bluuebunny/crossref_metadata_2025 huggingface dataset:
Sumoylation of Corepressor SAFB1 – Mechanism of Action and Biological Relevance in Breast Cancer.
Authors: Garee J. , Meyer R. , Oesterreich S.
Published: December 2010

Scaffold attachment factor B1 (SAFB1) was originally isolated due to its ability to bind Scaffold/Matrix attachment regions of genomic DNA. Subsequently, SAFB1 has also been shown to be involved in RNA processing, chromatin organization and stress response. Work from our laboratory has shown that SAFB1 can bind and modulate transcription of the HSP-27 promoter and repress activity of ERα. Repression of ERα occurs through an independent and transferable c-terminal repression domain. Further work has indicated that SAFB1 can interact with and repress transactivation of a host of nuclear receptors including PPARg and FXR, suggesting a general role for SAFB1 in transcriptional repression.Many proteins associated with transcriptional regulation are post translationally modified by small ubiquitin like modifier (SUMO). SUMO is a 98 amino acid mature polypeptide that is added post translationally to target proteins usually within a consensus sumoylation motif, ΨKXE (where Ψ is a large hydrophobic residue, K-lysine of modification and X any residue). The process of sumoylation is analogous to ubiquitination. SUMO modification of proteins associated with transcription has generally been associated with transcriptional repression. The mechanism by which this modification leads to transcriptional repression has yet to be defined.Work by our laboratory has indicated that SAFB1 is modified by sumoylation. In silico analysis indicated two consensus sumoylation motifs within SAFB1. Immunoprecipitation studies revealed SUMO modified forms of SAFB1. Mutating the lysine's within the sumoylation motifs to arginine (K231,294R) lead to a loss of sumo modified SAFB1 indicating these as the sites of modification. Transient ERE-Tk-Luciferase assays showed a loss of transcriptional repression with over expression of SAFB1 K231,294R compared to SAFB1 wildtype. Loss of transcriptional repression was also seen utilizing transient Gal4DBD reporter assays indicating an effect of sumoylation on SAFB1's intrinsic repressive function. Co-immunopreciptation studies have indicated interaction between SAFB1 and SUMO E3 ligases suggesting these may be important for modifying SAFB1. Additionally, over expression of members of the sumoylation machinery can enhance SAFB1 repressive activity indicating this modification is important for SAFB1 transcriptional repression. Loss of SAFB1 sumoylation does not alter protein half life indicating the loss of transcriptional repression is not through this mechanism.Our studied have indicated that sumoylation of SAFB1 is important for its repressive activity. Additional studies within the lab are aimed at addressing the mechanism and the complete extent of functional consequences of sumoylation of SAFB1. A better understanding of post translational modification of SAFB1 may provide additional insights into the diverse functions of SAFB1 and its role in breast cancer.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 4142.
HER2-Associated Breast Cancer Signature Using a 3D Culture Model.
Authors: Chaluvally-Raghavan P. , Zeisel A. , Koestler W. , Jacob-Hirsch J. , Rechavi G. , Domany E. , Yarden Y.
Published: December 2010

Background: Carcinomas of the breast account for one third of all cancers occurring in woman and responsible for approximately one quarter of cancer-related deaths in females. The HER2/Neu -oncogene is amplified in 20-25% percent of human primary breast cancers and this alteration is associated with disease behaviour. Whereas signalling pathways emanating from HER2 have been characterized, much less is known about the transcriptionally regulated genes that contribute to HER2 tumorigenic effects.Materials and Methods: Normal and HER2 overexpressing mammary epithelial cells (MCF10A) were grown in extracellular matrix to form 3D structures, which allow epithelial cells to organize into structures that resemble their in vivo architecture. RNA was isolated from the 3D structures and hybridized to an Affymetrix HuGene 1.0 ST oligonucleotide array.Results: Upon HER2 overexpression, mammary epithelial cells lost their polarity and formed disorganized structures in matrigel. Using microarrays we analyzed transcriptional events responsible for the morphological changes and found that several sets of genes such as integral proteins, transcription factors, matrix proteases and chemokines were highly altered in the HER2 overexpressing group. Using gene annotation we defined molecular-pathways which are highly altered in HER2 overexpressing cells. More generally, our study proposes a mechanistic description of the processes underlying the HER2 transcriptional network.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 4146.
Expression of Androgen Receptor in Breast Cancer.
Authors: Park S. , Park H. , Choi S. , Lee J. , Park B. , Lee K.
Published: December 2010

