http://breast-cancer-research.com/ The latest research articles published by Breast Cancer Research 2012-05-14T00:00:00Z IntroductionExperimental and clinical evidence points to a critical role of progesterone and the nuclear progesterone receptor (PR) in controlling mammary gland tumorigenesis. However, the molecular mechanisms of progesterone action in breast cancer still remain elusive. On the other hand, micro RNAs (miRNAs) are short ribonucleic acids which have also been found to play a pivotal role in cancer pathogenesis. The role of miRNA in progestin-induced breast cancer is poorly explored. In this study we explored progestin modulation of miRNA expression in mammary tumorigenesis. Methods: We performed a genome-wide study to explore progestin-mediated regulation of miRNA expression in breast cancer. miR-16 expression was studied by RT-qPCR in cancer cell lines with silenced PR, signal transducer and activator of transcription 3 (Stat3) or c-Myc, treated or not with progestins. Breast cancer cells were transfected with the precursor of miR-16 and proliferation assays, Western blots or in vivo experiments were performed. Target genes of miR-16 were searched through a bioinformatical approach, and the study was focused on cyclin E. Reporter gene assays were performed to confirm that cyclin E 3'UTR is a direct target of miR-16. Results: We found that 9 miRNAs were upregulated and 7 were downregulated by progestin in mammary tumor cells. miR-16, whose function as a tumor suppressor in leukemia has already been shown, was identified as one of the downregulated miRNAs in murine and human breast cancer cells. Progestin induced a decrease in miR-16 levels via the classical PR and through a hierarchical interplay between Stat3 and the oncogenic transcription factor c-Myc. A search for miR-16 targets showed that the CCNE1 gene, encoding the cell cycle regulator cyclin E, contains conserved putative miR-16 target sites in its mRNA 3' UTR region. We found that, similar to the molecular mechanism underlying progestin-modulated miR-16 expression, Stat3 and c-Myc participated in the induction of cyclin E expression by progestin. Moreover, overexpression of miR-16 abrogated the ability of progestin to induce cyclin E upregulation, revealing that cyclin E is a novel target of miR-16 in breast cancer. Overexpression of miR-16 also inhibited progestin-induced breast tumor growth in vitro and in vivo, demonstrating for the first time, a role for miR-16 as a tumor suppressor in mammary tumorigenesis. We also found that the ErbB ligand heregulin (HRG) downregulated the expression of miR-16, which then participates in the proliferative activity of HRG in breast tumor cells. Conclusions: In this study, we reveal the first progestin-regulated miRNA expression profile and identify a novel role for miR-16 as a tumor suppressor in progestin- and growth factor-induced growth in breast cancer. http://breast-cancer-research.com/content/14/3/R77 Martin Rivas Leandro Venturutti Yi-Wen Huang Roxana Schillaci Tim Huang Patricia Elizalde Breast Cancer Research 2012, null:R77 2012-05-14T00:00:00Z doi:10.1186/bcr3187 /content/figures/bcr3187-toc.gif Breast Cancer Research 1465-5411 ${item.volume} R77 2012-05-14T00:00:00Z PDF IntroductionAssociations of hormone-receptor positive breast cancer with excess adiposity are reasonably well characterized; however, uncertainty remains regarding the association of body mass index (BMI) with hormone-receptor negative malignancies, and possible interactions by hormone replacement therapy (HRT) use. Methods: Within the European EPIC cohort, Cox proportional hazards models were used to describe the relationship of BMI, waist and hip circumferences with risk of estrogen receptor (ER)-progesterone receptor (PR)- (n=1,021) and ER+PR+ (n=3,586) breast tumors within five-year age bands. Among postmenopausal women, the joint effects of BMI and HRT use were analyzed. Results: For risk of ER-PR- tumors, there was no association of BMI across the age bands. However, when analyses were restricted to postmenopausal HRT never users, a positive risk association with BMI (3rd versus 1st tertile HR=1.47[1.01-2.15]) was observed. BMI was inversely associated with ER+PR+ tumors among women aged [less than or equal to]49 years (per 5kg/m2 increase, HR=0.79[95%CI 0.68-0.91]), and positively associated with risk among women [greater than or equal to]65 years (HR=1.25[1.16-1.34]). Adjusting for BMI, waist and hip circumferences showed no further associations with risks of breast cancer subtypes. Current use of HRT was significantly