bluuebunny/crossref_metadata_2025 huggingface dataset:
Awareness of breast cancer among women in an urban region of China.
Authors: Wang G , Tian F , Wang L , Lian R , Zhao W , Liang X , Gao J
Published: December 2010

Abstract #2104

Introduction: Even though breast cancer can be cured if detected early and treated appropriately, it remains the most common cause of cancer mortality for women living in some major cities in China. Moreover, the incidence of this disease has increased steadily in China over the last two decades. The aim of this cross-sectional study was to investigate the knowledge about breast cancer concerning risk factors, signs and symptoms, effective screening methods, treatment choices and prognosis among native Chinese women.  Method: Two hundred and seventy healthy women were invited to complete a self-administered questionnaire in Taiyuan city of China. Participants' characteristics, including age, education, occupation, and awareness of breast cancer diagnosis, treatment and prognosis were collected and analyzed.  Results: Two hundred and thirty nine women had returned the questionnaire. The response rate was 88.5%. The median age of women in this study was 44 years old (range: 25 – 72). Among them, more than one third had received tertiary education, 6.7% had breast cancer family history, and one quarter ever had benign breast disease. Most of the participants (60.3%) did not know how to perform breast self-examination (BSE), whereas 61.5% never did BSE. Only 2.9% of those who had performed BSE did so regularly. As for effective screening methods, more than half of the participants chose clinical breast examination (CBE), and 21.8% regarded mammography as a reliable mean. Most of women considered breast cancer a curable disease and assumed that therapeutic approaches should include surgery, chemotherapy, endocrine therapy, radiotherapy, and in some cases, traditional Chinese medicine. Only 8.8% of the participants identified two or more of the eight breast cancer risk factors. The factors mostly identified included breast cancer family history (33.5%), late birth (10%), and early age at menarche (9.6%). The breast cancer signs and symptoms mostly indicated were breast lump (54.4%), breast pain (26.4%), and nipple discharge (23.9%). Women who received high level education appeared to be more likely to perform BSE and possessed more knowledge regarding to breast cancer risk factors and treatment.  Conclusion: The study indicates that awareness of breast cancer among Chinese women needs to be improved, and that breast cancer early-detection measures needs to be enhanced in China.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2104.
The use of a mixed reality breast simulator with an innovative touch map feedback system to teach breast history-taking and examination skills.
Authors: Deladisma AM , Kotranza A , Shah H , Fox P , Rossen B , Imam T , Wang S , Gucwa A , Pugh C , Lok B , Lind DS
Published: December 2010

Abstract #2105

Introduction: Physical examination remains an important method of breast cancer detection. Unfortunately, many health care professionals express concerns about missing breast lesions and current methods of teaching this essential skill are limited. Through an interdisciplinary collaboration, we created an immersive virtual patient to teach health professions students history-taking and breast examination skills.  Methods: Fifteen physician's assistant (PA) and 13 medical students (MS) interacted with a mixed reality human (MRH, a computer avatar with a mannequin-based breast simulator) with a breast complaint (Figure 1).    Students spoke to and touched the MRH to take a history and examine a simulated breast with two masses of differing size and consistency. Subjects were surveyed regarding the usefulness of the virtual teaching tool and composed a patient note documenting pertinent history and physical examination findings. Students received feedback regarding the content of their patient note (number of 17 essential content items documented) and on the completeness of their breast examination (percentage area covered) using a color-coded touch map.  Results: Student feedback related to the utility of this virtual educational tool was positive. Students only documented a mean of 7.8±2.7 (range=4-15) essential content items in the breast history. The completeness of the breast exam was a mean of 82% (range=62% to 97%) of total breast area examined (Figure 2, green=area examined, red=area missed).    More clinically experienced students (MS 3 and 4, N=9) performed better than those with no clinical experience (MS 1 and PA 1, N=19) in both history-taking (58% vs. 40%, p<0.05) and completeness of exam (90% vs. 84%, p<0.05). Fifty percent of students were able to locate at least one mass but only 14% were able to correctly document the location of both lesions.  Conclusions: The simulated experience differentiated performance among students with varying levels of clinical experience and identified a need for improved teaching and greater practice among all students. MRH scenarios provide a less anxious learning environment for students to practice breast history-taking and examination.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2105.
Canadian pattern of care for anemia: comparison of chemotherapies in adjuvant breast cancer setting.
Authors: Quinn A , Wong C , Younus J , Dranitsaris G , Goel R , Trudeau M
Published: December 2010