Background: Androgen receptor (AR) is one of biomarkers and its role in breast cancer is still unclear. The aim of this study was to investigate the relationship between AR expression and clinicopathological factors in breast cancer patients.Patients and Methods: AR was consecutively evaluated in 413 primary breast cancers from whole sections of surgically resected specimens using immunohistochemical staining from January 2008 to March 2009. The associations between AR expression and clinicopathological parameters were analyzed. Tumors with 10% or more nuclear stained cells were considered as positive for AR expression. The differences between variables were calculated by chi-square test and Fisher's exact test was used when appropriate.Results: The median age at diagnosis was 49 years (range, 26-84). AR was found in 72.7% (48/66) of in-situ carcinoma and in 72.9% (253/347) of invasive carcinoma. Overall expression rates of AR were 72.9%, which were higher than those of ER and PR expression, 68.5% and 62.0%, respectively. AR was significantly expressed in patients with no elevated preoperative serum cancer antigen 15-3 (CA 15-3) levels (p = .042), smaller tumor size (p = .035), lower histologic grade (p < .001), ER-positive (p < .001), progesterone receptor-positive (p < .001), and non-triple-negative breast cancer (p < .001). Metaplastic, medullary, and mucinous types carcinomas showed less AR expression (p = .030). Although it was statistically not significant, patients with younger age (≤ 35 years), axillary lymph node involvements, and higher stage showed higher rates of AR negativity. In ER-negative tumors, AR expression was significantly correlated with HER-2 over-expression (p < .001). In ER-positive tumors, however, there was no relationship between AR expression and HER-2 over-expression (p > .05).Conclusions: AR is expressed in a significant number of breast cancers and is associated with favorable tumor differentiation and smaller tumor size. These results might suggest that AR may be an independent prognostic factor in breast cancer. AR may also be associated with growth factor signaling and be useful therapeutic target in ER-negative tumors.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 4156.
Tumor Volume Analysis and Tumor Doubling.
Authors: Gordon A. , Stebbing J. , Shousha M. , Coombes R.
Published: December 2010

Background: Volume is the most sensitive and accurate parameter to assess tumor response to treatment. We have proposed that data be expressed as volume in cubic centimetres TVA (Tumor Volume Analysis)1. When we applied TVA to a breast cancer trial and converted the data from diameter centimetres to volume in cubic centimetres the differences were statistically signifcant2.Three dimensional measurements: Volume measurements are most accurate if all three dimensions are measured to record any asymmetric growth[ref]. In Table 1 diameter permutations are shown of a 3cm mass together with percentage changes in each example. This demonstrates the extent that changes in the lesser two axes of a mass affect tumor volume. Accurate 3D volume measurement is essential to estimate tumor doubling.Tumor Doubling and Redoubling: Tumor doubling for a carcinoma is quoted as under 177 days [ref] or under 500 days[ref]. With a mean of 100 days[ref] or 117 days[ref]. Much of the work has been carried out on metastatic pulmonary nodules of various tumors. An early paper did quote doubling times as by diameter and provided a graph to convert data into volume8. A calculator has been described and patented to provide these functions2.Tumor Doubling Time (TDT): Tumor doubling can be regarded as an increase in volume over a unit of time[8, ref]. We propose this TDT be quantified with the period of growth expressed in days for example TDT 120 implies a tumor doubling time of 120 days. A formula that does allow for exponential growth assuming a constant growth is as follows: TDT = D x log(2)/log(1+ r/100). Where TDT = tumor doubling time; D = interval in DAYS between volume measurements; r = rate of growth; r/100 = (Vol 2-Vol 1)/Vol 1 x 100%. This option for measuring tumor doubling can be added as a fourth and further facility to the calculator already described [Patent PCT/GB07/003526]2Conclusions: 1. Tumor doubling is qualified by volume changes in cm3. Tumor doubling time (TDT) in days quantifies tumor doubling. 2. Measurement of TDT can be made as an extra facility on the calculator already described. The calculator can be used to estimate TDT using two measured volumes and the interval in days. 3. A similar formula can also be used to measure downsizing in response to neoadjuvant therapy as complete response time (CRT).References:1. SABCS 2007 Poster # 2021Tumor Volume Analysis. A better way than RECIST? [A B Gordon, J Stebbing, R C Coombes]2. SABCS 2008 Poster # 4023 Tumor Volume Analysis in Clinical Oncology. [A B Gordon, J Stebbing, R C Coombes] 8. V P Collins et al. Observations on Growth Rates of Human Tumours. American Journal of Roentgenology. V. 76 p. 988-1000 1956Table 1. Volume Calculation of Three Dimensional MassSHAPE    SphereOne Diameter3x3x3cm14.14cm3100%TangerineTwo Diameters3x3x2cm9.42cm367%EggTwo Diameters3x2x2cm6.28cm344%CigarTwo Diameters3x1x1cm1.57cm311%DateThree Diameters3x2x1cm3.14cm322% 

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5030.
Bilateral Breast Cancer: Analysis of Incidence, Characteristics and Outcome of the Disease.
Authors: Kheirelseid E. , Jumastapha H. , Miller N. , Curran C. , Sweeney K. , Newell J. , Kerin M.
Published: December 2010

Background: Despite the widespread adoption of breast conservation therapy in breast cancer, there is a dramatic increase in the rate of bilateral mastectomy in USA. The aim of this study was to address the incidence of bilateral breast cancer (BBC) and to investigate its characteristics, survival and outcome compared to unilateral breast cancer.Methods: Data was acquired from the prospectively maintained NUI Galway breast cancer database between 1988 and 2008. BBC were then categorized as synchronous (within 12 months) or metachronous cancer (after 12 months of first tumour). SPSS was used for data analysis.Results: The incidence of bilateral breast cancer in our population was 4.4% (112 of 2524). Of those 2.1% were synchronous while 2.3% were metachronous. Compared to unilateral cases, bilateral cancer patients were younger (0=0.021) and had smaller size (p=0.001) and earlier stage (p<0.001) tumours at diagnosis. There were no differences in histological type (p=0.630) and grade (p=0.054)). While there was no significant difference in survival for patients with bilateral compared to unilateral tumour (p>0.05), the synchronous bilateral tumour was associated with poor overall survival (p=0.010) and disease free survival (p=0.013) in comparison to metachronous bilateral tumour.Conclusion: We observed significant differences in bilateral tumour characteristics compared to unilateral disease. Although bilateral synchronous tumour was associated with poor outcome and overall survival, patients with metachronous disease do as well as patients with unilateral tumour. The increasing practice of bilateral mastectomy is not supported by this large single centre experience.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5033.
Use of BIBW 2992, a Novel Irreversible EGFR/HER1 and HER2 Tyrosine Kinase Inhibitor To Treat Patients with HER2-Positive Metastatic Breast Cancer after Failure of Treatment with Trastuzumab.
Authors: Hickish T. , Wheatley D. , Lin N. , Carey L. , Houston S. , Mendelson D. , Solca F. , Uttenreuther-Fischer M. , Jones H. , Winer E.
Published: December 2010

