Untangling the ATR-CHEK1 network for prognostication, prediction and therapeutic target validation in breast cancer

Background: ATR-Chk1 signalling network is critical for genomic stability. ATR-Chk1 may be deregulated in breast cancer and have prognostic, predictive and therapeutic significance. Patients and methods: We investigated ATR and phosphorylated CHK1Ser345 protein (pChk1) expression in 1712 breast cancers (Nottingham Tenovus series). ATR and Chk1 mRNA were evaluated in 1950 breast cancers (METABRIC cohort). Pre-clinically, biological consequences of ATR gene knockdown or ATR inhibition by small molecule inhibitor (VE-821) were investigated in MCF-7 and MDA-MB-231 breast cancer cell lines and in non-tumorigenic breast epithelial cells (MCF10A). Results: High ATR and high cytoplasmic pChk1 expression was significantly associated with higher tumour stage, higher mitotic index, pleomorphism and lymphovascular invasion. In univariate analysis, high ATR and high cytoplasmic pChk1 protein expression was associated with shorter breast cancer specific survival (BCSS). In multivariate analysis, high ATR remains an independent predictor of adverse outcome. At the mRNA level, high Chk1 remains associated with aggressive phenotypes including lymph node positivity, high grade, Her-2 overexpression, triple-negative phenotype and molecular classes associated with aggressive behaviour and shorter survival.. Pre-clinically, Chk1 phosphorylation at serine 345 following replication stress (induced by gemcitabine or hydroxyurea treatment) was impaired in ATR knockdown and in VE-821 treated breast cancer cells. Doxycycline inducible knockdown of ATR suppressed growth, which was restored when ATR was re-expressed. Similarly, VE-821 treatment resulted in a dose dependent suppression of cancer cell growth and survival (MCF7 and MDA-MB-231) but had no effect on non-tumorigenic breast epithelial cells (MCF10A). Conclusions: We provides evidence that ATR and Chk1 are promising biomarkers and rational drug target for personalized therapy in breast cancer

Prognostic factors in metaplastic carcinoma of the breast: A multi-institutional study

Background: Metaplastic breast carcinoma (MBC) is a rare type of breast cancer that has basal-like characteristics and is perceived to have poorer prognosis when compared with conventional no specific type/ductal carcinomas (ductal/NST). However, current data on MBC are largely derived from small case series or population-based reports. This study aimed to assess the clinicopathological features and outcome of MBC identified through an international multicentre collaboration. Methods: A large international multicentre series of MBC (no=405) with histological confirmation and follow-up information has been included in this study. The prognostic value of different variables and outcome has been assessed and compared with grade, nodal status and ER/HER2 receptor-matched ductal/NST breast carcinoma. Results: The outcome of MBC diagnosed in Asian countries was more favourable than those in Western countries. The outcome of MBC is not different from matched ductal/NST carcinoma but the performance of the established prognostic variables in MBC is different. Lymph node stage, lymphovascular invasion and histologic subtype are associated with outcome but tumour size and grade are not. Chemotherapy was associated with longer survival, although this effect was limited to early-stage disease. In this study no association between radiotherapy and outcome was identified. Multivariate analysis of MBC shows that histologic subtype is an independent prognostic feature. Conclusions: This study suggests that MBC is a heterogeneous disease. Although the outcome of MBC is not different to matched conventional ductal/NST breast carcinoma, its behaviour is dependent on the particular subtype with spindle cell carcinoma in particular has an aggressive biological behaviour. Management of patients with MBC should be based on validated prognostic variables

Simulations of Alfvénic modes in TJ-II Stellarator

Alfvénic modes are one of the subclass of instabilities prevalent in burning plasmas due to interaction of energetic particles with background plasma. In this paper we investigate the properties of these modes with 3D simulations using modeling tools STELLGAP [1] and AE3D [2] of Neutral Beam Injection (NBI) heated H-plasmas in TJ-II low-magnetic-shear flexible heliac (B0 = 0.95 T, = 1.5 m, = 0.22 m). These simulations focus on modelling the experimental observations [3] for prominent modes in TJ-II plasmas. Our simulations show consistency in frequency and radial location with the measured Alfvén Eigenmodes [3]. These simulations are performed for chirping and steady modes in TJ-II discharge # 29839 at t = 1150 and 1160ms respectively

