bluuebunny/crossref_metadata_2025_split huggingface dataset:
Overexpression of CD44 in acquired endocrine resistant breast cancer modulates erbB activity and promotes an invasive phenotype.
Authors: Baruah BP , Smith C , Jordan N , Nicholson RI , Robertson J , Barrett-Lee P , Hiscox S 
Published: December 2010

Abstract
Abstract #3027
Background: Acquired endocrine resistance in breast cancer cells is accompanied by altered growth factor receptor signalling and a highly migratory cell phenotype. In tamoxifen-resistant ('TamR') MCF7 cells, our microarray analysis has demonstrated elevated levels of CD44, a transmembrane glycoprotein, known to interact with and modulate the function of a number of growth factor receptors and plays a major role in tumour metastasis. Here, we have explored the role of CD44 as a modulator of erbB family activity in TamR cells and evaluated the expression of CD44 in a series of breast cancer patients on endocrine therapy.&#x2028; Methods: RT-PCR was performed to confirm overexpression of CD44 at the gene level in TamR cells. Western blotting and immunocytochemistry were used to investigate expression of CD44 at the protein level together with the activity of erbB signalling pathways following stimulation of CD44 by its ligand, hyaluronan (HA), or following stimulation with erbB ligands (TGFα, heregulin β1) after knockdown of CD44 expression using siRNA. Immunofluorescence microscopy was used to determine association between CD44 and erbB family members. Cell migration was assessed by seeding cells onto fibronectin-coated microporous inserts for 24 hours. CD44 expression levels were evaluated using paraffin-embedded tissue sections in a series of primary breast cancer tissues (n=103) from patients receiving endocrine therapy by immunohistochemical staining.&#x2028; Results: CD44 was overexpressed at the gene and protein level in TamR cells versus their endocrine sensitive counterparts. siRNA-mediated suppression of CD44 expression reduced basal EGFR/Her2 activity in TamR cells together with suppressing their migratory phenotype. siRNA-mediated inhibition of CD44 expression greatly attenuated the ability of erbB ligands to activate erbB receptors and inhibited TGFα and heregulin β1-induced migration. Conversely, stimulation of TamR, but not MCF7 cells, with HA promoted activation of erbB family members together with their downstream signalling intermediates ERK1/2, Src and AKT. Analysis of CD44 expression in clinical tissue revealed an association between elevated CD44 expression and a shortened response to endocrine therapy.&#x2028; Conclusions: These data demonstrate that overexpression of CD44 accompanies endocrine resistance where it enhances the sensitivity of these cells to factors (erbB ligands, HA) found within the tumour microenvironment. This may implicate CD44 in loss of endocrine response clinically and, therefore, provide a novel therapeutic target in endocrine resistant breast cancer.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3027.
Overexpression of L1CAM accompanies acquired endocrine resistance and is associated with the development of an aggressive cell phenotype.
Authors: Hiscox S , Jordan N , Crandon-Lewis A , Jiang W , Nicholson RI , Gee J 
Published: December 2010

Abstract
Abstract #3028
Background: Acquired resistance to endocrine therapies is associated with a gain in aggressive features in vitro which may impact on tumour spread in vivo. To identify potential candidate molecules which may promote this adverse cell behaviour, we performed microarray analysis on MCF-7 cell models of fulvestrant resistance ('FasR' cells) applying a commercial algorithm for signalling network discovery. We have subsequently investigated the significance of overexpression of L1CAM, a cell-surface adhesion molecule implicated in a MAPK signalling network and recently associated with tumour metastasis, in FasR cells and in clinical tissue.&#x2028; Materials and Methods: A network analysis algorithm was applied to t-test significant Affymetrix gene lists derived from FasR vs. MCF-7 cells (Almac Diagnostics). Subsequent to revealing a prominent L1CAM network, L1CAM gene and protein expression were assessed using RT-PCR, Western blotting and immunofluorescence. Inhibition of L1CAM was achieved using siRNA transfection. Cell invasion and migration were measured by seeding onto Matrigel- or fibronectin-coated microporous membranes respectively. After 48 hours, invasive/migratory cells were fixed, stained and counted. Analysis of cell signalling molecules was determined using Western blotting with phospho-specific antibodies. Clinical significance of L1CAM expression was determined by analysis of L1CAM expression in tumour (n=101) vs. normal (n=23) breast tissue using Q-PCR.