Background: BIBW 2992 (Tovok™*) is a novel, oral, irreversible inhibitor of the epidermal growth factor receptor (EGFR)/human epidermal growth factor receptor (HER)1 and HER2 inhibitor, with preclinical activity in trastuzumab-resistant cell lines overexpressing HER2 and Phase I clinical activity. A Phase II study of BIBW 2992 in patients with HER2-positive breast cancer who have progressed following treatment with trastuzumab was conducted in the US and the UK.Methods: A multi-institutional, open-label, single-arm Phase II study was conducted. Eligible patients had stage IIIB or IV HER2-positive metastatic breast cancer, with progression following trastuzumab treatment or intolerance of trastuzumab, received no prior EGFR-targeted therapy, had measurable disease, Eastern Cooperative Oncology Group performance status of 0–2 and adequate organ function. Patients received 50 mg BIBW 2992 once-daily until disease progression. Tumor assessments were performed every two treatment courses (one course is 28 days). The primary endpoint was objective response rate (Response Evaluation Criteria In Solid Tumors). Safety data were also collected.Results: A total of 41 patients started treatment on the trial. Patients had received a median of three lines of prior therapy (range, 0–15). Thirty-four patients are evaluable for response. Of these, four patients had a partial response (PR) and eight patients had stable disease maintained for at least four cycles. One patient with confirmed PR remained on treatment until progression at 63 weeks. The most frequently observed side-effects to date are rash (Common Terminology Criteria for Adverse Events [CTCAE] Grade 3 in four patients [9.8%]) and diarrhea (CTCAE Grade 3) in nine patients. There have been 21 dose reductions in 18 patients.Conclusion: BIBW 2992 was safe and induced promising early clinical responses in HER2-positive breast cancer patients who have progressed following treatment with trastuzumab. Manageable cutaneous adverse events and diarrhea were the main side-effects.*Trade name not FDA approved.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5060.
Addition of Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF) to Trastuzumab Stabilizes Disease in Patients with Trastuzumab-Resistant, HER2+ Metastatic Breast Cancer.
Authors: Cheng Y. , Cheng Y. , Valero V. , Davis M. , Hortobagyi G. , Ueno N. , Ueno N.
Published: December 2010

Introduction: Trastuzumab with or without chemotherapy is the standard of care for patients with HER2+ metastatic breast cancer. The proposed mechanism of trastuzumab-induced regression of HER2+ tumors includes inhibition of tumor cell proliferation, potentiation of chemotherapy, and facilitation of immune function through antibody-dependent cell-mediated cytotoxicity. GM-CSF (Leukine) is a cytokine that mediates antibody-dependent cell-mediated cytotoxicity. We studied the feasibility and efficacy of adding GM-CSF to trastuzumab in patients with trastuzumab-resistant, HER2+ metastatic breast cancer.Patients and Methods: Patients with measurable, HER2+ metastatic breast cancer that progressed after treatment with trastuzumab with or without chemotherapy were continued on trastuzumab alone at 2 mg/kg intravenous weekly. GM-CSF (250 μg/m2 subcutaneous daily) was added until the absolute neutrophil count (ANC) was greater than 10,000/mm3, then given every other day while the ANC was maintained below 10,000/mm3. Disease was restaged every 8 weeks. Treatment with trastuzumab and GM-CSF was continued until disease progression or intolerable toxicity.Results: Of 18 eligible patients with progressive HER2+ metastatic breast cancer, 17 (median age 48 yr, range 27–75 yr) were evaluable. Nine had hormone receptor–positive disease. The median number of metastatic sites was 2 (range 1–3); the most common site was the liver (n=10). The median number of prior regimens (trastuzumab with or without chemotherapy) for metastatic disease was 2 (range 1–5). One patient developed rapidly progressive disease 2 weeks after the start of study therapy and died soon after. The other 16 patients continued treatment until disease progression. No disease response was observed, but 5 patients (29%) had stable disease with a median duration of 15.8 weeks (range 10–53.9 weeks). Thirteen patients had grade 1 toxic effects; 6 patients, grade 2; and 2 patients, grade 3 (fatigue and muscle aches). The most common toxic effect was rash at the injection site, followed by skin rash, fatigue, and muscle aches. No grade 4 or irreversible toxic effect was seen.Conclusion: The addition of GM-CSF to trastuzumab alone in patients with trastuzumab-resistant, HER2+ metastatic breast cancer stabilizes the disease for a median duration of 15.8 weeks without causing any significant toxic effects in 29% of heavily pretreated patients. Its administration is simple, safe, and feasible. This regimen, trastuzumab and GM-CSF, needs further evaluation in combination with chemotherapy or other biological agents.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5103.
In Vivo Activity of ARRY-380, a Potent, Small Molecule Inhibitor of ErbB2 in Combination with Trastuzumab, Docetaxel or Bevacizumab.
Authors: Lee P. , Anderson D. , Bouhana K. , Garrus J. , Napier C. , Avrustkaya A. , White A. , Pheneger T. , Winkler J.
Published: December 2010

