bluuebunny/crossref_metadata_2025 huggingface dataset:
Nanoparticle albumin-bound (nab) IDN5404: a novel vascular disrupting agent with potent antitumor and antiangiogenic effects.
Authors: Trieu V , D'Cruz O , Piacente M , Huang T , Faxon S , Ahmed S , Desai N
Published: December 2010

Abstract #2138

Background: The tumor vasculature is an established target for the therapy of solid tumors. IDN5404 is a thiocolchicine dimer with antitubulin and topisomerase 1 inhibitor properties. In this study, nab-formulated IDN5404 was examined for antitumor effects in three xenograft models and for antiangiogenic as well as vascular disrupting activity (VDA) in the avian embryonic chorioallantoic membrane (CAM) assays.  Materials and Methods: Subcutaneous human breast (MX-1 and MDA-MB-231), colon (HT29), and prostate (PC3) tumors were grown in athymic nude mice and treated intravenously (IV) with nab-5404 alone at 10, 20, 30, or 40 mg/kg (q3dx4 or q4dx3) and in combination with IV or intraperitoneal (IP) nab-paclitaxel (Abraxane; 10, 15 or 30 mg/kg, qdx5 or q4dx3). The effect of dose, schedule, and sequence of combination regimen was tested. Nab-5404 was administered 24-hr before, concurrently or 24-hr after Abraxane administration. The effect of nab-5404 on embryonic angiogenesis was examined in vivo using the standard chick CAM assay with 3-day old embryos (n = 18). The quail CAM assay was used to determine the time course of VDA. Quail embryos (n = 36) were exposed to nab-5404 (1 to 16 µg/ml) on day 7 and digital CAM images were acquired over a 60 min period for scoring VDA.  Results: Significant dose-dependent tumor growth inhibition (TGI) was observed in all xenograft models. In the MX-1 and HT29 models, nab-5404 (30 mg/kg, q3dx4) exhibited potent antitumor activity (P < 0.0001, ANOVA, vs. saline) alone and in combination with Abraxane (TGI for MX-1: 65% and 76%; TGI for HT29: 44% and 74% respectively). Nab-5404 was similarly effective with the q4dx3 schedule. In addition, when Nab-5404 (20 mg/kg, q4dx3) was administered 24-hr before, concurrently or 24-hr after Abraxane (5 mg/kg, q4dx3), the combination resulted in TGI of >99%, 88%, and 72% respectively. Using the in vivo chick CAM assay, nab-5404 inhibited embryonic angiogenesis by over 90% at 5 µg/CAM without affecting viability. Furthermore, rapid collapse of blood vessels was observed within 30 min in the viable quail CAM assay when embryos were exposed to nab-5404 at 4-16 µg/ml.  Discussion: Nab-5404 alone and in combination with Abraxane demonstrated significant TGI in xenograft models of human breast, colon or prostate cancer. This effect was shown to be due to the rapid VDA of nab-5404. Maximum antitumor activity was seen when nab-5404 was administered 24 hr before Abraxane. Nab-5404, a microtubule-destabilizing vascular disruptive agent, potentiates the cytotoxic effects of nab-paclitaxel, a microtubule-stabilizing agent and the data suggest that these two agents may be utilized effectively in combination.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2138.
Daily injections of the glutathione disulfide mimetic NOV-002 ameliorates hematologic toxicities from neoadjuvant chemotherapy in breast cancer patients enrolled in the NEO-NOVO trial, and significantly increases circulating dendritic cells.
Authors: Diaz-Montero C , Paphitis N , Onicescu G , Zhu Y , Verma N , Garrett-Mayer E , Schuhwerk KC , Pazoles CJ , Glück S , Montero AJ
Published: December 2010

Abstract #2141

Background: Chemotherapy induced anemia is a common problem in breast cancer. Anemic events [Hgb<10 g/dL, use of erythropoietin stimulating agents (ESAs), or transfusion] are likely to occur in 30-40% of breast cancer patients (pts) receiving doxorubicin-cyclophosphamide (AC) and docetaxel (T). Owing to the increased risk of cancer mortality, tumor progression, and thrombovascular events associated with ESAs, other alternatives are needed. Febrile neutropenia with AC-T is a dose-limiting event and occurs in 10-20% of pts without prophylactic G-CSF. NOV-002 has been previously shown to stimulate hematopoiesis and reduce chemotherapy related hematologic toxicities, as well as have immunomodulatory properties. Methods: To determine the effect of daily NOV-002 on hematopoiesis and chemotherapy induced hematologic toxicities, weekly blood counts were obtained on all pts enrolled in the NEO-NOVO trial (Montero, et. al. SABCS 2008). Pts do not receive prophylactic G-CSF in the absence of prior febrile neutropenia with chemotherapy. To determine if NOV-002 is associated with induction of circulating dendritic cells (DCs), flow cytometry analysis is performed on whole peripheral blood prior to therapy and on day 1 of each cycle. DCs are defined as: lineage-/HLA-DR+; CD11b+/CD11c+. P-values were obtained from the signed rank test. Results: Thus far, 102 chemotherapy cycles have been administered to 16 pts. Baseline (BL) hemoglobin (Hgb), absolute neutrophil counts (ANC), and DCs are listed below.    No grade 3-4 anemia has been observed. ESAs have been used in 2 pts (12.5%), and no blood transfusions were required. One pt (6.25%) had febrile neutropenia (0.9% of all cycles). Grade 3 neutropenia on day 1 of cycles 1-8 occurred in 2 pts (12.5%) or 1.9% of all cycles. One pt had dose delay by 1 week due to neutropenia. Cycle 5 day 1 (C5D1) DCs were significantly higher relative to BL with a trend towards being higher at C8D1. Moreover, median ANCs were significantly higher on C8D1, while median Hgb remained stable between C5D1 and C8D1. Conclusions: Anemic events and febrile neutropenia were much lower than expected with the addition of NOV-002 to AC→T, likely due to its hematopoietic properties. DC induction may be an important anti-tumor mechanism of action of NOV-002.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2141.
Induction of apoptosis in vitro and in vivo effects of a synthetic nitroflavone derivative on murine mammary adenocarcinoma cells.
Authors: Ca´rdenas MG , Blank VC , Zotta E , Marder NM , Roguin LP
Published: December 2010

