary tumor cells. The contribution of XOR to tumor cell migration in vitro was investigated using MDA-MB-231 and MCF-7 cells and clonally selected derivatives of HC11 that exhibit either weak or strong migration in vitro. We observed that over-expression of an XOR cDNA in MDA-MB-231 and in HC11-C24, both possessing weak XOR expression and high migratory capacity, inhibited their migration in vitro. Conversely, pharmacological inhibition of XOR in MCF-7 and HC11-C4, both possessing high XOR expression and weak migratory capacity, stimulated their migration in vitro. Further experiments suggested that XOR derived ROS mediated this effect and also modulated COX-2 and MMP levels and function. These data demonstrate a functional link between XOR expression and MEC migration and suggest a potential role for XOR in suppressing BC pathogenesis.     

Uterine phenotype of young adult rats exposed to dietary soy or genistein during development

Dietary soy intake is associated with protection from breast cancer, but questions persist on the potential risks of the major soy isoflavone genistein (GEN) on female reproductive health. Here, we evaluated intermediate markers of cancer risk in uteri of cycling, young adult Sprague-Dawley rats lifetime exposed to one of three AIN-93G semipurified diets: casein (CAS), soy protein isolate (SPI+ with 276 mg GEN aglycone equivalents/kg) and CAS+GEN (GEN at 250 mg/kg). Postnatal day 50 (PND50) rats lifetime exposed to GEN or SPI+ had similar uterine luminal epithelium height, myometrial thickness, endometrial gland numbers, endometrial immunoreactive proliferating cell nuclear antigen (PCNA), and serum estrogen and progesterone, as CAS-fed rats. GEN-fed rats showed modestly increased apoptosis in uterine glandular epithelium, compared to those of CAS- or SPI+-fed groups. Diet had no effect on the uterine expression of genes for the tumor suppressors PTEN, p53 and p21, and the apoptotic-associated proteins Bcl2, Bax and progesterone receptor. Uterine tissue and serum concentrations of total GEN were higher in rats fed GEN than in those fed SPI+. Human Ishikawa endocarcinoma cells treated with GEN-fed rat serum tended to exhibit increased apoptotic status than those treated with CAS-fed rat serum. Exogenously added GEN (0.2 and 2 microM) increased, while estradiol-17beta (0.1 microM) decreased Ishikawa cell apoptosis, relative to untreated cells. Results suggest that lifetime dietary exposure to soy foods does not alter uterine cell phenotype in young adult rats, while GEN, by enhancing uterine endometrial glandular apoptosis in vivo and in vitro, may confer protection against uterine carcinoma. Given its limited influence on uterine phenotype of young adult females, GEN, when taken as part of soy foods or as supplement, should be favorably considered for other potential health benefits.     

Identification of proteins secreted from leptin stimulated MCF-7 breast cancer cells: a dual proteomic approach

Leptin is an adipocyte-derived hormone that regulates energy expenditure and food intake. A significant role for leptin in breast cancer has also been indicated by the resistance of leptin knockout mice in development of mammary tumors. In vitro, leptin induces proliferation of MCF-7 cells by activating cellular signaling pathways (1, 11, 12, 16, 17, 56). As leptin is emerging as an important factor for tumor growth, and hormones can exert their actions via autocrine/paracrine mechanisms, we hypothesized leptin may act by regulating epithelial-derived proteins. To test this hypothesis, leptin-regulated proteins secreted from MCF-7 mammary tumor cells were identified using proteomics techniques. Treatment of MCF-7 cells with 500 ng/ml leptin for 24 hours resulted in a 40% increase in cell number and a 5-fold increase in protein secretion as compared to controls. Establishing the significance of leptin-induced secreted factors, the addition of conditioned media from leptin-treated MCF-7 cells to synchronized MCF-7 cells resulted in 40% increase in cell number. Identification of leptin-regulated secreted proteins was done by 2D gel electrophoresis coupled with MALDI-TOF mass spectrometry. Proteins identified using Pro Found software and NCBI database included KF10 Collagen Precursor, Serologically Defined Breast Cancer Antigen NY-BR-62 and Cortactin Isoform a. A Human Cytokine Antibody Array system was used to identify low abundant proteins in the media of control and 500 ng/ml leptin-stimulated MCF-7 cells. In leptin treated cells, levels of FGF-9 were increased while IGFBP-3 and TGF-beta3 levels were decreased. Many previous studies have focused on the regulation of distinct cellular proteins by leptin during mammary tumor cell proliferation. However, ours is the first study to identify leptin-regulated secreted proteins, many of which are known to play important roles in cancer. Our data support that leptin can influence mammary tumor growth and progression through regulation of autocrine/paracrine factors and by modulating the extracellular matrix composition.     