ARRY-380 is an orally active, potent small molecule tyrosine kinase inhibitor targeting ErbB2. The compound is a reversible, ATP-competitive inhibitor with nanomolar potency against ErbB2 in both in vitro and in cell-based assays. This compound has very good in vivo and in vitro PK/ADME properties and has shown excellent activity in numerous mouse tumor models including breast (BT-474, MDA-MB-453), ovarian (SKOV-3) and gastric (N87) carcinoma models. Here we demonstrate excellent single agent activity and combinability with trastuzumab, docetaxel or bevacizumab in breast and ovarian carcinoma models. For the BT-474 studies, female SCID beige mice were implanted with tumor fragments. For the SKOV-3 tumor studies, female nude mice were inoculated with cells subcutaneously in the flank. Animals received: doses of ARRY-380 ranging up to 200 mg/kg/d, PO; and/or trastuzumab at 20 mg/kg, IP, Q3D or QW; and/or docetaxel at 10 mg/kg, IV, Q3D; and/or bevacizumab at 10 mg/kg, IP, Q4D x3. Tumor size was measured at regular intervals and subsets of animals were monitored for up to 90 days to determine tumor-free survival. In the BT-474 model, ARRY-380 demonstrated significant dose-related tumor growth inhibition (TGI; 50% at 50 mg/kg/d and 96% at 100 mg/kg/d) with numerous partial regressions (>50% reduction from baseline size) at the higher dose level in 9/12 animals. One complete response was observed at the higher dose. Trastuzumab alone provided a 45% TGI with no regressions. ARRY-380 (50 mg/kg/d) in combination with trastuzumab showed a 98% TGI with complete regressions in 9/12 animals and two partial regressions. At dose of 100 mg/kg/d of ARRY-380 in combination with trastuzumab, there was 100% TGI and all animals had complete responses. Docetaxel as a single agent produced a 55% TGI with no regressions. In combination with ARRY-380 (50 mg/kg/d), there was an 81% TGI and five partial regressions. In the SKOV-3 model, ARRY-380 demonstrated significant dose-related tumor growth inhibition (TGI; 39% at 50 mg/kg, BID and 96% at 100 mg/kg, BID) with partial regressions (>50% reduction from baseline size) at the higher dose level in all animals. Bevacizumab alone provided a 55% TGI with no regressions. ARRY-380 (50 mg/kg, BID) in combination with bevacizumab showed 80% TGI with partial responses in 7/8 animals and one stable disease. From this work we have demonstrated superb single agent activity for ARRY-380 in the BT-474 human breast carcinoma xenograft model and the SKOV-3 human ovarian carcinoma model. In addition, ARRY-380 has shown additive activity and tolerability with trastuzumab, docetaxel and bevacizumab. ARRY-380 has entered Phase I clinical trials in patients with advanced cancers.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5104.
Evaluation of Factors Associated with Recruitment in Breast Cancer Clinical Trials in a Specialized Breast Cancer Centre.
Authors: Lemieux J. , Forget G. , Brochu O. , Camden S. , Provencher L. , Robert J. , Cantin G. , Desbiens C. , Doyle C. , Morin J. , Poirier B.
Published: December 2010

Background: Recruitment of patients (pts) in cancer clinical trials has been reported to be between 3-5%. Very few data come from Canada. Methods: The objective was to measure the recruitment and its associated characteristics in breast cancer clinical trials for non-metastatic breast cancer pts. This was a retrospective cohort study at the Centre des Maladies du Sein Deschênes-Fabia, a specialized breast cancer centre in Québec City, Canada. Clinical trials opened between 2004 and 2008 were used. For each protocol, main criteria were used to define the population under study (e.g. triple negative breast cancer). Charts were identified from our database and reviewed in decreasing chronological order of diagnosis. Staging, hormone receptors status and Her2 status were available from the database. All charts were reviewed to assess eligibility criteria (inclusion and exclusion criteria). Also, information was sought in the notes about whether or not the protocol has been proposed to the pt, if the pt accepted and if not, reason(s) for refusal. Protocol and physicians characteristics were collected. Results: A total of 9 protocols were studied and 244 charts identified (relates to 704 pt-protocols because one pt could be assessed for more than one protocol if applicable). Eligibility could not be assessed for 19% (n=138) pt-protocols. Among all pt-protocols, 74.4% met all the eligibility criteria. There was a note in the chart, about protocol being proposed to a potentially eligible pt, in 13.8% of the pt-protocols. Overall, 9.7% of pt-protocols were recruited. Among patients who had a protocol offered, 74.2% accepted. Data on characteristics associated with recruitment will be presented at the meeting. Conclusion: Enrollment in clinical trials in a specialized breast cancer centre is slightly higher than the 3-5% generally reported but still low. Interventions should focus on finding ways to offer a clinical trial when available to every suitable eligible patient.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5122.
The Interaction of High Mobility Group Proteins with the p160 Coactivator SRC-1; a Role in Endocrine Resistance.
Authors: Redmond A. , Stafford A. , McIlroy M. , Hill A. , Young L.
Published: December 2010

