bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract P6-15-17: Synergy of Ixabepilone and Dasatinib in Triple Negative Breast Cancer Cell Lines
Authors: Ismail-Khan RR , Marichion DC , Cubitt CL , Lancaster JM. 
Published: April 2011

Abstract
Background: Treatment options are very limited for breast cancers that do not express HER2 or estrogen and progesterone receptors (Triple Negative Breast Cancer; TNBC). Dasatinib is an orally active small molecule inhibitor of the src kinase. It has been proposed that dasatinib inhibits growth of breast cancer cells by modulating epidermal growth factor receptor (EGFR) signaling. Approximately 70% of breast cancers over express EGFR family members as well as c-Src. Src is co-expressed with at least one of the EGFRs. Both src and EGFR1 expression have been associated with TNBC. Dasatinib has previously shown activity as a single agent in TNBC cell lines. However, dasatinib has limited single agent activity in a cohort of chemotherapy refractory TNBC patients according to a phase I/II study. We believe that dasatinib may sensitize breast cancer cells to anti-mitotic agents. Ixabepilone is a potent anti-mitotic agent that has shown significant response in TNBC patients, both in the neoadjuvant setting and in the metastatic setting, including those cancers that are taxane refractory. Therefore, combining these two drugs may have utility in this
setting and may lead to inhibition of tumor cell proliferation, invasion, metastasis, angiogenesis, and tumor cell survival. Objectives: To study the activity of dasatinib combined with Ixabepilone in breast cancer cell lines.
Methods: A panel of 8 breast cancer cell lines was used in these experiments. These included MCF-7(ER+), T-47D (ER +), SKBR3 (HER2+), and five triple negative cell lines: BT-549, HS578T, MDA-MB-157, MDA-MB-231, MDA-MB-436. First, these cell lines were evaluated for src expression by Western blot as well as sensitivity to dasatinib alone, ixabepilone alone and then the combination of these two drugs using a high-throughput, 384- well CellTiter-Blue cell viability assay. Percent viability was calculated from four replicate cell wells after 72-hour drug incubation and IC50s were determined. Experiments were repeated a minimum of three times. Synergistic, additive, or antagonistic effects were determined using the combination index (CI) method developed by Chou and Talalay.
Results: TNBC cell lines expressed higher levels of src protein than cell lines with positive expression of HER2, ER and/or PR by Western blot analysis. Viability assays suggested a correlation between src expression and dasatinib IC50. Combinations of dasatinib and ixabepilone were determined to have combinational indexes &lt;1 in all cell lines tested, suggestive of synergistic activity. Conclusion: This study demonstrates that in a range of breast cell lines with varying levels of src expression, dasatinib combined with ixabepilone is a synergistic combination and may prove to be beneficial to patients with very limited treatment options.
Figure available in online version.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P6-15-17.
Abstract PD02-02: Characterization of Mammary Stem Cells during Embryogenesis
Authors: Engle DD , Lin J , Spike B , Cheung S , Wahl G. 
Published: April 2011

Abstract
Mammary stem cells (MaSCs) play a key role in the development and maintenance of the breast. MaSCs self-renew and generate the different cell types of the breast such that a single MaSC can regenerate an entire functional mouse mammary gland. Similarly, tumors are heterogeneous, with some cells exhibiting potent tumor forming ability in transplantation studies. These cells exhibit the stem cell properties of self-renewal and multi-lineage differentiation in that they can serially regenerate the cellular complexity of the original tumor. These cells, referred to as cancer stem cells (CSCs), have in some cases been shown to be more resistant to cancer therapies. Therefore, if even a single CSC survives chemotherapy, it could result in cancer relapse. Consequently, an important goal is to be able to prospectively identify CSCs and elucidate critical growth control pathways to enable development of effective targeted therapies. Unfortunately, pure MaSCs or breast CSCs have not been isolated, precluding the generation of reagents to detect or eliminate them. My research takes a new direction to identify, isolate, and characterize MaSCs using properties central to mammary gland development and stem cell function. We isolated MaSCs during development and found that they are naturally enriched late in embryogenesis. We sorted the mammary cells at E18.5 using cell surface proteins we previously found to be expressed in the rudiment at E18.5 and were able to obtain MaSCs purified to a significantly higher degree than reported in the literature. These fetal MaSCs express several genes implicated in cancer, including erbB2, which is found in a subset of dangerous human cancers. In addition, like human cancers that over-express erbB2, fetal MaSC growth is inhibited by the erbB2 antagonist Lapatinib.
The MaSC population isolated during fetal development is enriched for epithelial cells expressing both myoepithelial and luminal keratin markers as well as epithelial cells expressing neither of these markers. It is possible that either of these sub-populations contains the stem cell activity. We have focused on using high throughput single cell QPCR analysis to cluster the fetal MaSCs by their keratin marker expression pattern and identify cell surface markers that can be used to isolate viable cells for stem cell activity assays. Altogether, these experiments enabled isolation of highly enriched MaSC populations for elucidation of key growth pathways and set the stage for future in vivo analyses.
Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr PD02-02.
Abstract PD02-10: Radiation Induces Notch-Dependent De Novo Generation of Breast Cancer Stem Cells
Authors: Lagadec C , Della Donna L , Vlashi E , Dekmezian C , Pajonk F. 
Published: April 2011

