bluuebunny/crossref_metadata_2025 huggingface dataset:
Capturing Changes in the Brain Microenvironment during Initial Tumor Cell Invasion.
Authors: Lorger M. , Felding-Habermann B.
Published: December 2010

Brain metastases occur mostly in patients with melanoma or primary tumors in the lung, breast or prostate. As therapies against primary tumors and extracranial metastases are improving and patients live longer, the incidence of brain metastases is increasing and brain lesions are seen in 20 to 40 % of patients with advanced metastatic disease. Unfortunately, no therapies are available to treat brain metastasis efficiently.Existing in vivo models of brain metastasis focus mainly on established macrometastases, while very little is known about the initial events of tumor cell brain colonization. In the present study, we used non-invasive bioluminescence imaging and detailed immunohistochemistry to capture the early events of tumor cell interaction with the cerebral microvasculature and invasion of the brain tissue. We used MDA-MB-435 metastatic cancer cells as a model because of their aggressive metastatic phenotype, and injected the cells into the internal carotid artery of immune deficient mice. Our results show that single cancer cells extravasate into the brain parenchyma 3 to 7 days after they arrest within brain capillaries and disrupt the vessel wall, bursting the microvasculature. Cancer cell arrest and extravasation occur exclusively in small vessels that are surrounded by pericytes and lack smooth-muscle cells. Early brain colonization consistently induces local activation of astrocytes, involving heterogeneous astrocyte populations. Reactive astrocytes express high levels of MMP-9 that may assist tumor cell invasion and provide first pro-angiogenic clues. Activated astrocytes consistently persist in the vicinity of growing metastases, and potentially contribute to metastatic proliferation. Occasionally, early host responses also include microglia with stellate or a reactive amoeboid morphology. Importantly, in addition to yielding actively proliferating macrometastases, tumor cell extravasation also leads to widespread single tumor cells dispersed throughout the brain tissue. The majority of these cells remains viable but stays quiescent and does not proliferate. We speculate that the diversity in early local responses of the brain microenvironment to initial cancer cell invasion contributes to the fate of extravasated tumor cells. An early involvement of diverse host cell types might provide distinct environmental clues and co-determine whether a single cell progresses to macrometastasis or remains dormant.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 4162.
Analysis of Primary Breast Tumour Stromal Cells and Their Potential Role in Disease Progression.
Authors: Hartmann M. , Dwyer R. , Kerin M.
Published: December 2010

It has emerged in recent years that interaction between breast cancer stromal and epithelial cells supports tumour progression through promotion of epithelial cell growth, migration and invasion. Although this is thought to be mediated through both secretion of paracrine factors and cell-cell contact, the precise mechanisms are poorly understood. The aim of this study was to further characterise stromal cells derived from primary breast tumours and potentially elucidate mechanisms through which they exert their action.Following written informed consent, specimens of human breast cancer were harvested from patients undergoing surgery. Cells were isolated from tumour (n=19) and tumour-associated normal (TAN, n=9) regions of breast tissue. Breast tissue obtained from reduction mammoplasty served as normal controls. Following tissue dissociation and digestion, stromal cells were isolated by differential centrifugation. Following culture of stromal cells, RNA was extracted, reverse transcribed and relative quantitative PCR performed using primers targeting Fibroblast Activation Protein (FAP), Transforming Growth Factor β (TGFβ), Transforming Growth Factor β Receptor II (TGFβRII), Matrix Metalloproteinase 3 (MMP3), and Vascular Endothelial Growth Factor A (VEGF A).There was a trend towards increased expression of VEGF A, TGFβRII and MMP3 in tumour compared to normal stromal cells, while the level of TGFβ expression was equivalent in all samples examined. There was a significant positive correlation between expression of the proangiogenic factor VEGF A and TGFβRII (Pearson correlation coefficient r=0.317, p<0.05). There was also a significant positive correlation between expression of FAP and TGFβRII (r=0.404, p<0.05).Tumour stromal cells have the potential to stimulate angiogenesis through secretion of paracrine factors such as VEGF. This proangiogenic factor is a known target of the TGFb signalling pathway. The elevated level of TGFβRII observed in this study may support TGFβ mediated upregulation of VEGF and thus promote tumour angiogenesis. Further characterisation of stromal cells and their impact on epithelial cell genotype and phenotype is critical to identify targets to inhibit initiation of the metastatic cascade.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 4163.
Coexpression and Synergistic Role of Multiple Atypical Chemokine Binders in Breast Cancer.
Authors: Ou Z. , Zeng X. , Li J. , Tang L. , Gu X. , Shen Z. , Shao Z.
Published: December 2010

