Estrogen Receptor Positive Lines Research Articles

Regulation of SLC6A14 trafficking in breast cancer cells by heat shock protein HSP90β

SLC6A14 is a plasma membrane transporter specific for neutral and basic amino acids, upregulated in many tumors. This study focused on breast cancer cell lines, showing the fully glycosylated band, known to be at the cell surface, in estrogen receptor positive lines. Inhibition of heat shock protein 90β (HSP90β) decreased the level of this band, what correlated with a decrease of SLC6A14 transport activity. A direct interaction between SLC6A14 and HSP90β was confirmed in proximity ligation assay, pointing to the role of HSP90 in folding control in endoplasmic reticulum and affecting farther transporter trafficking to the cell surface. Either inhibitor of SLC6A14 (α-methyltryptophan) or of HSP90 (radicicol) had the cytotoxic effect, when added alone, while treatment with both compounds had a synergistic effect. This points to SLC6A14 as a druggable target in breast cancer and a combination therapy being more efficient in killing cancer cells.

Inhibition of breast cancer cell proliferation in repeated and non-repeated treatment with zoledronic acid

BackgroundZoledronic acid is used to treat bone metastases and has been shown to reduce skeletal-related events and exert antitumor activity. The present in vitro study investigates the mechanism of action of Zoledronic Acid on breast cancer cell lines with different hormonal and HER2 patterns. Furthermore, we investigated the efficacy of repeated versus non-repeated treatments.MethodsThe study was performed on 4 breast cancer cell lines (BRC-230, SkBr3, MCF-7 and MDA-MB-231). Non-repeated treatment (single exposure of 168 hrs’ duration) with zoledronic acid was compared with repeated treatment (separate exposures, each of 48 hrs’ duration, for a total of 168 hrs) at different dosages. A dose–response profile was generated using sulforhodamine B assay. Apoptosis was evaluated by TUNEL assay and biomolecular characteristics were analyzed by western blot.ResultsZoledronic acid produced a dose-dependent inhibition of proliferation in all cell lines. Anti-proliferative activity was enhanced with the repeated treatment, proving to be statistically significant in the triple-negative lines. In these lines repeated treatment showed a cytocidal effect, with apoptotic cell death caused by caspase 3, 8 and 9 activation and decreased RAS and pMAPK expression. Apoptosis was not observed in estrogen receptor-positive line: p21 overexpression suggested a slowing down of cell cycle. A decrease in RAS and pMAPK expression was seen in HER2-overexpressing line after treatment.ConclusionsThe study suggests that zoledronic acid has an antitumor activity in breast cancer cell lines. Its mechanism of action involves the decrease of RAS and RHO, as in osteoclasts. Repeated treatment enhances antitumor activity compared to non-repeated treatment. Repeated treatment has a killing effect on triple-negative lines due to apoptosis activation. Further research is warranted especially in the treatment of triple-negative breast cancer.

P4-02-11: Insulin-Like Growth Factor I Promotes Estrogen Receptor Positive Breast Cancer Cell Proliferation, in Part, through CYP1A1 Signaling.

Abstract Activation of the transmembrane tyrosine kinase insulin-like growth factor 1 receptor (IGF-1R) contributes to breast cancer progression. Nonetheless, the mechanisms by which the IGF-1R contributes to breast cancer progression need to be better characterized to further develop therapeutic strategies that target this pathway. It is known that cytochrome P450 monooxygenases synthesize metabolites from endogenous and exogenous sources that promote breast cancer proliferation. Here we propose that activity of the IGF-1R modulates the activity of CYP1A1 to promote breast cancer cell proliferation. We have found that IGF-IR activation by treatment with IGF1 (5nM ) for 4hrs induces cytochrome P450 1A1 (CYP1A1) mRNA levels in estrogen receptor (ER) positive line T47D-CO, but not ER negative line MDA-MB-231. Treatment with IGF1 promotes the proliferation of ER positive lines T47D-CO and MCF7. Interestingly, we found that treatment with IGF1 only partially compensates the growth-inhibition induced by CYP1A1 knock down by siRNA, suggesting that IGF1 requires CYP1A1 for full promotion of cell growth. These preliminary data suggest that IGF-1 signaling functions, in part, through CYP1A1 to promote ER positive breast cancer cell proliferation. The mechanism of IGF-1 signaling through CYP1A1 is novel and may represent an effector mechanism for IGF-1. Further understanding the role of CYP1A1 downstream of the IGF-1R may allow novel approaches for inhibition of breast cancer cell proliferation downstream of the IGF-1R. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-02-11.