Breast cancer affects one woman in ten in the western world and despite the phenomenal advances in recent years, the mortality rate still remains at around 35%. Resistance to current endocrine therapies, including tamoxifen and aromatase inhibitors, is common with recurrence of disease in 30-40% of patients. The estrogen receptor (ER) coactivator protein SRC-1 plays an important role in this resistance. We examined SRC-1 protein expression in a large cohort of breast cancer patients (n=560) and found expression to be a strong predictor of disease recurrence in patients treated with adjuvant tamoxifen (hazard ratio: 5.032, p<0.0001). Identification of SRC-1 interacting proteins will provide key insights into the role of SRC-1 in the development of tumour recurrence. We employed a mass spectrometry-based screen to identify proteins which could differentially interact with SRC-1 in endocrine resistant compared with endocrine sensitive breast cancer cells. Selected SRC-1 interacting proteins relevant to endocrine resistance include the high mobility proteins HMGB1 and HMGB2, which have previously been reported to enhance binding of the estrogen receptor to its response elements in target gene promoters. Immunoprecipitation assays confirmed increased interactions between HMGB2 and SRC-1 following treatment of the endocrine resistant cell line LY2 with tamoxifen and estrogen. Steroid treatment also increased trafficking of the coactivator and the SRC-1 interacting protein to the nucleus of tumour epithelial cells. Treatment enhanced recruitment of SRC-1 to the promoter of the oncogene c-Myc and enhanced target gene expression. HMGB2 expression in the breast cancer patient TMA correlated with SRC-1 and poor disease-free survival. These findings would indicate an important role for HMGB2 in the development of endocrine resistance in breast cancer patients.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5138.
Prediction of Metastasis in Node-Negative, Hormone Receptor Positive, Tamoxifen Adjuvant Treated Primary Breast Cancer Patients Using Ki-67 (IHC) and RT-PCR Based 14-Gene Prognostic Signature.
Authors: Schlotter C. , Wassmann K. , Bosse U. , Kemming D. , Brandt B. , Vogt U.
Published: December 2010

Background: Risk estimation based on the recommendations of St. Gallen Consensus commonly decides for additional cytostatic therapy in node-negative (N-), hormone receptor positive (HR+) primary breast cancer patients. High proliferative activity in the HR+ subtype confers a 19-fold relative risk of relapse compared with HR+ tumors of low proliferative activity. Furthermore immunohistochemically determined Ki-67 is characterized as a Luminal B marker that identifies a high risk subgroup in HR+, N- breast cancer patients.Aim of this investigation was to compare risk estimation using Ki-67 (%) with the results of a RT-PCR based multi-gene prognostic signature. Methods: Tumor tissues of totally 321 unselected primary breast cancer patients were formalin fixed and routinely processed for immunohistochemical determination and scored for protein expression of ER, PgR and Ki-67. HER2 was determined using dd-PCR. Content of both hormone receptors and proliferation activity were evaluated counting positively coloured nuclei from at least 100 tumor cells. The median follow up was 61 months. To calculate the cut off for high proliferation Ki-67 values were subjected to a log rank CART analysis. High proliferation was defined by Ki-67 of 19% and more. With this cut off it was possible to distinguish significantly various patient cohorts (N-, pT1, postmenopausal) into two different risk groups regarding metastasis free survival. The definition of low risk was low proliferation and HER2 negativity, of high risk high proliferation or HER2 positivity. The risk for metastasis of 16 new primary breast cancer patients was estimated based on two surrogate risk groups and the multi-gene prognostic signature. Results: Tamoxifen adjuvant treated HR+, HER2-, low proliferating (Ki-67 <19%), N- low risk patients (N= 68) and HR+, HER2-, high proliferating/HR+, HER2+, N- high risk patients (N= 84) showed a recurrence rate of 2.94% and 27.4%, respectively (P< .000). In the ongoing observation study all 5 low proliferating HER2- tumors had a low risk gene prognostic signature. From 11 high proliferating HER2- patients one patient had a low risk gene signature (9%), 3 patients a moderate risk gene signature (27%) and 7 patients a high risk gene signature (64%). Conclusion: Despite the small number of patients investigated up to now (N= 16) the preliminary results appear to show that low proliferative activity is associated with a low risk gene prognostic signature, whereas high proliferation means only in part a high risk gene prognostic signature.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 6025.
Dose Dependent Effects of G-CSF on Ca27.29 in Early Stage Breast Cancer Patients.
Authors: Hepp P. , Rack B. , Schneeweiss A. , Schrader I. , Lorenz R. , Tesch H. , Soeling U. , Rezai M. , Gerber B. , Lichtenegger W. , Beckmann M. , Janni W.
Published: December 2010