Background: The expression and role of Duffy antigen receptor for chemokine (DARC), D6 and ChemoCentryx chemokine receptor (CCX-CKR) in breast cancer have already been studied independently in the past several years. However, the relationship and importance of these atypical chemokine binders (ACB) are still unclear. In this large scale clinical study, we invastigated the characteristics of multiple ACB simultaneously. Material and Methods: The protein levels of DARC, D6 and CCX-CKR were analyzed by immunohistochemistry in 517 patients with invasive breast cancer, 29 patients with noninvasive breast cancer. All of these cases come from the Cancer Hospital of Fudan University during 2001-2006, with complete clinicopathological data and follow-up data, including survival information. Results: In the invasive breast cancer patients, triple, double and single positive expression of these ACB were 135 cases (26.1%), 154 cases (29.8%) and 121 cases (23.4%), respectively. On the other hand, 107 cases (20.7%) were complete negative. The triple positive staining was significantly conversely correlated with lymph node metastasis, disease stage, relapse-free survival (RFS) as well as overall survival (OS). DARC, D6, and CCX-CKR were independent factors that influenced lymph node metastasis and their coexpression was favorable prognostic factor for RFS and OS. Discussion: Our results confirmed that DARC, D6, and CCX-CKR play a negative role in breast cancer. Furthermore, their triple or double high coexpression is more favorable for the patients, suggesting that multiple ACB may provide better protection against metastasis and relapse by synergistically regulating chemokine network in breast cancer.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 4166.
TrkA Overexpression Enhances Growth and Metastasis of Breast Cancer Cells, and Ku86 Is Crucial for TrkA Overexpression-Induced Breast Cancer Cell Invasion.
Authors: Lagadec C. , Meignan S. , Tastet C. , Com E. , Hondermarck H. , LeBourhis X.
Published: December 2010

The Trk family of neurotrophin tyrosine kinase receptors is emerging as an important player in carcinogenic progression in non-neuronal tissues. Here, we show that breast tumors present high levels of TrkA and phospho-TrkA compared to normal breast tissues. To further evaluate the precise functions of TrkA overexpression in breast cancer development, we have performed a series of biological tests using breast cancer cells that stably overexpress TrkA. We show that (1) TrkA overexpression promoted cell growth, migration and invasion in vitro; (2) overexpression of TrkA per se conferred constitutive activation of its tyrosine kinase activity; (3) signal pathways including PI3K-Akt and ERK/p38 MAP kinases were activated by TrkA overexpression and were required for the maintenance of a more aggressive cellular phenotype; and (4) TrkA overexpression enhanced tumor growth, angiogenesis and metastasis of xenografted breast cancer cells in immunodeficient mice. Moreover, recovered metastatic cells from the lungs exhibited enhanced anoikis resistance that was abolished by the pharmacological inhibitor K252a, suggesting that TrkA-promoted breast tumor metastasis could be mediated at least in part by enhancing anoikis resistance. Together, these results provide the first direct evidence that TrkA overexpression enhances the tumorigenic properties of breast cancer cells and point to TrkA as a potential target in breast cancer therapy.In a second step, we used a proteomic-based approach to identify proteins involved in TrkA overexpression-stimulated invasion of MDA-MB-231 breast cancer cells. Proteins from control and TrkA overexpressing cells were separated using a cup-loading two-dimensional electrophoresis system before MALDI and LC-MS/MS mass spectrometry analysis. Among several putative modified proteins, Ku86 was found to be the major protein which was increased in TrkA overexpressing cells. Moreover, Ku86 was co-immunoprecipitated with TrkA and its level at the cell surface was increased in TrkA overexpressing cells. Interestingly, inhibition with small-interfering RNA and neutralizing antibodies showed that Ku86 was required for TrkA-stimulated cell invasion. Together, these data allowed the identification of Ku86 as a new player involved in metastasis of breast cancer cells.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 4169.
FGFR1 and FGF Coamplification in Breast Cancer.
Authors: Albiges L. , Albiges L. , Quidville V. , Valent A. , Mathieu M. , Drusch F. , Job B. , Koscielny S. , Delaloge S. , Andre F. , Andre F.
Published: December 2010

Background:FGFR1 is a tyrosine kinase receptor involved in cell proliferation, migration and differentiation. FGFR1 gene is amplified in about 10% of breast cancer. Several studies have previously reported that FGFR1 inhibition could lead to a decrease in cell proliferation. Recent data obtained in prostate cancer have shown that FGFR1 activation could mediate epithelial to mesenchymal transition. In the present study, we evaluated the rational to target FGFR1 in clinical setting.Material and methods:We first assessed whether FGFR1 amplification was associated with a specific breast cancer phenotype. We assessed FGFR1 amplification by CGH array on 235 samples and by FISH (Z-2072, Spec FGFR1/Cen8 dual color probe, Zytovision®) on 260 samples, not included in the CGH part. We then assessed the functional effects of FGFR1 inhibitions in vitro. The antiproliferative effect of E3810 (FGFR1 inhibitor) was assessed both on FGFR1-non amplified (T47D/MCF7) and FGFR1 amplified (SUM44PE/MDA134) cell lines. Effect of FGFR1 inhibition was assessed on gene expression profiling.Results:When assessed by FISH, FGFR1 amplification was observed in 23/260 cases. Most of the samples (62%) exhibited an ER+/PR- phenotype, suggesting activation of intracellular kinase. FGFR1 amplification was observed in 24 out of 235 samples (10%). Interestingly, FGFR1 amplification was associated with gene gain in FGF3/FGF4/FGF19 (11q13) (55% versus 22%, p=0.05). FGFR1 inhibition by E-3810 inhibits MAPk phosphorylation, cell proliferation. A differential IC 50 for cell proliferation was observed between amplified cell lines and control cell lines. Preliminary data on gene expression profiling suggest that FGFR1 inhibition on breast cancer cells could lead to significant decrease in the Sox9 expression, a protein involved in epithelial to mesenchymal transition.Conclusion:FGFR1-amplified breast cancers present a discrete phenotype, including a high rate of ER+/PR- and high rate of FGF3/4/19 gains. Pharmacologic inhibition of FGFR1 leads to cell proliferation decrease and modulation of genes involved in EMT. These data suggest that there is a rational to target FGFR1 in clinical setting in patients with FGFR1-amplified breast cancer.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 4170.
Quantitative Analysis of HER2 Receptors Expression In Vivo by Near-Infrared Optical Imaging.
Authors: Capala J. , Chernomordik V. , Hassan M. , Zielinski R. , Gandjbakhche A.
Published: December 2010