Abstract #2145

Background: Flavonoids are polyphenolic compounds found in dietary components of vegetable origin. They exhibit multiple biological activities, such as antiproliferative, anxiolytic, antiallergic, antiangiogenic and antioxidant actions. We have previously demonstrated that the synthetic flavone 2'-nitroflavone (2'-NF) exerts a strong antimitogenic activity in various human and murine tumor cell lines, without disturbing the proliferation of non-neoplasic cells. In order to elucidate the mechanism of antitumor action, we first studied the in vitro ability of 2'-NF to induce apoptosis in a cell line derived from a murine mammary tumor (LM3), and then investigated its in vivo growth inhibitory effects.  Methods and results: Different experimental assays were performed to evaluate the in vitro apoptotic response of 2'-NF. Thus, when LM3 cells were treated in the presence or absence of 20 µM of 2'-NF, an increment in the proportion of hypodiploid cells was evident after 24 and 48 h of incubation. A typical DNA ladder fragmentation pattern was visualized by agarose gel electrophoresis and an increment in the total expression of the pro-apoptotic Bax protein was obtained by western blot after 24 hours of exposure to 2'-NF. Caspase 3 activity was also determined by fluorescence detection of the cleaved substrate Ac-DEVD-AMC after 24 hours of treatment.  In order to study the in vivo antitumor action, female BALB/c mice were first injected subcutaneously with 300000 LM3 cells. After 7 days, 1, 10 or 40 mg/kg of 2'-NF dissolved in 0.2 ml of vehicle (5% DMSO, 0.05% Tween-80, NaCl 0.15 M) were administrated i.p. twice a week for 3 weeks. During the treatment, tumor sizes were measured every 3 days and, at the end of the treatment, tumors were excised, measured and weighted. On average, treated-mice tumor volume was reduced between 70% (p<0.01) and 43% (p<0.05) with 10 and 40 mg/kg doses, respectively. No significant volume reduction was observed with 1mg/kg dose. Besides, tumors weight decreased approximately 50% (p<0.05) both with 10 and 40 mg/kg doses. TUNEL assay performed on tumor slices from treated (10mg/kg dose) and non treated mice showed that 2'-NF induces apoptosis on mammary tumors. In addition, western blot analyses of tumor lysate supernatants revealed an upregulation of the total levels of Bax and Fas receptor proteins in 2'-NF treated tumors with a concomitant downregulation of the antiapoptotic protein Bcl-2. Finally, toxicity studies performed in different organ-tissues from animals injected with 40 mg/kg of 2'-NF showed no histological alterations.  Conclusion: Taken together, our results suggest the potential therapeutic use of 2'-NF as an antitumor agent, capable of inducing apoptosis both in vitro and in vivo in breast cancer cells without generating systemic toxic effects.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2145.
Docetaxel and carboplatin followed by sequential capecitabine as first line treatment in patients with locally advanced or metastatic breast cancer.
Authors: Lorenzo A , Modolell A , Valero P , Murillo L , García-Bueno JM , Yubero A , Machengs I
Published: December 2010

Abstract #2155

Background: Anthracyclines and docetaxel (D) are the most active agents in metastatic breast cancer (BC). New regimens are being assesed to improve chemotherapy efficacy. D and carboplatin (C) has shown to be an active treatment in metastatic BC. Capecitabine (X) and D is also an active combination, despite the fact the toxicity profile needs to be improved. Sequential administration of D and X could lead to an improved toxicity profile compared with concomitant administration. We designed this study to evaluate activity and toxicity profile of treatment with D and C followed by sequential X in patients with locally advanced or metastatic BC.  Methods: Patients with locally advanced or metastatic BC, at least one measurable lesion, age ≥ 18 years, ECOG PS ≤ 2, and adequate bone marrow and hepatic functions, were included in the study. Prior chemotherapy or radiotherapy for advanced disease was not allowed. Patients received 6 cycles of D 75 mg/m2 iv D1 and C AUC5 iv D1, every 21 days followed by 12 cycles of X 2000 mg/m2/day o.r. D1-14 in a 21 days course.  Results: twenty patients have been analyzed. Median age was 55 years old (range 36-73), ECOG PS 0-1 94.4%; 90.0% of patients had infiltrating ductal carcinoma and 57.9% of patients had stage I-II at diagnosis. Main sites of metastatic disease were bone (35.0%), liver (25.0%) and lung (25.0%). Conservative surgery was performed in 35% of patients and mastectomy in 40%. Previous radiotherapy and chemotherapy was administered in 50.0% and 70.0% of patients, respectively. A total of 108 cycles of D and C (median 6, range 2-6) and 71 cycles of X (median 6, range 2-10) were administered. Median relative dose intensity was 96% for D and C and 95% for X. Over 17 evaluable patients for efficacy, 2 achieved complete response and 4 partial response; the ORR was 35.3% (95% CI:12.6-58.1). During D and C treatment, grade III/IV toxicities per patient were neutropenia (25.0%), leucopenia (10.0%), mucositis (5.0%), oedema (5.0%), febrile neutropenia (5.0%) and thrombophlebitis (5.0%). Most common grade II toxicities were nausea (20.0%), vomiting (15.0%) and asthenia (20.0%). During X treatment, grade III/IV toxicities per patient were neutropenia (7.1%) and hand-foot syndrome (7.1%). Main grade II toxicity was hand-foot syndrome (14.3%). Two patients finished treatment after 2 and 6 cycles of X due to diarrhea, oedema and anorexia, and hand-foot syndrome.  Conclusion: In this preliminary analysis, D and C followed by sequential X seems to be an active and well-tolerated regimen as first line treatment in patients with locally advanced or metastatic BC.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2155.
A multistage nanodelivery system for therapeutic applications and medical imaging: in vivo biodistribution and efficacy study in a murine orthotopic breast cancer model.
Authors: Tasciotti E , Bhavane R , Godin-Vilentchouk B , Serda R , Liu X , Robertson F , Ferrari M
Published: December 2010