associated with an increased risk of receptor-negative (HRT current use compared to HRT never use HR: 1.30[1.05-1.62]) and positive tumors (HR: 1.74[1.56-1.95]), although this risk increase was weaker for ER-PR- disease (Phet=0.035). The association of HRT was significantly stronger in the leaner women (BMI [less than or equal to]22.5kg/m2) than for more overweight women (BMI [greater than or equal to]25.9kg/m2) for, both, ER-PR- (HR: 1.74[1.15-2.63]) and ER+PR+ (HR: 2.33[1.84-2.92]) breast cancer and was not restricted to any particular HRT regime. Conclusions: An elevated BMI may be positively associated with risk of ER-PR- tumors, among postmenopausal women who never used HRT. Furthermore, postmenopausal HRT users were at an increased risk of ER-PR- as well as ER+PR+ tumors, especially among leaner women. For HR-positive tumors, but not for HR-negative tumors, our study confirms an inverse association of risk with BMI among young women of premenopausal age. Our data provide evidence for a possible role of sex hormones in the etiology of HR-negative tumors. http://breast-cancer-research.com/content/14/3/R76 Rebecca Ritte Annekatrin Lukanova Franco Berrino Laure Dossus Anne Tjonneland Anja Olsen Thure Overvad Kim Overvad Francoise Clavel-Chapelon Agnes Fournier Guy Fagherazzi Sabine Rohrmann Birgit Teucher Heiner Boeing Krasimira Aleksandrova Antonia Trichopoulou Pagona Lagiou Dimitrios Trichopoulos Domenico Palli Sabina Sieri Salvatore Panico Rosario Tumino Paolo Vineis Jose Ramon Quiros Genevieve Buckland Maria-Jose Sanchez Pilar Amiano Maria-Dolores Chirlaque Eva Ardanaz Malin Sund Per Lenner Bas Bueno-de-Mesquita Carla van Gils Petra Peeters Sanda Krum-Hansen Inger Gram Eiliv Lund Kay-Tee Khaw Nick Wareham Naomi Allen Timothy Key Isabelle Romieu Sabina Rinaldi Afshan Siddiq David Cox Elio Riboli Rudolf Kaaks Breast Cancer Research 2012, null:R76 2012-05-14T00:00:00Z doi:10.1186/bcr3186 /content/figures/bcr3186-toc.gif Breast Cancer Research 1465-5411 ${item.volume} R76 2012-05-14T00:00:00Z PDF IntroductionThe re-emergence of the tumour growth factor-beta (TGF-beta)-relatedembryonic morphogen Nodal has recently been reported in several different human cancers. In this study, we examined the expression of Nodal in a series of benign and malignant human breast tissues to determine the clinical significance of this expression and whether Nodal could represent a potential therapeutic target in breast cancer. Methods: Tissue sections from 431 therapeutically naive patients diagnosed with benign or malignant breast disease were stained for Nodal by immunohistochemistry and analysed in a blinded manner. The degree of Nodal staining was subsequently correlated with available clinical data, such as diagnoses and disease stage. These tissue findings were further explored in breast cancer cell lines MDA-MB-231 and MDAMB- 468 treated with a Nodal blocking antibody to determine biological effects for target validation. Results: A variable degree of Nodal staining was detected in all samples. The intensity of Nodal staining was significantly greater in undifferentiated, advanced stage, invasive breast cancer compared with benign breast disease or early stage breast cancer. Treatment of human breast cancer cells in vitro with Nodal blocking antibody significantly reduced proliferation and colony-forming ability in soft agar, concomitant with increased apoptosis. Conclusions: These data suggest a potential role for Nodal as a biomarker for disease progression and a promising target for anti-Nodal therapy in breast cancer. http://breast-cancer-research.com/content/14/3/R75 Luigi Strizzi Katharine Hardy Naira Margaryan David Hillman Elisabeth Seftor Beiyun Chen Xochiquetzal Geiger E Thompson Wilma Lingle Cathy Andorfer Edith Perez Mary Hendrix Breast Cancer Research 2012, null:R75 2012-05-11T00:00:00Z doi:10.1186/bcr3185 Nodal: prognostic breast cancer biomarker? The embryonic morphogen Nodal is associated with advanced stage invasive human breast cancer, and may have a role as both a prognostic biomarker and a therapeutic target. /content/figures/bcr3185-toc.gif Breast Cancer Research 1465-5411 ${item.volume} R75 2012-05-11T00:00:00Z PDF Identification of molecules and their effectors has led to new therapies designed to specifically inhibit pathways in molecularly defined breast cancer subtypes. An orphan nuclear receptor, estrogen-related receptor alpha, has been shown to be a downstream target of two tyrosine kinase growth factor receptors: human epidermal growth factor receptor 2 and the type I insulin-like growth factor