bluuebunny/crossref_metadata_2025_split huggingface dataset:
β1 integrin mediates tamoxifen resistance in breast cancer cells.
Authors: Pontiggia O , Bal de Kier Joffé E , Simian M 
Published: December 2010

Abstract
Abstract #1055
Even though it is now well established that the microenvironment plays a central role in modulating tumor progression and response to therapy, its function as a regulator of endocrine resistance in breast cancer has not yet been explored. To do so we have established a bicellular in vitro cell line, called LM05-Mix, from the estrogen dependent M05 mouse mammary tumor (BCRT, in press). The epithelial and fibroblastic cell populations of the LM05-Mix cell line were further separated to generate the LM05-E and LM05-F cell lines respectively, both of which are estrogen receptor (ER) and progesterone receptor positive. We have previously shown that treatment with 10-6M 4-OH-tamoxifen (TAM) induces apoptosis of the LM05-E cells, and that this effect is inhibited when the epithelial cells are co-cultured with the fibroblastic cell population. Immunofluorescence staining to the extracellular matrix component fibronectin (FN) was found to be especially associated to the fibroblastic cells in the co-culture. To investigate whether FN conferred TAM resistance, LM05-E cells and human MCF-7 cells were cultured on recombinant FN and treated for 48 hours with TAM. In both cases FN conferred resistance (P≤0.001). To understand what signaling pathways were involved, the experiments were repeated in the presence of specific inhibitors and both PD98059 and LY294002 reverted the protective effect, implicating the participation of both the MAPK/ERK and PI3K/AKT pathways. Next, to corroborate the activation of these pathways by FN, starved LM05-E cells were treated with FN (30 μg/ml). Phosphorylation of FAK, MAPK/ERK1/2 and AKT was detected by western blot between 3 and 15 minutes of treatment. To find out whether the activation of these pathways by FN could lead to ligand independent activation of the ER and thus account for the induction of TAM resistance, immunofluorescence staining was carried out to serine 118-phosphorylated ER. An increase in the phosphorylation of ER at this site was evident at 15 minutes of treatment both in LM05-E and MCF-7 cells. Finally, to confirm that the protective effect was mediated by the binding of β1 integrin to FN, cells were pre-incubated with either a specific β1 blocking antibody (AIIB2) or mouse IgG and seeded on FN. The protective effect of FN over TAM induced cell death was only reverted in the AIIB2 treated group (P≤0.01). Our results show that FN through its binding to β1 integrin confers TAM resistance and suggest that the mechanism involved would be through the modulation of the activity of ER. Targeting the interaction of breast cancer cells with the microenvironment could be a novel and future alternative for patients with breast cancer whose tumors progress to TAM resistance. This work is supported by a grant from the Susan G. Komen for the Cure Foundation to MS (BCTR0600341).
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 1055.
The microvasculature of invasive ductal breast carcinoma: a sterological approach.
Authors: Mahon S , Miller N , Curran C , Dockery P , Callagy G , Kerin MJ 
Published: December 2010

Abstract
Abstract #1057
Background:&#x2028; Microvessel density (MVD) is a commonly used parameter to describe tumour vascularity and has been widely studied as a possible prognosticator in breast cancer. Other descriptors of vascular beds have been developed and have been applied to a variety of systems including the reproductive and nervous systems. These stereological approaches are providing objective unbiased assessment of various vascular beds.&#x2028; Aim:&#x2028; The aim of this pilot study was to use stereological methods to describe the vasculature in invasive ductal breast carcinoma, to test the hypothesis that microvasculature anatomy effects Tumor Grade, Size and Nodal Status.&#x2028; Methods:&#x2028; A cohort of 51 breast cancer patients with pre-menopausal invasive ductal carcinoma and moderate NPI was identified.Sections were stained with CD34 using the automated Ventana Discovery staining platform. A number of stereological parameters were estimated in three tissue compartments (gland,stroma and adipose):Number per Unit area (Na), Length Density (Lv) and Radial Diffusion Distance. Data was analysed using Analyses of Variance methods using Minitab© Statistical software.&#x2028; Results:&#x2028; There was no significant difference in the radial diffusion difference when groups were analysed by Grade Size and Nodal Status. Similar results were found for Na and Lv&#x2028; Conclusion:&#x2028; This study suggests that there is no simple link between tumor vasculature and Grade, size and nodal status in Invasive Ductal Breast Carcinoma. This study illustrates the potential usefulness of the stereological approach in the study of vascular beds in breast cancer studies by providing access to potentially physiologically relevant anatomical parameters of microvasculature beds.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 1057.
Inhibition of Notch signaling reduces the stem-like cell population in T47D breast cancer cells and prevents mammosphere formation.
Authors: Grudzien P , Strack P , Golde T , Miele L , Foreman KE 
Published: December 2010