Abstract #2160

The ability to deliver therapeutic compounds specifically to diseased sites is crucial for effectively treating human illnesses. Nanotechnology is emerging as a tool for resolving challenges in the delivery of poorly administrable drugs by transforming them into nanometer-sized particles (nanotherapeutics). The progress thus far gave rise to a large number of nanotherapeutics, but the accomplishment of the original objective, which is to increase drug concentration at target sites, has not been fully realized. A series of biological barriers pose as insurmountable obstacles which limit or completely abolish the ability to selectively deliver a therapeutic agent. We hypothesized that a multi-stage system for systemic delivery (MSDS) could be designed to interact with and successfully overcome sequential biological barriers. We developed a MSDS using biodegradable, biocompatible silicon particles optimally sized and shaped to travel into the blood flow, avoid RES, marginate and adhere to tumor vasculature. The first stage carriers (FSCs) contain nanopores of specific size that can load, carry, release over time, and deliver multiple types of second stage nanoparticles (SSNs). The nanoparticles can be optimized to reach, through vessels fenestrations, the tumor environment and finally release their therapeutic payload into target cells.    The ease in chemically modifying the silicon surface permits the attachment of fluorescent and radiolabeled imaging molecules to the FSCs. Fluorescent and magnetic SSNs can be loaded into the pores of the FSCs. Once assembled, the resulting MSDS can be imaged using different in vivo imaging modalities from its administration to its final localization into the body. We further modified our FSCs with PEG molecules to increase their circulation time and with antibodies directed against the VEGFR2 membrane protein to increase the localization of the MSDS in tumor associated vessels. We monitored the biodistribution of all MSDSs and characterized their biocompatibility in the different tissues. Finally, we loaded the pores of the FSCs with a lipidic nanoformulation of the potent anti cancer drug doxorubicin (dox-liposomes and dox-micelles) and followed the efficacy of this treatment. Taken together, these studies provide first time evidence that silicon nanoporous particles can be used as effective carriers for the simultaneous delivery of different nanotherapeutics in vivo.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2160.
Can nipple-areola-sparing mastectomy be an alternative to mastectomy? Over 10 years of follow up at a Japanese institution.
Authors: Sakurai T , Umemura T , Jinta E , Shimizu S , Suzuma T , Yoshimura G , Sakurai T
Published: December 2010

Abstract #22

Introduction: Cosmetic outcome is one of the most important aspects of surgical treatment of breast cancer. Therefore, to maximize the cosmetic outcome for patients who require a mastectomy, what can be done from an oncologically safe perspective must be considered. Recent reports have suggested that nipple-areola-sparing mastectomy (NSM) is oncologically as safe as a classical mastectomy (MT) and provides a better cosmetic outcome than MT. However, little is known about the long-term outcome of NSM.  Aim: To determine the oncological and technical outcomes of NSM compared with MT using long term follow-up data.  Patients and Methods: We retrospectively analyzed 788 patients, median age 51 years, with primary breast cancer who underwent NSM from 1985 to 2004. The indication of NSM in this study was defined as being for any stage, any tumor size and any tumor-areola distance. Briefly, NAC was preserved when palpation and the outlook of the nipple was normal. Median tumor size and tumor-areola distance were 2.5cm (0-10.0) and 1.8cm (0-9.0), respectively. Our surgical notes for NSM were; (1) a thin flap (approximately 5 mm thick subcutaneous adipose tissue) was placed close to the tumor, and a thick flap (more than 1cm thick subcutaneous adipose tissue) was created more than 2cm away from the tumor to conserve breast volume using subcutaneous fat, and (2) tissue under the NAC was left at 5 mm in thickness, but the major ducts were removed from within its lumen. Eighty-one patients (10%) underwent immediate breast reconstruction (IBR) using implants. We also analyzed 144 patients with breast cancer who underwent MT in the same period as those who underwent NSM. No patient received radiotherapy. We compared the local recurrence rate (LRR), disease-free survival (DFS) and overall survival (OS) between 788 NSM and 144 MT cases.  Results: Median follow-up of NSM and MT was 92 (10-252) and 87 (0-231) months, respectively. To our surprise, no complications of NSM including nipple and areola necrosis were recorded (0/788). For NSM, there were 70 patients (8.9%) with local recurrence, including 23 (2.9%) at the nipple, 4 (0.5%) at the areola, 4 (0.5%) at both nipple and areola, and 39 at the skin flap (4.9%). This suggested that the rate of NAC relapse was quite low. There were no significant differences in LLR between NSM and MT, (10y, 8.9% vs. 7.6%, p=0.62), DFS (10y, 83% vs. 83%, p=0.34) and OS (10y, 88% vs. 86%, p=0.34), respectively. Cosmetic outcome of NSM was better than that of MT in the majority of patients.  Conclusion: These results suggested that NSM provides a good cosmetic outcome and is as safe oncologically as MT. Our long term follow-up data shows that NSM is a possible alternative to mastectomy.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 22.
GRB7-dependent pathways are potential therapeutic targets in triple-negative breast cancer.
Authors: Sparano JA , Gray R , Goldstein LJ , Childs BH , Bugarini R , Rowley S , Baker J , Shak S , Badve S , Baehner FL , Perez EA , Shulman LN , Martino S , Sledge Jr. GW , Davidson NE
Published: December 2010