Immunohistochemistry profiles of breast ductal carcinoma: factor analysis of digital image analysis data

<p>Abstract</p> <p>Background</p> <p>Molecular studies of breast cancer revealed biological heterogeneity of the disease and opened new perspectives for personalized therapy. While multiple gene expression-based systems have been developed, current clinical practice is largely based upon conventional clinical and pathologic criteria. This gap may be filled by development of combined multi-IHC indices to characterize biological and clinical behaviour of the tumours. Digital image analysis (DA) with multivariate statistics of the data opens new opportunities in this field.</p> <p>Methods</p> <p>Tissue microarrays of 109 patients with breast ductal carcinoma were stained for a set of 10 IHC markers (ER, PR, HER2, Ki67, AR, BCL2, HIF-1α, SATB1, p53, and p16). Aperio imaging platform with the Genie, Nuclear and Membrane algorithms were used for the DA. Factor analysis of the DA data was performed in the whole group and hormone receptor (HR) positive subgroup of the patients (n = 85).</p> <p>Results</p> <p>Major factor potentially reflecting aggressive disease behaviour (i-Grade) was extracted, characterized by opposite loadings of ER/PR/AR/BCL2 and Ki67/HIF-1α. The i-Grade factor scores revealed bimodal distribution and were strongly associated with higher Nottingham histological grade (G) and more aggressive intrinsic subtypes. In HR-positive tumours, the aggressiveness of the tumour was best defined by positive Ki67 and negative ER loadings. High Ki67/ER factor scores were strongly associated with the higher G and Luminal B types, but also were detected in a set of G1 and Luminal A cases, potentially indicating high risk patients in these categories. Inverse relation between HER2 and PR expression was found in the HR-positive tumours pointing at differential information conveyed by the ER and PR expression. SATB1 along with HIF-1α reflected the second major factor of variation in our patients; in the HR-positive group they were inversely associated with the HR and BCL2 expression and represented the major factor of variation. Finally, we confirmed high expression levels of p16 in Triple-negative tumours.</p> <p>Conclusion</p> <p>Factor analysis of multiple IHC biomarkers measured by automated DA is an efficient exploratory tool clarifying complex interdependencies in the breast ductal carcinoma IHC profiles and informative value of single IHC markers. Integrated IHC indices may provide additional risk stratifications for the currently used grading systems and prove to be useful in clinical outcome studies.</p> <p>Virtual Slides</p> <p>The virtual slide(s) for this article can be found here: <url>http://www.diagnosticpathology.diagnomx.eu/vs/1512077125668949</url></p

Relevance of the MicroRNA (Mirna) Processor DICER Expression in the Biological Behaviour and Pathological Response of Nigerian Breast Cancer Tissues

Background: Breast cancer (BC) among Nigerians is characterised by high grade, triple negative, basal-like phenotype tumours with high proliferation indices and poor prognosis. The loss of Dicer expression has been speculated to play a key role in BC with similar features among the women in the Western countries. Objectives: To demonstrate the role of the Dicer expression in relation to pathological response in BC, in order to determine the biological behaviour and its prognostic significance in BC among Nigerian women using immunohistochemistry and Tissue microarray (TMA). Methods: This study investigated the immune profiles of the Dicer in 241 tissue microarray of breast cancer tissue of Nigerian women and correlated the protein expression with the pathological response and the other biomarker expressions to determine the functional significance in Nigerian women. Results: Protein expression of Dicer as compared with other biomarkers expression showed there was significant association between the loss of Dicer expression and the down-regulators of Breast Cancer Associated Gene-1 (BRCA1), metastasis tumour antigen-1(MTA 1) (p = 0.004), Inhibitor differentiation-4 (ID4) (p = 0.002), ubiquitin conjugating enzyme-9 (UBC9) (p = 0.008) and protein inhibitor of activated signal transducer gamma PIASγ (p = 0.002). Other relevant Homologous repair pathway markers included poly (ADP-ribose) polymerase-1(PARP1) (p < 0.001) and RAD51 (p < 0.001), cell cycle regulator protein-27(p27) (p = 0.024), the proliferation kinetic protein ( Ki-67) (p = 0.003) and epidermal growth factor receptor (EGFR) expression (p = 0.013). Survival analysis also showed that there was no significant correlation between tumours negative for Dicer and patient outcome. Conclusion: This study demonstrated that the loss of Dicer is associated with intermediate to higher grade tumour, discrepant MI/Ki-67 expression, p27 loss, homologous recombination response dysregulation, high EGFR and Ki-67 expression. Therefore, Dicer expression appears to play a major role in the biology of BC among Nigerian women. A targeted therapy on Dicer expression would enhance the management of BC among Nigerian women