receptor. Identifying the mechanistic actions of orphan nuclear receptors could lead to new biomarkers and molecular targets in malignancy. http://breast-cancer-research.com/content/14/3/309 Aleksanda Ochnik Douglas Yee Breast Cancer Research 2012, null:309 2012-05-10T00:00:00Z doi:10.1186/bcr3124 /content/figures/bcr3124-toc.gif Breast Cancer Research 1465-5411 ${item.volume} 309 2012-05-10T00:00:00Z XML IntroductionCT10 regulator of kinase (Crk) adaptor proteins (CrkI, CrkII and CrkL) play a role in integrating signals for migration and invasion of highly malignant breast cancer cell lines. This has important implications, as elevated CrkI/II protein levels were observed in a small cohort of breast cancer patients, which identified a potential role for Crk proteins in breast cancer progression. Numerous in vitro studies identified a role for Crk proteins in cell motility, but little is known about how Crk proteins contribute to breast cancer progression in vivo. Methods: The clinical significance of Crk proteins in human breast cancer was assessed by analyzing published breast cancer datasets using a gene expression signature that was generated following CrkII over-expression, and by examining Crk protein expression in tissue microarrays of breast tumors (n=254). Stable knockdown of Crk (CrkI/CrkII/CrkL) proteins was accomplished using an shRNA-mediated approach in two basal breast cancer cell lines, MDA-231 1833TR and SUM1315, where the former have a high affinity to form bone metastases. Both in vitro assays (cell migration, invasion, soft agar growth) and in vivo experiments (intra-cardiac, tibial and mammary fat pad injections) were performed to assess the functional significance of Crk proteins in breast cancer. Results: A gene signature derived following CrkII over-expression correlated significantly with basal breast cancers and with high grade and poor outcome in general. Moreover, elevated Crk immunostaining on tissue microarrays revealed a significant association with highly proliferative tumors within the basal subtype. RNAi-mediated knockdown of all three Crk proteins in metastatic basal breast cancer cells established a continued requirement for Crk in cell migration and invasion in vitro and metastatic growth in vivo. Furthermore, Crk ablation suppressed anchorage independent growth and in vivo orthotopic tumor growth. This was associated with diminished cell proliferation and was rescued by expression of non-shRNA targeted CrkI/II. Perturbations in tumor progression correlated with altered integrin signaling, including decreased cell spreading, diminished p130Cas phosphorylation, and Cdc42 activation. Conclusions: These data highlight the physiological importance of Crk proteins in regulating growth of aggressive basal breast cancer cells, and identify Crk-dependent signaling networks as promising therapeutic targets. http://breast-cancer-research.com/content/14/3/R74 Kelly Fathers Emily Bell Charles Rajadurai Sean Cory Hong Zhao Anna Mourskaia Dongmei Zuo Jason Madore Anie Monast Anne-Marie Mes-Masson Andree-Anne Grosset Louis Gaboury Michael Hallett Peter Siegel Morag Park Breast Cancer Research 2012, null:R74 2012-05-08T00:00:00Z doi:10.1186/bcr3183 /content/figures/bcr3183-toc.gif Breast Cancer Research 1465-5411 ${item.volume} R74 2012-05-08T00:00:00Z PDF IntroductionThe chromodomain helicase DNA binding protein 5 (CHD5) has recently been identified as a tumor suppressor in a mouse model. The CHD5 locus at 1p36 is deleted, and its mutation has been detected in breast cancer. We therefore evaluated whether CHD5 plays a role in human breast cancer. Methods: We screened mutations in 55 tumors, determined promoter methylation in 39 tumors, measured RNA expression in 90 tumors, analyzed protein expression in 289 tumors, and correlated expression changes with clinicopathological characteristics of breast cancer. Functional effects of CHD5 on cell proliferation, invasion and tumorigenesis were also tested. Results: Although only one mutation was detected, CHD5 mRNA expression was significantly reduced, accompanied by frequent genomic deletion and promoter methylation, in breast cancer. The extent of methylation was significantly associated with reduced mRNA expression, and demethylating treatment restored CHD5 expression. Lower CHD5 mRNA levels correlated with lymph node metastasis (P = 0.026). CHD5 protein expression was also reduced in breast cancer, and lack of CHD5 expression significantly correlated with higher tumor stage, ER/PR-negativity, HER2 positivity, distant metastasis and worse patient survival (P [less than or equal to] 0.01). Functionally, ectopic expression of CHD5 in breast cancer cells inhibited cell proliferation and invasion in vitro and tumorigenesis in nude mice. Consistent with the inhibition of invasion, CHD5 downregulated mesenchymal markers vimentin, N-cadherin and ZEB1 in breast cancer cells. Conclusion: Downregulation of CHD5, mediated at least in part by promoter methylation, contributes to the development and progression of human breast cancer. http://breast-cancer-research.com/content/14/3/R73 Xiao Wu Zhengmao Zhu Weidong Li Xiaoying Fu Dan Su Liya Fu Zhiqian Zhang Ang Luo Xiaodong Sun Li Fu Jin-Tang Dong Breast Cancer Research 2012, null:R73 2012-05-08T00:00:00Z doi:10.1186/bcr3182 /content/figures/bcr3182-toc.gif Breast Cancer Research 1465-5411 ${item.volume} R73 2012-05-08T00:00:00Z PDF IntroductionCentromere protein A (CENP-A), an essential centromere protein, has been associated with high grade cancers. This study was undertaken to determine if CENP-A is a prognostic factor for breast cancer patients not receiving systemic therapy or predictive of response to tamoxifen or neoadjuvant chemotherapy. Methods: mRNA levels of CENP-A and CENP-B, a centromere protein that binds independently of CENP-A, were measured in breast cancer specimens from 484 patients receiving no systemic therapy, 276 patients receiving tamoxifen, and 233 patients treated with neoadjuvant chemotherapy. Associations between CENP-A, CENP-B, Ki-67, relapse, and chemotherapy response were determined. Results: CENP-A but not CENP-B was higher in estrogen receptor (ER)-negative tumors than ER-positive tumors and positively correlated with Ki-67 expression. Among patients with ER-positive disease who received no systemic therapy or tamoxifen, higher levels of CENP-A were associated with lower rates of 5-year distant relapse free survival (DRFS). On multivariate analyses including Ki-67, high CENP-A expression had a hazard ratio of 10.9 for relapse in patients with ER-positive disease not receiving systemic therapy (95% CI, 2.86 to 41.78; P = 0.00047) and 1.64 for patients with ER-positive disease receiving tamoxifen (95% CI, 0.99 to 2.71; P = 0.054). CENP-A was not an independent prognostic marker in ER-negative tumors. For both ER-positive and ER-negative tumors, CENP-A was not a significant independent predictor of chemotherapy response. Conclusions: CENP-A was a significant independent prognostic marker for patients with ER-positive breast cancer not treated with systemic therapy but had limited predictive value in tamoxifen treated patients and was not predictive of response to neoadjuvant chemotherapy. http://breast-cancer-research.com/content/14/3/R72 Susan McGovern Yuan Qi Lajos Pusztai William Symmans Thomas Buchholz Breast Cancer Research 2012, null:R72 2012-05-04T00:00:00Z doi:10.1186/bcr3181 CENP-A marker for breast cancer relapse The essential centromere protein, CENP-A, is a strong prognostic marker for distant relapse in estrogen-receptor positive breast cancer not treated with systemic therapy, and may contribute to disease progression. /content/figures/bcr3181-toc.gif Breast Cancer Research 1465-5411 ${item.volume} R72 2012-05-04T00:00:00Z PDF IntroductionIncreasing evidence supports that the detection of circulating tumour cells (CTCs) predicts outcomes of nonmetastatic breast cancer patients. CTCs differ genetically from the primary tumour and may contribute to variations in prognosis and response to therapy. As we start to understand more about the biology of CTCs, we can begin to address how best to treat this form of disease. Methods: 98 nonmetastatic breast cancer patients were included in this study. CTCs were isolated by immunomagnetic techniques using magnetic beads labelled with a multi-CK-specific antibody (CK3-11D5) and CTCs detection through immunocytochemical methods. Estrogen receptor, Progesterone receptor and epidermal growth factor receptor (EGFR) were evaluated by immunofluorescence experiments and HER2 and TOP2A by Fluorescence in situ Hybridization. We aimed to characterize this set of biomarkers in CTCs and correlate with clinical-pathological characteristics. Results: Baseline detection rate was 46.9% [greater than or equal to]1 CTC/30 ml threshold. CTCs-positive were more frequent in HER2-negative tumours (p=0.046). In patients younger than 50 years old, HER2 amplified and G1-G2 tumours had a higher possibility of being nondetectable CTCs. Heterogeneous expression of hormonal receptors (HR) in samples from the same patients was found. Discordances between HR expression, HER2 and TOP2A status