miR-29b regulates migration of human breast cancer cells

microRNAs (miRNAs) are short non-coding RNAs that regulate gene expression by targeting mRNAs, inhibiting the expression of the associated proteins. Although a role for aberrant miRNA expression in cancer has been postulated, the pathophysiologic role and relevance of aberrantly expressed miRNAs in tumor biology has not been established. We evaluated the expression pattern of miRNAs in human breast cancer cells by qPCR, finding out an up-regulated miRNA miR-29b and studying its biological effect by migration assay. We defined a target gene PTEN by bioinformatics approach and western blot. In breast cancer cell line MDA-MB-231 cell, which migrate faster than MCF-7, we observed that miR-29b was highly over-expressed. Inhibition of miR-29b in cultured cells increased the expression of the phosphatase and tensin homolog (PTEN) tumor suppressor, promoting apoptosis, decreasing migration, and decreasing invasion. In contrast, enhanced miR-29b expression by transfection with pre-miR-29b decreased the expression of PTEN and impaired apoptosis, increasing tumor cell migration and invasion. Moreover, PTEN was shown to be a direct target of miR-29b and was also shown to contribute to the miR-29b-mediated effects on cell invasion. Modulation of miR-29b altered the role of PTEN involved in cell migration and invasion. Aberrant expression of miR-29b, which modulates PTEN expression, can contribute to migration, invasion, and anti-apoptosis.     

Exercise-induced muscle-derived cytokines inhibit mammary cancer cell growth

Regular physical activity protects against the development of breast and colon cancer, since it reduces the risk of developing these by 25-30%. During exercise, humoral factors are released from the working muscles for endocrinal signaling to other organs. We hypothesized that these myokines mediate some of the inhibitory effects of exercise on mammary cancer cell proliferation. Serum and muscles were collected from mice after an exercise bout. Incubation with exercise-conditioned serum inhibited MCF-7 cell proliferation by 52% and increased caspase activity by 54%. A similar increase in caspase activity was found after incubation of MCF-7 cells with conditioned media from electrically stimulated myotubes. PCR array analysis (CAPM-0838E; SABiosciences) revealed that seven genes were upregulated in the muscles after exercise, and of these oncostatin M (OSM) proved to inhibit MCF-7 proliferation by 42%, increase caspase activity by 46%, and induce apoptosis. Blocking OSM signaling with anti-OSM antibodies reduced the induction of caspase activity by 51%. To verify that OSM was a myokine, we showed that it was significantly upregulated in serum and in three muscles, tibialis cranialis, gastronemius, and soleus, after an exercise bout. In contrast, OSM expression remained unchanged in subcutaneous and visceral adipose tissue, liver, and spleen (mononuclear cells). We conclude that postexercise serum inhibits mammary cancer cell proliferation and induces apoptosis of these cells. We suggest that one or more myokines secreted from working muscles may be mediating this effect and that OSM is a possible candidate. These findings emphasize that role of physical activity in cancer treatment, showing a direct link between exercise-induced humoral factors and decreased tumor cell growth.     

Thymoquinone up-regulates PTEN expression and induces apoptosis in doxorubicin-resistant human breast cancer cells

The use of innocuous naturally occurring compounds to overcome drug resistance and cancer recalcitrance is now in the forefront of cancer research. Thymoquinone (TQ) is a bioactive constituent of the volatile oil derived from seeds of Nigella sativa Linn. TQ has shown promising anti-carcinogenic and anti-tumor activities through different mechanisms. However, the effect of TQ on cell signaling and survival pathways in resistant cancer cells has not been fully delineated. Here, we report that TQ greatly inhibits doxorubicin-resistant human breast cancer MCF-7/DOX cell proliferation. TQ treatment increased cellular levels of PTEN proteins, resulting in a substantial decrease of phosphorylated Akt, a known regulator of cell survival. The PTEN expression was accompanied with elevation of PTEN mRNA. TQ arrested MCF-7/DOX cells at G2/M phase and increased cellular levels of p53 and p21 proteins. Flow cytometric analysis and agarose gel electrophoresis revealed a significant increase in Sub-G1 cell population and appearance of DNA ladders following TQ treatment, indicating cellular apoptosis. TQ-induced apoptosis was associated with disrupted mitochondrial membrane potential and activation of caspases and PARP cleavage in MCF-7/DOX cells. Moreover, TQ treatment increased Bax/Bcl2 ratio via up-regulating Bax and down-regulating Bcl2 proteins. More importantly, PTEN silencing by target specific siRNA enabled the suppression of TQ-induced apoptosis resulting in increased cell survival. Our results reveal that up-regulation of the key upstream signaling factor, PTEN, in MCF-7/DOX cells inhibited Akt phosphorylation, which ultimately causes increase in their regulatory p53 levels affecting the induction of G2/M cell cycle arrest and apoptosis. Overall results provide mechanistic insights for understanding the molecular basis and utility of the anti-tumor activity of TQ.     

