bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract P4-01-06: The Role of the PI3K/AKT, RAS/MAPK and Novel Signaling Pathways in Resistance to HER2-Targeted Therapy
Authors: O'Brien NA , Wilcox K , McDermott M , Crown J , O'Donovan N , Slamon DJ. 
Published: April 2011

Abstract
Despite the proven clinical benefits of HER2-targeted therapy, resistance to trastuzumab and lapatinib exists. Understanding the molecular mechanisms responsible for resistance is critical to improving the survival rates of patients with this aggressive subtype of breast cancer. Currently, there are no clinically validated biomarkers of resistance to therapy. To address this issue, we measured the responses of a panel of 18 HER2-amplified breast cancer cell lines to both trastuzumab and lapatinib and evaluated the activity of PI3K/AKT/mTOR and RAS/MAPK/ERK signaling in resistance. Eight of the cell lines were identified as resistant to trastuzumab and seven as resistant to lapatinib by both two and three dimensional response assays. Five of the trastuzumab resistant cell lines remained sensitive to lapatinib (IC50 &lt; 1 μM), indicating that these two agents have at least partially non-overlapping mechanisms of resistance. The combination of low PTEN or the presence of activating mutations of PI3K, and thus increased activity of PI3K/AKT signaling, is a strong predictor of resistance to trastuzumab. However, this is not the case with lapatinib, two of the cell lines with activating mutations of PI3K (MDA-361 and HCC-202) were resistant to lapatinib whereas two other PI3K mutant cells lines (SUM-190 and HCC-1954) were lapatinib sensitive, indicating that lapatinib resistance can occurindependently of increased PI3K/AKT signaling. To further investigate these mechanisms, we measured the effects of trastuzumab and lapatinib treatment on AKT/mTOR and ERK activation. Cell lines sensitive to trastuzumab or lapatinib showed a marked reduction in pAKT, pS6K and pERK in response to either agent, whereas the resistant cells maintained high pAKT, pS6K and pERK levels in the presence of trastuzumab and lapatinib. In cell lines resistant to trastuzumab yet sensitive to lapatinib, treatment with lapatinib significantly reduced the phosphorylation of AKT, S6K and ERK. Thus in a subpopulation of cell lines lapatinib can overcome trastuzumab resistance via continued deactivation of PI3K/AKT/mTOR and MAPK/ERK signaling. However, there still remains a subgroup of HER2-positive cells that are resistant to both trastuzumab and lapatinib. We used global expression analysis by both mRNA microarray and proteomic screening to identify a set of genes and proteins that are consistently altered in these innately resistant cells relative to the sensitive cell lines. In vitro validation of these targets will reveal which are playing a functional role in resistance and are thus potentially druggable targets to be used in improving the survival of patients with HER2-positive disease.
Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-01-06.
Abstract P4-01-07: Endoplasmic Reticulum-Associated Calcium Regulates Resistance of the Anti-Apoptotic Protein Bcl-2 to Taxol-Induced Apoptosis in Breast Cancer Cells
Authors: Pan Z , Gollahon L 
Published: April 2011

Abstract
Background: The unique anti-apoptotic function of Bcl-2 results in cellular resistance to many chemotherapeutic agents used in solid tumor treatments. However, there is a controversy regarding the relationship between Bcl-2 resistance and the mechanism of action for Taxol, an anticancer agent commonly used in breast cancer treatment. Although previous studies showed both Bcl-2 and Taxol may have effects on calcium homeostasis independently, no studies have addressed whether calcium is the keystone in the relationship between Bcl-2 resistance and Taxol action. Objective: Since most internal calcium is stored in the endoplasmic reticulum (ER), this study focused on the role of ER calcium changes in Bcl-2-activated resistance to Taxol-induced apoptosis. The ER calcium channel IP3R and its relationship with Bcl-2 and Taxol were investigated in order to clarify the mechanism underlying calcium regulation.