Abstract
Abstract #106
Breast cancer has been proposed as a stem cell-derived disorder based on studies identifying cancer stem cells (CSCs) in breast tumors and established cell lines. These self-renewing CSCs are thought to be responsible for breast cancer formation, progression and recurrence; therefore, a deeper understanding of the signaling pathways regulating CSC survival will benefit development of novel therapeutic strategies. Notch signaling, which is known to be dysregulated in breast cancer and has been implicated in mammary stem cell self-renewal, can be effectively blocked by gamma-secretase inhibitors (GSIs). While GSIs are currently in clinical trials for breast cancer, it is unclear how these compounds will affect CSCs or if CSCs from different breast cancer phenotypes (estrogen receptor-alpha (ER)-positive and negative) will be differentially affected. Here, we demonstrate stem-like cells derived from the T47D breast cancer cell line show elevated levels of Notch signaling, with an ER-negative subclone (T47D-C42) having higher intrinsic levels of Notch activity compared to an ER-positive subclone (T47D-A18). Blockade of Notch signaling with three structurally distinct GSIs reduced the number of stem-like cells, and completely abolished secondary mammosphere formation, which is a measure of stem cell self-renewal. Importantly, this effect was irreversible with MRK-003, but not with the two other GSIs. Moreover, our results suggest that the ER-negative cell line was more sensitive to the GSIs compared to the ER-positive line. This is consistent with our recent report that loss of estrogen signals results in Notch activation. These novel findings support a role for Notch signaling in CSC self-renewal and proliferation, and they suggest Notch inhibition may have clinical benefits in targeting CSCs.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 106.
Brain metastases (BM) in patients presenting with HER-2 amplified advanced breast cancer (ABC).
Authors: Sweldens C , Huget P , Weytjens R , Prove A , Vermeulen P , Van Dam P , Dirix L 
Published: December 2010

Abstract
Abstract #1060
Background : The HER-2 amplified breast cancer phenotype is suggested to have an increased propensity for CNS metastasis. The introduction of trastuzumab might be related to this association. In order to elucidate this relationship we analyzed our prospectively collected database of all patients presenting with metastatic HER-2 amplified breast cancer in the time frame between Jan 2002 until Dec 2004.&#x2028; Material and Methods : The AZ-Augustinus Breast Cancer Tissue Bank was initiated in 1999. It ensures the prospective entry of all clinical and pathological features of patients presenting in our Breast Clinic clinical database. The data entry is performed by dedicated Datamanagers and is linked to the Biological Repository Collection (tissue, serum, plasma, bone marrow). Patients presenting with primary metastatic, HER-2 amplified ( FISH +) between 2002 and 2004 were analyzed for this study. Since Jan 2002, all patients presenting with metastatic disease underwent a routine CT-scan of the brain as initial staging procedure.&#x2028; Results : A total of 33 patients presented with HER-2 +, unpretreated metastatic breast cancer. Mean age: 53 years (range 30-69). Sites of metastases at diagnosis ( % of patients) : liver: 71 %, lung: 36 %, bone: 86 %, brain (BM): 15 % (5), lymph nodes and others: 20%. In total 5/33 (15%)&#x2028; presented with brain only metastasis. During the follow-up period 17 more patients developed BM, amounting to 22/33 (67%). The median time between diagnosis of BM after diagnosis of ABC was 22 months (0 – 98). Mean time to developing BM was 18 months. In total 62 lesions in the CNS were counted on MRI in these 22 patients. 23 lesions (38%) had a cerebellar localisation.&#x2028; Conclusions: Assuming that the presentation with metastasis is a reflection of the unperturbed biology of disease, our data suggest that the CNS is not an exceptionally preferred site for metastasis in untreated HER-2 + advanced breast cancer. A preference for a cerebellar localisation is suggested.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 1060.
P53 functional assay in yeast: evaluation in 1856 patients in a large prospective clinical trial (EORTC 10994/BIG 00-01).
Authors: Bonnefoi H , Zimmer AS , Piccart M , Mauriac L , Campone M , Tubiana-Hulin M , Petit T , Rouanet P , Welnicka-Jaskiewicz M , Cufer T , Blot E , Becette V , Bogaerts J , André S , Cameron D , Bergh J , Iggo R 
Published: December 2010

