bluuebunny/crossref_metadata_2025 huggingface dataset:
P3-14-22: Response to Neoadjuvant Chemotherapy in Elderly Patients with Locally Advanced Breast Cancer.
Authors: Vanderhaegen J , Wildiers H , Christiaens M-R , Smeets A , Leunen K , Neven P , Vergote I , Vanlimbergen E , Weltens C , Janssen H , Moerman P , Brouckaert O , Brouwers B , Paridaens R
Published: June 2012

Background

Inoperable, locally advanced breast cancer (LABC) is common in the elderly. Studies have shown good responses with neoadjuvant chemo- and endocrine therapy. However, studies targeting the elderly population are rare.

Aim: To assess complete pathological response rate (pCR) following neoadjuvant chemotherapy (NACT) and to evaluate the effectiveness of this therapeutic option in elderly women with LABC.

Patients: Women (≥70 years) diagnosed at our University Hospital with LABC between 1999 and 2010 and treated with neoadjuvant chemotherapy. After neoadjuvant systemic treatment pCR was assessed if surgery was performed.

Results: 44 women with a mean age of 74.9 (70-93) years were included. Most tumors were ductal (IDA 88.6%), of grade 3 (79.5%) and hormone-receptor (HR) negative (61.4%). Among the 21 tumors (47.7%) that were HER2+ (immunohistochemistry (IHC) 3+ or 2+ and FISH+), all but 3 received neoadjuvant Trastuzumab. Only 4 patients had no surgery (clinical decision or personal choice) while the remaining 40 patients underwent either breast-conserving surgery (17.5%) or mastectomy (82.5%). 23 patients (52.3%) had pathological partial response, 14 patients (31.8%) pCR and 3 patients no response. pCR was more frequently achieved in HR-negative (44.4%) than in HR- positive patients (11.8%). In both groups HER2 positive tumors had better pathological response than HER2 negative tumors. After a mean follow-up of 41 months (5-120) 29 patients (65.9%) had no evidence of disease, 6 patients (13.6%) had (loco)regional recurrence, 12 patients (27.2%) developed metastatic disease and 6 patients (13.6%) died because of breast cancer. Relapse was more frequently seen in patients with a pPR (47.8%) versus patients with a pCR (14.3%).

Conclusion: In a selected population of elderly patients with LABC and treated with neoadjuvant chemotherapy, a reasonable percentage of pCR can be achieved. Concordant with younger populations, this percentage is higher in HR-/HER2 positive breast cancer. Neoadjuvant chemotherapeutic treatment can thus be as beneficial for the older patient as for younger ones. In fit elderly patients, neoadjuvant chemotherapy should therefore not be omitted solely on the basis of advanced age.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P3-14-22.
P3-15-02: The Change of Bone Turnover Markers during Neoadjuvant Anastrozole Versus Exemestane: A Randomized Single-Center Study.
Authors: Jinno H , Shimada K , Takahashi M , Hayashida T , Hirose S , Ikeda T , Kitagawa Y
Published: June 2012

Background: Non-steroidal aromatase inhibitors including anastrozole and letrozole have been reported to increase the rate of bone turnover, accelerate loss of bone, and increase the incidence of fractures. Although steroidal inactivator exemestane has similar inhibitory effect on the aromatization, animal studies have shown a weak but potentially important anabolic effect of exemestane metabolites, which might lead to decreased bone resorption. This randomized trial was conducted to compare the effect of exemestane and anastorozole on bone turnover markers.

Patients and Methods: Fifty-two postmenopausal women with ER-positive, HER2−negative, invasive, nonmetastatic, and operable breast cancer were randomly assigned to neoadjuvant exemestane (25 mg daily) or anastrozole (1 mg daily) for 4 months. The primary endpoint was change in bone turnover markers including resorption markers urinary and serum N-telopeptide (NTX) and the formation markers serum bone alkaline phosphatase (BAP). The secondary endpoint was tumor objective response (OR) assessed by both caliper and ultrasound. Comparisons were also made of breast conservation rate and adverse events.

Results: The changes in serum NTX from baseline to week 16 were not statistically different between anastrozole and exemestane. The changes in urinary NTX from baseline to week 16 were also not statistically different between both groups .BAP did not show any significant increase in the exemestane group (10.8%; p=0.21), whereas the increase in BAP from baseline to week 16 in the anastrozole group was 15.0% and marginally significant (p=0.05). There were no significant differences in OR in the intent-to-treat population between patients receiving anastrozole or exemestane (45.8% vs. 39.2%; p=0.63). The OR was similar between the patients with a baseline Ki67 index of ≥15% and <15% in exemestane group (37.5% vs. 41.2%; p=0.86). In anastrozole group the OR in patients with a baseline Ki67 index of ≥15% and <15% was 44.4% and 53.8%, respectively(p=0.66). Breast conservation rate was similar between anastrozole and exemestane (62.5% vs. 67.9%; p=0.68). Treatment was well tolerated and much the same for both groups.