Abstract 4129: Effects of the metabolism of omega-3 fatty acid EPA by CYP1A1 in breast cancer proliferation and survival

Abstract Omega-3 fatty acids are widely known for their importance in heart disease. However, the impact of omega-3 fatty acids in breast cancer is not fully understood. Through these studies we seek to understand the role of the metabolism of the omega-3 fatty acid known as eicosapentaenoic acid (EPA) in breast cancer and determine whether it may affect breast cancer progression. It is known that high levels (60uM) of EPA have anti-proliferative effects in various cancer lines. Consistent with these reports, we found that EPA treatment (60uM) of the estrogen receptor positive line MCF7 and the estrogen receptor negative line MDA-MB-231 decreases cell proliferation. Interestingly, lower dosing of EPA (15uM-40uM) significantly stimulated the proliferation of these lines. It has been shown that cytochrome P450 1A1 (CYP1A1) selectively epoxygenates EPA to 17(18)-epoxyeicosatetraenoic acid (17,18-EpETE). Measurement of cell growth by MTT assay shows that 17,18-EpETE promotes the proliferation of the MCF7 and MDA-MB-231 lines in a dose- (≥ 1uM) and time-dependent (max effect at 48-72hrs) manner. Together, these results led us to hypothesize that CYP1A1 metabolizes EPA into 17,18-EpETE, thereby promoting breast cancer proliferation and survival. To better understand this epoxygenase mechanism, we studied the impact of CYP1A1 and the 17,18-EpETE metabolite of EPA on cell proliferation by MTT assay, cell cycle progression by propidium iodine staining, and apoptosis by Annexin V flow cytometry. We found that CYP1A1 is expressed in MCF7 and MDA-MB-231 lines and siRNA-mediated knock down significantly inhibits the proliferation of these lines. CYP1A1 knock down also blocks the proliferative effect of treatment with 15uM EPA. Our preliminary data also indicate that 17,18-EpETE promotes growth of the MDA-MB-231 line, in part, by decreasing apoptosis and increasing the S- and G2/M cell populations. Consistently, knock down of CYP1A1 in MDA-MB-231 line increases the G0/1 cell population and decreases number of cells in the S and G2/M phases, consistent with a G1 block. The mechanism of action in MCF7 cells is yet to be determined. Better understanding of the mechanism by which EPA metabolism affects breast cancer may impact our understanding of breast cancer prevention and therapeutics. We propose that CYP1A1 may be of significant importance in breast cancer progression through novel mechanisms that depend on biosynthetic pathways promoting EPA epoxygenation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4129.

Epidermal growth factor receptor gene expression in estrogen receptor-positive and negative human breast cancer cell lines.

Expression of epidermal growth factor (EGF) receptor by human breast cancer tissues has an inverse relationship with expression of the estrogen receptor and may be associated with a poor clinical response. We have studied the regulation of EGF receptor expression in a series of human breast cancer cell lines with varying degrees of estrogen responsiveness. Three estrogen receptor-positive lines, MCF-7, ZR-75-1, and T47D, were found to have less than 70,000 EGF binding sites per cell by radioreceptor assay and were growth stimulated in vitro by EGF. Four estrogen receptor-negative lines, MDA-MB-231, Hs578T, EVSA-T, and BT-20, contained greater than 70,000 EGF binding sites per cell and showed no in vitro growth stimulation by EGF. In all cell lines EGF receptor number was correlated with the amount of EGF receptor protein and RNA. Differences in EGF receptor expression between the cell types was not due to amplification of the EGF receptor gene. Rather, variations in EGF receptor expression between lines were due, at least in part, to differences in the rate of EGF gene transcription as determined by nuclear run-off studies. Our data confirm the previously described inverse relationship between expression of EGF and estrogen receptors. We show here that the absence of estrogen receptor expression in human breast cancer cell lines is associated with higher levels of functional EGF receptor protein and mRNA.