Docosahexaenoic Acid Induces Apoptosis in MCF-7 Cells In Vitro and In Vivo via Reactive Oxygen Species Formation and Caspase 8 Activation

Background

The present study sought to further investigate the in vitro and in vivo anticancer effects of a representative omega-3 fatty acid, docosahexaenoic acid (DHA), with a focus on assessing the induction of oxidative stress and apoptosis as an important mechanism for its anticancer actions.

Methodology/Principal Findings

In vitro studies showed that DHA strongly reduces the viability and DNA synthesis of MCF-7 human breast cancer cells in culture, and also promotes cell death via apoptosis. Mechanistically, accumulation of reactive oxygen species and activation of caspase 8 contribute critically to the induction of apoptotic cell death. Co-presence of antioxidants or selective inhibition or knockdown of caspase 8 each effectively abrogates the cytotoxic effect of DHA. Using athymic nude mice as an in vivo model, we found that feeding animals the 5% fish oil-supplemented diet for 6 weeks significantly reduces the growth of MCF-7 human breast cancer cells in vivo through inhibition of cancer cell proliferation as well as promotion of cell death. Using 3-nitrotyrosine as a parameter, we confirmed that the fish oil-supplemented diet significantly increases oxidative stress in tumor cells in vivo. Analysis of fatty acid content in plasma and tissues showed that feeding animals a 5% fish oil diet increases the levels of DHA and eicosapentaenoic acid in both normal and tumorous mammary tissues by 329% and 300%, respectively.

Conclusions/Significance

DHA can strongly induce apoptosis in human MCF-7 breast cancer cells both in vitro and in vivo. The induction of apoptosis in these cells is selectively mediated via caspase 8 activation. These observations call for further studies to assess the effectiveness of fish oil as a dietary supplement in the prevention and treatment of human breast cancer.     

Inhibition of mycobacterial infection by the tumor suppressor PTEN

The tumor suppressor PTEN is a lipid phosphatase that is frequently mutated in various human cancers. PTEN suppresses tumor cell proliferation, survival, and growth mainly by inhibiting the PI3K-Akt signaling pathway through dephosphorylation of phosphatidylinositol 3,4,5-triphosphate. In addition to it role in tumor suppression, the PTEN-PI3K pathway controls many cellular functions, some of which may be important for cellular resistance to infection. Currently, the intersection between tumorigenic signaling pathways and cellular susceptibility to infection is not well defined. In this study we report that PTEN signaling regulates infection of both noncancerous and cancerous cells by multiple intracellular mycobacterial pathogens and that pharmacological modulation of PTEN signaling can affect mycobacterial infection. We found that PTEN deficiency renders multiple types of cells hyper-susceptible to infection by Mycoplasma and Mycobacterium bovis Bacillus Calmette-Guérin (BCG). The lipid phosphatase activity of PTEN is required for attenuating infection. Furthermore, we found mycobacterial infection activates host cell Akt phosphorylation, and pharmacological inhibition of Akt or PI3K activity reduced levels of intracellular infection. Intriguingly, inhibition of mTOR, one of the downstream components of the Akt signaling and a promising cancer therapeutic target, also lowered intracellular Bacillus Calmette-Guérin levels in mammary epithelial cancer MCF-7 cells. These findings demonstrate a critical role of PTEN-regulated pathways in pathogen infection. The relationship of PTEN-PI3K-Akt mTOR status and susceptibility to mycobacterial infection suggests that the interaction of mycobacterial pathogens with cancer cells may be influenced by genetic alterations in the tumor cells.     