Abstract #25

Background: Breast cancer lacking expression of the estrogen and progesterone receptor and overexpression of HER2/neu (ie, "triple-negative” disease) accounts for about 10-15% of all breast cancer and is characterized by a higher risk of recurrence, early recurrence, resistance to cytotoxic therapy, and lack of any specific targeted therapy.  Methods: We extracted RNA from primary tumor samples of 246 patients with stage I-III triple-negative breast cancer (confirmed in a central lab) treated with 4 cycles of adjuvant doxorubicin (60 mg/m2) plus cyclophosphamide (600 mg/m2) or docetaxel (60 mg/m2) who were enrolled on trial E2197, and correlated RNA expression (by quantitative RT-PCR using a panel of 371 rationally selected genes) with recurrence. There was no difference in recurrence between the two treatment arms in the entire study population, nor in the 246 patients in this analysis (of whom 59 recurred) after a median followup of 76 months.  Results: Higher expression of GRB7 was the only gene significantly associated with an increased risk of recurrence (nominal p value 0.0000853, Korn's adjusted p value controlling false discovery at 10% (KP10) p=0.0359), but did not correlate with any clinicopathologic features except age (low expression associated with age > 65 years, p=0.03). In a Cox proportional hazards model adjusted for age, nodal status, tumor size, and grade, higher GRB7 expression was associated with an increased risk of recurrence when evaluated as a continuous variable (hazard ratio 3.41; p = 0.001) or as a dichotomous variable (hazard ratio 2.24 above vs. below median; p=0.006). The 5-year recurrence rates were 10.5% (95% C.I.7.8%, 14.1%) in the low and 20.4% (95% C.I. 16.5%, 25.0%) in the high GRB7 groups. There were only six genes whose expression correlated with GRB7 (r> 0.4), including ERBB2 (r=0.70), DDR1 (discoidin domain receptor tyrosine kinase 1; r=0.53), KRT19 (keratin 19; r=0.49), ERBB3 (r=0.48), GPR56 (G protein-coupled receptor 56; r=0.48) and PHB (prohibitin; r=0.42).  Conclusions: GRB7 is a calmodulin-binding protein which has an SH2 (Src homology 2) domain that binds to phosphorylated tyrosine residues and other specific protein targets, and which plays a critical role in signaling (EGFR, HER2), motility (eprhins), migration (focal adhesion kinase), and cell-matrix/cell-cell interactions (integrins). Higher GRB7 RNA expression is associated with a significantly higher risk of recurrence in triple-negative breast cancer, indicating that GRB7 or GRB7-dependent pathways are potential therapeutic targets in triple-negative disease.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 25.
Atypical and columnar cell lesions in breast needle biopsies for indeterminate microcalcifications: predictors of higher risk findings requiring surgical excision.
Authors: Albarracin CT , Sigauke E , Whitman G , Yang WT , Resetkova E , Johnson MM , Nguyen CV , Sneige N
Published: December 2010

Abstract #3004

Background: Columnar cell lesions (CCL) are frequently identified in biopsies performed for calcifications. The primary objective of this study is to identify histologic features of CCL in directional vacuum-assisted needle biopsies (DVAB) that could predict a higher risk lesion requiring excision.  Design: From June 2002 to January 2008, we retrieved the materials for 88 DVABs and their subsequent segmental resections. We excluded carcinomas, intraductal papillomas, fibroadenomas, adenosis, or radial scars. Using World Health Organization criteria, CCLs were classified as CCL with no atypia (CCL-NA), CCL with cytologic atypia (CCL-CA; flat epithelial atypia), or architectural atypia/atypical ductal hyperplasia (CCL-ADH) in the background of CCL. In cases with atypia, the involved terminal duct lobular units/ducts (TDLUs) were quantified. Atypical cases were separated two groups as follows: minimal ADH (≤2 TDLU and minimal cytologic atypia) and extensive ADH/borderline DCIS (≥3 TDLU and/or significant cytologic atypia bordering on DCIS).Radiographs were evaluated to assess amount of calcifications removed. Biopsy results were correlated with the subsequent surgical excision results.  Results: Eight of 88 biopsies were excluded because of overdiagnosis or because the lesions were not associated with microcalcifications. The remaining 80 biopsies included 5 cases of CCL-NA, 4 pure CCL-CA, 20 mixed CCL-CA + ADH and 51 pure CCL-ADH. Microcalcifications were identified in association with the targeted lesions. There were no higher risk findings in any of the resection specimens with CCL-NA and pure CCL-CA. Ten of the atypical lesions showed carcinoma on excision (9 DCIS and 1 invasive cancer). Twenty-five percent (9/36) extensive ADH/borderline DCIS showed carcinoma on excision. In contrast, only 2.5% (1/39) of minimal ADH cases showed carcinoma on excision. Radiologic review revealed residual abnormality in this one case.  Conclusion: In summary, the extent of lobular involvement by atypia (CCL-CA or CCL-ADH) correlated with the presence or absence of higher risk lesions upon subsequent excision. These results suggest that additional surgery may not be necessary for CCL-CA or CCL-ADH that is limited to ≤ 2 foci provided sampling of the microcalcifications is adequate.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3004.
MUC1 expression in early-stage triple-negative breast cancer.
Authors: Miedler J , Abdul-Karim F , Wang N , Baar J
Published: December 2010