Background:13% of 2556 patients (pts) examined in the SUCCESS trial showed elevated levels of Ca27.29 only after chemotherapy (SABCS2008). Early data indicate a possible relationship between the administration of G-CSF and a rise in the tumor marker. This analysis focuses on the dose dependency of this effect.Methods:The SUCCESS Trial is a phase III trial comparing FEC-Docetaxel vs. FEC-Doc-Gemcitabine regime and 2 vs. 5 years of treatment with zoledronate in patients with primary breast cancer in 3754 pts (BC) (N+ or high risk). Blood samples for this analysis are drawn before and after chemotherapy (CHT). Ca27.29 has been measured with ST AIA-PACK Ca27.29 reagent using MUC-1 for AIA-600II (Tosoh Bioscience, Tessenderlo, Belgium). The cutoff for Ca27.29 is >=32 U/ml. For chi² analysis patients were grouped to increasing or not increasing values of Ca27.29 and 1 to 6 cycles with G-CSF or no G-CSF at all. The absolute difference of Ca27.29 values before and after CHT was correlated with the number of G-CSF cycles administered.Results:The analysis of Ca27.29 is based on the data of 2556 pts. 1252 pts (49%) received at least one course of G-CSF. 338 pts (13%) exceeded the threshold for CA27.29 only after CHT. In this group 209 pts (62%) received G-CSF and 129 (38%) did not. 1043 pts with stable or decreased CA27.29 received G-CSF (47%) and 1175 did not (53%). This difference was highly significant (p<0.0001). Correlating the number of G-CSF cycles received during CHT with the absolute difference in Ca27.29 levels showed a highly significant positive correlation of 0.13 (Spearman-Rho; p<0.0001[two-sided]).Discussion and Conclusion:This analysis gives strong evidence that there is a dose dependent correlation between elevated levels of CA27.29 post CHT and the application of G-CSF. This might be attributed to an illegitimate expression of MUC-1 in leucocytes during leucopoiesis. Whether this effect is also related to the destruction of remaining disseminated tumor cells needs to be further evaluated.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 6030.
No Clinical Evidence for Increase in Tumour Malignant Potential in Patients (Pts) with Metastatic Breast Cancer (mBC) Treated with Bevacizumab (BV) and Docetaxel (D) in the Phase III AVADO Study.
Authors: Harbeck N. , Chan A. , ten Bokkel Huinink D. , Chollet P. , Gimenes D. , Fabiani C. , Salvagni S. , Pérez-Michel L. , Schneeweiss A. , Chlistalla A. , Miles D.
Published: December 2010

Background: The monoclonal antibody BV is directed against soluble VEGF and has shown significant clinical benefit when combined with chemotherapy in a range of tumour types, including in three phase III trials in mBC. Recent preclinical reports suggest that some anti-angiogenic agents (VEGFR tyrosine kinase inhibitors [TKIs] or VEGFR antibodies) increase the malignant potential of tumours (Pàez-Ribes; Ebos: Cancer Cell 2009:15). Although BV has a distinctly different mechanism of action, exploratory analyses to investigate those preclinical findings were performed on data from AVADO, a placebo (PL)-controlled study in first-line mBC.Methods: Pts received D (100mg/m2) q3w for up to 9 cycles, plus PL or BV (7.5 or 15mg/kg) q3w until disease progression (PD) or unacceptable toxicity. Mortality rates were calculated at 30-day intervals up to day 210 after discontinuation of PL or BV for any reason. For pts discontinuing PL or BV for toxicity, PFS was analysed using Kaplan-Meier methods. In the overall population, the proportion of pts with new metastatic lesions was analysed at PD.Results: As of 30 April 2009, 91 pts had discontinued PL or BV for toxicity; median PFS from discontinuation was longer in the BV arms than the PL arm. Mortality rates in pts stopping BV or PL for any reason (n=339) were similar or lower in the BV arms than the PL arm at all 30-day intervals for the first 210 days after discontinuation, the timeframe over which the analyses were performed. At PD, fewer BV than PL pts had developed new lesions.  PL + DBV 7.5mg + DBV 15mg + DITT population, n241248247 Pts with PD, n (%)208 (86)212 (85)210 (85) Pts with PD & new lesion, n (%)160 (77)*154 (73)*138 (66)*All pts discontinuing BV or PL, n108118113 Mortality, n (%)    Day 9028 (12)26 (11)15 (7) Day 15041 (18)38 (16)32 (15) Day 21053 (23)52 (23)45 (22)Pts discontinuing BV or PL for toxicity, n292735 PFS from discontinuation of BV or PL    Median, months3.36.46.8 HR vs PL 0.710.73 [95% CI] [0.40–1.27][0.42–1.24]*% of pts with PDDiscussion: Although recent preclinical studies, carried out using agents targeting VEGF receptors, suggest that anti-angiogenic therapy increases tumour malignant potential, exploratory data from a large clinical trial of BV do not support this theory. PFS in AVADO was longer after discontinuation of BV than after discontinuation of PL. Mortality rates up to day 210 after PL/BV discontinuation were similar. The proportion of BV pts with new metastatic lesions at PD was lower than that of PL pts, suggesting that metastatic spread was not increased.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 6086.
Different Biological Processes Are Associated with Prognosis and Chemotherapy Sensitivity in the Different Molecular Subtypes of Breast Cancer.
Authors: Iwamoto T. , Iwamoto T. , Bianchini G. , Coutant C. , Shiang C. , Matsuoka J. , Symmans W. , Hortobagyi G. , Simon R. , Pusztai L.
Published: December 2010