Purpose: HER2 overexpression has been associated with a poor prognosis and resistance to therapy in breast cancer patients. However, up to now, quantitative estimates of this important characteristic have been limited to ex vivo ELISA essays of tissue biopsies and/or PET-based analysis. We aim at developing a novel approach in optical imaging, involving specific probes that do not interfere with the binding of the therapeutic agents, thus, excluding competition between therapy and imaging. Affibody-based molecular probes seem to be ideal for in vivo analysis of HER2 receptors using near-infrared optical imaging. To achieve quantification of HER2 receptors and overcome experimental uncertainties, we analyze post-injection temporal variations of the fluorescence intensity in the tumor area.Experimental design: Quantitative in vivo near-infrared imaging uses HER2-specific albumin-binding domain-fused-(ZHER2:342)2 Affibody molecules, labeled with AlexaFluor750 dye, as targeting agents. Subcutaneous tumor xenografts, expressing different levels of HER2 (BT474, MDA-MB361, MCF7, U251), are imaged at different times post-injection. Mathematical modeling allowed assessment of correlation between measured characteristics and HER2 expression levels from ex-vivo assays of the same tumors.Results: Affibody-Alexa Fluor conjugates may be used as a specific near-infrared probe for the non-invasive imaging of HER2-positive tumors, while analysis of the imaging parameters, directly estimated from the sequence of optical images, provides quantitative insight on the HER2 overexpression, as suggested by observed correlation between tumor cell amplification/overexpression of HER2, and these parameters.Conclusions: Method might potentially be employed in clinic for noninvasive monitoring of the HER2 expression variation in response to therapeutic intervention.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5002.
The Role of FDG PET/CT in Patients with Locoregional Breast Cancer Recurrence and a Comparison to Conventional Imaging.
Authors: Aukema T. , Rutgers E. , Vogel W. , Teertstra H. , Oldenburg H. , Vrancken Peeters M. , Wesseling J. , Russell N. , Valdés Olmos R.
Published: December 2010

Background: The prognosis and clinical management of patients with locoregional recurrence of breast cancer depends on the extent of the disease. Conventional work-up, consisting of bone scintigraphy, liver ultrasound, X-ray or CT of the thorax and MRI of the thoracic wall, may underestimate the local extent or miss the presence of distant metastases. The aim of this study was to evaluate the impact of 18F-fluorodeoxyglucose positron emission tomography/computed tomography (FDG PET/CT) on clinical management in patients with locoregional breast cancer recurrence amenable for locoregional treatment and compare the results with conventional imaging.Material and Methods: We performed a retrospective analysis of all patients referred for FDG PET/CT with locoregional breast cancer recurrence between January 2006 and August 2008. PET/CT findings were compared with results of additional imaging, pathology and follow-up. The impact of FDG PET/CT findings on clinical management of patients, was evaluated on the basis of clinical decisions obtained from patient files.Results: 56 patients were included. In 93% of the patients FDG PET/CT confirmed the locoregional recurrence. In 32 patients (57%) FDG PET/CT revealed additional tumour localisations. Distant metastases were detected in 11 patients on conventional imaging and in 23 patients on FDG PET/CT images (p<0,01). In 25 patients (45%), FDG PET/CT detected additional lesions not visible on conventional imaging. FDG PET/CT had an impact on clinical management in 27 patients (48%) by detecting more extensive locoregional disease or distant metastases. In 20 patients (36%) unnecessary mutilating surgery was prevented and treatment was changed to palliative systemic treatment. The sensitivity, specificity, accuracy, positive and negative predictive values of FDG PET/CT were respectively 97%, 92%, 95%, 94% and 96%.Discussion: FDG PET/CT, in addition to conventional imaging, plays an important role in staging patients with locoregional breast cancer recurrence since its result changed the clinical management in almost half of the patients. FDG PET/CT could potentially replace conventional staging imaging in patients with a locoregional breast cancer recurrence.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5011.
Mammosphere Culture of Established Cell Lines Does Not Enrich for a More Tumorigenic Breast Cancer Stem Cell Population.
Authors: Iorns E. , Clarke J. , Dean S. , Lippman M.
Published: December 2010