in CTCs and their primary tumour were found in the sequential blood samples. Less that 35% of patients switched their CTC status after receiving chemotherapy. EGFR-positive CTCs were associated to Luminal tumours (p=0.03). Conclusion: This is the largest exploratory CTCs biomarker analysis in nonmetastatic BC patients. Our study suggests that CTCs biomarkers profiles might be useful as a surrogate marker for therapeutic selection and monitoring since heterogeneity of the biomarkers distribution in CTCs and the lack of correlation with the primary tumor biomarker status were found. Further exploration of the association between EGFR-positive CTCs and Luminal tumours is warranted. http://breast-cancer-research.com/content/14/3/R71 Rosa Nadal Ana Fernandez Pedro Sanchez-Rovira Marta Salido Maria Rodriguez Jose Garcia-Puche Marta Macia Josep Coromines Miguel Delgado-Rodriguez Lucas Gonzalez Joan Albanell Monica Fernandez Francesc Sole Jose Lorente M. Jose Serrano Breast Cancer Research 2012, null:R71 2012-05-03T00:00:00Z doi:10.1186/bcr3180 /content/figures/bcr3180-toc.gif Breast Cancer Research 1465-5411 ${item.volume} R71 2012-05-03T00:00:00Z PDF IntroductionPre-clinical data suggest p53-dependent anthracycline-induced apoptosis and p53-independent taxane activity. However, dedicated clinical research has not defined a predictive role for TP53 gene mutations. The aim of the current study was to retrospectively explore the prognosis and predictive values of TP53 somatic mutations in the BIG 02-98 randomized phase III trial in which women with node-positive breast cancer were treated with adjuvant doxorubicin-based chemotherapy with or without docetaxel. Methods: The prognostic and predictive values of TP53 were analyzed in tumor samples by gene sequencing within exons 5-8. Patients were classified according to p53 protein status predicted from TP53 gene sequence, as wild type (no TP53 variation or TP53 variations which are predicted not to modify p53 protein sequence) or mutant (p53 non-synonymous mutations). Mutations were subcategorized according to missense or truncating mutations. Survival analyses were performed using Kaplan-Meier method and log-rank test. Cox-regression analysis was used to identify independent predictors of outcome. Results: TP53 gene status was determined for 18% (520/2887) of the women enrolled in BIG 02-98. TP53 gene variations were found in 17% (90/520). Non-synonymous p53 mutations, found in 16.3% (85/520), were associated with older age, ductal morphology, higher grade and hormone-receptor negativity. Of the non-synonymous mutations, 12.3% (64/520) were missense and 3.6% were truncating (19/520). Only truncating mutations showed significant independent prognostic value, with an increased recurrence risk compared to patients with non-modified p53 protein (Hazard Ratio=3.21, 95% confidence interval =1.740-5.935, P=0.0002). p53 status had no significant predictive value for response to docetaxel. Conclusions: p53 truncating mutations were uncommon but associated with poor prognosis. No significant predictive role for p53 status was detected.Trial registration: NCT00174655 http://breast-cancer-research.com/content/14/3/R70 Lynnette Fernandez-Cuesta Catherine Oakman Priscila Falagan-Lotsch Ke-seay Smoth Emmanuel Quinaux Marc Buyse M Dolci Evandro De Azambuja Pierre Hainaut Patrizia Dell'Orto Denis Larsimont Prudence Francis John Crown Martine Piccart-Gebhart Giuseppe Viale Angelo Di Leo Magali Olivier Breast Cancer Research 2012, null:R70 2012-05-02T00:00:00Z doi:10.1186/bcr3179 /content/figures/bcr3179-toc.gif Breast Cancer Research 1465-5411 ${item.volume} R70 2012-05-02T00:00:00Z PDF Randomized adjuvant trials continue to show significant reductions in distant recurrence and death for early-stage women treated with adjuvant trastuzumab. BCIRG-006 showed superior disease-free and overall survival of two trastuzumab-containing regimens in comparison to a non-trastuzumab-containing regimen. The rates of disease-free and overall survival were not statistically different for the two trastuzumab-containing arms. Ongoing study is needed to identify markers of resistance to trastuzumab and incorporate newer agents in the adjuvant setting in order to further decrease rates of distant recurrence and death from HER2-positive breast cancer. http://breast-cancer-research.com/content/14/2/308 Rachel Jankowitz Adam Brufsky Breast Cancer Research 2012, null:308 2012-04-30T00:00:00Z doi:10.1186/bcr3120 /content/figures/bcr3120-toc.gif Breast Cancer Research 1465-5411 ${item.volume} 308 2012-04-30T00:00:00Z XML