Abstract
Abstract #1067
Background:Although mutations in the p53 gene can occur in 20-30% of sporadic breast cancer, and are associated with other poor prognostic factors, there is no clear evidence as to whether p53 status should influence therapy. Immunohistochemistry has been demonstrated to be an inferior method compared with sequencing, but the optimal technique to assess the functional p53 status is not known. The yeast test is a functional assay which tests the transcriptional competence of p53, and detects biologically important mutations. Here we present the results of p53 evaluation using this method in a prospective study.&#x2028; Material and methods: EORTC 10994/BIG 00-01 trial is a randomized trial comparing the neoadjuvant use of an anthracycline-based regimen with a taxane-containing regimen in patients with large operable or locally advanced breast cancers. Fresh frozen tumour biopsies were mandatory before starting chemotherapy. Frozen sections were examined centrally and the p53 test was done if the invasive tumour cell content was above 20%. RNA extract was used to assess p53 status: p53 wild type tumours were defined as tumours whose cDNA transfected in yeast gave less than 20% red colonies (background); p53 mutated tumours gave 20% or more red colonies.&#x2028; Results: The trial accrual was completed in November 2006 after inclusion of 1856 patients. There was no frozen sample available in 70 patients. In 276 patients the % of tumour cells was &lt;20. The p53 test failed in 62 patients, generally because the quality or quantity of RNA was too low for RT-PCR. P53 was successfully assessed in 1458 patients: 661 (45%) tumours are p53 mutated and 797 (55%) are p53 wt. p53 mutated and wild type tumours were grade III in 40% (265/661) and 19% (153/797), respectively, and estrogen receptor negative in 41% (270/661) and 14.5% (116/797), respectively. We did not see any correlation between p53 status and tumour size or nodal status at diagnosis. HER2 status data are being collected.&#x2028; Conclusion: P53 status has been successfully evaluated using a yeast assay in 78.5% (1458/1856) of cases in the context of a multicenter trial. In indirect comparisons the frequency of patients with functionally altered p53 status is higher than with sequenced based methods. Our results confirm an association between p53 mutation and high grade or estrogen receptor negative status, but not with age, tumour size or clinical nodal status. More data regarding the quality and quantity of RNA collected will be presented at the meeting.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 1067.
Stem cell and epithelial-mesenchymal transition markers on circulating tumor cells in patients with metastatic breast cancer.
Authors: Aktas B , Tewes M , Hauch S , Fehm T , Kimmig R , Kasimir-Bauer S 
Published: December 2010