Conclusions: There is no significant differences of the change in bone turnover markers between anastrozole and exemestane. These results indicate exemestane had no anabolic effects which would lead to fewer adverse effects on bone.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P3-15-02.
P3-16-01: Safety Profile and Clinical Activity of Single-Agent BKM120, a Pan-Class I PI3K Inhibitor, for the Treatment of Patients with Metastatic Breast Carcinoma.
Authors: Rodon J , Bendell JC , Abdul Razak AR , Homji N , Trandafir L , Quadt C , Graña-Suárez B , Siu LL , Di Tomaso E , Demanse D , Massacesi C , Hirawat S , Burris III HA , Baselga J
Published: June 2012

Background

Phosphatidylinositol 3-kinase (PI3K) is critical to cancer cell growth, survival, and metabolism. BKM120 is an oral pan-class I (α, β, γ, δ) PI3K inhibitor that has demonstrated in vitro and in vivo tumor cell growth inhibition in a range of cancer types including breast cancer.

Materials and methods: The Phase I study CBKM120X2101 investigating single-agent daily BKM120 in patients (pts) with advanced solid tumors has been recently completed with the maximum tolerated dose established at 100 mg/day. Here, we report the analysis of metastatic breast carcinoma (MBC) pts enrolled in this study.

Results: Overall, 83 pts have enrolled, 21 of whom have MBC. At the cut-off date of 25th February 2011, 20 MBC pts were evaluable: 1 pt at 80 mg, 1 pt at 150 mg and 18 pts at 100 mg. Patient characteristics were as follows: median age 55 years (range 37–71); performance status ECOG 0/1/2 for 7/12/1 pts, respectively; visceral disease was reported in 16 pts, including liver, 10 pts (50%); lung, 9 pts (45%); and pleura, 5 pts (25%); all pts had >3 lines of systemic therapy (3-12). The median time from last treatment and study entry was 46 days (29-235). The median duration of BKM120 treatment administered was 7.5 weeks (1.0−96.4). The most frequent grade 3 drug-related adverse events (AEs) were: transaminases increase, 4 pts; psychiatric disorders, 3 pts, consisting of anxiety, affective disorder, and mood alteration (1 pt each); diarrhea, 2 pts; fatigue, 2 pts; and hyperglycemia, 1 pt. The only grade 4 drug-related AE was hyperglycemia, reported in 1 pt at 150 mg. Most AEs were manageable with treatment interruption and dose reductions. Eighteen pts were evaluable for objective tumor response by RECIST. Two pts (11%) exhibited partial responses, which were confirmed in a triple-negative MBC pt, and unconfirmed in an ER+ HER2− MBC pt. For these 2 pts, the treatment duration was 29+ (ongoing) and 6 months, respectively. An additional 9 pts (50%) had stable disease, lasting >4 months in 7 pts (35%).

Conclusions: This preliminary analysis showed that BKM120 has single-agent activity in heavily pretreated pts with MBC, and an acceptable safety profile. Molecular profiling and updated pharmacokinetic results will be presented at the meeting.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P3-16-01.
P3-16-03: Enhancing Anti-IGF Directed Therapeutics by Co-Targeting Autophagy.
Authors: Oh AS , LaPara K , Yee D
Published: June 2012

Autophagy (macroautophagy) is a cellular mechanism that maintains homeostasis and cell growth by regulating the turnover of damaged proteins. Autophagy has a critical role in maintaining cellular viability during nutrient starvation and metabolic stress by recycling intracellular proteins for nutrient reallocation. Whether the autophagy pathway utilized by breast cancer cells has a survival mechanism or is a feature of cell death is not fully understood. Since the IGF/insulin signaling maintains cellular growth, nutrient utilization, and survival, increased autophagy or autophagic flux might be induced with IGF-I inhibitors as a compensatory survival mechanism. In breast cancer cells treated with anti-IGF1R kinase inhibitors, we observed an increase in autophagic flux; similar to levels induced by nutrient deprivation.

Objective: In this study, we examined whether IGF1R inhibitor -induced autophagy is cytoprotective or cytotoxic. If autophagy has a protective role in nutrient-depleted conditions it could reduce the long term efficacy of IGF1R inhibition by making cells refractory to blockade of this signaling system. In this study, we hypothesized that blocking autophagy in combination with anti-IGF1R therapies (antibodies or small molecules) would further enhance growth inhibition in a panel of breast cancer cells types (luminal and basal-like/triple negative).

Results: We initially examined whether autophagy inhibitors (chloroquine or bafilomycin A1) had a differential growth effect on breast cancer cells that varied in their sensitivity to IGF1R stimulated growth. We found that low-to-moderate IGF growth dependent MDA-MB-231 and MDA-MB-435A breast cancer cells were more sensitive to chloroquine alone. However, sensitivity to bafilomycin A1 differed between breast cancer cell lines compared to chloroquine. In conditions of cellular stress when autophagy is normally triggered, co-treatment with IGF inhibitor (OSI-906) and chloroquine, increased growth inhibition in all breast cancer cell lines. Further, basal-like breast cancer cells (i.e. low-to-moderate IGF growth dependent cells) had a higher baseline autophagic flux compared to luminal MCF-7L breast cancer cells (i.e. highly IGF-I dependent). Using MCF-7L cells selected for long-term resistance to IGF1R kinase inhibitor (AEW-541), we found that these cells were not sensitivite to chloroquine or Bafilomycin A1 compared to wild type MCF-7Ls. Inhibiting autophagy in combination with anti-IGF therapies could offer benefit over single therapy alone if autophagy inhibitors are given together with IGF1R inhibitors, but not in IGF1R inhibitor resistant cancers. Additionally, targeting autophagy as a single agent therapy may be more beneficial in triple negative breast cancer compared to other subtypes of breast cancer.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P3-16-03.
P3-16-16: Overcoming EGFR Resistance Using Dasatinib in Combination with Cetuximab and Cisplatin in Triple Negative Breast Cancer Cell Lines.
Authors: Kim EMH , Gartner E , Choi L , Boerner J
Published: June 2012