Methods and Materials: For this study, MDA-MB-468, a Bcl-2 negative breast carcinoma cell line, its Bcl-2 stable transfectant and MCF-7, a Bcl-2 positive breast carcinoma cell line were used. The dynamic calcium changes induced by Taxol in these cells were determined by measuring free cytosolic calcium fluctuations using Fluo4-AM and ER calcium changes by the calcium cameleon, D1ER. Taxol-induced apoptosis was measured using the Annexin V-FITC assay. The calcium interfering agents BAPTA-AM and 2-APB were applied to evaluate whether Taxol-induced calcium changes were related to this apoptotic event. Western blot was used to detect changes induced by Taxol treatment in Bcl-2 and IP3R protein levels.
Results: Taxol induced a rapid ER calcium release followed by a gradual ER calcium depletion in both Bcl-2 positive and negative cell lines, indicating that ER calcium store provides a direct target for Taxol, independently of Bcl-2 expression. Interfering with these calcium changes inhibited the Taxol-induced apoptosis, suggesting ER calcium release promotes Taxol-induced apoptosis. Our results also showed that Bcl-2 inhibited the basal ER calcium release. Taxol stimulated ER calcium release and thus attenuated Bcl-2-initiated resistance to apoptosis. Relevance: This research addressed the question of whether ER calcium changes play a critical or marginal role in mediating resistance of Bcl-2 to Taxol-induced apoptosis. Elucidating the role of calcium in this process will not only help to clarify the mechanism of drug resistance associated with Bcl-2, but also aid in more effective application of Taxol in breast cancer treatment.
Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-01-07.
Abstract P4-02-03: Gene Copy Number Alterations Related to mRNA and miRNA Expression in Endocrine Resistant Breast Cancers
Authors: Larionov A , Reis-Filho JS , Lambros MBK , Sproul D , Dixon JM. 
Published: April 2011

Abstract
Background
Endocrine resistance is a major clinical issue in breast cancer. Multiple studies provide data relating molecular features of primary tumours on their response to endocrine treatment. However, there is a paucity of data on molecular profiles of breast tumours at time when endocrine resistance has already manifested.
Aim
Provide multilayer molecular profiling of a series of breast tumours at time of relapse or progression on endocrine treatment.
Results
High density microarray-comparative genomic hybridisation (aCGH) analysis was performed in 39 endocrine treated breast tumours at time of progression or relapse. 25 of these tumours were also profiled using whole genome mRNA and miRNA arrays (HT-12 mRNA array and DASL Sentrix Array Matrix micro-RNA array, Illumina).
Overall the aCGH profiles of resistant tumours were similar to aCGH profiles of un-selected luminal breast tumours and cell lines published earlier. The most common gene copy gain areas (&gt;10M of length) were located in 1q23.1-44, 5p13.1-33, 8p11.23-q24.3, 16p12.1-13.3, 17q21.33-24.1 and 20q13.2-13.33, while the most common losses were in 1p33-36.32, 8p12-23.3, 11q22.3-24.1, 16q12.2-24.3 and 17p11.2-13.3. Unsupervised hierarchical clustering by gene copy number profiles split of the resistant tumours into 3 groups. As expected, on average mRNAs and microRNAs were higher expressed in tumours carrying gains of their genes and were decreased in tumours carrying the losses. Copy number gains in this series were associated with the upregulation of multiple RNAs, including C8orf76, TATDN1, CYC1, ZBTB10, C16orf63, THUMPD1, TFB2M, miR-454, miR-301a and miR-296-3p/5p. Losses were associated with the downregulation of RNAs including ACADVL, BNIP3L, UBE4A, REXO2, ATP5L, PPP2CB, NBL1 and PCM1. Of the above genes UBE4A and ATP5L are closely co-localised in 11q23.3; miR454-miR301a are located closely in 17q22. Such associations suggest that changes in expression of these genes in breast cancers may be driven by alterations in the gene copy number. Analysis of the recurrent amplification regions on this dataset is underway.
Conclusions
aCGH profiles of endocrine resistant tumours at time of relapse or progression are similar to those found in un-selected and non-treated luminal tumours. The study provides the first dataset integrating gene copy number with mRNA and microRNA gene expressions in clinical specimens of endocrine resistant breast cancers.
Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-02-03.
Abstract P4-02-06: Beyond CYP2D6 Genotyping: Personalized Dosing of Tamoxifen (Tam) Based on Low Serum Endoxifen Levels
Authors: Barginear MF , Jaremko M , Yu C , Peter I , Kasai Y , Kemeny M , Adelson K , Desnick R , Raptis G. 