A non-IGF binding mutant of IGFBP-3 modulates cell function in breast epithelial cells

We demonstrated previously that IGFBP-3 alone had no effect on cell death, but dramatically modulated apoptosis in Hs578T IGF non-responsive cells. We investigated whether a non-IGF binding mutant of IGFBP-3 retained its intrinsic actions in this cell line, prior to investigating its actions in IGF-responsive cells (MCF-7 and MCF-10A). In the Hs578T cells, the ceramide analogue, C2-induced apoptosis, non-glycosylated, glycosylated or mutant IGFBP-3 alone had no effect but on co-incubation with C2, all forms of IGFBP-3 markedly accentuated triggered apoptosis. In MCF-7 cells, IGFBP-3 was unable to modulate C2-induced death. In the MCF-10A cells, IGFBP-3 acted as a potent survival factor. IGFBP-3 also affected cell growth in the MCF-10A cells (inhibiting at low doses but increasing growth at higher concentrations). These actions of IGFBP-3 in the MCF-10A cells were independent of IGF-1. IGFBP-3 has differential IGF-independent effects on cell death and growth in normal breast and breast cancer cells.     

Induction of Endothelial Cell Apoptosis by Solid Tumor Cells

The mechanisms by which tumor cells extravasate to form metastasis remain controversial. Previous studies performedin vivoandin vitrodemonstrate that the contact between tumor cells and the vascular wall impairs endothelium integrity. Here, we investigated the effect of breast adenocarcinoma MCF-7 cells on the apoptosis of human umbilical vein endothelial cells (HUVEC). TUNEL labeling, nuclear morphology, and DNA electrophoresis indicated that MCF-7 cells induced a two- to fourfold increase in HUVEC apoptosis. Caspase-3 activity was significantly enhanced. Neither normal cells tested (mammary epithelial cells, fibroblasts, leukocytes) nor transformed hematopoietic cells tested (HL60, Jurkat) induced HUVEC apoptosis. On the contrary, cells derived from solid tumors (breast adenocarcinoma, MDA-MB-231 and T47D; fibrosarcoma, HT 1080) had an effect similar to that of MCF-7 cells. The induction of apoptosis requires cell-to-cell contact, since it could not be reproduced by media conditioned by MCF-7 cells cultured alone or cocultured with HUVEC. Our results suggest that cells derived from solid tumors may alter the endothelium integrity by inducing endothelial cell apoptosis. On the contrary, normal or malignant leukocytes appear to extravasate by distinct mechanisms and do not damage the endothelium. Our data may lead to a better understanding of the steps involved in tumor cell extravasation.     

Increased expression of MDM2, cyclin D1, and p27Kip1 in carcinogen-induced rat mammary tumors

It is thought that environmental pollutants, such as polycyclic aromatic hydrocarbons (PAH), contribute to human breast tumorigenesis, yet their roles remain incompletely elucidated. The prototypical PAH 7,12-dimethylbenz(alpha)anthracene (DMBA) specifically and effectively induces mammary tumor formation in rodent models. In an attempt to explore the molecular mechanisms by which PAH initiates and promotes mammary tumorigenesis, we examined the expression of several cell cycle regulators in rat mammary tumors induced by DMBA. Expression of cyclin D1, murine double minute-2 (MDM2), and Akt was up-regulated in tumors in comparison to normal mammary glands, as indicated by RT-PCR, Western blot analysis, and immunohistochemical staining. Expression of p27Kip1 protein was also elevated in the tumors with increased cytoplasmic localization. However, RB protein remained hyperphosphorylated. To directly test the effects of DMBA, the MCF-7 human breast cancer cells were treated. DMBA induced MDM2 expression in a dose- and time-dependent fashion in the MCF-7 cells, and this activation appeared to be p53 dependent. These data suggest that activation of cyclin D1, MDM2, and AKT as well as increased expression and cytoplasmic localization of p27Kip1 may play a role in this model of environmental pollutant-induced mammary tumorigenesis.     