Background: Patients presenting with triple negative breast cancers (TNBC's) have a poorer prognosis compared to those with other subtypes of breast cancer, and effective therapeutic targets have yet to be identified. The majority of TNBC's overexpress EGFR, however studies have demonstrated that EGFR inhibition as a monotherapy is ineffective. Interestingly, adding the EGFR inhibitor, cetuximab to cisplatin chemotherapy doubled both the response rate and time to progression compared with patients only on cisplatin. c-Src tyrosine kinase plays a critical role in signal transduction downstream of growth factor receptors, and has been found to contribute to the resistance of TNBC cell lines through the phosphorylation of EGFR. We hypothesize that dasatinib, a c-Src inhibitor, may help overcome EGFR resistance and in combination therapy enhance the cytotoxic activity.

Material and Methods: A panel of breast cancer cell lines, including 7 triple negative cell lines, were tested using increasing doses of dasatinib (1 nM to 10 μM), cetuximab (1 μg/ml to 100 μg/ml), and cisplatin (10 nM to 10 μM) alone and in combination. Growth inhibition was determined after 3 days of treatment using the MTT colorimetric assay, and apoptosis was assessed through Caspase-3 activation and TUNEL assays.

Results: All cell lines were generally resistant to cetuximab, with 100 μg/ml required to decrease cell viability. Two TNBC lines, SUM149 and SUM229, showed an additional decrease in cell viability with dasatinib added to cetuximab, while SUM102 and BT20 cells showed an additional decrease in viability using the dasatinib and cisplatin combination, compared to single agents. In 4 out of 7 TNBC lines, the combination of dasatinib, cisplatin, and cetuximab showed a significant decrease in cell viability compared to the combination without dasatinib (p-value < 0.02). The decrease in proliferation using the combination of dasatinib, cisplatin, and cetuximab was due to an increase in apoptosis. In contrast, this combination did not increase the loss of cell viability or apoptosis in ER/PR+ and HER2+ breast cancer cell lines.

Discussion: Targeting c-Src family kinases with dasatinib may help overcome the resistance to EGFR inhibition in triple-negative breast cancer. This may have significant clinical implications in treating TNBC patients, and hence further investigation into the mechanism of this effect is warranted.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P3-16-16.
P3-17-11: Dovitinib (TKI258), a Dual Inhibitor of FGFR and VEGFR, Induces Tumor Growth Suppression in Xenograft Models of Primary Human Breast Cancer.
Authors: Shi MM , Linnartz R , Versace R , Graus Porta D , Kay A , Dugan M
Published: June 2012

Introduction: The objective of this study was to evaluate the efficacy of dovitinib (TKI258), a small-molecule dual inhibitor of fibroblast growth factor receptor (FGFR) and vascular endothelial growth factor receptor (VEGFR) in mouse models of human breast cancer. Molecular epidemiological studies have implicated that FGFR1 and FGFR2 gene amplification plays a critical role in breast cancer tumorigenesis. FGFR1 gene amplification correlates with gene expression and is associated with poor prognosis in patients with estrogen receptor-positive (ER+) tumors. Dual FGFR/VEGFR inhibitory activities of dovitinib make it an attractive molecule for targeting breast cancer.

Experimental Design: Dovitinib was evaluated in 3 primary human breast carcinoma xenograft models with different histological and molecular profiles: a triple-negative breast cancer model (HBCx-15), a HER2+ model (HBCx-5), and an ER+ model (HBCx-3). Dovitinib was administered orally at 20- and 40-mg/kg dose levels (n = 8–10/group), once daily for 21 consecutive days.

Results: At a dose of 40 mg/kg, dovitinib demonstrated strong antitumor efficacy as a single agent, particularly in the ER+ HBCx-3 and the HER2+ BCx-5 tumor models (ΔT/T0 = −26.42% and ΔT/ΔC = 5.09%, respectively), whereas the response was less significant in the triple negative breast cancer HBCx-15 model (ΔT/ΔC = 31.7%).

Conclusion: Dovitinib showed significant antitumor effect in human breast cancer xenograft mouse models. This study provides a strong rationale for clinical investigation of dovitinib in patients with advanced breast cancer, particularly in patients with ER+ and HER2+ disease.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P3-17-11.
P3-18-01: cMET Inhibitor and the Inhibition of Growth of Breast Cancer Cells in Bone Marrow Matrix Environment.
Authors: Ye L , Mason MD , Bramble P , Jiang WG
Published: June 2012

Background: Hepatocyte growth factor (HGF) is a cytokine that has a diverse but potential role in cancer including breast cancer. HGF, which action is mediated by its specific receptor, cMET, stimulate the aggressiveness of cancer cells by increasing the invasiveness and cellular migration. HGF is also a potent angiogenic factor. Small inhibitors to the HGF receptor are currently investigated in clinical trials of various cancers. In solid tumours which have potency of bone metastasis, HGF and particularly the HGF receptor, cMET, have been found to be particularly over-expressed in tumour cells metastasised to bones.