Abstract #3009

Background. Triple-negative breast cancer (TNBC) comprises ∼10-15% of newly-diagnosed early-stage breast cancer. Most TN tumors are high grade and have a high incidence of recurrence and distant metastases. Patients with early stage TNBC usually do not receive any maintenance systemic therapy beyond standard cytotoxic chemotherapy and may therefore benefit from novel experimental therapies, including immunotherapy. MUC1 is a tumor antigen expressed on differentiated tumor cells, including BC, and therefore represents an ideal target for MUC1-based vaccination. In this study, we evaluated TNBC for the expression of MUC1 by immunohistochemical (IHC) analysis.  Material and Methods. Fifty-three cases of ER(-) PR(-) HER-2/neu(-) early-stage BC were evaluated for expression of the MUC1 glycoprotein by IHC stain analysis. Immunoreactivity was graded based on the intensity and percent of tumor cells staining for MUC1. Intensity of staining was graded as negative (0), positive (1) or strongly-positive (2), and the percent of tumor cells staining positively was graded from 0 to 100%. A specimen was considered positive if immunoreactive cells were found in at least 5% of tumor cells. An overall score of 0 to 2.0 was then calculated for each case by multiplying the intensity of staining by the percent of tumor cells staining positively.  Results. Forty nine of the fifty three cases of TNBC (92%) were MUC1+ by IHC stain analysis. The mean score was 0.90, with a range of 0 to 1.9. Cases were evenly distributed over this range, and the majority of cases (66%) exhibited moderate to strong expression of MUC1 (score 0.5 to 1.90). A minority of cases (26%) demonstrated weak expression, and 8% of cases lacked expression of MUC1 by study criteria.  Discussion. MUC1 expression has been documented in the epithelium of secretory tubules, acini, and intralobular ducts of normal breast tissue and in breast carcinoma of undefined subtype. However, this is the first study to specifically evaluate TNBC for the expression of MUC1. The results of our study suggest that the vast majority of cases of early stage TNBC express MUC1. Since other MUC1+ tumors such as prostate, pancreatic and ER+ breast cancer have been successfully vaccinated with MUC1-based peptide vaccines, this study suggests that consideration may be given to patients with early stage TNBC as possible candidates for such novel MUC1-based vaccines after having completed standard therapies for TNBC.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3009.
Intra operative determination of axillary node metastasis by RT PCR: experience on 50 breast cancer patients.
Authors: Baranzelli M , Chauvet M , Verhults P , Giard S , Révillion F , Peyrat J , Robin Y , Bonneterre J
Published: December 2010

Abstract #3012

Background: Sentinel lymph node (SLN) status is highly predictive of overall axillary lymph node involvement. SLN positive patients(pts)with macro or micrometastasis require delayed axillary clearance. Intraoperative imprints have a good sensitivity for detection of macrometastasis but micrometastases are seldom diagnosed. Thus, we tested the intraoperative determination of axillary node metastasis according to the Rapid Gene Search Breast Lymph Node (BLN) Assay (Veridex) based on RT-PCR.  Patients and methods:50 pts operated between September 17 and March 20 ,2008, had a SLN procedure. Lymph nodes were cut in 2 mm slices, and 1 out of 2 was examined with Rapid BLN Assay; the other one had intraoperative imprints and definitive histological assessement in paraffin-embedded sections including serial slices with hematoxilin eosine safran coloration and Cytokeratine AE1-AE3 immunochemistry.  Results : 103 lymph nodes were examined with a median number of 2 SLN by pt. With Rapid BLN Assay, 12(9 pts) were positive and confirmed by histological interpretation on the other half for 11 (8 pts). 89 lymph nodes were negative (41 pts); in 1 (1 pt), isolated tumor cells (CTI) were detected in the other half at histological interpretation. Intraoperative imprints detected 4 macrometastases. One node declared positive at the intra-operative examination imprint corresponded to a granulomatous lesion after histological interpretation and Rapid BLN Assay was negative. Sensitivity was 31% for intraoperative imprints, 92% for Rapid BLN Assay and the histological examination of paraffin--embedded sections. The median time for intraoperative determination was 34 minutes for 1 SLN compared with 20 minutes for imprints.  Conclusion: Given the high sensitivity of the rapid BLN assay, the excess duration of the procedure is acceptable for the surgeon. Furthermore, this procedure avoids a secondary axillary clearance in pts without intraoperative examination or with negative imprints and with metastases in SLN.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3012.
Quality improvement analysis of HER2 FISH testing in breast cancer in response to the new ASCO/CAP guidelines.
Authors: Fang M , Thalody V , Tannenbaum S
Published: December 2010