Abstract
Like in normal tissues, the self-renewal of cancer stem cells (CSCs) might be also under tight control of developmental pathway like the Notch, Wnt, Sonic Hedgehog or TGFβ pathways. The Notch pathway plays an important role in normal breast development, cell fate, and normal stem cell self-renewal, and its deregulation has been shown to play a role in cancer. Aberrant Notch signaling has been implicated in the development and progression of both preinvasive ductal carcinomas in situ and invasive. Interestingly, in breast cancer, the Notch pathway plays major role for CSCs maintenance. We previously published that BCSCs (Breast Cancer Stem Cells) are more resistant to radiation. Here we demonstrate a link between BCSCs radio-resistance and the Notch pathway. We show that irradiation of MCF-7 cells increases CSC numbers and that this correlates with an induction of Notch signaling proteins expression in a dose and time specific manner. Jagged 1 was quickly (1h) increased 28-fold after 2Gy, DLL1 was increased 15-fold after 3 to 6h of 2 and 4 Gy irradiation, and Notch 2 was increased 16-fold 6h after 2, 4, 6 or 8 Gy, while DLL3 was increased 10-fold after the highest doses (6 to 12 Gy). Inhibition of Notch signaling pathway by the γ-secretase inhibitor prevents enrichment for CSCs and reduces radiation-induced overexpression of Notch proteins. More interestingly, we demonstrated that radiation-induced Notch signaling contributes to the phenotype plasticity of BCSCs and their progeny. Therefore, ionizing radiation induces de novo generation of BCSCs from non-tumorigenic cells. Moreover, we also identified, by FACS analysis, an induction of polyploid CSCs. expressed a relatively higher level of Oct4 and Sox2 than the unirradiated non-CSCs (two key transcription factors involved in stemness maintenance). In summary, we provide evidence that radiation-induced Notch signaling contributes to the plasticity of the BCSC phenotype, thereby generating BCSCs de novo from non-tumorigenic cells. Our data suggest that stochastic as well hierarchical CSC models apply to breast cancer.
Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr PD02-10.
Abstract PD03-02: A Randomized Pre-Surgical Trial of Metformin in Breast Cancer. Preliminary Feasibility and Safety Results
Authors: Bonanni B , Cazzaniga M , Puntoni M , Pruneri G , Serrano D , Lazzeroni M , Guerrieri-Gonzaga A , Macis D , Luini A , Veronesi P , Galimberti V , DeCensi A. E.O. 
Published: April 2011