Materials and methods. A panel of breast cancer cell lines were used. Cell growth was determined using a colorimetic method. Cell adhesion and migration were investigated using a ECIS model. Bone matrix proteins were prepared from fresh bones. A small inhibitor to human cMET, PF02341066 was used in the present study in all the cell models.

Results. Depending on cell types, cancer cells tended to grow at a faster rate in bone marrow microenvironment than under normal conditions (12% higher for MDA MB 231 cells, for example). While PF02341066 have a concentration dependent effect on the growth of breast cancer, this effect became more marked when cells were growing in the bone marrow matrix. Breast cancer cells migrated rapidly in the presence of bone marrow environment, in comparison with controls. This was also inhibited by the cMET inhibitor. The study further showed that cells over-expressing a molecule linked to bone metastasis, namely ALCAM (1,2), responded more vigorously to cMET inhibitor in the matrix adhesion and cellular migration.

Conclusions. The present study has shown that the HGF receptor inhibitor has an inhibitory effect on breast cancer cells. This is particularly so when cells are grown in bone matrix microenvironment. It is concluded therefore that inhibitors to the HGF receptor may have a particular role in the management of bone metastasis in breast cancer.

References: 1. Davies S, Jiang WG. ALCAM, activated leukocyte cell adhesion molecule, influences the aggressive nature of breast cancer cells, a potential connection to bone metastasis. Anticancer Res. 2010, 30(4):1163–8; 2. King JA, et al. Activated leukocyte cell adhesion molecule in breast cancer: prognostic indicator. Breast Cancer Res. 2004;6(5):R478–87

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P3-18-01.
P4-01-10: The Role of the Steroid Receptor Coactivator SRC1 and Its Functional Partner HOXC11 in the Development of Endocrine Resistant Breast Cancer.
Authors: Walsh CA , McCartan D , Hill ADK , Young LS
Published: June 2012

Background: The steroid receptor coactivator; SRC1, has been well described in the development of endocrine resistant breast cancer. SRC1 associates with clinically aggressive tumours and promotion of distant metastasis. It directly interacts with the developmental transcription factor, HOXC11 and together they are found to strongly predict poor disease-free survival in breast cancer patients (hazard ratios: 5.79; P < 0.0001). In this study, we investigate the mechanism of SRC1 and HOXC11 action in tumour adaptability and subsequent resistance to endocrine therapy.

Materials and Methods: Cells which are resistant to tamoxifen (LY2s) have enhanced SRC1 and HOXC11 mRNA and protein expression in comparison to their endocrine sensitive parent cells (MCF-7s). ChIP-sequencing data for SRC1- and HOXC11- DNA interactions in conjunction with DNA microarray and RNA-sequencing data identified potential signalling targets at play in the LY2 model of endocrine resistance. Real-time analysis and flow cytometry confirmed these interactions at a transcriptional and protein level. These observations were further confirmed in primary breast cancer cultures using flow cytometry.

Results: SRC1 and HOXC11 interactions are driven in tamoxifen treated LY2 resistant cells. Combined SRC1 ChIP-sequencing and expression array data analysed in conjunction with HOXC11 ChIP-sequencing and RNA-sequencing data reveal that the SRC1/HOXC11 transcriptional process can orchestrate the loss of luminal cell markers such as ERα, CD24 and PTCH1 whilst concomitantly upregulating mediators of tumourigenicity such as CD44 and MSI2. Primary breast cancer cultures confirm the loss of CD24 in tamoxifen resistant patients. In these patients, loss of CD24 was accompanied by loss of steroid receptor expression (ERα and PR) and by a gain of the basal marker CD44.

Discussion: Here, we describe a new signalling pathway where SRC1 and HOXC11 regulate two distinct but complementary mechanisms to drive tumour adaptability. Silencing of luminal cell markers and a concomitant increase in a basal cell phenotype has the potential to alter the survival mechanism of breast cancer cells to evade targeted therapy.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-01-10.
P4-02-04: Androgen Receptor (AR) Expression in a Cohort of Patients (pts) with Triple Negative Breast Cancer (TNBC).
Authors: Gucalp A , Gupta G , Patil S , Wen YH , Akram M , Brogi E , Powell SN , Ho AY , Hudis CA , Traina TA
Published: June 2012

Background

TNBC, defined by the absence of ER, PR, and HER2, is associated with higher risk of recurrence and BC-related mortality, earlier age at diagnosis, menarche, and 1st pregnancy, increased parity, higher BMI, and African-American/Hispanic race. TNBC is a heterogeneous group. Using gene expression analysis, our group described a subset of AR+ ER/PR- BC that exhibits androgen-dependent growth. In vitro studies confirmed the functional role of AR and showed that growth could be abrogated by antiandrogens.(Doane et al 2006) We translated this work into a phase II trial of bicalutamide in pts with AR+ ER/PR- metastatic BC (MBC). (NCT00468715) We now describe the prevalence and clinicopathological characteristics of AR+TNBC in primary disease in a single-institution retrospective cohort.