&#x2028; Results: Development of fulvestrant resistance in MCF7 cells was associated with ER loss and a gain in migratory and invasive capacity in vitro. L1CAM was overexpressed at both gene (p&lt;0.05) and protein (p&lt;0.01) level in FasR cells whereas little or no L1CAM was detectable in MCF7 cells confirming microarray findings. Network and ontological analysis predicted L1CAM lies upstream of an ERK2 network (26 genes; p&lt;0.0001) impinging on transcription factors/regulators which may promote aggressive cell behaviour. In agreement, siRNA-mediated inhibition of L1CAM suppressed the activity of nuclear MAP kinase and reduced the intrinsic migratory and invasive nature of FasR cells in vitro. Preliminary analysis indicated L1CAM expression was inversely associated with ER status (p&lt;0.01) and reduced overall survival (p=0.01).&#x2028; Conclusions: These data suggest that L1CAM overexpression associates with emergence of ER-negative fulvestrant resistance in breast cancer, where it may confer a highly motile and invasive phenotype. Additionally, our clinical studies suggest L1CAM may also contribute to inherently aggressive, ER-negative breast cancers. L1CAM signalling may thus provide a new therapeutic target to subvert such aggressive states in vivo.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3028.
Control of luminal type A intrinsic subtype enriched transcription factor network by insulin: implications of diabetes on breast cancer classification.
Authors: McCune KR , Bhat-Nakshatri P , Thorat M , Badve S , Nakshatri H 
Published: December 2010

Abstract
Abstract #3029
Background: Luminal type A and type B represent estrogen receptor alpha (ERalpha)-positive breast cancers with luminal type A expressing higher levels of ERalpha and is associated with better prognosis. Recent studies have identified a specific functional transcription factor network comprising GATA-3, FOXA1 and ERalpha in normal luminal cells as well as in luminal type A breast cancer that dictates their hormone dependence. Signaling molecules that may disrupt this network and force these cells to acquire hormone-independence are not known. T-bet (Tbx21) has been described as a major negative regulator of GATA-3 activity. As the expression and/or activity of some of the above factors are controlled by insulin, the objective of this study was to investigate whether elevated level of insulin, as evidenced in type II diabetes, alters gene expression pattern in luminal type A breast cancers by interrupting GATA-3:FOXA1:ERalpha network and thus forcing these cancers to acquire non-luminal phenotype and/or hormone-independence.&#x2028; Methodologies: The effect of insulin on the expression of ERalpha, FOXA1, GATA-3 and T-bet was measured in ERalpha-positive MCF-7 cells by Western blot analysis. The effect of T-bet on estrogen-regulated gene expression was measured by stable overexpression of T-bet in MCF-7 cells and subsequent qRT-PCR analysis. Publicly available Oncomine database was used to determine the expression pattern of T-bet and its relation to ERalpha status in primary breast cancers. A proliferation assay was used to determine sensitivity of T-bet overexpressing cells to tamoxifen in the presence and absence of insulin.&#x2028; Results: Insulin induced the expression of T-bet, which was partially reversed by estrogen. ERalpha and GATA-3 levels were reduced in MCF-7 cells stably overexpressing T-bet suggesting that T-bet reduces GATA-3-dependent ERalpha expression. Estrogen-inducible expression of estrogen target genes GREB-1 and Myb was lower in T-bet overexpressing cells compared to parental cells, although basal expression was elevated in T-bet overexpressing cells. Treatment of T-bet overexpressing cells with insulin decreased tamoxifen sensitivity. Although T-bet expression was generally higher in ERalpha-negative breast cancers compared to ERalpha-positive breast cancers, a subpopulation of ER-positive breast cancers express elevated levels of T-bet.&#x2028; Conclusions: Insulin may change the gene expression pattern through T-bet-mediated disruption of master cell-type-specific transcriptional network including GATA-3, ERalpha and FOXA1 that dictates the phenotype of hormone-dependent luminal type A breast cancer. T-bet may serve as a marker to identify ERalpha-positive breast cancers that express low levels of GATA-3 and have progressed to hormone-independence.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3029.