Background: Breast cancer includes multiple different molecular subtypes. We hypothesized that different biological processes and molecular markers may be associated with prognosis and chemotherapy sensitivity in the different subtypes.Methods: We performed gene set analysis in ER-positive/HER-2-normal (ER+) and ER-negative/HER-2-normal (ER-) cancers separately to identify gene sets that are associated with prognosis and chemotherapy sensitivity. We tested 5295 different, functionally annotated gene sets (GS) that collectively represent almost all known biological and metabolic pathways in human cells. Significance was estimated with permutation test. Three separate cohorts on untreated HER2-normal patients (n=234, n=175, n=170) and 3 cohorts of patients with neoadjuvant therapy (n=198, n=85, n=61) were included in this analysis. We performed analysis of each data set separately and also as pooled data.Results: When data sets were analyzed individually, 753 to 938 GS were associated with prognosis (P≤0.05) in ER+ cancers including 186 common to all, and 408 to 579 GS were associated with prognosis in ER- cancers including 1 common to all and 141 common to at least two data sets. Within each data set, the number of overlapping GS between the ER+ and ER- prognostic lists ranged from 33 to 112. In the neoadjuvant data sets, 555 to 674 GS were associated with response in ER+ cancers (195 common to at least two, 4 common to all) and 543 to 730 in ER- cancers (268 common to two, 22 common to all). When the predictive GS for ER+ cancers were compared to those for ER- cancers, there were only 10 GS in common. A meta analyses of the combined prognostic data sets yielded 384 GS with p≤0.0001 in ER+ cancers and 47 in ER- cancers, respectively, with 12 GS common to both. A meta analyses of the combined neoadjuvant data sets yielded 92 GS associated with pCR in ER+ cancers and 110 GS in the ER- cancers. Only 2 GS were common to both. Functional analysis indicated that DNA replication, mitotic spindle checkpoint and plasma cell function were the most common prognostic pathways in ER+ cancer. T cell differentiation, glycolipid metabolism and immune functions were the most commonly prognostic pathways in ER- cancer with a notable absence of proliferation related gene sets. DNA replication, spindle and microtubule activity and cell cycle regulation were associated with chemotherapy response in ER+ cancers; oxidative stress, blood vessel formation in ER- cancers.Conclusion: These data indicate that prognosis and chemotherapy response are associated with different gene sets in ER+ and ER- cancers.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 6124.
Instability in Chromosomes 9, 7, and 5 Correlate with Lymph Node Metastasis at Presentation in Basal Phenotype Carcinoma.
Authors: Oakley III G. , Brufsky A. , Chivukula M.
Published: December 2010

Introduction: Basal phenotype breast carcinoma has been described as an aggressive variant with comparatively poor prognosis; however, increasing experience suggests that only a subset show increased mortality. We performed microarray analysis of DNA from 11 basal phenotype carcinomas (from 10 patients) and paired normal breast tissue with copy number and loss of heterozygosity analysis with hierarchical clustering to see if characteristic genetic changes were present that correlated to lymph node metastasis at presentation.Materials and Methods: Electronic search of the medical records from 2006-2008 of our institution by an honest broker for all cases of basal phenotype carcinoma (based on medullary-like histology and immunohistochemistry positive for at least two of CK14, CK17, CK5/6 and EGFR) with known lymph node status at presentation and archived snap frozen fresh and tumor samples returned 10 cases (one with bilateral basal phenotype carcinoma), which were de-identified. 5 of these cases had lymph node metastasis at presentation. DNA isolation by column extraction was performed on 0.25 g of paired tumor and normal samples (confirmed by pathologist review before extraction). An Affymetrix 5.0 mapping microarray was performed on isolated DNA per manufacturer's protocols, and analyzed with Partek Genomics Suite software using copy number/LOH workflows using hidden Markov model and genomic segmentation in separate analyses with unsupervised hierarchical clustering.Results: Copy number and LOH analysis using hidden Markov model and genomic segmentation methods gave equivalent hierarchical clustering for our data set. Adjusting the obtained cluster analysis for known lymph node metastasis showed that changes in chromosomes 9, 7, and 5 that differentially clustered 4 of 5 cases which presented with lymph node metastasis (and 2 closely related initially lymph node negative cases).Discussion: Our results suggest that changes on chromosomes 9, 7 and 5 may correlate with the most important prognostic indicator in breast cancer, lymph node status, in basal phenotype carcinomas. Globally suboptimal amplification prevented accurate quantitation of copy number changes or precise loss of heterozygosity localization on these chromosomes. Additional studies to confirm these results using separate molecular techniques are underway, as well as correlation with 5 year survival when the cohort reaches that milestone.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 6133.
HOXB9 Promotes the Acquisition of Tumorigenic Phenotypes in Mammary Epithelial Cells.
Authors: Hayashida T. , Hayashida T. , Takahashi F. , Chiba N. , Brachtel E. , Jinno H. , Kitagawa Y. , Maheswaran S.
Published: December 2010

Background:The class I HOX gene family consists of 39 members with a shared highly conserved 61-amino acid homeodomain motif. HOX genes are important regulators of developmental processes, and their role in neoplastic transformation and tumor progression is increasingly recognized (Abate-Shen et al., Nat Rev Cancer, 2002; Cantile et al., Eur J Cancer, 2003). However, the molecular mechanisms by which HOX proteins promote tumorigenesis is not well understood. We recently, observed that HOXB9, a 9th HOX gene paralogue involved in mouse mammay gland development, is deregulated in breast cancer and enhanced expression correlated with high tumor grade. A role for elevated HoxB9 expression in breast tumor progression is demontrated by its ability to activate the ErbB and TGF-B pathways which influence tumor-associated phenotypes in cells.Methods and results:Overexpression of HOXB9 was found in 43% of primary breast cancer by RT-PCR and in situ hybridization (Figure 1A) and correlated with high tumor grade. Ectopic expression of HOXB9 in MCF10A mammary epithelial cells induced EMT, cell migration, invasion (Figure 1B, and 1C). It also increased the expression of angiogenic factors, which enhance the formation of new vessels in mouse dorsal air sac model. Conversely, genetic ablation of endogenous HOXB9 in MDA-MB-231 breast cancer cells suppresses their motility and angiogenic potential. Further, we confirmed that HOXB9-induced tumor phenotypes arise through the activation of both ErbB-AKT and TGFß signaling pathways. Finally, in mouse xenograft model, we observed that HOXB9 cooperates with activated H-Ras to transform mammary epithelial cells leading to large, vascularized and invasive tumors (Figure 2).Discussion:Our findings imply that overexpression of HOXB9 in human breast cancer contributes to tumor progression through activation of signaling pathways that alter both tumor-specific cell fates and tumor-stromal microenvironment, leading to increased invasion and metastasis. It is suggested that combined suppression of ErbB and TGFß signaling pathways to target breast cancers overexpressing HOXB9 may be effective in tumor inhibition.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 6145.
Heparanase: A Critical Determinant of Breast Cancer Metastasis to Brain.
Authors: Marchetti D. , Zhang L.
Published: December 2010