Published: April 2011

Abstract
Background: The efficacy of Tam varies widely. Tam is biotransformed to endoxifen and 4-OH-Tam by the cytochrome P450 2D6 enzymes. Studies on 2D6 variants as a predictor of Tam's efficacy show conflicting results. Compared to Tam, endoxifen and 4-OH-Tam have a 100-fold greater affinity for the estrogen receptor (ER) and in suppressing estrogen-dependent cell proliferation. However, endoxifen is present at concentrations up to 20-fold higher and is pharmacologically distinct from 4-OH-Tam. Endoxifen's effect on ERα-degredation and transcription is concentration dependent; serum endoxifen levels ≥40nM block ERα-mediated transcriptional activation and inhibit estrogen-induced breast cancer cell proliferation.
Methods: Pts taking Tam 20 mg/day, for any reason, were potentially eligible. Serum Tam, Tam metabolites and nineteen 2D6 alleles were analyzed. 2D6 genotyping was performed by PCR-based procedures and metabolites were assessed on Liquid Chromatography/Tandem Mass Spectrometry. Pts were stratified by 2D6 function into 3 groups: extensive metabolizers (EM) who had normal enzyme activity; intermediate metabolizers (IM) who had reduced enzyme activity; and poor metabolizers (PM) who had no enzyme activity. Tam was increased to 30 mg/day if serum endoxifen &lt; 40nMol. Serum Tam and Tam metabolites were measured every 2 weeks for 90 days.
Results: 121 patients have enrolled to date. Age, ethnicity, BMI, concomitant medications, and adherence were recorded. 25/117 (21%) pts had low serum endoxifen; all of these pts were increased to Tam 30 mg/day. Of this cohort, 36% were PMs (PM/PM and PM/IM), 32% were IMs (EM/PM and IM/IM) and 32% were EMs (EM/EM and EM/IM). All patients’ serum endoxifen increased to &gt;40nMol by day 30 (Diagram below).
Conclusions: 1) Sub-therapeutic serum endoxifen can be found in any 2D6 phenotype. 2) It is possible to achieve therapeutic serum endoxifen/Tam metabolite levels by optimizing the dose of Tam. 3) Subsequently, it is likely that other gene variants are involved in Tam metabolism hence why 2D6 testing may lead to conflicting results. 4). Further investigation is warranted as personalizing the dose of Tam is feasible and may potentially improve the outcome of pts taking Tam regardless of 2D6 genotype. Figure available in online version.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-02-06.
Abstract P4-04-01: Proliferation, Migration, Stem Cell Activity and the Role of Cyclin D1 in Breast Cancer
Authors: Lamb RM , Lehn S , Tobin N , Berglund P , Landberg G. 
Published: April 2011

Abstract
Background: Proliferation, migration, and stem cell (SC) activity have all been implicated in the development and progression of breast cancer. Growing evidence suggests a relationship between these cellular characteristics, which are tightly regulated by complex signalling pathways. The oncogene cyclin D1 is an essential cell cycle protein that regulates G1 to S phase transition and is overexpressed in many breast cancers. Despite the proliferation activating properties of cyclin D1 it has been linked to a less malignant phenotype.
Aim: The aim is to investigate the relationship between migration, proliferation and SC activity in breast cancer and to determine the molecular mechanisms that control these processes focusing on cyclin D1. Methods and Results: To establish a link between proliferation and migration two breast cancer cell lines (MDAMB231 and MDAMB468) were quiesced and proliferation assessed by Ki67 and migration by transwell migration assays. G0/G1 cells displayed an increased migratory capacity (2.9 ±0.15 fold) compared to cycling cells. Next we downregulated cyclin D1 in actively cycling cells using siRNA which resulted in significantly decreased proliferation and the novel observation of increased migration (1.7±0. 19 fold). Having established a link between proliferation and migration the next aim was to investigate relationships with SC activity. To achieve this, cells were sorted following Hoechst 33342 and 7AAD labelling into three populations G0/G1, S, and G2/M. SC-like activity was assessed using a non-adherent mammosphere culture system where single cells are seeded at a density of 500 cells per cm2 for cultured for 7 days. The data clearly demonstrated that G0/G1 cells had increased migratory capacity and also SC-like activity (1.5±0.2 fold) compared to actively cycling cells. Interestingly, down-regulation of cyclin D1 increased both migratory and SC-like activity with most significant effects observed in G0/G1 and S phase of the cell cycle indicating that cyclin D1 is a key protein in the regulation of these cellular processes. When downregulating the cyclin D1 associated kinases CDK4/6 no effect on migration or SC activity was observed suggesting that cyclin D1 affects these cellular processes independent of CDK activation. Finally, when analysing a large set of premenopausal breast cancers we observed an inverse, proliferation-independent link between cyclin D1 and cancer recurrence as well as tumour size (p=0.03), validating that the protein might abrogate infiltrative and malignant behaviour in vivo.