Abstract
Abstract #107
Background: Stem cell like tumor cells have been suggested to be the active source of metastatic spread in primary tumors. Furthermore, these cells may undergo phenotypic changes, known as epithelial-mesenchymal transition (EMT), which allows them to travel to the site of metastasis formation without getting affected by conventional treatment. Assuming that metastasis requires a dissemination of tumor stem cells or tumor cells showing EMT, it was the purpose of this study to identify these cells among circulating tumor cells (CTC) in blood samples of 28 metastatic breast cancer patients.&#x2028; Materials and Methods: 5 ml blood was analyzed for EpCAM, MUC-1 and HER-2 transcripts with the AdnaTest BreastCancer (AdnaGen AG). The recovered c-DNA was additionally multiplex tested for three EMT markers [Twist, Akt2, PI3K (TAP)] and separately for the tumor stem-cell marker ALDH1. The analytical sensitivity was determined by the detection of a low number of target cells (5 IGROV cells spiked into 5 ml blood of healthy donors) using the AdnaTest BreastCancer procedure. The identification of EMT markers was considered positive if at least one marker was detected in the sample. Healthy donor samples without spiked tumor cells were used to determine the specificity of the test. The resulting PCR fragments were separated and quantified using the Agilent Bioanalyzer 2100.&#x2028; Results: Applying an amplicon cut-off value of 0.2 ng/μl for Akt2, 0.15 ng/μl for Twist, 0.25 ng/μl for PI3K and 0.15 ng/μl for ALDH1, 97% of 30 healthy donor samples investigated were negative for TAP and 95% for ALDH1 transcripts. The spiking experiments revealed 80% recovery of the IGROV cells. CTC were detected in 12/28 (43%) breast cancer samples. All samples were further examined for tumor stem cell or TAP markers. In the CTC (+) group 50% were positive for at least one of the TAP markers and 42% for ALDH1. In the CTC (-) group the percentages were 19% and 12%, respectively. The expression of TAP and/or stem cell markers in CTC was compared with the clinical follow up results. In CTC (+) patients diagnosed as non-responders the expression of TAP and ALDH1 was found in 5/12 (40%) samples. In the CTC (+) responder group only one sample expressed TAP and no positive case was found for ALDH1. In the CTC (-) non-responder group 1/14 samples was positive for TAP and ALDH1. 2/14 (14%) of CTC (-) responders were positive for TAP and 1/14 for ALDH1, respectively.&#x2028; Conclusion: A major proportion of CTC found in the blood of metastatic breast cancer patients shows EMT and tumor stem cell characteristics supporting the hypothesis that these markers are an indicator for therapy resistant tumor cell populations and, therefore, for an inferior prognosis.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 107.
Molecular breast cancer prognosis in node negative early breast cancer.
Authors: Schmidt M , Tramm T , Böhm D , Kyndi M , Lehr H , Alstner J , Kölbl H , Stropp U , von Törne C , Weber K , Hennig G , Gehrmann M , Overgaard J 
Published: December 2010

Abstract
Abstract #1071
Background: A number of molecular signatures have been published to aid breast cancer prognosis and therapy response prediction. However, most signatures rely on measurements of dozens of genes on an array platform. Here we report the identification and validation of a prognostic algorithm based on a small number of expressed genes reflecting dominant prognostic motifs like steroid hormone receptor expression, proliferation, matrix degradation and B-cell infiltration. The test is designed for mRNA extracted from formalin fixed paraffin embedded tumour tissue (FFPE).&#x2028; Materials and Methods: 200 fresh frozen tumours from node negative patients, who did not receive systemic therapy after surgery, were profiled on HG-U133a arrays. Primer and probes were designed for genes identified as prognostic relevant and used to profile the corresponding paired FFPE tumour tissue of our identification cohort by kinetic PCR. A genomic algorithm was constructed and tested in FFPE tumor tissue of an independent validation cohort of 240 node negative patients that remained untreated in the adjuvant setting.&#x2028; Results: Based on the Prognostic-score of our algorithm we classified the patients of the validation cohort into three classes: low risk (25% of patients), intermediate risk (25%) and high risk (50%). The probability to die after having a recurrence within 10 years was 4.7% for the low risk group, 11.9% for the intermediate risk group and 33.1% for the high risk group. Kaplan Meier analysis revealed that the trend for survival (Logrank test for trend P = 0.0002) was highly significant. Adjuvant! Online calculated probabilities to die of cancer were also used to classify patients into three risk groups of comparable size (25/25/50 percent of patients). The observed risk was 9.1% in the low risk, 26.7% in the intermediate and 23.6% in the high risk group. The area under the ROC curve for Adjuvant! Online was 0.619 (95% CI, 0.52 to 0.718; P = 0.031) while for our algorithm it was 0.755 (95% CI, 0.6628 to 0.8473; P &lt; 0.0001).&#x2028; Discussion: We identified a prognostic algorithm for node negative breast cancer based on kinetic PCR analysis of FFPE tumor tissue. Validation in an independent cohort showed the ability of the algorithm to classify patients into clinically relevant risk classes. Preliminary analysis shows a better prognostic performance than current clinical guidelines.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 1071.
Significantly higher expression of vascular endothelial growth factor (VEGF) and shorter survival after recurrences in premenopausal node negative patients with triple negative breast cancer.
Authors: Linderholm BK , Klintman M , Grabau D , Bendahl P , Fernö M , Per M 
Published: December 2010