Background:The cancer stem cell model suggests that tumors arise from and are maintained by a relatively small proportion of cells, termed cancer stem cells. According to this model, only these cells have the capacity for self-renewal and the ability to repopulate the tumor bulk by giving rise to the phenotypically heterogeneous populations of cells that comprise the primary tumor. Evidence to support the cancer stem cell model has been provided by isolation of subpopulations of cells on the basis of differential expression of cell surface markers, followed by functional transplantation into animal models. These studies reveal that only a small subset of CD44+/CD24-/low cancer cells retain the ability to form new tumors after transplantation in immunodeficient mice, consistent with the cancer stem cell model. These results indicate that it may be necessary to target and eliminate cancer stem cells in order to eradicate tumors.Material and Methods:Previous studies have suggested that only breast stem cells have the capacity to proliferate in suspension as spherical colonies termed mammospheres, and that mammosphere derived breast cancer cells are CD44+/CD24-/low and able to form tumors that reproduce the heterogeneity of the original primary tumor when injected into the mammary fat pad of immunodeficient mice. This indicates that mammospheres are comprised of breast cancer stem cells. To identify and characterize breast cancer stem cells more completely, the mammosphere culture system was used to isolate and propagate breast cancer stem cells from three basal breast cancer cell lines (BT549, MDA231, MDA436) and three luminal breast cancer cell lines (MCF7, SKBR3, T47D). The tumorigenicity of the cultured mammospheres and parental cell lines was examined using dilution studies in immunodeficient mice and gene expression profiles were studied by microarray analysis.Results and Discussion:No significant differences in tumorigenicity were observed between breast cancer cells cultured as mammospheres versus standard culture conditions. Furthermore, no consistent differences in gene expression profiles were observed between mammosphere and parental cells across the panel of cancer cell lines. These results suggest that mammosphere culture is not capable of significantly enriching for a more tumorigenic breast cancer stem cell population from established breast cancer cell lines. This may indicate that cancer subpopulations that exist in primary tumors are lost during the long term culture of breast cancer cell lines and that these systems may not be a suitable model for breast cancer stem cell studies.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 502.
Pathological and Patient Factors Affecting the Accuracy of Ultrasound Combined with Fine Needle Aspiration Cytology in Pre-Operative Staging of the Axilla in Breast Cancer.
Authors: Hogan B. , Peter M. , Langlands F. , White J. , Dall B. , Horgan K.
Published: December 2010

Introduction:Ultrasound combined with fine needle aspiration cytology is effective in pre-operative staging of the axilla in breast cancer. Accurate pre-operative diagnosis of lymph node metastases allows for a one stage axillary operation and may also influence decisions regarding neo-adjuvant chemotherapy and breast reconstruction. The aim of this study is to identify pathological and patient factors that influence the accuracy of ultrasound and FNAC in determining the status of the axilla pre-operatively.Methods:Three hundred patients with primary operable invasive breast cancer had an axillary ultrasound pre-operatively. If the ultrasound was normal the patient was offered a sentinel node biopsy. If it identified equivocal or pathological nodes a fine needle aspirate cytology was performed. If the cytology was malignant then an axillary node clearance was performed.Results:Ultrasound combined with FNAC correctly determined the status of the axilla pre-operatively in 78% of cases. Sensitivity for the detection of metastases was 32% with 100% specificity. Factors affecting the accuracy of ultrasound and FNAC were the pathological size of tumour (p<0.05) and the size of the metastases (p<0.05). Age of the patient, pathological grade and type of tumour did not significantly affect the accuracy of the procedure.Conclusion:Ultrasound combined with fine needle aspirate cytology can be used effectively to determine the status of the axilla pre-operatively. All patients regardless of age or their particular tumour characteristics should be offered this procedure pre-operatively.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5020.
Utility of Breast CT in the Management of Breast Cancer – Results from a Prospective Multi-Institutional Study.
Authors: Akashi-Tanaka S. , Sato N. , Ohsumi S. , Kimijima I. , Inaji H. , Teramoto S. , Akiyama F.
Published: December 2010

Background:In order to obtain negative margins after breast conserving surgery (BCS), even repeated surgery is widely accepted. Before surgery, it is important to conduct a precise assessment of the extent of the breast cancer so that each patient can receive individualized surgery. To evaluate the utility of breast computed tomography (CT) as a tool to manage BCS, a multi-institutional prospective study was conducted in Japan.Patients and Methods:Patients were eligible to participate in this study if they had histologically proven breast cancer and were determined to be BCS candidates based on palpation, mammography (MMG) and ultrasonography (US). Written informed consent was obtained from each patient. First, the surgeon marked the line of planned excision on the skin using information from the MMG and US. Next, an expired angiographic catheter was placed on the mark to show the original surgical margin on the CT image. Breast CT was scanned 60 seconds after the bolus injection of the contrast material in the supine surgical position. The surgeon determined the extent of surgery based on the breast CT results. Surgical specimens were serially sectioned in 5-mm slices.Results:Three hundred and two patients were enrolled in this study. The CT scanners used in this study varied from a single helical CT to a 64-row multidetector CT. The results of the breast CT changed the extent of resection in 14.7% of patients. Among the 5 patients who were recommended to undergo a mastectomy, 4 patients had multicentric tumors pathologically and 1 patient had a widely spread intraductal component. The other patients were recommended to have a quadrantectomy based on the extent of breast cancer that was visualized by CT. Three patients (1%) who required conversion from a lumpectomy to quadrantectomy resulted in overexcision. In short, breast CT correctly changed the extent of surgery in 13.7% of the examined patients.Conclusion:This prospective study suggested that breast CT is useful for hospitals equipped with any type of CT and can be used to provide patients with individualized surgery.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5021.
Optimizing Breast Cancer Follow-Up: Diagnostic Value and Costs of Additional Routine Breast Ultrasound.
Authors: Wojcinski S. , Hirschauer E. , Degenhardt F. , Schmidt W.
Published: December 2010