Abstract #2118

Background: Anemia is a common side effect of chemotherapy in adjuvant breast cancer patients (pts) but its treatment may be inconsistent. Erythropoiesis-stimulating agents (ESAs) are the only alternative to blood transfusion in the treatment of chemotherapy-induced anemia, yet their appropriate use continues to be defined. The purpose of this study was to determine the frequency of anemia associated with adjuvant chemotherapy in Canadian breast cancer pts and to explore the corrective interventions which were undertaken.  Methods: A retrospective chart review was conducted at 11 cancer centres in Ontario, Canada. Beginning on September 1, 2004 data were collected for 60 consecutive cases of adjuvant breast cancer at each centre, giving a total of 620 patients. Chemotherapy consisted mainly of FEC100, AC-T, AC, CEF, FEC50, and AC-T Dose Dense. Overall, 8% of pts were enrolled in a clinical trial and 76% received adjuvant radiotherapy. Hemoglobin level (Hb), supportive therapy (antibiotics, GCSF, ESA, and transfusion) and type of chemotherapy were recorded at each cycle.  Results: The median pt age was 53. The mean baseline Hb was 132.8 (92.0-163) g/L, 11.6% of pts had Hb<120g/L. During the course of chemotherapy, the percentage of subjects with Hb<120 at least once was 55.8%, Hb<110 was 37.8%, Hb<100 was 21.2 % and Hb<90 was 11.5%, with a median of 36.0, 52.5, 71.0, and 75.0 days to develop anemia respectively. For pts whose Hb dropped below 100 g/L, 45.4% occurred after the 1st cycle, 68.8% after the 2nd, 83.7% after the 3rd, and 92.2% after the 4th chemotherapy cycle. Overall, 13.2% of subjects received prophylactic antibiotics, 26.3% were supported by G-CSF, 9.0% received an ESA (epoetin alfa) during their chemotherapy and 7.4% were transfused. The rates of anemia as well as means of intervention are summarized in Figure 1. ESAs were not used prophylactically in this setting.  Conclusions: In Canada, breast cancer patients receiving adjuvant chemotherapy commonly become anemic during their course of treatment. This retrospective study demonstrated that ESAs play an important role in the management of anemia in the adjuvant setting. Canadian treatment guidelines recommend that ESA therapy be initiated when Hb <100 g/L. Our finding shows that ∼42% of patients whose Hb dropped below 100g/L were treated with ESAs, suggesting that ESAs were not overused in Canadian practice.   

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2118.
Targeting of breast cancer with non-oncology drugs – possible novel therapeutic option for triple-negative breast cancer.
Authors: Liedtke C , Yan K , Wu Y , Hortobagyi GN , Symmans WF , Valero V , Goette M , Kiesel L , Pusztai L
Published: December 2010

Abstract #2119

Introduction: Triple-negative breast cancer (TNBC) is defined by lack of ER, and PR expression and normal HER2 expression. It is characterized by aggressive biological presentation and unfavorable outcome. It is associated with frequent lack of response to current chemotherapy agents. Recently, 392 drugged genes that serve as targets for 1557 both oncologic and non-oncologic drugs were identified from the pharmacologic database DrugBank (Lauss et al. Pharmacogenomics 2007). In this study, we compared expression levels of these known drug targets between TNBC and receptor-positive cancers. Methods: Gene expression profiles were obtained from fine needle biopsies of newly diagnosed early stage breast cancer before any therapy (n=133, MDACC dataset). Differential expression of these genes was validated in a second independent data set (n=286, Rotterdam dataset) and in a panel of cell lines (n=19). In order to assess the functional relevance of known drug targets overexpressed in TNBC we performed in vitro experiments. Cell lines were treated with various concentrations of selected drugs alone or in combination with paclitaxel. Results: We mapped 675 U133A probe sets representing 347 unique drug targets to the Affymetrix U133A Genechip. 44 drug targets were overexpressed in TNBC compared to non-TNBC in the MDACC dataset. Thirty-three of these (75%) were also overexpressed among TNBC compared to non-TNBC in the Rotterdam dataset. Glutathione-S-transferase pi (GSTpi, target of clomipramine) was the most highly and consistently overexpressed target in human TNBC and in estrogen- and HER2-receptor-negative breast cancer cell lines. The GSTpi overexpressing (and triple negative) human mammary cell line HBL-100 showed dose-dependent inhibition of growth after 144 hrs of incubation with clomipramine, whereas growth of the GSTpi-low expressing (non-triple negative) breast cancer cell lines MCF-7 and SKBR3 was not inhibited by this drug. Treatment with clomipramine did not alter sensitivity to paclitaxel in either cell line in vitro. Conclusion: Gene expression analysis indicates that targets for several known drugs are over expressed in TNBC relative to other types of breast cancers. We hypothesize that some of these drugs may influence the behavior of TNBC and may represent future therapeutic options. Further experiments are needed to fully explore the functional implications of these findings; however, our preliminary results suggest that inhibition of GSTpi with clomipramine leads to inhibition of cell growth of TNBC cell lines in vitro.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2119.
Triple negative breast cancer: a phase 2, multi-center, open-label, randomized trial of gemcitabine/carboplatin (G/C), with or without BSI-201, a PARP inhibitor.
Authors: O'Shaughnessy J , Yoffe M , Osborne C , Blum J , Rocha C , Ossovskaya V , Sherman B , Bradley C
Published: December 2010