Expression of ER-a36, a novel variant of estrogen receptor a, and resistance to tamoxifen treatment in breast cancer.
Authors: Shi L , Dong B , Li Z , Lu Y , Ouyang T , Li J , Wang T , Fan Z , Fan T , Lin B , Wang Z , Xie Y 
Published: December 2010

Abstract
Abstract #3037
Background: Recently, a 36 kDa variant of estrogen receptor a (ER-a66), ER-a36, has been identified and cloned. ER-a36 predominantly localizes on the plasma membrane and in the cytoplasm and mediates a membrane-initiated “nongenomic” signaling pathway. In this study, we investigated the association between ER-a36 expression and tamoxifen resistance in breast cancer patients.&#x2028; Methods: ER-a36 protein expression in tumors from 710 breast cancer patients with a median follow-up of 7.9 years was assessed using immunohistochemistry (IHC) assay. Survival curves were compared using the log-rank test and multivariate analysis was performed using Cox model. All statistical tests were two-sided.&#x2028; Results: Among the patients with ER-a66 positive tumors who received tamoxifen treatment (n=307), overexpression of ER-a36 was associated with poorer disease-free survival (DFS) and disease-specific survival (DSS) and remained as an unfavorable independent factor of survival in multivariate analyses (DFS: HR=2.27; 95% CI= 1.40 to 3.68; P=. 001; DSS: HR=2.42; 95% CI= 1.37 to 4.28; P= .002). In contrast, among patients with ER-a66 positive tumors who did not receive tamoxifen (n=129), ER-a36 expression was not associated with survival, indicating a correlation between ER-a36 expression and tamoxifen resistance. Furthermore, ER-a36 expression was not associated with survival in ER-a66 negative tumors whether the patients received tamoxifen (n=73) or not (n=149). Our in vitro experiments with MCF7/ER36 cells also confirmed that high ER-a36 expression resulted in tamoxifen resistance.&#x2028; Conclusions: Patients with ER-a66 positive tumors that also express high levels of ER-a36 are less likely to benefit from tamoxifen treatment. ER-a36 is an important predictive marker for tamoxifen therapy in ER-a66 positive breast cancer patients.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3037.
Intermittent treatment of hormone-dependent breast cancer using exemestane: translational research from lab bench to bedside.
Authors: Masri S , Phung S , Chung C , Somlo G , Chen S 
Published: December 2010

Abstract
Abstract #3040
Background and Rationale: Endocrine therapy agents such as aromatase inhibitors (AIs) have shown good clinical efficacy in the treatment of post-menopausal breast cancer patients. Currently, the major clinical hindrance with endocrine therapy is disease progression or acquired resistance. Intermittent treatment of endocrine therapy agents has been proposed in several clinical trials to address whether this treatment strategy can delay the onset of acquired resistance in hormone-dependent breast cancers. To support these clinical trials, we have performed a pre-clinical study using the steroidal AI exemestane (EXE) to understand potential molecular features that differentiate continuous treatment of EXE from intermittent administration.&#x2028; Methods: The ER+ and aromatase-overexpressing cell culture model system (MCF-7aro) was used in this pre-clinical study. The cells were treated either continuously (T+EXE Cont) or intermittently (T+EXE Interm) for 2 weeks with EXE and 1 week without EXE, in the presence of testosterone (T). In addition, a non-steroidal AI letrozole, was used in an identical intermittent manner.&#x2028; Results: Based on cell culture studies, acquired resistance was reached at 14 weeks in the T+EXE Cont treated cells, while T+EXE Interm cells were still responsive to EXE. At 34 weeks, acquired resistance developed in the T+EXE Interm cell lines, which demonstrates a delay in time to progression. The non-steroidal AI letrozole did not demonstrate a delay in acquired resistance using this intermittent model. Based on microarray analysis, we observed that gene expression of certain estrogen-responsive genes was elevated in the T+EXE Cont cells, while select genes showed lower levels of up-regulation in T+EXE Interm lines.&#x2028; Discussion: Based on our previous studies, up-regulation of a key estrogen-responsive gene, amphiregulin (AREG), is responsible for activating EGFR and mediating resistance to EXE. Expression of genes such as AREG are decreased in the T+EXE Interm cell lines, suggesting the suppression of growth factor signaling implicated in driving resistance. In addition, we have reported that EXE acts as an aromatase destabilizer at the protein level. Unlike letrozole or anastrozole, EXE-mediated protein degradation of aromatase may play a key role in reducing estrogen levels that are responsible for driving breast cancer survival and proliferation. Since these pre-clinical studies provide an important mechanistic basis for intermittent endocrine therapy, a clinical trial to evaluate the intermittent use of exemestane has been discussed and is planned to initiate in the near future.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3040.