Tumor suppression by a proapoptotic calcium-activated chloride channel in mammary epithelium

Little is known of the roles played by ion channels in cancer. Here we describe a pair of closely related calcium-activated chloride channels whose differential regulation in normal, apoptotic, and transformed mouse cells suggests that channel function is proapoptotic and antineoplastic. While mCLCA1 predominates over mCLCA2 under normal physiological conditions, this relationship is reversed by apoptotic stress both in developing mammary gland and in cultured HC11 mammary epithelial cells. Consistent with an apoptosis-promoting role, splicing of mCLCA2 is disrupted in apoptosis-resistant tumor cell lines and in HC11 cells selected for resistance to detachment-induced apoptosis (anoikis). Unexpectedly, mCLCA1 message is also down-regulated in these cells by at least 30-fold. These results suggest that both genes antagonize survival of mammary tumor cells by sensitizing them to anoikis. When MCF7 or HEK293 tumor cells were transfected with plasmids encoding either mCLCA1 or mCLCA2, colony formation was greatly reduced relative to a vector-transfected control, demonstrating that calcium-sensitive chloride channel (CLCA) expression is deleterious to tumor cell survival. Furthermore, mammary epithelial cells overexpressing mCLCA2 had twice the rate of apoptosis of normal cells when subjected to serum starvation and formed multinuclear giants at a high frequency in normal culture, suggesting that mCLCA2 can promote either apoptosis or senescence.     

Lipotropes regulate bcl-2 gene expression in the human breast cancer cell line,MCF-7

Lipotropes, a methyl group containing nutrients, including choline, methionine, folic acid, and vitamin B(12), are essential nutrients for humans. They are important methyl donors that interact in the metabolism of one-carbon units and are essential for the synthesis and methylation of deoxyribonucleic acid. The purpose of this study was to examine the effects of excess lipotropes on the growth of a human breast cancer cell line, MCF-7, and normal mammary cells, MCF-10A, in culture. Both cell lines were grown in basal culture medium for 24 h and then switched to medium supplemented with 50 times the amount of each lipotrope as basal culture medium (control). Although there were no significant differences in growth between treatments in either cell line, gene array and Northern analysis revealed that expression of bcl-2 was decreased in lipotrope-treated MCF-7 cells. The ability to induce tumor cell death could have many uses in the prevention and treatment of cancer. Bcl-2 regulates apoptosis and has been shown to directly affect the sensitivity of cancer cells to chemotherapy agents, and it is suggested that strategies designed to block Bcl-2 might prove useful in sensitizing tumor cells to chemotherapy-induced apoptosis. This study shows that although excess lipotropes do not inhibit the growth of breast cancer cells, they can down-regulate the bcl-2 gene, suggesting that lipotropes may increase the susceptibility of breast cancer cells to anticancer drugs.     

Epithelial-Stromal Interactions in Human Breast Cancer: Effects on Adhesion,Plasma Membrane Fluidity and Migration Speed and Directness

Interactions occurring between malignant cells and the stromal microenvironment heavily influence tumor progression. We investigated whether this cross-talk affects some molecular and functional aspects specifically correlated with the invasive phenotype of breast tumor cells (i.e. adhesion molecule expression, membrane fluidity, migration) by co-culturing mammary cancer cells exhibiting different degrees of metastatic potential (MDA-MB-231>MCF-7) with fibroblasts isolated from breast healthy skin (normal fibroblasts, NFs) or from breast tumor stroma (cancer-associated fibroblasts, CAFs) in 2D or 3D (nodules) cultures. Confocal immunofluorescence analysis of the epithelial adhesion molecule E-cadherin on frozen nodule sections demonstrated that NFs and CAFs, respectively, induced or inhibited its expression in MCF-7 cells. An increase in the mesenchymal adhesion protein N-cadherin was observed in CAFs, but not in NFs, as a result of the interaction with both kinds of cancer cells. CAFs, in turn, promoted N-cadherin up-regulation in MDA-MB-231 cells and its de novo expression in MCF-7 cells. Beyond promotion of “cadherin switching”, another sign of the CAF-triggered epithelial-mesenchymal transition (EMT) was the induction of vimentin expression in MCF-7 cells. Plasma membrane labeling of monolayer cultures with the fluorescent probe Laurdan showed an enhancement of the membrane fluidity in cancer cells co-cultured with NFs or CAFs. An increase in lipid packing density of fibroblast membranes was promoted by MCF-7 cells. Time-lapsed cell tracking analysis of mammary cancer cells co-cultured with NFs or CAFs revealed an enhancement of tumor cell migration velocity, even with a marked increase in the directness induced by CAFs.Our results demonstrate a reciprocal influence of mammary cancer and fibroblasts on various adhesiveness/invasiveness features. Notably, CAFs'' ability to promote EMT, reduction of cell adhesion, increase in membrane fluidity, and migration velocity and directness in mammary cancer cells can be viewed as an overall progression- and invasion-promoting effect.     