Background: Standard means in breast cancer follow-up include clinical examination and mammography for the detection of local recurrences. Additional test will only be conducted if there are unclear or suspicious findings. Ultrasound has not yet been established as a routine method for breast cancer follow-up care. Our study evaluates the diagnostic efficacy of breast ultrasound in detecting occult malignancies. Moreover, we analyzed the effect of false positive findings resulting in additional examinations and biopsies, and we conducted a benefit-cost-analysis and a questioning of the affected patients.Materials and Methods: A total of 879 eligible patients who were seen at the University Hospital of Saarland for the purpose of breast cancer follow-up were recruited between October 2004 and January 2006. All patients received clinical examination and mammography and were then divided into 2 groups. Group 1 (n=703) were clinically and mammographically asymptomatic patients. Group 2 (n=176) had abnormalities either in palpation or mammography. All patients received additional ultrasound in order to detect occult malignancies. Both groups were analyzed with respect to the diagnostic value of additional ultrasound, the rate of additional examinations and biopsies, the extra costs and the individual stress as experienced by the patient.Results: In group 1 (asymptomatic patients) 10 occult malignancies were detected due to the additional ultrasound examination, i.e. one diagnosed malignancy in 70 examined patients (detection rate 1.4%). By reason of false positive ultrasound results, 65 additional control-ultrasounds, 14 additional MRI examinations and finally 22 biopsies of benign lesions were unnecessarily conducted. Weighing in the total costs for ultrasound, MRI and biopsy, an amount of 4150€ (5290$) was spent to find one additional cancer.In group 2, apart from the symptomatic lesions, 3 occult malignancies were exclusively identified due to the ultrasound examination (detection rate 1.7%). On the other hand, false positive ultrasound results caused 7 additional control-ultrasounds, 14 additional MRIs and 11 biopsies of benign lesions. Hence, an amount of 4410€ (5620$) was spent to identify one additional malignancy by the means of additional ultrasound in this group.Overall, 50 patients (5.7%) received biopsy due to false positive results either in ultrasound or mammography, but these patients claimed on questioning not to feel “troubled and anxious”, but “saver and more carefully looked after”.Discussion: Regarding our results, we strongly recommend the routine use of ultrasound in breast cancer follow-up for both, asymptomatic and symptomatic, women, as occult malignancies can be identified cost-efficiently, although additional ultrasound may also result in higher false positive findings.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5027.
A Prospective Study of Computerized Digital Infrared Image Analysis (NoTouch BreastScanTM) in Biopsy Proven Breast Cancers.
Authors: Nair N. , Thakur M. , Hawaldar R. , Nadkarni M. , Parmar V. , Badwe R.
Published: December 2010

Introduction: Early detection of breast cancer is known to have a more favourable outcome. Currently clinical breast examination and imaging modalities, primarily mammography are used for screening purposes. In India, more than 85% of the population is below the age of 50 years, wherein the sensitivity of mammography is at best 64%. Additional drawbacks of the procedure are physical discomfort and ionizing radiation dose to the patient. So newer techniques have been investigated which detect cancer induced neovascularity with digital thermal imaging. The purpose of this clinical study was to determine the efficacy of a software assisted thermal image analysis tool to distinguish between benign and malignant lesions of the breast.Methods: A prospective study was conducted in women who presented to the breast clinic with clinically or mammographically suspicious breast lesions. They also underwent thermal imaging of the breast. All mammographically suspicious lumps were subjected to histopathological confirmation. The mammography and infrared (IR) reports were compared to the histopathology.Results: In 90 women, 180 breasts were independently analyzed by both digital IR software analysis and mammography. Eighty five out of these 180 had suspicious lesions on mammography or clinical examination which were subjected to pathological confirmation. Mammography being the present diagnostic gold standard, all normal mammograms in clinically normal breast were considered as non-malignant. The sensitivity and specificity of digital thermography in detecting malignant lesions was 88.24% and 70.52% respectively with NPV of 87.01% and PPV of 72.82%. While for mammography the sensitivity and specificity were 96.25% and 96.7% with NPV of 96.7% and PPV of 96.25%. In women below 50 years of age (62/90) the sensitivity and specificity of digital thermography was 89.83% and 64.61% with NPV of 87.5% and PPV of 69.74%. Further, in the same subset no statistically significant difference was detected in the sensitivity of digital thermography to that of mammography (p = 0.7263).Conclusion: Our initial experience shows that the detection rate by digital thermal imaging is comparable to mammography in clinically palpable breast tumors. We also note that there is no significant difference in sensitivity of thermal imaging in women on either side of 50 making it a potentially testable tool for screening in younger women.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5028.
Isolation of Tumor Initiating Cells with Metastatic Potential from Human Primary Invasive Ductal Carcinoma.
Authors: Marsden C. , Marsden C. , Wright M. , Wright M. , Carrier L. , Carrier L. , Pochampally R. , Pochampally R. , Pochampally R. , Rowan B. , Rowan B.
Published: December 2010