Abstract
Background: Metformin has been associated with decreased breast cancer (BC) incidence in diabetic patients in epidemiological studies. Moreover, this drug results in initiation of an AMPK-dependent energy stress response which can adversely affect survival of breast cancer cell lines and inhibition of PI3K/Akt/mTOR signaling leading to reduced proliferation of BC cell lines.
Methods: We conducted a randomized, pre-surgical, phase IIb, placebo-controlled, biomarker trial in women with stage I-III BC candidate to elective surgery. The primary endpoint was the change in cell proliferation in malignant, dysplastic and hyperplastic tissue as measured by Ki-67 labeling index (LI). With 150 subjects, the study was 80% powered to test for the interaction between metformin activity and ER status. Two interim analyses are being planned after 100 and 200 women enrolled. Results: As of June 18, 2010, 175 subjects have been randomized and 162 have completed treatment. Here we report data on the first 100 women enrolled. At the symposium, full data on the first 200 women enrolled will be available. As of December 31, 2009, a total of 163 women were screened, 26 were not eligible and 40 refused to participate, 6 dropped out during the study for AEs (n=2) or refusal to continue treatment (n=4), thus leaving 95 subjects assessable for the primary endpoint. The main subject and tumor characteristics blinded as to the allocated arm were mean age 52 (31-77), Pre/postmenopause, 54/41, mean BMI, 23.6 (18.0-40.2). At baseline, median Ki-67LI was 18% (range 4%-65%) at biopsy and 19% (4%-70%) after 4 weeks at surgery. All adverse events except for 1 SAE were grade 1 or 2, consisting of G2 nausea and G2 diarrhea in 4% and 6% of the cases, respectively. The prevalence of ductal intraepithelial neoplasia was 91% (median Ki-67 LI, 10%) in samples both adjacent and distant from the tumor, whereas the prevalence of ductal hyperplasia was 77% (median Ki-67LI, 2%) in samples distant from the tumor. Conclusions: Our preliminary results show the feasibility, high compliance and safety of a metformin trial in breast cancer patients. Assessment of tissue and circulating biomarkers is currently ongoing to characterize the whole spectrum of metformin activity in malignant and dysplastic tissue.
Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr PD03-02.
Abstract PD03-09: Metformin Induces Apoptosis in Triple Negative Breast Cancer Cells Via Inhibition of Stat3 Activity
Authors: Deng XS , Wang S , Deng A , Liu B , Edgerton SM , Thor AD. 
Published: April 2011

Abstract
Background: Metformin inhibits breast cancer cell proliferation and colony formation, with S phase arrest and the induction of apoptosis in triple negative breast cancer cells in vitro. In these cells, metformin down-regulates Cyclin D1 and Cyclin E, as well as EGFR, p-EGFR, p-AKT, p-MAPK, p-Src and p-mTOR, whereas it up-regulates p-AMPK (Liu, et al. Cell Cycle, 2009). Non-triple negative breast cancer cells are resistant to metformin induced apoptosis, although they show similar changes in p-AMPK induction with metformin (Alimova, et al. Cell Cycle, 2009). We hypothesized that there are unique signaling intermediates associated with metformin responsivity in triple negative cells and have identified the signal transducer and activator of transcription 3 (Stat3) as a potential target. Stat3 is constitutively activated in a wide range of tumors, including breast cancer (up to 60%). It reportedly promotes cancer cell proliferation and survival.
Methods: Human triple negative breast cancer cell lines (MDA-468, MDA-231, BT20 and HCC70) were used to evaluate interactions between metformin and Stat3 signaling. Activation of Stat3 was examined by Western blot analysis with phosphorylation-specific antibodies. Cell proliferation was determined by MTS assay. Apoptosis was quantitated by an apoptosis ELISA assay and Western blots for PARP and caspase cleavage. In these 4 cell lines, Stat3-over-expressing clones were obtained via transfection of a constitutive active (CA) construct of Stat3. Specific knock-down of Stat3 expression was achieved by using a lentiviral system containing Stat3 small hairpin RNA.
Results: In a dose and time dependent manner, metformin inhibits Stat3 phosphorylation at sites Tyr705 and Ser727, Cyclin D1 and E protein expression, cleavage of PARP and the pro-caspases 3, 8, and 9. Overexpression of the CA-Stat3 attenuates the aforementioned meformin-associated PARP and caspase cleavage as well as apoptosis, and it suppresses metformin induced cell cycle arrest and changes in cyclin D1 and E. In contrast, specific knock-down of Stat3 expression sensitizes the triple negative breast cancer cells to metformin-mediated apoptosis and cell cycle arrest, enhancing the signaling changes we describe above. Conclusion: These data indicate that Stat3 is a critical intermediary for metformin action in triple negative breast cancer cells. Our studies suggest that targeting Stat3 activation may be a useful strategy to treat breast cancer patients with triple negative phenotype.
Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr PD03-09.
Abstract PD04-01: Does Analysis of Disseminated Tumor Cells in Bone Marrow Give Additional Prognostic Information in Primary Breast Cancer? Analysis of Disseminated Tumor Cells in Bone Marrow — Report of a Prospective Study with 5 Years Follow-Up
Authors: Falck A-K , Bendahl P-O , Ingvar C , Lindblom P , Lövgren K , Rennstam K , Fernö M , Rydén L. 
Published: April 2011