Abstract #2120

Background: PARP is a critical enzyme of cell proliferation and DNA repair and BSI-201 has been shown to be a potent inhibitor of PARP-1 in humans. Triple negative breast cancer (TNBC) shares important features with BRCA1-related breast cancer, a validated target for PARP inhibition. Our studies demonstrate that PARP-1 gene expression is statistically significantly upregulated in TNBC compared with normal breast tissue. The primary objective of this study is to assess the Clinical Benefit Rate (CBR=CR+PR+SD > 6 months) of Gemcitabine/Carboplatin with or without BSI-201 in patients with TNBC.  Methods: Eligible subjects are ≥ 18 years old who had received ≤ 2 prior chemotherapies for metastatic disease with histologically documented breast cancer that is ER-negative, PR-negative, and HER2-negative. Patients were randomized (1:1 ratio) to one of 2 study arms: a) arm 1: G/C alone; b) arm 2: BSI-201 + G/C. G/C was given on days 1 and 8; G = 1000 mg/m2, C at an AUC=2. In study arm 2, BSI-201 was administered I.V. twice weekly (days 1, 4, 8 and 11) at a dose of 5.6 mg/kg. Cycles were defined as being 21 days in duration. Modified RECIST criteria are used to assess tumor response every 6 weeks (every 2 cycles). Subjects who have progressive disease may crossover to receive BSI-201 and continue G/C. Archived tissue samples were retrieved for the assay of cancer related genes including PARP-1 by multiplex quantitative RT-PCR.  Results: To date, 50 subjects have been enrolled and treated for up to 8 cycles of therapy. Overall 39 of 50 (78%) subjects experienced at least 1 adverse event. The frequency and nature of the AE's do not differ between the two treatment groups. Gene expression profiling results from the first 28 patients enrolled confirm that the cancers of patients enrolled have uniformly low hormone receptor expression, variable HER2 receptor expression and significant upregulation of PARP-1 compared with normal breast tissue.  Conclusions: This is the first randomized study of a PARP inhibitor in cancer patients. BSI-201 is safe and well tolerated when given in combination with G/C and adverse events observed were consistent with the known safety profiles of G / C regimens. An updated evaluation of toxicity profile and PARP-1 expression analysis will be presented.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2120.
Combination or sequential single agent for the treatment of metastatic breast cancer (MBC) patients (pts). Impact of further chemotherapy (CT) in overall survival (OS) in the Alamo registry.
Authors: Lluch A , Ruiz A , Martín M , Alba E , Pastor M , de la Haba J , Llombart A , Ramos M , Martínez del Prado P , Escudero M
Published: December 2010

Abstract #2121

Introduction: Combinations can improve OS in MBC; however sequential single agent is sometimes preferred. In a recent report, pts receiving paclitaxel monotherapy in first line had a significantly worse OS if they were not receiving post study CT (PSC); no difference was seen in pts receiving paclitaxel+gemcitabine (Llombart, EBCC 2008). In this trial only 58% of pts received PSC.  Purpose: To assess, outside the context of a clinical trial, the amount of MBC pts receiving CT beyond first line, as well as the impact of combination use and further CT lines in their OS.  Methods: Alamo 1-2 is a breast cancer patient registry run by the GEICAM. 15482 pts diagnosed from 1990-1997 in 54 sites were included in the database. 4668 pts were stage IV; 778 (16.7%) metastatic at diagnosis and 3890 (83.3%) have had a recurrence.  Results: 3045 (65%) pts received CT in first line, 83% were combinations (with anthracyclines 42%, CMF 16%, taxane+anthracyclines 9.4% or +other agents 8%) and 16% were monotherapy. Among other variables studied, only previous treatment (in early stage) was influencing the choice of a combination (pts without previous CT received more combinations than pts receiving CMF, than pts receiving anthracyclines). Median survival for pts receiving single agent was significantly shorter compared to pts receiving combination, 16.2 and 21.85 months (m) respectively, HR=1.37 (IC 95%: 1.21-1.55; p<0.0001). Several covariates showed significant prognostic value in survival in the Cox multivariate model: monotherapy treatment in first line, anthracyclines pre-exposure, age > 65, negative hormonal receptor status, hepatic disease, Grade 3 and ≥ 3 disease sites. Half of the pts never received further CT after first line treatment. Only age and number of disease sites were influencing this decision in the Cox multivariate model. Median survival was 24.9 m in pts receiving further CT and 14.5 m in the ones not receiving it. In pts not receiving further CT, median OS was significantly shorter if they were treated with single agent in first line in comparison to those receiving previous combination: HR= 1.59 (IC 95%: 1.33-1.89; p< 0,00001). This difference was not significant in pts receiving further CT: HR= 1.15 (IC 95%: 0.97-1.37; p=0.101).  Conclusion: Our data show that only half of the MBC pts receive further CT after first line; they have longer survival. We found significant evidence that further CT is impacting the OS in pts treated with single agent in first line, but not in those receiving previous combination. Those facts should be taken into consideration when selecting single agent or a combination in first line.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2121.
Potential utility of angiotensin II receptor blockers in the treatment of estrogen receptor positive infiltrating ductal carcinoma.
Authors: Jin S , Coopersmith R , Byrnes J , White D
Published: December 2010

Abstract #2125

Despite significant advances in diagnosis and treatment, breast cancer remains the world's leading cause of cancer mortality in women. In this study we employed differential expression analysis tools to mine the BioExpress®(1) database of transcriptional profiles and found that Angiogtensin II receptor 1 (AT1-R) mRNA expression was highly correlated with expression of estrogen receptor alpha (ERα) in biopsies from patients with infiltrating ductal carcinoma (IDC). Specifically, AT1-R transcript was up-regulated 12.5 fold in ER+ primary IDC relative to ER- primary IDC (P = 0.0001).  Experiments using ER+ (T47D) and ER– (HCC1143) breast cancer cell lines derived from human mammary gland ductal carcinoma demonstrated that only the ER+ line responded to angiotensin II-mediated growth stimulation, and this effect was dramatically suppressed in a concentration dependent manner by AT1-R blockers (ARBs). Additional cell culture experiments evaluating eight cell lines derived from IDC further validated the concept that co-expression of both ERα and AT1-R was required for responsiveness to angiotensin II-induced cell proliferation. We have also found that ARBs significantly enhanced the anti-proliferative activity of both tamoxifen, an ER modulator, and formestane, an aromatase inhibitor. Most importantly, we have shown in a T47D tumor xenograft model that ARBs, candesartan and irbesartan, dose-dependently inhibited tumor progression in NODScid mice. Currently, we are evaluating the synergistic/additive effects of ARBs with tamoxifen in additional T47D xenograft models.  In conclusion, our data provides evidence that ARBs may have utility in the prevention and treatment of ER+ IDC. These observations further validate reports that ARBs may suppress tumor cell proliferation, angiogenesis, and cancer progression. To our knowledge our findings represent the first example of a potential means to stratify a patient population for ARB-based tumor therapy using a clinically accepted biomarker, findings which may have significant clinical implications for the treatment of IDC.  1) BioExpress® is a knowledgebase of human disease biology comprising over 20,000 individual patient samples profiled over the entire human transcriptome representing > 400 disease states including approximately 200 human breast cancer samples (169 diagnosed as IDC) and 68 normal breast tissue controls.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2125.
Doxorubicin pharmacokinetics following two different liposomal formulations (CAELYX™ vs MYOCET™) in advanced breast cancer patients.
Authors: Schueller J , Czejka M , Krexner E , Aigner K , Gruenberger B
Published: December 2010