Abstract #3013

Background: The new ASCO/CAP guidelines for HER2 testing in breast cancer redefined the interpretation of HER2 FISH testing result. A new category of equivocal result is defined as FISH ratio of 1.8-2.2. Patients with equivocal results constitute a poorly studied subgroup with uncertain association of test scores to benefit from HER2-directed therapy. Additional quality control and quality assurance recommendations were also detailed in the guideline. As part of our institutional effort for quality improvement of HER2 testing, we reviewed the FISH HER2 testing performed in our institution for the past four years.  Methods: The FDA-approved PathVision HER2 FISH kit (Abbott Molecular, CA) was used for all FISH testing. The Herceptest ParmDx antibody (Dako Cytomation) and the Neu-Her2/Cerb-2 antibody (Vision Biosystems) were used for all IHC testing. The new ASCO/CAP interpretation criteria were applied to the FISH enumeration results and compared with the IHC results and interpretation. Correlation with clinical decision of trastuzumab therapy and clinical outcome was reviewed.  Results: A total of 554 samples between 2004 and 2008 were tested for HER2 FISH. According to the new interpretation criteria, 12.8% of the samples had amplified values and 3.8% of the samples had equivocal values. IHC was negative for Her2 for all the samples with equivocal FISH results. Only one patient was treated with trastuzumab. In addition to this equivocal category, some samples showed genetic heterogeneity with some of the tumor showing cells with HER2 amplification whereas cells at different areas of the same tumor were not amplified. Half of these patients were treated and the remaining 50% were not. The discordant rate between FISH and IHC was 3.6% overall, and aneusomy 17 may explain some of the discordance. Of patients with discordant HER2 results by FISH and IHC, physicians' decision to treat with trastuzumab or not appeared to be based on either FISH or IHC with equal frequencies.  Conclusion: Due to small sample size and retrospective nature of this review, outcome data was uninformative in these small cohorts. As suggested by the ASCO/CAP expert panel, pooling data from all adjuvant trastuzumab trails on subjects with equivocal results would be helpful to provide an evidence base for physicians to make treatment decisions. Alternatively, we are beginning to explore molecular expression profiles to see whether some of these equivocal or discordant cases behave more like HER2 positive or negative patients.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3013.
Effects of short-time tamoxifen therapy on breast cancer apoptosis and cell cycle regulators.
Authors: Madeira M , Millen EC , Vivian F , Logullo [F , Gebrim LH
Published: December 2010

Abstract #3017

Background: Tamoxifen is the most used endocrine therapy in patients with estrogen and/or progesterone positive receptors breast cancer. Tamoxifen acts in a complex manner as estrogen antagonists in tumor tissue, and its interaction with growth factors may lead to inhibition of cell proliferation.  Objectives: The aim of this study was to evaluate the expression of estrogen receptor (ER), progesterone receptor (PR), COX-2, Ciclyn D-1, Bcl-2, BAK, BAX and nitric oxide synthase (NOS I, II and III) in breast carcinoma cells using immunohistochemical essay on tumor samples before and after short-time (14 days) exposure to tamoxifen.  Material and Methods: A randomized double-blind study was performed with 29 stage II and III breast cancer patients divided according to the following groups: Group C (n= 15 / control) and Group T (n=14 / 20mg of TAM/day for 14 days). Immunohistochemical expression of ER, PR, COX-2, Ciclyn D-1, Bcl-2, BAK, BAX and NOS I, II and III were evaluated before (diagnosis biopsy) and after treatment. The results were classified according Allred score (semi quantitative), based on the intensity and fraction marked cells. The scores of each reaction were compared pre and post treatment, and also to the control group.  Results: The mean scores of each reaction before and after treatment for Group T and Group C are seen in the table 1.    No significant difference was observed with short-time tamoxifen therapy (table 2 - Mann Whitney test).    Discussion: Although tamoxifen has been in clinical use for over 30 years in oncology, its complex manner of action is not totally elucidated. Our results did not show a significant change in breast cancer apoptosis and cell cycle regulators, but bring new data about the short-time exposure to tamoxifen.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3017.
Computational inference of gene expression signatures differentiating between breast cancers responsive or resistant to letrozole.
Authors: Georgieva A , Evans DB , Miller J , Sing T , Dixon J , Miller WR
Published: December 2010