Conclusion: These novel functions of cyclin D1 illustrates the interplay between tumor proliferation and migration and SC activity and may explain the lessened malignant behaviour in cyclin D1 high breast cancers.
Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-04-01.
Abstract P4-04-03: Transcriptome Analyses of Mouse and Human Mammary Cell Subpopulations Reveals Multiple Conserved Genes and Pathways
Authors: Lim E , Wu D , Pal B , Bouras T , Asselin-Labat M-L , Vaillant F , Yagita H , Lindeman GJ , Smyth GK , Visvader JE. 
Published: April 2011

Abstract
INTRODUCTION: Molecular characterization of the normal epithelial cell types that reside in the mammary gland is an important step toward understanding pathways that regulate self-renewal, lineage commitment, and differentiation along the hierarchy. We previously reported distinct stages in human mammary epithelial cell development and linked them to breast tumor subtypes previously defined by gene expression profiling. Here we determined the gene expression signatures of four distinct subpopulations isolated from the mouse mammary gland. The epithelial cell signatures were used to interrogate mouse models of mammary tumorigenesis and to compare with their normal human counterpart subsets to identify conserved genes and networks.
METHODS: RNA was prepared from freshly sorted mouse mammary cell subpopulations (mammary stem cell (MaSC)-enriched, committed luminal progenitor, mature luminal and stromal cell) and used for gene expression profiling analysis on the Illumina platform. Gene signatures were derived and compared with those previously reported for the analogous normal human mammary cell subpopulations. The mouse and human epithelial subset signatures were then subjected to Ingenuity Pathway Analysis (IPA) to identify conserved pathways.
RESULTS: The four mouse mammary cell subpopulations exhibited distinct gene signatures. Comparison of these signatures with the molecular profiles of different mouse models of mammary tumorigenesis revealed that tumors arising in MMTV-Wnt-1 and p53-/- mice were enriched for MaSC-subset genes, whereas the gene profiles of MMTV-Neu and MMTV-PyMT tumors were most concordant with the luminal progenitor cell signature. Comparison of the mouse mammary epithelial cell signatures with their human counterparts revealed substantial conservation of genes, whereas IPA highlighted a number of conserved pathways in the three epithelial subsets.
CONCLUSIONS: The conservation of genes and pathways across species further validates the use of the mouse as a model to study mammary gland development and highlights pathways that are likely to govern cell-fate decisions and differentiation. It is noteworthy that many of the conserved genes in the MaSC population have been considered as epithelialmesenchymal transition (EMT) signature genes. Therefore, the expression of these genes in tumor cells may reflect basal epithelial cell characteristics and not necessarily cells that have undergone an EMT. Comparative analyses of normal mouse epithelial subsets with murine tumor models have implicated distinct cell types in contributing to tumorigenesis in the different models.
Figures available in online version.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-04-03.
Abstract P4-04-05: Histone Deacetylase Inhibitors Increase Markers of the Dedifferentiated Cancer Stem Cell Phenotype in Human Breast Cancer Cells
Authors: Debeb BG , Xu W , Lacerda L , Hortobagyi GN , Robertson F , Ueno NN , Lucci A , Reuben JM , Buchholz TA , Woodward WA. 