Abstract #2128

Background: Liposomal encapsulation of Doxorubicin (DOX) was designed to minimize healthy tissue distribution by altering pharmacokinetics (PK) thus reducing cardiotoxicity while preserving antitumour efficacy. Study objective was to compare the PK behaviour of the two main liposomal formulations of DOX, i.e. pegylated (Caelyx, C) versus non pegylated (Myocet, M), in a single center study including 17 MBC patients, all Anthracyclin pretreated with additional risk faktors.  Methods: Cohort 1 consisted of 10pts (mean age 62, 49-77) treated by C as 1HR infusion of 50mg/m²/4 wks. Plasma samples were collected at timepoints min 0, 30, 60, 120, 180 and d1, 7, 14, 28. In cohort 2, 7 pts (mean age 58, 51 – 70) received M 75mg/m² 1H Infusion / 3wks, plasma levels determined at min 0, 30, 60, 90, 120, 180 and d1, 2, 3. Total amount of DOX was determined following the 1st infusion respectively, analytic procedure included solid phase extraction quantification by HPLC and PK analysis by Win Nonlin Pro. Doxorubicinol was detected after M only.  Results: in Cohort1 after C, mean Cmax of 6.8 µg/ml surprisingly occured at a mean tmax of 2.3 d, log conc time curve showed linear decrease of DOX conc suggesting continuous release into the tissue (mean conc d 7 3.3, d 14 1.0, d 21 0.3, d 28 0.09 µg/ml) . No metabolites were found, AUC (does not represent reliable drug exposure and bioavailability of DOX) expressed as µg/ml.d was 54.6. In contrast AUClast of M was calculated as 35µg ml.H and mean Cmax by 7.5 µg/ml at infusion end. DOXol was measured in all M treated patients at a slower metabolisation rate and late appearance compared to conventional DOX. t/2 el of C was 6 times longer than that of M (HR 90 vs 15). Differences in plasma disposition and PK of M vs C are depicted in table 1 and compared to pooled data of conventional DOX derived from former studies (50 mg/m² 30min inf, n=40). Cltot of conv. DOX was 10 fold higher than M and 500 fold higher than C, Vss about 12 times that of M and 70 times that of C. These PK differences underlines a different toxicity profile between PEGL (C, PPE syndrome) and NPEGL (M, myelotox, nausea), both reducing cardiac tox by avoiding peak plasma levels due to prolonged circulation time.   

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2128.
The thiazolidinedione peroxisome proliferator-activated receptor gamma (PPARγ) agonist troglitazone alters histone post-translational modifications in MCF7 breast cancer cells.
Authors: Davies GF , Ross AR , Juurlink BH , Troy HA
Published: December 2010

Abstract #2140

Troglitazone (TRG), a thiazolidinedione (TZD) PPARγ ligand, is a potent antiproliferative agent in many different cancer cell lines. We have previously demonstrated in multiple drug resistant (MDR) human K562 leukemia cells that TRG, in combination with the anthracycline antibiotic Doxorubicin (DOX), is an effective anticancer agent in MDR cells. The killing of drug resistant leukemia cells was accompanied by increased global histone H3 acetylation. In this report, we investigated the effects of TRG on both histone metabolism and killing of human MCF7 breast cancer cells. MCF7 cells were treated with TRG, as well as additional TZD's, such as Rosiglitazone, Ciglitazone and Pioglitazone. In all cases, a dose-dependent increase in H3K9 acetylation, total H2AX protein levels, and cell killing were noted. Striking similarities in histone modification profiles were also noted between TRG and histone deacetylase inhibitor (HDACi) treatment. A wide spectrum of common histone post-translational modifications (PTMs) were specifically induced by TRG and HDACi treatment, as determined by mass spectrometry.    These findings suggested that TRG may possess HDACi activity. In support of this, in vitro HDACi activity assays showed TRG, but not other tested TZDs, possessed HDACi-like activity. Global H3K9 acetylation was also increased by inhibition of the PI3K/AKT pathway (but not the MEK/ERK pathway), raising the possibility that TRG and HDACi's may act via inhibition of AKT. Consistent with this idea, MCF7 cells treated with TRG or HDACi's exhibit reduced phosphorylated AKT. Our data supports a mechanistic model linking TRG with inhibition of histone de-acetylation in human breast cancer cells.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2140.
Development of a targeted topoisomerase I inhibitor for anticancer therapy.
Authors: Cao YZ , Yang J
Published: December 2010