Abstract #3024

Introduction:Genes which have been shown either to be differentially expressed or differentially changed between responders and non-responders to the neoadjuvant treatment with the aromatase inhibitor letrozole have been described. However, their network structure and interconnectivity have not been formally analysed.Bayesian network (BN) inference algorithms have demonstrated promise in the analysis of genomics data to recover fragments of biological pathways due to their ability to capture many types of relationships between variables and to handle noisy data.An inference score (4), normalized to be between -1 and 1, is a measure of the strength of the interaction (either positive or negative) between different nodes in the recovered network.  Materials and methods:This computational study uses BN methodology to elucidate any possible network structure and refine a gene expression signature distinguishing responders from non-responders groups to letrozole treatment. It has been applied to 205 covariables (69 baseline expression, 45 day 14 expression and 91 changes in expression with treatment), identified via Random Forests as being differentially expressed between 37 tumours responding to neoadjuvant letrozole and 15 non-responders. The number of false positive relationships was limited by considering 4 different sub-sets of the original data 1) all 205 variables, but allowing for limited connectivity between them; 2) only the baseline variables 3) only the change variables with treatment and 4) only day 14 post-treatment variables. It was conjectured that the variables linking significantly to response status would appear in the overlap between lists 1) and 2), lists 1) and 3) and lists 1) and 4).  Results:The analysis of the overall dataset highlighted the importance and the interconnectivity of ribosomal proteins. More specifically, the BN algorithm found two connected networks of 20 genes each with 13 and 14 ribosomal proteins and an average inference score of 0.65. In addition, BN links patient type (responder vs. non-responder) directly to genes that have been implicated in the natural history of breast cancer such as BCL-2 and KIAA0101 with inference scores of 0.4 and 0.45, respectively. The inferred network also contained a link with inference score of 0.4 between patient type and phosphatidylinositol glycan, whose oncogenic role has been suggested by others. The preliminary network analysis of the baseline covariables did not yield any meaningful pattern that differentiates the two patient types, except for the involvement of the cyclin E2 pathway with a “composite” inference score of 0.33.  Conclusions: With initial encouraging results, further refinement of the BN analysis, as well as enrichment of the data should facilitate the discovery of a predictive baseline gene expression pattern.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3024.
Prognostic relevance of circulating tumor cells (CTCs) in peripheral blood of breast cancer patients before and after adjuvant chemotherapy – translational research program of the German SUCCESS-trial.
Authors: Schindlbeck C , Rack B , Jueckstock J , Schneeweiss A , Thurner-Herrmanns E , Schneider A , Rezai M , Beckmann M , Pantel K , Lichtenegger W , Sommer H , Friese K , Janni W , Chatsiproios D
Published: December 2010

Abstract #303

Background:  The detection of Disseminated Tumor Cells in the Bone Marrow (BM) of Breast Cancer patients has shown prognostic significance in all stages of the disease. An alternative towards BM examination could be the analysis of peripheral blood for Circulating Tumor Cells (CTCs), which has demonstrated prognostic significance in metastatic disease. We analyzed peripheral blood samples from pts. Before and after adjuvant taxane-based chemotherapy as part of the translational research program of the German SUCCESS-trial.  Methods: Peripheral blood samples (23 ml each) from 1500 N+ and high risk N- breast cancer pts were analyzed for the presence of CTCs with the CellSearchSystem (Veridex, USA) before and after adjuvant chemotherapy. After automated immunomagnetic enrichment with an anti-Epcam-antibody, cells are labelled with anti-cytokeratin (8,18,19) and anti-CD45 antibodies to distinguish between epithelial cells and leukocytes. Samples are screened automatedly and positive events are visualised on a screen for re-evaluation.  Results: In143 pts (10 %) more than 1 CTC was detected before the start of systemic treatment (mean 14, range 2-827). 2 CTCs were found in 4 % of pts, 3 % had 3-5 CTCs and 1 % 6-10 and >10 CTCs, resp. The presence of CTCs did not correlate with tumor size (p=.32), grading (p=.36), hormonal status (p=.28) or Her2 status of the primary tumor (p=.82), but with the presence of lymph node metastases (p=.003). As negative control, three of 74 healthy individuals showed more than 1 CTC.  After completion of chemotherapy, 9 % of pts (n=130) presented with >1 CTC (mean 6, range 2-124). Of those initially CTC positive, 10% remained positive (n=15), whereas of those initially CTC negative, 8 % turned positive after chemotherapy (n=115, p=.42).  21 recurrences have been reported during a median follow-up time of 12 months, and 7 pts have died of their disease. While detection of CTCs before systemic treatment did not show prognostic relevance for DFS (p=.89) and OAS (p=.71), presence of CTCs after chemotherapy was a significant predictor for both reduced DFS (p=.04) and OAS (p=.03).  Conclusions:  Our preliminary results demonstrate prognostic relevance of the presence of CTCs after adjuvant chemotherapy for the first time. Even in the adjuvant setting as well as during follow up, the examination of CTCs could be a valuable tool for monitoring the disease and the effectiveness of therapies. Longer follow-up of the SUCCESS-trial will have to be awaited to confirm these findings.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 303.
Comparative effect of tibolone and its metabolites on sulfatase activity in normal and cancerous breast tissues.
Authors: Pasqualini JR , Chetrite GS , Cortes-Prieto J
Published: December 2010

Abstract #3035

Background: Tibolone (Org OD-14, active substance of Livial®) is a synthetic steroid with a 19-nor-testosterone derivative structure. This compound has weak estrogenic, progestagenic, and androgenic properties and is used to prevent climacteric symptoms and postmenopausal bone loss. Tibolone is extensively metabolized in 3a- and 3ß-hydroxy-derivatives and their 4-en isomer. Previous studies in this laboratory demonstrated that tibolone and its metabolites are anti-sulfatase agents in the hormone-dependent breast cancer cells: MCF-7 and T-47D, while the present study explores their effect on estrone-sulfatase activity in normal and cancerous breast tissues.  Materials and Methods: : Two regions of the mammary tissue from each post-menopausal patient with breast cancer (n=8) were selected : the tumoral tissue and a distant zone which was considered as normal. Slices of tumoral or normal breast tissues (25-95 mg) were incubated in buffer (20 mM Tris-HCl, pH 7.2) with physiological concentrations of [3H]-estrone sulfate (E1S) (5x10-9M) alone or in the presence of tibolone or its metabolites (5x10-5 or 5x10-7M) during 14h at 37°C. E1S, estrone and estradiol were characterized by thin layer chromatography and quantified using the corresponding standard.  Results: In breast cancer tissues the sulfatase activity was twice as high as in normal breast tissue. At concentrations of 5x10-7 and 5x10-5M tibolone inhibits sulfatase activity in breast cancer tissue by 25 and 33% respectively. Using the same concentrations for the 3a-hydroxy these were 32 and 57%, for the 3ß-hydroxy 22 and 32%, and for 4-en derivatives 0 and 48% respectively. In the normal tissues the values were: for tibolone, 3 and 20%, for the 3a-hydroxy, 16 and 29 %, for the 3ß-hydroxy, 13 and 25%, and for the 4-en derivative 0 and 22% respectively. In these experiments, it was observed that estrone does not convert to estradiol.  Conclusion: The present data indicates that the estrone sulfatase activity is higher in breast cancer tissue and that the inhibitory effects of tibolone and its metabolites are significantly higher in this tissue than in the normal breast. It is suggested that the present results can open new possibilities in the knowledge of the mechanism of action of tibolone and its metabolites in normal and cancerous breast.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3035.
MicroRNA-206 expression is down-regulated in estrogen receptor a-positive human breast cancer.
Authors: Kondo N , Toyama T , Sugiura H , Fujii Y , Yamashita H
Published: December 2010