Methods: We identified 1,032 pts with resectable, TNBC (ER/PR<1%; HER2<2+/FISH<2.2) who had surgery at MSKCC from 1998–2006. Exclusion criteria: neoadjuvant chemotherapy, prior radiation, inflammatory/MBC. IRB approval was obtained. We constructed tissue microarrays (TMA) from 210 primary tumors (> 1 cm) with each tumor represented by three 0.6mm cores. AR was tested with DAKO antibody (Clone AR441; dilution 1:500). TMAs were digitized with a Mirax scanner. MetaMorph image analysis software was used to quantify the ratio of DAB staining to hematoxylin signal. A ratio >1 SD above mean was defined as AR+. AR+ cores were manually reviewed; false positives due to core artifact were excluded. To evaluate clinicopathological variables and differences in recurrence-free survival (RFS) and overall survival (OS) by AR status we used chi-square/t-tests and Kaplan-Meier methods/log-rank test, respectively.

Results: 169 pts had adequate cores for image analysis/quantification of AR. 10% of pts tested AR+ (17/169). Median (med) followup: AR+=6 years (yr), AR-=5.6yr. Demographic/clinicopathological variables: Table 1 (ages in med yr). Overall med age=54yr (29-84). Adjuvant chemotherapy received: AR+ 82%, AR- 87%, p =0.40; 77% received anthracycline/taxane-based therapy. Med time to distant metastasis (DM)=2.1yr (0.2−6.2yr). We were unable to demonstrate a difference in 5yr RFS (69% vs. 77%; p=0.37) or OS (68% vs. 84%; p=0.25) between AR+ and AR- TNBC.

Conclusions: Consistent with our prospective study, AR is expressed in ∼10% of TNBC tumors in this retrospective cohort. The pts in our dataset may be older, postmenopausal, more likely to self-report white race and have T1-2/N0-1 BC. No statistically significant differences were observed in demographic/clinicopathological variables or survival outcomes between AR+ and AR- TNBC. Additional TMA data from our database will be presented.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-02-04.
P4-02-05: The Regulation of Artemin Signalling by IGF-1 in Mammary Carcinoma Cells.
Authors: Yip C , Liu D , Lobie PE , Perry JK
Published: June 2012

Artemin is a neurotrophic signalling factor which belongs to the glial-derived neurotrophic factor (GDNF) family of ligands. Artemin acts as a survival, proliferation and migration factor for a number of neurological cell types, by signalling through the RET (rearranged during transfection) receptor and, in most cases, the GDNF receptor (GFR)-a3 co-receptor. Recently, a number of published studies have implicated Artemin as a potential oncogene in several cell types, including mammary carcinoma cells. Other studies further indicate that Artemin may influence cancer progression and tamoxifen resistance in some breast cancers. Available clinical data has demonstrated that increased Artemin expression is correlated with decreased overall patient survival in breast cancer patients and a poor outcome in tamoxifen treated breast cancer patients. Here we investigate interaction between the Artemin and the insulin-like growth factor-1 (IGF-1) signal transduction pathways. Using mammary carcinoma cell lines, we demonstrate that IGF-1 treatment increases the endogenous expression of both Artemin and its endogenous receptors, RET and GFRa3. Semi-quantitative RT-PCR assays demonstrated that IGF-1 stimulated mRNA expression of Artemin as well as RET and GFRa3 in wild-type MCF-7 and ZR-75-1 cells in a time-dependent and dose-dependent manner. The same effect was not observed in wild-type T47D cells where IGF-1 did not increase Artemin mRNA expression.

We also demonstrated that forced expression of Artemin in MCF-7 cells consistently enhanced the response of these cells to IGF-1 in a number of cell function assays. Forced expression of Artemin significantly enhanced IGF-1-mediated stimulation of total cell number in MCF-7 cells. Consistent with this, Artemin enhanced IGF-1-mediated stimulation of S-phase entry and cell survival. In a soft agar assay, forced expression of Artemin also enhanced IGF-1-mediated stimulation of colony formation. Conversely, depletion of Artemin expression using siRNA abrogated the response to IGF-1 stimulation in MCF-7 cells. Artemin depletion significantly decreased IGF-1-stimulated increase in total cell number by decreasing IGF-1-stimulated cell proliferation and protection from apoptotic cell death. In addition, forced expression of Artemin in MCF-7 cells reduced cell sensitivity to the IGF-1 receptor small molecule inhibitor, AG1024. In conclusion, we have demonstrated that IGF-1 increases Artemin mRNA and protein expression in the breast cancer cell lines MCF-7 and ZR-75-1 and have identified potential cross-talk between the Artemin and IGF-1 signalling pathways in MCF-7 cells.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-02-05.
P4-02-06: Progesterone Receptor Expression Predicts Poor Outcome in Estrogen Receptor Positive, Lymph Node Negative Breast Cancer – A Population Based Study.
Authors: Purdie CA , Quinlan P , Jordan LB , Ashfield A , Baker L , Dewar JA , Thompson AM
Published: June 2012

Background: Estrogen receptor (ER) positive, lymph node (LN) negative breast cancer usually carries a good prognosis; endocrine therapy is often the only adjuvant treatment. However, a small proportion of such patients do badly (possibly representing those with luminal B cancers identified by gene expression profiling). A clinically applicable method for identifying this subgroup of poor prognosis ER+, LN- patients is required to offer them more intensive adjuvant therapy.