Abstract
Introduction: Disseminated tumor cells (DTC) in bone marrow of patients with breast cancer has, beyond the formerly known and clinically used prognostic markers, in recent publications been identified as an independent predictor of poor prognosis in patients with non-metastatic disease. Detection of DTC could therefore be a way to identify patients with an increased risk of recurrence among patients with a “good” standard prognostic profile. The present prospective study aimed to evaluate the presence and prognostic value of DTC in bone marrow in women undergoing surgery for primary breast cancer.
Material and methods: From 1999 to 2003 bone marrow aspirates were obtained from the sternal crest in 569 patients during surgery for primary invasive breast cancer. Tumor cells in bone marrow were identified with cytokeratins (4,5,6,7,8,10,13,14,15,16,18,19) against epithelial cells and analyzed with immunofluorescence and immunohistochemistry. The results were related to clinical follow up for 5 years using distant disease free survival (DDFS) and breast cancer specific survival (BCSS) as end-points by uni-and multivariate analysis including standard prognostic factors.
Results: Analysis of DTC were performed in 445 patients and cytokeratin positive cells were found in 159 of these (36%). The presence of DTC in bone marrow was not related to lymph node metastases (N+ 38% vs N-35%, p= 0.4), ER+/ER-(38% vs 30%, p= 0.2) or histological grade (40% grade 1; 37% grade 2; 30% grade 3, p= 0.18). BCSS was related to: lymph node engagement (+ vs -; HR 3.5, p&lt; 0.001), ER (+ vs -; HR 0.55, p= 0.046), histological grade (2 vs 1: HR 4.9, p= 0.03; 3 vs 1: HR 7.4, p= 0.008), age (HR 1.02/year, p= 0.04) and tumor size (&gt;20 mm vs &lt;20 mm: HR 1.8, p= 0.05) in Cox multivariate analysis. Presence of DTC did not correlate either with DDFS (p= 0.97) or BCSS (p= 0.75). In the subgroup N0/ER-, a significantly higher rate of distant metastases was observed in patients with micrometastases in bone marrow (2/12 vs 0/25; p= 0.04; log rank test) and BCSS in the same subgroup was significantly related to the presence of DTC in bone marrow (2/12 vs 0/25; p= 0.04).
Conclusion: Detection of micrometastases in bone marrow in primary breast cancer has previously been shown to be a significant independent predictor of poor prognosis in different studies. In a consecutive cohort including unselected patients we were not able to confirm these results. Our data suggest that it is too early to implement the method of bone marrow aspiration and diagnosis of DTC in clinical routine.
Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr PD04-01.
Abstract PD04-04: Cytokeratin-19 mRNA Positive Circulating Tumor Cells during Follow-Up of Patients with Early Breast Cancer: Prognostic Relevance for Late Relapse
Authors: Saloustros ES , Perraki M , Apostolaki S , Politaki E , Agelaki S , Kallergi G , Xyrafas A , Georgoulias V , Mavroudis DA. 
Published: April 2011