Abstract
Abstract #1077
Background: Triple-negative breast cancer (TNBC) amounts to approximately 15% of all breast cancer and is defined as lacking expression of oestrogen- and progesterone receptors (ER and PgR) and HER2 leaving chemotherapy as the only treatment option. The aggressive nature of TNBC is reflected by high histological grade, early onset of relapses, and more commonly visceral metastases. Further exploration of biological features in this group could (would) help to find potential targeted therapies. Aim: To compare intratumoural levels of VEGF and survival in patients with TNBC compared with HER2 positive patients and ER and/or PgR positive but HER2 negative patients. Patients and methods: 210 premenopausal patients with lymph-node negative breast cancer included in a prospective clinical trial were studied. (Ref Malmström et al JCO 19:2010-2019, 2001). Levels of VEGF were determined by an enzyme-linked immuno-sorbent assay (ELISA) on lysates from freshly frozen breast tumor material. HER2 amplification was determined by in situ hybridization of tumor with Her2 + or 3+ reactivity with IHC (determined by IHC followed by confirmatory FISH in 2+ or 3+). Results: Thirty-six out of 210 (17%) patients were classified as having a TNBC. The median level of VEGF in the total patient population was 81 ng/mg total protein (range 2.0- 3610). Significantly higher VEGF levels were found in TNBC compared with HER2 pos. or HER2 neg. ER/PgR pos. patients; median levels 250, 157 and 53 ng/mg total protein respectively (p&lt;0.001). Similarly, 84% of TNBC had VEGF above the median compared to 62% in HER2 pos and 41% in the HER2 neg. ER/PgR pos. group (p&lt;0.001). TNBC was significantly associated to high histological grade (89% grade III) (p&lt;0.001), and high S-phase fraction (p=0.001). The highest rate of distant metastases was (seen or) recorded for HER2 pos patients while TNBC and HER2 neg. ER/PgR pos. had similar relapse rates (p&lt;0.001; three-group comparison). After adjustment for chemotherapy, patients with TNBC survived significantly shorter after relapse compared with ER/PgR pos/HER2 neg patients (HR=7.2;95%CI=2.1-25; p=0.002) but not compared to HER2 pos patients (HR=2.3;95%CI=0.73-7.0; p=0.16). Conclusion: We here for the first time show significant higher levels of an angiogenesis stimulating factor, VEGF in TNBC. Whether blockade of angiogenesis can be an effective new targeted treatment option for this poor prognosis group will be elucidated in ongoing trials.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 1077.
Breast cancer molecular subtypes and the risk of second malignancies.
Authors: Radecka B , Hudala-Klecha J , Badzio A , Szwiec M , Drosik K , Jassem J 
Published: December 2010

Abstract
Abstract #1078
Background: Particular molecular subtypes of breast cancer (BC) have been shown to manifest various clinical behaviors. In a large consecutive series of pts we compared treatment outcomes of four major BC subtypes, with particular focus on the risk of second malignancies.&#x2028; Materials and methods: Triple negative breast cancer (TNBC) was characterized by negativity of hormone receptors (HR) and HER2 (0-2+ by immunohistochemistry). Other subtypes included HR+/HER2-, HR+ /HER2+ and HR-/HER2+. All pts underwent curative surgery from January 2003 to December 2007, followed by radiation and/or systemic therapy according to current standards. Univariate risks of relapse and death were estimated using Kaplan-Meier survival curves and compared with log-rank test, and the multivariate analysis used stratified Cox model.&#x2028; Results: Of the 783 tumors, 520, 50, 48 and 165 were classified as HR+/HER2-, HR+ /HER2+, HR-/HER2+ and TNBC, respectively. The TNBC and HR-/HER2+ subtypes were associated with a higher tumor grade (p&lt;0.001), higher T stage (p&lt;0.001) and higher proportion of medullary type (p&lt;0.001) compared to other two subgroups. Other characteristics, such as age and menopausal status showed no difference. The probability of relapse-free survival (RFS) and overall survival (OS) in TNBC and HR-/HER2+ groups were similar, and lower than in other two types (TNBC vs. HR+/HER2- and HR+ /HER2+; p=0.002 and 0.03 for RFS and OS). Impaired prognosis of TNBC and HR-/HER2+ pts remained significant after correction by T and N stage. TNBC was associated with almost doubled risk of second malignancy (12.7 vs. 6.5%) compared to all other types (p=0.008). This risk was significantly increased for contralateral BC and endometrial cancer (p=0.002 and 0.024, respectively).&#x2028; Conclusion: TNBC and HR-/HER2+ subtypes are associated with worse prognosis. Additionally, TNBC carries an increased risk of second malignancies, most likely due to higher proportion of hereditary BC.
Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 1078.