Elevated expression of STK3 mRNA and protein is associated with poor outcome in invasive breast cancer

Purpose of the study: The mammalian sterile 20-like kinase (MST2/STK3) and its close homologue MST1(STK4) are members of the germinal centre kinase group II (GCK II) family of mitogen-activated protein kinases (MAPK). High STK3 expression is known to be correlated with poor prognosis in various cancers playing a role in cell migration and invasion. This study aimed to determine correlations of STK3 expression with clinicopathological variables in BCs. Methods: STK3 mRNA expression was investigated in the METABRIC BC cohort (n=1980) and externally validated using online BC expression datasets [bc-GenExMiner v4.0]. STK3 protein expression was studied in a well characterised series of primary invasive BCs (n=1024) using immunohistochemistry including correlations with clinicopathological parameters, other biomarkers and patient outcome. Results: Copy number (CN) gain of STK3 was correlated with adverse prognostic features: higher grade and poor NPI (p<0.0001) High STK3 expression was also associated with poor prognostic factors, including high grade, younger age, larger tumour size, poorer NPI and negative ER/PR status (p<0.001). In PAM50 subtypes, high STK3 expression was associated with Luminal B/basal like tumours. Cytoplasmic STK3(c-STK3) protein expression was associated with increased mitotic index, poorer NPI (p<0.001) and basal-like markers CK5/6 and EGFR (p<0.05). In univariate analysis, high c-STK3 expression showed poorer outcome in the whole cohort and ER+ subgroups (p<0.05). Pooled STK3 gene expression data in the external validation cohort confirmed association with poor outcome (p<0.0001, HR = 1.60, 95% CI 1.28–2.01). Conclusions: Results suggest c-STK3 as a poor prognostic marker in invasive BC including ER+ subgroups warranting further functional studies

Cell division cycle 25C (CDC25C) expression confers poor prognosis in invasive breast cancer

Background: CDC25C, belonging to the Cdc25 phosphatase family, plays a major role in cell cycle control, impacting on DNA repair and apoptosis. It has been shown that poor prognosis/copy number high Luminal A breast cancers (BCs) are enriched for the Aurora kinase pathway including CDC25C leading to CDK1 activation (Ciriello et al, Breast Cancer Research Treatment, 2013:409). This study examined the associations of CDC25C with clinicopathological and molecular features in BCs including the low grade ER positive cohort. Methodology: CDC25C mRNA expression was studied in the METABRIC BC cohort (n=1980) and externally validated using online expression datasets [bc-GenExMiner v4.0]. CDC25C protein expression level was assessed immunohistochemically on a large annotated series of BC (n= 1330) and correlations made with clinicopathological parameters and patient outcome. Results: High CDC25C expression was significantly associated with poor prognostic factors including high grade, large tumour size, medullary like tumours, poorer NPI, ER-/PR- Her2+ status (p<0.001) and was differentially expressed in poor prognosis integrative clusters 5 and 10 (p<0.001). Cytoplasmic CDC25C (c-CDC25C) protein showed positive association with non-NST and non-medullary tumour subtypes while nuclear CDC25C (n-CDC25C) negatively associated with tumour stage (p<0.05). There was no association with ER, PR status, NPI and lymph nodes. However, high c-CDC25C resulted in poor survival at 20 years in the Grade 1 ER+ cohort (p=0.007), while high n-CDC25C showed better long term survival (p<0.001). Pooled CDC25C expression data in the external validation cohort showed an association with poor outcome (p<0.0001, HR = 1.45, 95 % CI 1.28—1.64). Conclusion: CDC25C appears to be associated with poor prognosis in BC including the Grade 1 ER+ cohort, indicating the importance of further functional analyses