Abstract #2146

Background:  DNA topoisomerase I (Top1) has been identified as the molecular target of a variety of drugs, due to the important roles that they play in biological systems. Studies show that breast cancer cell lines are sensitive to Top1 agents such as camptothecin. Recent clinical trials have suggested that breast cancer patients may respond to drugs which target Top1. This suggests that some breast cancers may have elevated expression of the enzyme.  Thus, development of Top1 inhibitors may be a good target for anticancer therapy. To better target cancer cells, the drugs can be linked to a pH-sensitive linker such as the N-Ethoxybenzylimidazoles (NEBIs) developed in our lab. The NEBIs are designed to undergo hydrolysis in slightly acidic aqueous solutions at physiological temperature, comparable to the environment found in endosomes, lysosomes, and the extracellular environment of tumors. The hydrolysis of the NEBI causes the dissociation of the drug from a carrier (e.g. Herceptin), and the rates can be tuned by modifying the electron withdrawing and donating groups on the NEBI ring. It was demonstrated that the rates can be tuned to give half-lives ranging from 36 minutes to 9 months.  Materials and Methods:  We have developed a new class of imidazole-containing drugs which may inhibit Top1 activity. These compounds have a 4-ring, indenoisoquinoline structure and were synthesized by a simple condensation reaction which gives good yield. Ring substituents were modified to achieve molecules of optimal potency. The resulting drugs were subsequently linked to the NEBI, and hydrolysis studies were performed to determine the rate of active drug release.    Cytotoxicity of cleaved drugs was screened using the sulforhodamine B (SRB) assay and topoisomerase inhibition assays will be performed in the near future.  Results:  Cytotoxicity studies showed that the hydrolyzed drugs are toxic to the 10μM range in cancer cells. One of the compounds has recently been approved for the National Cancer Institute screen, in which the activity will be evalutated with 60 different cancer cell lines of diverse tumor origins. Ongoing studies will assess the topoisomerase I inhibition activity of these compounds.  Discussion:  We have developed a novel prodrug which comprises of a toxic Top1 inhibiting drug attached to a pH-sensitive linker. This may prove to be an effective approach to anticancer therapy. Future studies will be pursued in using folic acid as the drug carrier to target breast cancer cells. Studies show that folate receptor overexpression is associated with poor outcome in women with breast cancer. Furthermore, it has been shown that folate receptors may undergo endocytosis, which will trigger the release of the active drug from the linker.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2146.
The safety of dose-dense liposomal-encapsulated doxorubicin in association with docetaxel (MyTax) in breast cancer.
Authors: Mancini M , Cannita K , Santomaggio A , Tudini M , De Galitiis F , Morelli M , Rispoli A , Martella F , Porzio G , Pelliccione M , Cocciolone V , Lanfiuti Baldi P , Penco M , Romano S , Fratini S , Stifani G , Marchetti P , Ficorella C , Ricevuto E
Published: December 2010

Abstract #2157

Background: Liposomal-Encapsulated Doxorubicin (LED) shows equivalent efficacy, better cardiac tolerability at higher cumulative dose than conventional anthracyclines in breast cancer treatment.  Methods: Sixteen pts were enrolled in a dose-finding study of LED (TLC-D99 Myocet ®) associated to Docetaxel (TXT). Twelve pts were treated with a fixed TXT dose (50 mg/m2) and TLC-D99 at three dose levels, 40-45-50 mg/m2, days 1 and 15 every 2 weeks using an intra- and inter-patient approach; four pts wrere treated at the TLC-D99 recommended dose (50 mg/m2). Cardiac monitoring of LVEF was performed every two cycles; Precursor Brain Natriuretic Peptide (proBNP) and cardiac Troponin (c-TnI) before and after 24 h chemotherapy was evaluated.  Results: Breast cancer (BC) disease extension: metastatic (MBC), 8; locally advanced BC, 5; T2-T3 BC, 3. Previous chemotherapy: untreated, 11 pts; adjuvant, 5 pts. Enrolled pts for each dose-level: I, 7; II, 9; III, 14. Newly treated pts: I dose-level, 7; II dose-level, 3; III dose-level, 6. Valuable cycles for each dose-level in a total 77 cycles: I, 14; II, 21; III, 42. DLTs were observed in 3 pts, 21%, and 3 cycles, 4%: 2 cardiac, characterized by a 19% LVEF decrease and a symptomatic arrhythmia; one G4 hematologic resistant to G-CSF. DLTs for each dose-level by pts and cycles, respectively: I, 14% (1/7 pts) and 7% (1/14 cycles); II, no DLT in 9 pts and 21 cycles; III, 14% (2/14 pts) and 5% (2/42 cycles). Cumulative G3-4 toxicities by pts and cycles, respectively: cardiac arrhythmia 6% and 1,3%, cardiac general (symptomatic LVEF decrease), 6% and 1,3%; alopecia 81% and 65%; neutropenia resistant to G-CSF, 6% and 1,3%. Cardiac DLTs were observed in 2 elderly pts (>65 y). The 2 cardiac DLTs were observed in 2 out of 3 pts with pre-existing diastolic dysfunction. No pathologic increase of c-TnI levels was detected. Seven pts showed increased pro-BNP after chemotherapy; 1 of these with increased pro-BNP after chemotherapy, persistent the day 1 of each subsequent chemotherapy showed a DLT;  G2 toxicities by patients and cycles, respectively: asthenia 37% and 18%, stomatitis/mucositis 12% and 5%, nausea 31% and 12%. Median rDI of TLC-D99 was 25 mg/m2/w and TXT 25 mg/m2/w for pts, respectively. Preliminary efficacy in 16 assessable pts: LA-BC and MBC, 1 CR (pCR) 7 PR (OR 62%), 4 SD and 1 PD; T2-T3 BC, 2 PR and 1 SD.  Conclusion: dose-dense TLC-D99/Docetaxel association can be safely recommended at the dose of 50 mg/m2 for each drug. Docetaxel intensification is ongoing.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2157.
Aberrent hedgehog signaling is an early event in breast cancer development.
Authors: O'Toole S , Swarbrick A , Millar E , McLeod D , McNeil C , Qiu M , Lopez-Knowles E , Caldon E , Oakes S , Ormandy C , Morey A , Musgrove E , Henshall S , Sutherland R
Published: December 2010