Material and methods: A consecutive, unselected series of 1072 new, operable breast cancer patients diagnosed between 2000 and 2004 was studied for ER and progesterone receptor (PR) expression, HER2 status and standard pathological and demographic parameters. ER and PR expression were scored on immunohistochemistry (IHC) on preoperative, diagnostic core biopsies using the “Quickscore” method. This ensured optimal fixation and tissue processing avoiding the variable fixation of resection specimens and the well-described sampling error of tissue micro-array (TMA) methodologies. PR expression is frequently heterogeneous resulting in false negative scores in at least 12% of cases by TMA analysis. HER2 status was assessed using IHC with dual color FISH for cases scoring 2+. Adjuvant therapy was prescribed using standard protocols; all patients with ER+ disease received adjuvant endocrine therapy. Follow up data were obtained from the oncology database, and the registrar of deaths for the date and cause of death. All deaths not attributable to breast cancer were censored at the date of death. Accordingly, the primary end-point was breast cancer-specific overall survival. Survival analysis was carried out by Kaplan-Meier survival curves analysed by the Log-Rank test. Multivariate analysis was carried out using Cox's regression.

Results: Overall, PR- cancers had a worse prognosis than PR+ (p<10−12, Hazard Ratio 3.40), even in the ER+ (p=0.006, HR 1.86), LN- (p=10−8, HR=5.33) and LN+ (p=10−11, HR=3.26) sub-groups. In the ER+ LN- group, the absence of PR expression predicted worse prognosis (88% vs 96% 8 year survival, p=0.0003) with a hazard ratio of 3.75. This is considerably more significant than Ki67 IHC scoring reported for a similar group of patients in other studies (HR 2.22). Multivariate analysis demonstrated that PR expression was an independent prognostic variable second only to LN status and more powerful than ER.

Discussion: Patients with ER+, LN-, PR- breast cancers have a significantly worse prognosis than those with ER+, LN-, PR+ cancers. Unlike Ki67, PR IHC uses a simple, cost effective, standard methodology (as for ER) and should identify patients who may require chemotherapy in addition to endocrine therapy in this group of otherwise good prognosis patients.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-02-06.
P4-02-11: Insulin-Like Growth Factor I Promotes Estrogen Receptor Positive Breast Cancer Cell Proliferation, in Part, through CYP1A1 Signaling.
Authors: Rodriguez M , Becker M , Yee D , Potter D
Published: June 2012

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

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-02-11.
P4-02-12: Plasma Estradiol Levels and Degree of Estrogen Receptor Positivity by Image Analysis in a Large Cohort of Breast Cancer Cases: Results from the Nurses' Health Study.
Authors: Collins LC , Frieling GW , Ahern TP , Hu R , Hankinson SE , Tamimi RM
Published: June 2012

Background

Breast cancer risk is strongly associated with endogenous reproductive hormones. There is also good evidence to suggest that endogenous estradiol levels are more strongly associated with the development of estrogen receptor (ER) positive breast cancers than with ER negative cancers. However, the degree of ER positivity in those cancers that develop is not yet established. Therefore, we sought to evaluate the proportion and intensity of tumor cells demonstrating ER positivity using image analysis in a large cohort of breast cancers for which we also had plasma estradiol levels.

Design: Tissue microarray (TMA) sections of breast cancers from women enrolled in the Nurses’ Health Study were immunostained for ER (1D5, Dako Corporation, Carpinteria, CA). Slides were then scanned using the ScanScope slide scanning system (Aperio Technologies Inc., Vista, CA) and ER staining was assessed for both intensity and percent of nuclei staining positively using a specific algorithm, Nuclear v9, which evaluates nuclear size, intensity, roundness, curvature, compactness, and elongation. Plasma samples were collected prior to the date of diagnosis for all women in the study and estradiol levels were determined by radioimmunoassay following extraction and celite column chromatography.

Result: Image analysis data for ER was available for 691 breast cancers. The mean percent of tumor cell positivity was approximately uniformly distributed, with a mean of 44.5%, (median=41.3%, max=99.6%). The proportion of tumor cells staining for ER from the TMA cores was positively associated with patient age (p<0.0001), and postmenopausal status (p=0.008), while inversely associated with tumor grade (p<0.0001). Among women with image analysis data for ER, plasma estradiol data were also available for 320 postmenopausal women. Estrogen receptor positivity (by proportion or intensity) did not vary according to plasma estradiol levels (p=0.17).