Abstract
PURPOSE: We reported previously that the detection of CK-19 mRNA-positive circulating tumor cells (CTCs) after adjuvant chemotherapy in patients with early breast cancer is associated with poor clinical outcome. Reliable prognostic markers for late disease relapse are not available. In this study we investigated the prognostic value of CTCs’ detection during the first 5 years of follow-up in predicting late disease relapse. PATIENTS AND METHODS: Blood was obtained from 281 patients with early breast cancer who had not experienced disease relapse during the first two years of follow up. CTCs were detected using a real-time reverse transcriptase polymerase chain reaction for CK 19 mRNA after the completion of adjuvant chemotherapy and every six months thereafter for a 5-year follow up period. Patients were classified into four groups based on the CTCs’ status during the first two years and the subsequent three years of follow up (persistently negative, persistently positive, negative turned to positive and the opposite).
RESULTS: The vast majority of patients (92.9%) received some kind of hormonal therapy in addition to chemotherapy. Sixty-nine (24.6%) patients remained CTCs-free throughout the 5-year period. A change in CK-19 mRNA status was observed in 126 (44.9%) patients; 60 (21.4%) patients with initially CK-19 mRNA-positive CTCs during the first 24 months turned CTC-negative afterwards while 66 (23.5%) patients who were initially CTC-negative became CTC-positive. Eighty-six (30.6%) patients remained persistently CK-19 mRNA-positive for the entire 5 years. After 94 months of median follow up (range: 51-147), the relapse rate for each of the above groups of patients is illustrated in the following table.
Table 1: CTCs’ detection status and risk of late disease repapse.
Compared to the persistently negative, only the group of CK-19 mRNA-persistently positive patients had a statistically significant higher risk of relapse (38.4% versus 7.2%, p&lt; 0.001), shorter progression-free survival (not reached vs 135 months, P&lt;0.001) and significant difference in overall survival (not reached vs 146 months, p=0.045).
CONCLUSION: Persistent detection of CK-19 mRNA-positive CTCs in the blood during the first 5 years of follow up after adjuvant chemotherapy is associated with an increased risk of late disease relapse indicating the presence of chemotherapy-and hormonetherapy-resistant residual disease. This prognostic evaluation may be useful when deciding on subsequent adjuvant systemic therapy.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr PD04-04.
Abstract PD04-05: Prognostic Value of Circulating Tumor Cells According to Immunohistochemical Defined Molecular Subtype in Advanced Breast Cancer Patients
Authors: Munzone E , Botteri E , Nolè F , Esposito A , Adamoli L , Zorzino L , Sciandivasci A , Cassatella MC , Goldhirsch A , Rotmensz N , Sandri MT. 
Published: April 2011

Abstract
Background: Breast cancer (BC) is not a single disease with variable morphologic features and biomarkers but, rather, a group of molecularly distinct neoplastic disorders. Four main molecular classes of BC have been distinguished by gene expression profiling, called luminal-A, luminal-B, HER2-positive and triple-negative (TN) cancers. These subgroups correspond reasonably well to clinical characterization on the basis of estrogen receptor (ER) and HER2 status, as well as proliferation markers or histologic grade. Correlating circulating tumor cells (CTCs) with molecular subtypes might identify subgroup of patients (pts) at different risks and groups that might benefit more from prognostic evaluation by CTCs. Patients and methods: We retrospectively analyzed 187 consecutive metastatic BC pts who had baseline CTCs enumeration performed between March 2005 and December 2009 by CellSearch. Pts were categorized in 3 prognostic groups in accordance to the number of CTCs, 0 CTC, &lt;5 CTCs and ≥5 CTCs, and in 4 categories based on the tumour biology: luminal-A with ER &gt;1%, any progesterone receptor (PgR), G1/G2, Ki67&lt;30%; luminal-B with ER &gt;1%, any PgR, G3, Ki67&gt;30%; HER-2 positive with HER-2 overexpressed or FISH amplified; TN with both ER and PgR 0% and HER-2 not overexpressed/FISH not amplified. Kaplan-Meier method for survival analysis was used.
Results: Median age at baseline was 56 years (range 31-78). CTCs were a strong predictive factor for overall survival (OS) in all pts (Log-Rank P&lt;0.001), in pts with bone metastases (MTS) only (n=43; p=0.045), visceral MTS only (n=42; borderline significant p=0.08) and both bone and visceral MTS (n=74; P&lt;0.001). The same figure was not observed in the few pts with only soft tissue MTS (n=28; p=0.315). Sixty-one (33%) pts had Luminal-A tumour, 58 (31%) Luminal-B, 53 (28%) HER-2 positive and 15 (8%) TN. CTCs were a significant prognostic factor in all molecular subtypes as shown in Figure 1. Conclusion: This large series confirms CTCs as an important prognostic factor for metastatic BC in all different molecular subtypes, but pts with 0 CTCs at baseline and TN or luminal B tumors seem to perform better as compared to other categories. Furthermore, pts with metastatic disease including bone and/or visceral sites benefited most from CTCs prognostic evaluation. Further studies with larger series and longer follow-up time could help identify metastatic BC subgroups in which CTCs analysis would be particularly useful and should be recommended. Figure available in online version.

Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr PD04-05.