Published: April 2011

Abstract
It has been suggested that differentiated cancer cells can de-differentiate into the cancer stem cell phenotype (Meyers et al., 2009; Gupta et al., 2009) and that Histone Deacetylase (HDAC) inhibitors enable efficient induction of pluripotent stem cells from adult fibroblasts (Huangfu et al., 2009). Moreover, we have recently demonstrated that the HDAC inhibitor valproic acid radiosensitized breast cancer cells grown on plastic with serum while it radioprotected breast cancer cells grown in stem cell promoting culture conditions (Debeb et al., 2010). Based on these data, we hypothesized that HDAC inhibitors increase the cancer stem cell population via dedifferentiation of differentiated cancer cells. To examine our hypothesis, Sum159 breast cancer cells were FACS-sorted based on ALDH activity and subsequently treated with one of two HDAC inhibitors, valproic acid or SAHA (suberoylanilide hydroxamic acid). After sorting, ALDH-negative cells were treated either with valproic acid, SAHA, or vehicle. After a week, the percentage of ALDH-positive cells (passage 0, P0) was examined with flow cytometry while the remaining cells were passaged and incubated with and without valproic acid or SAHA for a week and the percentage of ALDH-positive cells again evaluated (P1). This was repeated for the third time (P2). Samples from each generation were also collected to examine the protein expression. On average, a 3-fold increase in ALDH positive cells was seen in valproic acid-treated cells (35.6% vs.12.6%) and a 1.5-fold increase in SAHA-treated cells (41% vs. 28%) compared to vehicle-treated controls. This effect was maintained through multiple passages. Moreover, the expression of Beta-catenin and EMT associated genes like vimentin, fibronectin, n-cadherin, which have been implicated in generating cancer stem cells was significantly increased withtreatment in initial and passaged cells. Further functional endpoint studies are needed to validate these in vitro marker-based findings.
Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-04-05.
Abstract P4-04-06: Histone H2A Inhibits Breast Cancer Stem Cell Growth
Authors: Tourville Poirier AM , Balicki D. CRCHUM 
Published: April 2011

Abstract
Introduction: Recent studies have identified epigenetic abnormalities as the underlying cause of many breast cancers. In particular, the aberrant acetylation of core histones may be a pivotal source of dysregulated expression of both oncogenes and tumour suppressor genes. Histone deacetylase inhibitors are known to restore histone acetylation and to induce cellular differentiation and growth arrest in breast cancer cells and are the object of extensive research for novel anticancer treatments. We hypothesize that acetylated proteins, in particular histone H2A, could also serve as an alternative substrate for histone deacetylase, thereby promoting cellular histone acetylation and arrest of cancer cell growth. Indeed, our previous results indicate that exogenous calf thymus histone H2A penetrates cells and their nuclei, and induces differentiation and senescence in human breast cancer cells. In contrast, human recombinant histone H2A and H2AX, which lack post-translational modifications, fail to yield these results in spite of the fact that human and bovine histone H2A are virtually identical in their amino acid sequences. However, these recombinant molecules were obtained from bacteria, which lack the intracellular machinery that is necessary to generate post-translational modifications. Thus, post-translational histone modifications are pivotal to the inhibition of breast cancer cell proliferation. Objective: We have extended our study to establish if exogenous calf thymus histone H2A could also induce growth arrest of breast cancer stem cells, and subsequently to determine which portion (s) of the histone H2A molecule is responsible for this action, using overlapping synthetic peptides.
Materials and Methods: Breast cancer stem cells were incubated with exogenous calf thymus histone H2A and seven synthetic peptides corresponding to overlapping portions of the entire protein and include all of its post-translational modifications. Cellular morphology was observed and viable cells were counted using trypan blue exclusion. Control experiments were performed with normal breast cells, cells from fibrocystic breast tissues, and breast cancer cells that were not significantly enriched in cancer stem cells.
Results and Discussion: Histone H2A inhibits breast cancer stem cell growth and increases their adhesion. Stem cells have recently been identified in many malignant tumours, including breast cancer. Since cancer stem cells are thought to be an important source of therapeutic resistance, they provide a novel research focus for alternative treatment strategies. Our results are very encouraging and may lead to the development of new pharmacological molecules targeting epigenetic abnormalities in breast cancer stem cells.
Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-04-06.