Abstract #24

Background: The Hedgehog (Hh) signaling pathway is a highly conserved system which regulates development and cell fate. It also contributes to the pathogenesis of a number of human malignancies, including basal cell, prostate and pancreatic carcinoma as well as medulloblastoma. There is emerging evidence that Hh may play an important role in breast cancer.  Methods: We examined the immunohistochemical expression of key Hh proteins, sonic hedgehog (Shh), Patched (Ptch) and Gli1 in 3 cohorts of patients; 2 independent cohorts of the histological progression model of breast cancer and a cohort of 292 patients with a median follow up of 5.5 years. We also investigated expression of Shh in the C3/SV40T mouse model of mammary carcinogenesis. We then investigated the effects of recombinant Shh in the MCF10A in vitro model of early breast cancer development. Finally, a mammary reconstitution graft model was employed where donor mammary epithelial cells were briefly cultured and transduced with a retroviral construct carrying a Shh-GFP construct, then transplanted into the cleared mammary fat pad of a recipient mouse.  Results: Overexpression of Shh was an early event in human premalignant lesions, with progressive increases (p<0.05) with the development of hyperplasia, atypia and malignancy in both test and validation progression cohorts. Similar results were observed in the mammary gland lesions of the C3/SV40T mouse. In human breast cancer, overexpression of Shh was associated with a poor prognosis (recurrence HR=2.0, p=0.003 and breast cancer specific death (HR=2.4, p=0.002) and also associated with a poor prognosis in chemotherapy (HR=2.4, p=0.03) and endocrine therapy (HR 3.5, p=0.03) treated patients. There was a highly significant (p=0.004) association by chi squared analysis with the basal-like subtype of breast cancer. Based on these data, we investigated the effects of Shh in 2 experimental systems – Shh caused disorganization of MCF-10A acini and was associated with the development of an unusual discohesive phenotype. In the mammary graft model, expression of Shh was associated with ductal epithelial hyperplasia in 58% of ducts, compared to 5% of controls.  Conclusion: These experimental data suggest that excessive Shh may cause hyperplasia and dysplasia of normal human and murine mammary gland epithelium. Together with our observations in human breast preinvasive and malignant lesions, these results support a role for aberrant Hh signaling in breast carcinogenesis.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 24.
A UK NEQAS ring study evaluating observer variation in the diagnosis of HER2 amplification using the Kreatech™ HER2 FISH probe.
Authors: Bartlett JM , Campbell FM , Ibrahim M , Thomas J , Wenyck P , Ellis IO , Kay E , Connolly Y , O'Grady A , Cunningham P , Barnett S , Starczynski J , Miller K
Published: December 2010

Abstract #3005

Background: Whilst HER2 testing is well established as a key requirement for all newly diagnosed breast cancers the quality and consistency of HER2 testing remains a point of controversy. Most countries use FISH or ISH based testing for HER2 gene amplification in equivocal (IHC 2+) cases. Data on reproducibility and portability of novel and even established HER2 methods is sparse. We report on a multicentre evaluation of intra and inter-observer/site variation in scoring HER2 results from 45 breast cancers measured in 8 laboratories using the Kreatech™ HER2 FISH probe.  Methods: A commercial tissue micro-array (TMA; Stretton Scientific UK) containing 2 replicate cores from 45 breast cancers was circulated to 8 laboratories with varying experience in FISH methodology. Five laboratories (1-5) were UK NEQAS reference labs, 3 further experienced routine diagnostic labs (A-C) participated in the scheme designed to test the site to site reproducibility of the Kreatech™ FISH probe. All laboratories FISHed a 90 core TMA and 7/8 reported results for all assessable cores, 1 lab reported 1 result per evaluable cases (as per the original study design). Results were collated centrally and analyzed for intra and inter-observer between both “reference” laboratories (labs 1-5) and non-reference laboratories (A-C).  Results: Overall 81% of cores (range 70-94%) were successfully analysed by Kreatech™ FISH. Intra-observer variation was lower than expected at 4.7% (range 3.5-8.4%) for HER2 ratio. Mean intersite variation in HER2:Chr 17 ratio was significantly lower (8.5%) for reference vs non-reference (13.4%) laboratories (p=0.0013). Of 546 reported cores only 7 (1.3%) were discordant (with 4 borderline). Analysis of site to site variation highlighted possible systematic differences in scoring approaches in 3/8 laboratories including a “reference laboratory”.  Conclusions: Intra- and inter-observer variation of absolute HER2 ratios appears to be tightly controlled across 8 laboratories using Kreatech™ HER2 FISH probes. Detailed comparisons of scoring results suggests some systematic differences between a few laboratories in the interpretation of FISH. However the majority of laboratories show extremely tight reproducibility of numeric HER2 FISH results. This study highlights the potential for extremely robust and quantitatively reproducible FISH results from both “diagnostic” and “reference” laboratories. However it also suggests continued quality assessments are essential to good performance.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3005.
“Polysomy 17” by FISH for HER2 in breast cancer is a misnomer: results of chromosome 17 analysis by array comparative genomic hybridization.
Authors: Yeh I , Martin MA , Robetorye RS , Gunn S
Published: December 2010