Conclusion: While high concentrations of plasma estradiol are more strongly associated with the development of estrogen receptor positive breast cancer versus estrogen receptor negative cancers, the degree of estrogen receptor positivity in the breast cancers that develop is not influenced by this variable.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-02-12.
P4-04-01: Identification of a Unique Mammary Cell Type Expressing Mesenchymal Markers, but Capable of Multilineage Epithelial Differentiation.
Authors: Landua JD , Lewis MT
Published: June 2012

We have identified a previously uncharacterized cell population within the epithelial compartment of the post-pubertal mouse mammary gland. These cells can be found as singlets or as like pairs in both the cap and body cell layer of terminal end buds (TEB) of developing ducts, and are ultimately localized in a suprabasal position distributed throughout the mature mammary ductal tree. In differentiated ducts, these cells typically extend long processes that contact multiple surrounding luminal epithelial and myoepithelial cells suggesting a function in cell-cell communication. In younger animals, identified cells do not express epithelial markers including the major cytokeratins (CK) 5, 6, 8, and 14, or E-cadherin, nor do they express myoepithelial cell markers such as smooth muscle actin or p63. However, in 20 week old non-parous females, these cells can give rise to epithelial cell types expressing CK 5, 6, 8, or 14. In transplantation assays, epithelial fragment and whole mammary gland transplantation did not indicate a major role in regenerative growth of mammary ducts. Transplantation of dissociated epithelial cells and wound-healing assays are in progress. Taken together, these observations suggest that this unique cell population possess stem cell-like features in that they are capable of self-renewal, and of giving rise to all major differentiated cell types within the mouse mammary gland.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-04-01.
P4-07-04: Nomogram Including Circulating Tumor Cells (CTC) Count before and during Chemotherapy for Individual Survival Prediction of Metastatic Breast Cancer Patients.
Authors: Bidard F-C , Hajage D , Bachelot T , Delaloge S , Brain E , Campone M , Wolp-Diniz R , Dieras V , Mathiot C , Asselain B , Pierga J-Y
Published: June 2012

Background

CTC count before a new line of treatment and CTC count early changes under chemotherapy have been reported as an independent prognostic marker in metastatic breast cancer in a recent pooled analysis in 841 pts (Liu M. ASCO 2011). The aim of this study was to build a prognostic tool including CTC and other parameters to assesse its predictive value for progression-free survival (PFS) and overall survival (OS).

Methods: Data from the IC 2006–04 study were used. This prospective multicentre study included 267 metastatic breast cancer patients treated by first line chemotherapy with or without targeted therapy, in whom appropriate pre-treatment prognostic variables (age, performance status, number of metastatic sites, disease-free interval, ER, PR and HER2 status, tumor grade, LDH, serum markers, CTC count by CellSearch technique before treatment and before cycle 2) were available for statistical analysis. We constructed a multivariate Cox regression model for PFS and OS prediction. A stepwise selection process was applied to achieve the most informative and parsimonious models. Performance was measured with the C-index statistic. Internal validation was performed using leave-two-out technique.

Results: Four nomograms have been obtained, in two clinical settings: at inclusion (before the start of any treatment) taking into account the initial CTC count, and during treatment (before cycle 2) taking into account CTC changes under treatment. Their accuracy was good for PFS and OS prediction, with C-index ranging from 0.72 to 0.88. Internal validations allow considering a good accuracy of the models in an external population.

Conclusion: These clinically relevant nomograms are a simple tool for a personalized prognostic assessment including CTC assessment. Validation on independent series of patients are ongoing.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-07-04.
P4-07-13: Prognostic Impact of Circulating Tumor Cells Assessed with the Cell Search Assay and Adna Test Breast in Metastatic Breast Cancer Patients – The DETECT Study.
Authors: Mueller V , Riethdorf S , Rack B , Wolfgang J , Fasching PA , Solomayer E , Aktas B , Kasimir-Bauer S , Mury D , Pantel K , Fehm T
Published: June 2012

Background

Over the last decade circulating tumor cells (CTC) were established as a prognostic factor in breast cancer patients. However, there are very limited studies comparing different test methods, although several are available.The DETECT trial for metastatic breast cancer patients was designed to investiate the prognostic impact of CTC. Here, we report on the prognostic relevance of CTC testing with different detection methods.

Material and Methods: Patients with primary metastatic breast cancer or metastatic recurrence were prospectively enrolled in this multicenter trial. CTC were detected using the FDA-approved Cell Search® assay applying immunocytochemistry and the RNA-based Adna Test Breast Cancer™. After a median follow-up of 11 months the first survival data are now presented.

Results: Both methods could be performed in 221 patients. Using the CellSearch™ assay 116 of 221 patients were CTC-positive based on the cut-off level of 5 cells. Presence of CTC was associated with the site of metastatic disease. The OS was 15.4 months in CTC positive pts. (95%-CI: 13.5−17.1 mths) compared to 20.4 mths. in CTC negative pts. (19.1−21.9 mths.; p<0.001). In the multivariate analysis presence of CTC was the only independent predictor for overall survival (HR: 3.4, 95%-CI: 1.7−6.3) including tumorbiological factors, menopausal status, number and sites of metastatic disease. The progression-free survival was not correlated with CTC status in our cohort receiving different types of systemic treatment (p=0.197). When the AdnaTest Breast was performed, 88 of 221 (40%) patients were CTC positive. Except for HER2 status, no correlation could be observed between CTC positivity and any of the clinicopathological factors. CTC positivity assessed by the AdnaTest Breast has no impact on PFS and OS. A multivariate analysis was therefore not performed.