Abstract #3042

There are large-scale molecular differences between estrogen receptor (ER)-positive and ER-negative breast cancers. ERa is essential for estrogen-dependent growth, and its level of expression is a crucial determinant of response to endocrine therapy and prognosis in ER-positive breast cancer. Multiple mechanisms involved in altering ERa gene expression in breast cancer have been identified, including ERa gene amplification as well as transcriptional silencing by DNA methylation of CpG islands within the ERa promoter and mutations within the open reading frame of ERa. However, expression levels of ERa in breast cancer tissues differ widely among patients, and frequently change during disease progression and in response to systemic therapies. Recent evidence has shown that microRNA (miRNA) mutations or mis-expression correlate with various human cancers, and miR-206 is reported to decrease endogenous ERa mRNA and protein levels in human MCF-7 breast cancer cells via two specific target sites within the 3'-untranslated region of the human ERa transcript. In this study, we show that miR-206 expression is markedly decreased in ERa-positive breast cancer tissues assayed by quantitative RT-PCR analysis. We observe that miR-206 expression is inversely correlated with ERa but not ERb mRNA expression and the expression levels of miR-206 are gradually decreased as the score for ERa protein expression increases in breast cancer tissues. Moreover, transfection experiments revealed that introduction of miR-206 into estrogen-dependent MCF-7 cells inhibits cell growth and induces a dose-dependent repression of ERa mRNA levels. Transfection with miR-206 produces a dose-dependent decrease of mRNA expression of ERa-target genes, such as progesterone receptor, cyclin D1 and pS2 in MCF-7 cells. Our results suggest that miR-206 is a key factor for the regulation of ERa expression in development and progression of human breast cancer and could be a novel candidate for endocrine therapy that targets only ERa.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3042.
Insulin like growth factor in breast cancer subtypes.
Authors: Yerushalmi R , Gilks B , Nielsen T , Leang S , Cheang M , Woods R , Gelmon K , Kennecke H
Published: December 2010

Abstract #3048

Background: Insulin Like Growth Factor -1 Receptor (IGF-1R) is an important new therapeutic target expressed in all cancer types. IGF-1R supports cell survival pathways implicated in resistance to cancer therapy. Knowledge of the pattern of IGF-R1 expression among breast cancer subtypes and its impact on prognosis may enhance development of therapeutics targeting this pathway.  Methods: Patients with early breast cancer cases, stage I-III, referred to the BC Cancer Agency from 1986 to 1992 were included. Archival paraffin tissue blocks were used to construct a tissue microarray. Among 4,046 patients with early stage on the TMA, 1,238 patients (30.6 %) were excluded due to missing subtype biomarkers, IGFR staining or both. Breast cancer subtypes were defined as Luminal A (ER/PR+, HER2- and Ki67 <19%), Luminal B (ER/PR+,and HER2- and Ki67 >19%), Luminal HER2+ (HER2+ and ER/PR+), HER2 (HER2+ and ER-and PR-), and Basal {HER2-,ER-PR- and (CK 5/6+ and/or EGFR+)}. IGF-1R staining was done with Santa Cruz antibody and was scored negative if there was no or weak staining and positive if staining was moderate or strong. Chi-square and Kaplan-Meier Survival analysis were done to compare IGF-1R expression among subtypes and determine impact on Breast Cancer Specific (BCSS) and Overall Survival (OS).  Results: A total of 2,808 evaluable cases were included with a median follow-up of 12.5 years. IGF-1R staining was scored positive in 86.4%, and negative in 13.6%. Cases with Luminal A (1,676), Luminal B (426), Luminal HER2+ (199), HER2 (206 ) and Basal (301 ) had an IGF-1R+ rate of 89.9%, 94.4 %, 83.4%, 59.2% and 76.1%, respectively (p<0.0001). 10 year BCSS was 68% (95% CI 66.0%-70.0%) in IGF-R1+ and 63% (95% CI 59.1%-66.9%) in IGF-R1 - group. Among subtypes, IGF-1R positivity was associated with improved BCSS only in Luminal A patients (p=0.015 ) and was not prognostic in other subtypes.  Conclusion: Luminal breast cancer subtypes are associated with high rates of IGF1-R expression, while non-luminal groups have lower rates of expression. The prognostic impact of IGF-R1 expression supports the role of this pathway as a therapeutic target particularly among hormone receptor positive breast cancer.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3048.