Abstract #3008

Background: The term “polysomy 17” has been widely used to refer to breast cancer cases in which fluorescence in situ hybridization (FISH) assays for HER2 were performed and found to contain >3 copies of the chromosome 17 centromere (probe CEP 17, D17Z1). The proportion of breast cancers that are reported to have polysomy 17 varies, but some studies report over 40% incidence. In these cases, the centromere is assumed to be representative of the entire chromosome. In this study, we analyzed chromosome 17 in breast cancers using bacterial artificial chromosome (BAC) array comparative genomic hybridization (array CGH) to assess the incidence of whole chromosome 17 polysomy.  Design: Fresh frozen tissue from 95 breast cancers was analyzed, including 84 invasive ductal carcinomas, 10 invasive lobular carcinomas, and one case with mixed ductal-lobular features. Frozen sections were reviewed to ascertain tumor content, and genomic DNA was isolated from 25-50 mg of tissue from each case. Samples were then analyzed by array CGH using an array that included whole genome coverage as well 63 probes for chromosome 17. A subset of 29 cases was also analyzed by FISH using the Vysis Pathvysion HER2 assay, which includes a chromosome 17 centromere probe and a HER2 probe.  Results: Multiple abnormalities were identified on chromosome 17, including losses and gains in both the p and q arms. Examination of the centromeric region by array CGH showed 6 cases with gains, and 10 cases with losses. Three of six cases with centromeric gains also showed HER2 amplifications. Two cases showed polysomy by FISH. Although other chromosomes in our study exhibited whole chromosome polysomy using array CGH, no cases showed complete polysomy of chromosome 17.  Conclusions: A review of the literature for chromosome 17 polysomy in breast cancer reveals that this finding almost exclusively relies on FISH analysis using chromosome 17 centromere probes. Our analysis of 95 breast cases by array CGH did not reveal any gains of whole chromosome 17. These results suggest that whole chromosome 17 polysomy is extremely uncommon in breast cancer. Those cases that are identified to have chromosome 17 polysomy by FISH are better termed “chromosome 17 centromere amplified.”

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3008.
Comparison of fluorescence in situ hybridization and dual color chromogenic in situ hybridization for the assessment of HER2 status on core-needle breast cancer biopsies.
Authors: Suárez-Gauthier A , García-García E , Conde E , Muñoz A , Torres M , Miró C , García-Estevez L , López-Ríos F
Published: December 2010

Abstract #3015

Background: Core-needle biopsies have been shown to be extremely useful for evaluation of prognostic and predictive markers. In breast cancer patients, this is particularly important in the setting of neoadjuvant chemotherapy. However, it is well known that immunohistochemistry can give conflictive results on small biopsies or core-needle biopsies. This study evaluates the concordance between HER2 gene amplification determined by fluorescence in situ hybridization (FISH) and a new dual color cromogenic in situ hybridization (CISH) in a series of core-needle breast biopsies.  Material and Methods: A total of 37 invasive breast carcinomas diagnosed on core-nedle breast biopsies were selected from our files. Dual color FISH (Dako, Glostrup) and dual color CISH (Dako, Glostrup) were performed in each case. Scoring of the FISH and CISH slides was identical, counting HER2 and CEN-17 signals from 60 tumour nuclei per case. Finally, the gene to CEN-17 ratio was calculated using the cut-off value of HER2/CEN-17 ratio >2 as amplified.  Results: All 37 specimens were analyzed successfully by CISH and FISH. A high concordance was found between FISH and CISH in the assessment of HER2 status. 12 cases were amplified and counted easily with both techniques, showing similar ratios. Polysomy was also detected with both techniques in 5 cases with no amplification.  Discussion: Given the presumed heterogeneous expression of HER2 immunohistochemistry on core-needle biopsies, dual color CISH might be an alternative to FISH in this setting. Moreover, CISH allows for a better concurrent analysis of morphology, which is particularly important when studying small samples.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3015.
Notch and breast cancer: effects of a γ-secretase inhibitor on cell cycle.
Authors: Luraguiz NS , Yong TH , Sun AW
Published: December 2010