Dietary organic isothiocyanates are cytotoxic in human breast cancer MCF-7 and mammary epithelial MCF-12A cell lines

Organic isothiocyanates (ITCs) are dietary components present in cruciferous vegetables. The purpose of this investigation was to examine the cytotoxicity of 1-naphthyl isothiocyanate (NITC), benzyl isothiocyanate (BITC), beta-phenethyl isothiocyanate (PEITC), and sulforaphane in human breast cancer MCF-7 and human mammary epithelium MCF-12A cell lines, as well as in a second human epithelial cell line, human kidney HK-2 cells. The cytotoxicity of NITC, BITC, PEITC, and sulforaphane, as well as the cytotoxicity of the chemotherapeutic agents daunomycin (DNM) and vinblastine (VBL), were examined in MCF-7/sensitive (wt), MCF-7/Adr (which overexpresses P-glycoprotein), MCF-12A, and HK-2 cells. Cell growth was determined by a sulforhodamine B assay. The IC50 values for DNM and VBL in MCF-7/Adr cells were 7.12 +/- 0.42 microM and 0.106 +/- 0.004 microM (mean +/- SE) following a 48-hr exposure; IC50 values for BITC, PEITC, NITC, and sulforaphane were 5.95 +/- 0.10, 7.32 +/- 0.25, 77.9 +/- 8.03, and 13.7 +/- 0.82 microM, respectively, with similar values obtained in MCF-7/wt cells. Corresponding values for BITC, PEITC, NITC, and sulforaphane in MCF-12A cells were 8.07 +/- 0.29, 7.71 +/- 0.07, 33.6 +/- 1.69, and 40.5 +/- 1.25 microM, respectively. BITC and PEITC can inhibit the growth of human breast cancer cells as well as human mammary epithelium cells at concentrations similar to those of the chemotherapeutic drug DNM. Sulforaphane and NITC exhibited higher IC50 values. The effect of these ITCs on cell growth may contribute to the cancer chemopreventive properties of ITCs by suppressing the growth of preclinical tumors, and may indicate a potential use of these compounds as chemotherapeutic agents in cancer treatment.     

Cell cycle arrest by the PTEN tumor suppressor is target cell specific and may require protein phosphatase activity

PTEN, a tumor suppressor commonly targeted in human cancer, possesses phosphatase activities toward both protein and lipid substrates. While PTEN suppresses gliomas through cell cycle inhibition which requires its lipid phosphatase activity, PTEN's effects on other tumor types and the role of its protein phosphatase activity are controversial or unknown. Here we show that exogenous wild-type PTEN arrests some, but not all human breast cancer cell lines in G1, in a manner independent of endogenous PTEN. Unexpectedly, the G129E mutant of PTEN selectively deficient in the lipid phosphatase activity still blocked the cell cycle of MCF-7 cells, while the G129R and H123Y mutants lacking both phosphatase activities were ineffective. These results suggest that PTEN's protein phosphatase activity likely contributes to its tumor suppressor function in subsets of tumors and that elucidation of downstream targets which dictate cellular responses to PTEN may have important implications for future cancer treatment strategies.     

BRCA1 accumulates in the nucleus in response to hypoxia and TRAIL and enhances TRAIL-induced apoptosis in breast cancer cells

A major contributing factor to the development of breast cancer is decreased functional expression of breast cancer susceptibility gene 1, BRCA1. Another key contributor to tumorigenesis is hypoxia. Here we show that hypoxia increased the nuclear localization of BRCA1 in MCF-7 and MDA-MB-468 human breast cancer cell lines without changing its steady-state expression level. Nuclear accumulation of BRCA1 was not evident in MCF-12A or HMEC (human mammary epithelial cell) nonmalignant mammary epithelial cells under the same conditions. Hypoxia also increased the cell surface expression of TRAIL on MDA-MB-468 cells. Neutralization of TRAIL precluded the hypoxia-induced accumulation of BRCA1 in the nucleus, whereas exogenously administered TRAIL mimicked the effect. Treatment of MDA-MB-468 cells with TRAIL resulted in a dose- and time-dependent increase in apoptosis. Furthermore, TRAIL-induced apoptosis in HCC1937 cells, which harbor a BRCA1 mutation, increased synergistically when wild-type BRCA1 was reconstituted in the cells, and downregulation of BRCA1 expression in MDA-MB-468 cells reduced the apoptotic response to TRAIL. These data provide a novel link between hypoxia, TRAIL and BRCA1, and suggest that this relationship may be especially relevant to the potential use of TRAIL as a chemotherapeutic agent.