Identification of mechanisms responsible for brain-metastatic breast cancer (BMBC) is imperative for the development of novel therapies. Heparanase (HPSE) is the only functional mammalian endoglycosidase degrading heparan sulfate (HS), the main polysaccharide component of growth factor - binding cell surface and extracellular matrix proteoglycans. HPSE relevance in cancer invasion and metastasis has been established. HPSE overexpression correlates with cancer metastasis, tumor vascularity, and the reduced post-operative survival of cancer patients, making it a promising target for anti-cancer therapeutics.Her-2 is a member of the epidermal growth factor receptor (EGFR) family; Her-2 overexpression and EGFR positivity are key characteristics of BMBC. EGFR is important for Her-2 signaling via Her-2/EGFR heterodimer formation. However, mechanisms linking Her-2 function and pathways to precise targets and BMBC onset have not been investigated.We hypothesize that Her-2/EGFR activation augments BMBC by inducing HPSE. We examined HPSE expression, intracellular trafficking, and activity in human Her-2 expressing BMBC models. We demonstrate that: 1) HPSE is expressed in human BMBC cells and expression is augmented according to their metastatic propensities (231parental > 231Br > 231Br2 > 231Br3); 2) EGF induces HPSE expression and nucleolar localization in Her-2 expressing BMBC cells in a dose- and time-dependent fashion; 3) DNA Topoisomerase I (Topo I) is a HPSE target in nucleoli; and 4) HPSE affects Topo I activity and function. Topo I is essential for transcription, and abnormal levels of the enzyme have been observed in a variety of tumors, including brain tumors.These novel findings are relevant to BMBC mechanisms. HPSE – Her-2 interactions will be elucidated to decipher how Her-2 signaling regulates HPSE functionality and mechanistic linkages between these two oncogenic proteins in BMBC cell lines and pre-clinical models.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 6156.
Aberrant Expression of ARP2/3 in Breast Cancer and the Association with Disease Progression.
Authors: Moazzam M. , Ye L. , Kynaston H. , Mansel R. , Jiang W.
Published: December 2010

Background: Increased motility is an important characteristics of neoplastic cells, a cell function mediated through actin polymerization. During this process, aside from creation of new branches and lengthening of pre-existing actin molecule, Actin-Related Proteins (ARP)- 2 and ARP-3 work as a complex promotes polymerization through production of new nuclei for actin polymerization. This regulation is orchestrated by other intracellular regulators including and WAVE and WASP proteins, which have been shown to be aberrant in breast cancer (1,2). In this study we determined the differential expression of ARP-2 and ARP-3 and correlated the expression with various prognostic factors.Methods: Expression of ARP-2 and ARP-3 was examined in a cohort of mammary tissues (n=33 normal breast tissue and n=127 primary breast tumor tissue samples). Transcript levels of ARP2 and ARP3 were then determined using quantitative real time PCR (Q-PCR) and protein levels were assessed using immunohistochemical (IHC) staining. Results were analyzed by Mann-Whitney U test.Results: Cytoplasmic staining for both ARP-2 and ARP-3 was noted along with strong epithelial staining as compared to stromal cells. Quantitative real time PCR (Q-PCR) data analysis showed lower expression of both ARP -2 & -3 in tumour tissue as compared to normal but without statistical significance. ARP-2 expression was significantly reduced in tumour samples from patients with poor prognosis (p=0.037) and patients who died of breast cancer (p=0.0265). Primary breast tumor tissue samples from patients classified as TNM stage 3 and 4 showed statistically significant lower expression of ARP-3 as compared to normal tissue (p= 0.019 and 0.020, respectively). ARP-3 expression was also significantly lower in patients who developed local recurrence of breast cancer (p=0.027). Using a Spearman correlation analysis, ARP-3 transcripts were find to be significantly correlated with the WAVE-2 transcript (r=0.42, p<0.01).Conclusions: Breast cancer shows aberrant expression of ARP-2 and ARP-3, a pattern linked to the prognosis of the patients. ARP-3 and WAVE-2 may have an intimate interplay in this association which warrants further investigation.References1. Fernando HS et al. Expression of the WASP verprolin-homologues (WAVE members) in human breast cancer. Oncology. 2007;73:376-3832. Martin TA, et al. N-WASP is a putative tumour suppressor in breast cancer cells, in vitro and in vivo, and is associated with clinical outcome in patients with breast cancer. Clin Exp Metastasis. 2008;25:97-108

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 6169.