Disseminated breast cancer cells may be present at distant sites at the time of primary diagnosis of breast cancer in patients that exhibit no outward signs of clinical metastasis. It is hypothesized that early breast cancer metastasis is initiated by a small population of breast tumor initiating cells (bTICs) within the primary tumor that are inherently resistant to chemotherapy and hormonal therapy. To better understand the biological pathways that permit bTICs to metastasize and to evade current breast cancer therapies, bTICs have been isolated from breast cancer biopsies from patients diagnosed with primary invasive ductal carcinoma. Breast cancer needle biopsies from primary tumors were obtained during the routine care of patients with consent and IRB approval. Core biopsies were mechanically and enzymatically dissociated, yielding a single cell suspension which was subsequently cultured under non-adherent and serum-free conditions to obtain tumorspheres. Injection of about 500 bTICs into the mammary fat pad of female nude mice resulted in tumor formation at the site of injection within two months that was maintained as a small palpable mass for at least six months. H+E staining of sections of the primary tumors revealed complex cellular organization and both microvascular and macrovascular structures. Serial transplantation of the primary tumors from the 6/7 samples resulted in tumor formation at the site of injection. Upon primary tumor formation in the mammary fat pad, metastatic human breast cancer cells were detected within the lungs, liver, brain and bone marrow (femur) from 5/7 tumors after six months. Metastatic cells were detected by PCR for an alpha-satellite sequence in the centromeric region in human Chromosome 17, and by in situ hybridization using an human specific Alu DNA oligonucleotide probe. Metastatic bTICs were mostly detected as single cells or small clustering of cells present throughout the metastatic organ. These data demonstrate that cells with the phenotype of “tumor initiating cells” can be isolated from primary biopsies of human invasive ductal carcinoma and cultured in vitro. Unlike direct heterotransplant tissues from primary tumor biopsies, the majority of bTICs samples form tumors when injected into immunodeficient mice and further exhibit a highly metastatic phenotype.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 504.
Identifying Symptom Clusters in Breast Cancer: Implications on Patient Quality of Life.
Authors: Lis C. , Birdsall T. , Stark J. , Cain L. , Campbell K. , Gilbert K. , Gupta D.
Published: December 2010

Background: Patients undergoing treatment for advanced cancer often experience multiple symptoms. These symptoms occur with varying frequency, intensity, and impact. Pain, fatigue, insomnia, anorexia and dyspnea are consistently among the 10 most prevalent symptoms associated with cancer and its treatment. We conducted a multivariate analysis to investigate the impact of cancer symptoms on patient quality of life (QoL) in breast cancer.Material and Methods: A consecutive series of 226 breast cancer patients treated at the Cancer Treatment Centers of America at Southwestern Regional Medical Center between Jan 2001 and Nov 2006. Cancer symptoms were assessed using the symptom subscales of European Organization for Research Treatment of Cancer - Quality of Life Questionnaire (QLQ-C30). Symptom scores range from 0-100 with lower scores indicating better while higher scores indicating worse symptoms. QoL was assessed using Ferrans and Powers Quality of Life Index (QLI). QLI measures overall QoL and QoL in 4 major subscales: health/physical, social/economic, psychological/spiritual, and family. QLI scores range from 0-30 with higher scores indicating better QoL. The relationship between QLQ-C30 symptoms and QLI was evaluated using multivariate linear regression.Results: Of 226 patients, 98 were newly diagnosed while 128 had failed prior treatment before coming to our hospital. The mean age was 52.5 years (range = 27-82 years). We found that every 10 unit increase in fatigue (p=.004) and pain (p=.001) was significantly associated with 0.65 and 0.66 unit decline in QLI health/physical subscale after controlling for age, treatment history, stage at diagnosis and other cancer symptoms. Similarly, every 10 unit increase in insomnia (p=.02) was significantly associated with 0.35 unit decline in QLI family subscale. Finally, every 10 unit increase in pain (p=.01) and insomnia (p=.03) was significantly associated with 0.34 and 0.21 unit decline in overall QoL.Discussion: We found fatigue, pain and insomnia to be independent predictors of QoL after controlling for each other, age, treatment history and stage at diagnosis. Recognition and timely treatment of symptom clusters could result in improved QoL in patients with breast cancer.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5043.
Medical Ozone (O3) Oil or Gas Applications Heal Osteonecrosis of the Jaw (ONJ) in Patients Treated with Bisphosphonates (BPs). Preliminary Results of a Single Arm Study.
Authors: Ripamonti C. , Maniezzo M. , Ghiringhelli R. , Cislaghi E. , Mariani L.
Published: December 2010

Treatment of bone metastasis with BP is aimed at preventing skeletal related events (SREs) and rarely leads to ONJ (<1%) (Hoff, JBMR. 2008). Preventive dental care is a simple and effective way to reduce the risk reduction of ONJ in 75% of the cases (Ripamonti, Ann Oncol. 2009). To date, there are still no standardized therapy for the treatment of ONJ. We report the effect of localized of O3 oil or gas application in cancer patients (12 breast cancer, 4 prostate cancer, 2 lung cancer, 1 NHL, 3 multiple myeloma) with ONJ observed in our institution. Patients had previously received nitrogene-BPs treatment in the absence of odontoiatric preventive measures. All the patients were in had stable disease without progression; 10 of them had ONJ lesions ≤ to 2.5 cm.O3 oil suspension applications on ONJ lesions ≤ 2.5 cm was carried out with localized applications directly on the lesions; patients with larger lesions (≥ 2.5 cm) were treated with the ozone gas locally applied. All the patients received treatment of O3 oil every third day and all of them were treated with antibiotic therapy (azithromycin, 500 mg/day) 10 days prior the initiation of the treatment with O3 oil. The statistical analysis is based on a Simon two-stage design, the second stage is on-going. In this preliminary analysis we focus on 10 patients treated with O3 oil with a medium follow up of 8 months, and 12 patients on medical gas ozone.73% of the patients (n=16) showed complete response in terms of radiological lesion disappearance with complete reconstruction of oral tissue. Among them, fourteen patients developed spontaneous sequestrum with expulsion of the necrotic bone whereas in 2 patients with large extension of bone involvement surgical intervention was necessary.Seventy percent of the patients treated with O3 oil experienced a complete response after 4 applications whereas patients treated with gas needed 4 to 16 applications (depending on the severity of the lesion). No patients presented adverse events related to the use of ozone treatments. Six patients are still on treatment and are improving as well.According to these results few application of O3 oil suspension in patients with smaller lesions and gas medical ozone for wider lesions following antibiotic therapy can rapidly lead to complete healing of ONJ. Further cases and complete follow-up data are required. These data indicate that ONJ is a manageable condition which can be not only prevented by means previously dental examination but also healed with medical ozone applications.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5046.
Pharmacological Inhibition of Signaling through the Activin Receptor Type IIa Prevents Primary Tumor Growth and Skeletal Metastases in Mouse Models of Breast Cancer.
Authors: Mulivor A. , Barbosa D. , Kumar R. , Underwood K. , Ucran J. , Seehra J. , Pearsall R.
Published: December 2010