Abstract #3016

Background:  Notch ligands and receptors are type I membrane proteins that regulate cell fate during cell-cell contact. Notch activation is oncogenic in many instances. Within the mammary gland, aberrant activation of the Notch pathway leads to adenocarcinoma. In human, high level of Notch1 expression in breast tumor is associated with reduced patient survival while better survival was observed where tumors expressed higher level of Notch2. In a recent report, it was observed that accumulation of Notch1 NICD increase Notch signaling in a wide variety of human breast carcinomas. Breast cancer is one of the most common malignancies in women with a cumulative lifetime risk of developing the disease as high as one in every eight women. Current research focused on treatment that disrupt the molecular mechanisms leading to breast carcinogenesis, which may be a more effective in treating breast cancer. GSI (g-secretase inhibitor) which blocks the activation of Notch is presently undergoing Phase I clinical trial conducted by Merck in breast cancer patients. We have investigated the different effects of GSI treatments on ER- and ER+ breast cancer cells and the results are discussed.  Material and Methods:  The human breast cancer cells lines MCF-7 and T47D (ER+), MDA-MB-435S and MDA-MB-231 (ER-) were treated with GSI, a Notch inhibitor, Tamoxifen, a selective estrogen receptor modulator (SERM) and combination of both drugs. Cell proliferation was determined by BrDU ELISA assay (Roche). Apoptosis following treatment was measured with TUNEL Assay Kit (Roche). The Cellular DNA Flow Cytometric Analysis Kit was used to determine the treatment on cell cycle. Trizol Reagent was use to extracted the RNA and Real Time PCR and RT Profiler PCR Array System were perform to detected the expression of genes.  Results and Discussion:  GSI alone is very effective on inhibiting the proliferation and promoting apoptosis of ER- cells but not ER+ cells. When combined with Tamoxifen, however, a significant killing effect was seen in ER+ cells. Cell cycle analysis revealed S phase block. The fold repression of Hes1 and Bcl2 was confirm after treatment with GSI in both cell lines. Together, these data strengthen the rationale for applying Notch inhibitors therapeutically. It also indicates the differences of the GSI treatment on ER+ vs ER- cells. In ER+ cells, both ER and Notch signaling pathways need to be targeted to effectively inhibit tumor cell growth and induce apoptosis. In ER- cells, targeting Notch signaling pathway is sufficient in achieving the same effects. The information will be critical when translating such research into clinic on deciding treatments for the breast cancer patients.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3016.
Kruppel-like transcription factor 5 promotes breast cell survival through pERK-mediated MKP-1 protein stabilization.
Authors: Liu R , Zhou Z , Chen C
Published: December 2010

Abstract #3019

Kruppel-like Factor 5 (KLF5) is a transcription factor promoting cell survival and tumorigenesis in multiple cancers. High level of KLF5 mRNA is associated with a shorter survival for breast cancer patients. However, the role of KLF5 and mechanism of KLF5 actions in breast cancer remain unclear. In this study, we found that KLF5 knockdown by siRNA in two basal type breast cell lines MCF10A and BT20 induces apoptosis, as indicated by loss of cell viability and cleaved PARP and caspase 3. Interestingly, a survival phosphotase, MKP-1, is downregulated at protein level by KLF5 ablation. Consistently, KLF5 over-expression increases the MKP-1 protein expression. However, MKP-1 is not a KLF5 direct target gene because the MKP-1 mRNA level is not regulated by KLF5. By the cycloheximide chase assays, we found that KLF5 decreases the MKP-1 protein degradation. However, KLF5 does not decrease the MKP-1 E3 ubiquitin ligase SKP2 expression. The ERK inhibitor U0126 specifically blocks the KLF5 induced MKP-1, suggesting that KLF5 up-regulates MKP-1 through activating ERK signaling. Finally, we demonstrated that MKP-1 over-expression blocks KLF5 knockdown induced apoptosis in MCF10A cells. These findings suggest that KLF5 is a survival factor which promotes breast cell survival partially through pERK-mediated MKP-1 stabilization. The KLF5-pERK-MKP-1 signaling axis may provide new therapeutic targets for invasive breast cancer.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3019.
Activated c-Src (Y215) kinase expression predicts for early relapse on tamoxifen in human breast cancer.
Authors: Tovey SM , Tan BA , Campbell J , Elsberger B , Brunton V , Mallon E , Cooke TC , Edwards J
Published: December 2010

Abstract #3025

Recent studies have demonstrated activation of the non-receptor tyrosine kinase c-Src in tamoxifen resistant breast cancer cell lines, which when treated with Src inhibitors demonstrate a reduction in motility and invasiveness. However, there are little published translational studies confirming a role for c-Src in tamoxifen resistance in vivo. We aim to determine if activated c-Src expression correlates clinically with increased recurrence on tamoxifen.  Methods  Tissue microarrays were constructed from a patient cohort of 402 Tamoxifen treated ER positive patients. Immunohistochemistry was performed using commercially available antibodies to Total Src as well as to activated (phosphorylated) c-Src at 2 sites (Y416 and Y215) (pSrc416 and pSrc215 respectively). Expression was assessed by 2 independent scorers (weighted histoscore method). HER1-3 status and expression of membranous phosphorylated ER (sites serine 118 and 167) had previously been determined. All statistical calculations were performed using SPSS 15, including Kaplan-Meier life table analysis with log rank testing of differences in breast cancer related relapse whilst on tamoxifen.  Results  Membranous pSrc215 was observed in 20.3% of the cases but was more frequently observed in the cytoplasm (85.9%) and nucleus (90.5%). Overexpression of cytoplasmic pSrc215 was associated with an earlier time to recurrence (p = 0.037).    Cox regression analysis confirmed this to be independent of grade, nodal and HER1-3 status (p=0.028, HR 3.17 (1.14–8.85)). Cytoplasmic pSrc215 was also strongly correlated with membranous phospho-ER, EGFR and HER3 status (Mann Whitney; p<0.001, p=0.026, p<0.001 respectively). Membranous and nuclear pSrc215 staining were not associated with time to relapse.  Over expression of Total Src and pSrc416 did not predict for relapse.  Discussion  Our findings provide support to in vitro studies linking c-Src with tamoxifen resistance. The phosphorylation site pSrc215 is a putative site of activation by HER2 and PGFR resulting in up to 50 fold activation of Src in vitro. The correlations demonstrated with HER status and membranous ER lead us to speculate that this interplay between non genomic ER and Src activation may provide a mechanism for the lack of response to tamoxifen in these patients.  Further studies are required to determine if activated pSrc215 represents activated c-Src alone or may also represent activation of other Src family members as the Y416 and Y215 sequences are highly conserved amongst the Src kinases with homologous activation sites. If confirmed, future use of novel combination therapies with Src inhibitors may have an important role in combating tamoxifen resistance.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3025.