Conclusions: Currently, several different tests are available for CTC detection. Only a few tests have been approved by the FDA and been validated in large clinical trials. Therfore, it will be important to compare new techniques with the Cell Search assay.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-07-13.
P4-07-20: Apoptosis in Circulating Tumor Cells (CTCs) of Early and Metastatic Breast Cancer Patients.
Authors: Kallergi G , Konstantinidis G , Papadaki M , Agelaki S , Mavroudis D , Stournaras C , Georgoulias V
Published: June 2012

Background: Metastasis has been associated with the presence of circulating (CTCs) and disseminated (DTCs) tumor cells in peripheral blood and bone marrow of breast cancer patients, respectively. Recent studies have confirmed that the detection of CTCs and DTCs represents a strong and independent prognostic factor for decreased disease-free and overall survival. However, it is not clear so far whether all CTCs are capable to generate metastasis or some of them are destined to die. The aim of the present study was to analyze the presence of apoptotic CTCs in patients with early and metastatic breast cancer. Patients and methods: Double staining immunofluorescent (IF) expreriments were performed in peripheral blood mononuclear cells (PBMC) cytospins of patients with early (n=23) and metastatic breast cancer (n=29), prior to the initiation of adjuvant and first-line chemotherapy, respectively. Pancytokeratin A45-B/B3 antibody (as a marker of CK-positive cells) was coupled with either M30 (marker of apoptotic cells) or Ki67 (marker of proliferating cells) antibodies. Apoptotic CTCs were also evaluated using a polycaspase detection kit. Results: Significantly lower proportions of apoptotic CTCs were detected in metastatic compared to early breast cancer patients using the polycaspase kit (3% vs 49%, p=0.0001). These results were confirmed with M30 staining (32% vs 67%, in metastatic and early disease, respectively, p=0.023). The median percentage of apoptotic CTCs per patient was also lower in advanced compared to early disease patients (0% vs 100% with the polycaspase kit and 15% vs 70% with M30 staining). Ki67 positive CTCs were identified in 56% of early and metastatic patients, while the ratio of Ki67-positive/Total CTCs was 15% and 36% respectively. Conclusions: Apoptotic CTCs are more commonly observed in early compared to metastatic breast cancer, whereas, Ki-67 positive CTCs are more frequently encountered in metastatic disease. The differential incidence of apoptotic and proliferative CTCs in early and metastatic disease is probably related to disease progression. The evaluation of these markers in CTCs from patients with breast cancer may provide useful prognostic information and could be used to monitor the effectiveness of therapy.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-07-20.
P4-08-04: Preoperative Thrombin Pathway Activation Identifies Node Positive Patients in Early Breast Cancer.
Authors: Shaker H , Kirwan C , Landberg G , Bundred NJ
Published: June 2012

Activation of the thrombin pathway (TP) is seen in 50% of cancers and plays a significant role in promoting metastasis. Cancer-induced TP activation is exacerbated by surgery, and this activation may be greater in the presence of metastases.

Our aim was to determine if pre-operative plasma d-dimer, a marker of TP activation, correlated with the presence of LN metastases in early breast cancer. We also examined whether surgery-induced TP activation is affected by tumour-associated factors.

Methods

Plasma d-dimer was measured using automated ELISA pre-operatively and at days 1 and 42 following surgery in 103 prospectively recruited patients undergoing surgery for early breast cancer(87 invasive,13 DCIS). D-dimer values were log-transformed and correlated with LN metastases, tumour size and grade and hormone receptor status using parametric statistical tests.

Results

Median age was 68 years (range 46–82). Mean tumour size was 14.6 mm (range 7.5-40). Seventy invasive cancer patients had wide local excision(WLE) and sentinel node biopsy(SNB) and 17 had mastectomy(MX) +/− axillary clearance(ANC).

Twenty-two patients had positive lymph nodes at primary surgery (MX 8,WLE 14). Of these 3(14%) were ER-ve and 19(86%) were ER+ve.

Mean pre-operative d-dimer was higher (p<0.01) in LN positive (704ng/ml, 95% confidence interval, CI, 455–1088) versus LN node negative invasive cancers (443ng/ml, CI 304–627) or DCIS (366ng/ml, CI 284–471). D-dimer was also higher in patients with lymphovascular invasion (612ng/ml, CI 423–886) than in those without (454ng/ml, CI 385–534) and this approached significance (p=0.084).

Post-operative d-dimer was higher (p<0.05) after MX compared to WLE at all timepoints. Following WLE + SNB for invasive cancer, post-operative d-dimer rose and was higher at day 1 and day 42 in LN positive vs LN negative and in ER negative versus ER positive cancers (see table). Repeated measures ANOVA analysis showed that the change in d-dimer over time was significantly different between subjects according to LN status (p=0.037) or ER status (p=0.015). Only one patient in the ER negative group had LN metastases. There was no association between pre or postoperative d-dimer and tumour size, tumour grade or PR status.

Conclusion

Pre-operative TP activation and the thrombotic response to surgery (as measured by d-dimer) is greater in the presence of LN metastases regardless of tumour size.

ER negative cancers (7/8 of which were LN negative) also resulted in a greater d-dimer rise, suggesting a difference in the thrombotic response to surgery between different cancer phenotypes.

Ongoing work will determine if peri-operative TP activation can act as an independent predictor of node status and cancer recurrence.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-08-04.