Breast cancer cells have a strong propensity to metastasize to bone, resulting in debilitating complications such as intractable bone pain, spinal compression, pathological fractures, or loss of mobility and function. Anti-resorptive agents, such as bisphosphonates, are commonly used to halt the loss of bone, but do not stimulate the formation of new bone. Previous data demonstrated that treatment with an activin antagonist, a soluble form of the extracellular domain of the activin receptor type IIA (ActRIIA) fused to a murine IgG-Fc fragment (RAP-011), can restore bone mass in ovariectomized mice with established bone loss and prevent bone loss in a murine model of multiple myeloma.The goal of the current studies was to establish the ability of RAP-011 to directly inhibit breast cancer cell proliferation, delay primary tumor growth and prevent skeletal metastases. Bioluminescent breast cancer cells (MCF-7-luc and MDA-MB-231-luc-D3H2LN) were administered orthotopically to establish a primary tumor model or by intracardiac injection to establish a metastatic breast cancer model. Both studies were performed using athymic nude mice that had been pretreated for 2 weeks with RAP-011 (10 mg/kg S.C. twice weekly) or vehicle (VEH).In vitro studies demonstrated that activin A stimulated proliferation of some tumor cell lines, while the addition of soluble ActRIIa blocked the stimulation. In the orthotopic models of breast cancer RAP-011 treated mice had significantly smaller primary tumors as early as 3 weeks post-inoculation. RAP-011 treated mice had tumors that remained significantly smaller for the duration of the 10 week study (-60%, p<0.02).In the model of metastatic breast cancer, RAP-011 treated mice displayed lower levels of hind limb, mandible and lung metastases compared to VEH-treated mice. RAP-011 treated mice had reduced tumor burden (-70%, p<0.001), fewer osteolytic lesions (-45%, p<0.05) and increased overall survival by 11 days compared to VEH-treated mice. DXA scanning of the mice revealed RAP-011 treated mice had a significant increase in bone mineral density compared to the VEH treated mice (+10%, p<0.05), reflecting the decreased levels of osteolytic metastases. Histological analysis of the femur and tibia confirmed increased levels of tumor growth and osteolytic lesions, as well as decreased levels of trabecular bone volume in VEH treated mice.These data support the role of Activin in the establishment and growth of tumors and that the use of a soluble ActRIIA receptor can prevent primary tumor growth and inhibit metastases in patients with breast cancer.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5058.
Everolimus (Afinitor®), Trastuzumab (H), and Paclitaxel (P) or Vinorelbine (V) in the Treatment of HER-2+ Metastatic Breast Cancer (MBC) Patients (Pts) Pre-Treated with Lapatinib-Containing Therapy: Pooled Analysis of 2 Phase I Studies.
Authors: Cardoso F. , Dieras V. , Campone M. , Massacesi C. , Manlius C. , Thevenaz P. , Zhang Y. , Jerusalem G. , Sahmoud T. , Andre F. , Gianni L.
Published: December 2010