Co-targeting leptin and insulin-like growth factor I signaling: dramatic effects of epidermal growth factor receptor inhibitors on triple negative breast cancer cells.
Authors: Saxena NK , Smith LT , Knight BB , Sharma M , Sharma D 
Published: December 2010

Abstract
Abstract #3044
Introduction: Obesity is an independent risk factor for breast cancer and obese breast cancer patients exhibit a higher risk for larger tumor burden, increased metastasis and poor response to endocrine therapy. Obesity affects breast carcinogenesis by autocrine and paracrine effects of adipocytokine leptin and IGF-1. We have previously shown that leptin induces growth stimulation of breast cancer cells by recruiting histone acetyltransferases and mediator complex to cyclin D1 promoter via activation of Stat3. In the present study, we found a novel bidirectional crosstalk between IGF-1 and leptin signaling that promotes invasion and migration of triple-negative (ER, PR, HER2 negative) breast cancer cells; MDA-MB-231, MDA-MB-468, MDA-MB-435 and HCC-1806.&#x2028; Methods and Results: IGF-1 induced significant tyrosine phosphorylation of leptin receptor (Ob-Rb) and leptin induced tyrosine phosphorylation of IGF-1 Receptor (IGF-1R). Combined treatment of leptin and IGF-1 induced synergistic activation of both Ob-Rb and IGF-1R along with activation of Akt and ERK. Furthermore, IGF-1 and leptin synergistically transactivated epidermal growth factor receptor (EGFR) and induced proliferation of triple negative breast cancer cells. We also found that tyrosine phosphorylation of EGFR induced by combined treatment of leptin and IGF-1 was significantly inhibited not only by EGFR tyrosine kinase inhibitor, AG1478 but also with a broad-spectrum matrix metalloproteinase inhibitor, GM6001 indicating that leptin and IGF-1 induced EGFR transactivation is dependent on proteolytic release of EGFR ligands. Intriguingly, we also found that combined treatment of leptin and IGF-1 potently induced invasion of triple negative breast cancer cells in Matrigel invasion and quantitative Electric Cell-Substrate Impedance Sensing invasion assays. Inhibition of leptin and IGF-1 mediated EGFR activation using a highly potent, reversible inhibitor of HER1/EGFR tyrosine kinase, erlotinib or dual epidermal growth factor receptor (EGFR)/human EGFR-2 (HER2) kinase inhibitor, lapatinib effectively inhibited leptin and IGF-1 induced invasion. Also, inhibition of EGFR activation using erlotinib and lapatinib reduced leptin- and IGF-1-induced migration of triple negative breast cancer cells.&#x2028; Discussion: Taken together these data indicate a novel bidirectional crosstalk between leptin and IGF-1 signaling that transactivates EGFR and promotes metastatic properties, invasion and migration of triple negative breast carcinoma cells. Our novel findings indicate the possibility of using EGFR inhibitors erlotinib and lapatinib to counter the pro-cancerous effects of leptin and IGF-1 signaling in triple negative breast cancers.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3044.
Interactive effects of low-dose estradiol and tamoxifen on the postmenopausal primate endometrium and mammary gland.