Triple-negative breast cancer with brain metastases: a comparison between basal-like and non-basal-like biological subtypes

The aim of this study was to divide the group of triple-negative breast cancer patients with brain metastases into basal-like and non-basal-like biological subtypes in order to compare clinical features and survival rates in those two groups. A comprehensive analysis of 111 consecutive triple-negative breast cancer patients with brain metastases treated in the years 2003–2009 was performed. In 75 patients, immunohistochemistry was used as a surrogate of microarray in order to evaluate the expression of three basal markers: cytokeratin 5/6 (CK 5/6), EGFR/HER1 and c-KIT. The basal-like (ER/PgR/HER2-negative, CK5/6positive and/or HER1-positive) and non-basal-like (ER/PgR/HER2-negative, CK5/6-negative, HER1-negative) subsets were selected. Clinical features and survivals were compared in both groups. In the group of 111 triple-negative breast cancer patients, median DFS, OS and survival from brain metastases were 20, 29 and 4 months, respectively. In 75 patients who were evaluable for basal markers, median DFS, OS and survival from brain metastases were 18, 26 and 3.2 months, respectively. In the basal-like subtype, the survival rates were 15, 26 and 3 months, respectively, and in the non-basal-like subtypes, they were 20, 30 and 2.8 months, respectively. No statistically significant differences in survivals were detected between the basal-like and non-basal-like biological subtypes. Factors influencing survival from brain metastases were: Karnofsky performance status (KPS), the status of extracranial disease and age. Biological markers differentiating triple-negative group into basal-like and non-basal-like subtype (CK 5/6, HER1, c-KIT) had no influence on survival. In patients with triple-negative breast cancer and brain metastases, well-known clinical, but not molecular, features correlated with survival

Clinicopathological and prognostic significance of RECQL5 helicase expression in breast cancers

RECQL5 is a member of the RecQ family of DNA helicases and has key roles in homologous recombination, base excision repair, replication and transcription. The clinicopathological significance of RECQL5 expression in breast cancer is unknown. In the current study we have evaluated RECQL5 mRNA expression in 1977 breast cancers, and RECQL5 protein level in 1902 breast cancers [Nottingham Tenovus series (n=1650) and ER- cohort (n=252)]. Expression levels were correlated to aggressive phenotypes and survival outcomes. High RECQL5 mRNA expression was significantly associated with high histological grade (p=0.007), HER2 overexpression (p=0.032), ER+/HER2-/high proliferation genefu subtype, integrative molecular clusters (intClust 1and 9) and poor breast cancer specific survival (BCSS) (ps<0.0001). In sub-group analysis, high RECQL5 mRNA level remains significantly associated with poor BCSS in ER+ cohort (p<0.0001) but not in ER- cohort (p=0.116). At the protein level, in tumours with low RAD51, high RECQL5 level was significantly associated with high histological grade (p<0.0001), higher mitotic index (p=0.008), de-differentiation (p=0.025), pleomorphism (p=0.027) and poor BCSS (P=0.003). In sub-group analysis, high RECQL5/low RAD51 remains significantly associated with poor BCSS in ER+ cohort (p=0.010), but not in ER- cohort (p=0.628). In multivariate analysis, high RECQL5 mRNA and high RECQL5/low RAD51 nuclear protein co-expression independently influenced BCSS (p=0.022) in whole cohort and in the ER+ sub-group. Pre-clinically, we show that exogenous expression of RECQL5 in MCF10A cells can drive proliferation supporting an oncogenic function for RECQL5 in breast cancer. We conclude that RECQL5 is a promising biomarker in breast cancer