Introduction: The re-introduction of H-based therapy after progression on a lapatinib-containing regimen has not yet been supported by clear evidence. The aim of this analysis is to describe the safety and efficacy of the regimens everolimus (E) + H, and either P or V in patients pre-treated with lapatinib (L) in relation to those who did not receive previous L (N-L). Methods: Data from pts enrolled in two multicenter phase I studies (NCT00426556 and NTC00426530), with similar study design, were pooled and analyzed according to previous treatment with L. Regimens were: oral E daily (5 or 10mg) or weekly (20 or 30mg) + weekly H 2mg/kg IV, and either P 80mg/m2, IV D1-8-15 q4w (33 pts) or V 25 mg/m2, IV D1-8 q3w (46 pts). Results: Twenty-six (33%) of the overall 79 pts enrolled, received previous L (equally distributed among E dose cohorts). Baseline characteristics of the L and N-L groups, respectively, were: median age: 49 and 53 yo; visceral disease: 85% and 75% of pts; median number of previous chemotherapy lines for metastatic disease: 3 (range: 1-17) and 1 (range: 0-7); prior-taxanes and anthracyclines: 96% and 81% of the L pts, and in 96% and 85% of the N-L pts, respectively; 78 of 79 pts were H-resistant. The 26 pts pretreated with L were all resistant to L and had a median treatment duration of 24 wks (range: 6-123). In the L and N-L groups, the most frequently reported severe toxicities were G3/4 neutropenia in 58% and 75%, febrile neutropenia in 8% and 2%, G3 asthenia/fatigue in 19% and 13%, G3 stomatitis in 19% and 11%, G3 diarrhea in 4% and 6%, respectively. G1/2 rash was reported in 15% and 34% of L and N-L pts, respectively. Mean left ventricular ejection fraction was similar at baseline and at end of treatment (EOT) in L and N-L pts: 63% (+/- 7%) in both groups at baseline, and 60% (+/- 7%) and 64% (+/- 6%) at EOT, respectively. Table 1 summarizes the efficacy observed in the 74 evaluable pts to date. Time to progression (TTP) appears longer in the N-L group and may be a result of less pre-treatment in this group. Conclusions: In our experience patients pre-treated with lapatinib are not at a higher risk of toxicity and do not seem to have a substantially different outcome when re-treated with trastuzumab-containing regimens in combination with everolimus.Table 1. Overall response and time to progression (Kaplan-Meier based) Lapatinib-pretreated (n=25)Not Lapatinib-pretreated (n=49)Best Response, n (%)  CR–3 (6)PR5 (20)12 (25)SD16 (64)27 (55)PD4 (16)7 (14)TTP, median (weeks)29 (21-41)38 (29-44)CR, complete response; PD, progressive disease; PR, partial response; SD, stable disease; TTP, time to progression.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5064.
Targeting Cancer Stem Cells and Vasculature by a Novel Anti-Delta-Like 4 Ligand (DLL4) Antibody for Treatment of Triple Negative Breast Cancer.
Authors: Yen W. , Fischer M. , Lewicki J. , Gurney A. , Hoey T.
Published: December 2010

Triple negative (ER-, PR- Her2-) breast cancer accounts for 10% of all types of breast cancer and is known to particularly aggressive and refractory to current therapies. Chemotherapeutic agents, such as taxanes, are currently the only treatment option for this type of breast cancer, however, these treatments often result in high rates of local and systemic relapse. It has been suggested that cancer stem cells drive tumor growth and progression and are preferentially resistant to many current therapies. Delta-like 4 ligand (DLL4) is an important component of the Notch signaling pathway which is known to mediate stem cell self-renewal and vascular development. We hypothesized that targeting cancer stem cells and the tumor vasculature by interfering with the DLL4/Notch pathway will improve treatment outcome. We developed anti-DLL4 antibodies that recognize either the human or murine protein and have potent binding and antagonist activities. We have used these antibodies to investigate the role of DLL4 in triple negative breast cancer and to probe the mechanism in tumor and stromal cells in xenograft models derived from primary breast tumors. These studies showed that anti-DLL4 was efficacious as a single agent and in combination with paclitaxel (Taxol) against triple negative breast tumors including a tumor that developed paclitaxel resistance. Gene expression analysis showed that anti-DLL4 affected vascular-related genes in the stroma and Notch target genes in the tumor and stroma. Furthermore, inclusion of anti-DLL4 delayed breast tumor recurrence following termination of paclitaxel treatment. A tumorigenicity study indicated that treatment with anti-DLL4 decreased cancer stem cell frequency as measured by in vivo limiting dilution assay, whereas paclitaxel alone was ineffective. The combination of anti-DLL4 and paclitaxel further decreased cancer stem cell frequency compared to single agents. Our findings provide a rationale for targeting cancer stem cells and tumor vasculature through inhibition of the DLL4/Notch signaling for breast cancer treatment.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5071.
Growth Inhibition of Human Breast Cancer Cells with an Iron Chelator (Deferasirox) and Docetaxel.
Authors: Lipton A. , Witters L. , Nick H.
Published: December 2010

Treatment of cells with an iron chelator results in G1/S arrest and apoptosis. Tumor cells appear to be more sensitive to iron chelation probably because they express higher levels of the iron containing enzyme, ribonucleotide reductase. Previously, our laboratory has shown that an iron chelator (deferoxamine) inhibited the growth of a variety of cancer cell lines and also prolonged survival in tumor bearing mice. Docetaxel is an antineoplastic agent belonging to the taxane family. Docetaxel induces bcl-2 phosphorylation and subsequent apoptosis and is effective in the treatment of patients with breast cancer. The purpose of this study was to assess the in vitro effects of combining an iron chelator, deferasirox (Exjade: Novartis Pharma AG), and docetaxel on the growth of MCF-7 and HER-2/neu-transfected MCF-7 human breast cancer cells. Cells were exposed to deferasirox or docetaxel alone or in combination for 3 days (9 different dose combinations were tested). Cell growth was determined using the colorimetric MTT tetrazolium dye assay. Dose-dependent growth inhibition was observed in both the MCF-7 and the HER-2/neu-tranfected MCF-7 cells after 3 days exposure to deferasirox (IC50: 1.5 and 2.3 μM, respectively) or docetaxel (IC50: 0.9 nM and 2.0 nM, respectively). Exposure to the combination of deferasirox (0.25 – 1.5 μM) and docetaxel (0.1 – 0.5 nM) resulted in synergistic growth inhibition in both the MCF-7 and HER-2/neu-transfected MCF-7 human breast cancer cells. Sequencing experiments with the HER-2/neu-transfected MCF-7 cells showed that simultaneous exposure of the cells to deferasirox and docetaxel was superior to pretreating the cells with either of the agents 6 or 24 hours prior to addition of the second agent. These results suggest this combination warrants further investigation as a therapeutic strategy for breast cancer.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5075.