Authors: Wood CE , Cline M 
Published: December 2010

Abstract
Abstract #3047
Background: Long-term use of combined estrogen+progestin hormone therapies increases breast cancer risk in postmenopausal women. Selective estrogen receptor modulators (SERMs) are promising alternatives to progestins, particularly for breast cancer survivors and other women at high-risk for breast cancer considering hormone therapy. The purpose of this study was to evaluate interactive effects of low-dose estradiol (E2) and the SERM tamoxifen on the postmenopausal endometrium and mammary gland.&#x2028; Materials and Methods: Ovariectomized adult female cynomolgus monkeys were given placebo (n=6), oral estradiol (E2, 0.25 mg/1800 kcal) (n=6), tamoxifen (TAM, 20 mg/1800 kcal) (n=5), or E2+TAM (n=6) for 4 months. Doses of E2 and TAM were scaled to approximate those given to women.&#x2028; Results: Endometrial thickness and uterine weight were significantly higher in E2, TAM, and E2+TAM groups compared to placebo (P &lt; 0.01 for all). In contrast, endometrial gland and stromal proliferation determined by Ki67 immunolabeling was significantly greater only in the E2 group (P &lt; 0.01 for both). Endometrial Ki67 gene expression was also higher only in the E2 group (P &lt; 0.01). Gene markers of estrogen receptor alpha (ER) activity such as TFF1 and STC2 were significantly increased in E2, TAM, and E2+TAM groups (P &lt; 0.01 for all). On endometrial histology TAM and E2+TAM induced stromal fibrosis and occasional simple glandular hyperplasia with cystic change. In the mammary gland, TFF1 gene expression was increased by all treatments (P &lt; 0.05 for all), while no statistically significant treatment effects were seen for epithelial proliferation or progesterone receptor expression.&#x2028; Discussion: Previous studies have suggested that the weak estrogen-like effects of tamoxifen in the uterus contribute to increased endometrial cancer risk. Our findings reveal a more complex pattern of tamoxifen action involving increased ER activity, antagonism of E2-induced proliferation, and distinctive stromal remodeling. These results suggest that tamoxifen does not exhibit simple weak estrogen agonist / antagonist effects in the primate endometrium but instead acts through pathways independent of classical ER signaling to alter cancer risk. Future gene profiling should aid in the identification of these alternative pathways.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3047.
Normal breast tissue cells from breast cancer patients express epidermal growth factor receptor HER1-4 protein concentrations characteristic for cancer cells.
Authors: Olsen DA , Oestergaard B , Bokmand S , Jakobsen EH , Brandslund I 
Published: December 2010

Abstract
Abstract #3052
Background: The epidermal growth factor receptor HER2 is expressed to higher than normal concentrations in 25%-30% of human breast cancers and in approximately 90% of these cases the overexpression is a result of gene amplification. The mechanism behind HER2 amplification is unknown but may be caused by continuous stimulation and activation of the HER1 system. Consequently we have mapped the concentrations of all four epidermal growth factor receptors HER1-4 in breast cancer tissue, in autologous non-cancer breast tissue and normal breast tissue from healthy women.&#x2028; Material and Methods: Tissue samples of malignant and adjacent normal breast tissue were collected from 118 consecutive women admitted for surgical treatment of breast cancer. In addition 26 samples of normal breast tissue from healthy women were collected. The tissue samples weighing between 10 and 40 mg were homogenized and the proteins were extracted. The total protein amount in the samples were measured and the protein concentrations of HER1-4 were determined quantitatively using ELISA expressed as pg/mg total protein.&#x2028; Results: HER1 demonstrated lower than normal concentrations both in autologous non-cancer tissue and cancer tissue. HER2 showed 4 times higher than normal concentrations in autologous reference tissue and 24 times higher in cancer tissue, while the corresponding figures for HER3 was factor 8 and 13 and for HER4 factor 10 and 8. These differences were significant at p&lt;10-4. Further it was observed that correlations between the receptors were maintained from normal breast tissue to autologous reference breast tissue but were lost in cancer tissue. Adjusting the concentrations of receptors by differences in cell number content did not change differently in the cancer patients.&#x2028; Discussion: We propose these findings to indicate that breast cancer is a systemic disease where also non-cancer tissue cells in the cancer patients are continuously activated thus favouring the subsequent development of cancer. This may explain the observation that occurrence of new cancers are increased in patients previously treated for cancer. Measurement of decreased HER1 and increased HER2-4 in breast cells from healthy women may in the future be used as an indicator of increased risk for cancer development.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3052.