Pleiotropic effects of FGFR1 on cell proliferation, survival, and migration in a 3D mammary epithelial cell model

Members of the fibroblast growth factor (FGF) family and the FGF receptors (FGFRs) have been implicated in mediating various aspects of mammary gland development and transformation. To elucidate the molecular mechanisms of FGFR1 action in a context that mimics polarized epithelial cells, we have developed an in vitro three-dimensional HC11 mouse mammary epithelial cell culture model expressing a drug-inducible FGFR1 (iFGFR1). Using this conditional model, iFGFR1 activation in these growth-arrested and polarized mammary acini initially led to reinitiation of cell proliferation, increased survival of luminal cells, and loss of cell polarity, resulting in the disruption of acinar structures characterized by the absence of an empty lumen. iFGFR1 activation also resulted in a gain of invasive properties and the induction of matrix metalloproteinase 3 (MMP-3), causing the cleavage of E-cadherin and increased expression of smooth muscle actin and vimentin. The addition of a pan MMP inhibitor abolished these phenotypes but did not prevent the effects of iFGFR1 on cell proliferation or survival

The Genome Sequence DataBase: towards an integrated functional genomics resource

During 1998 the primary focus of the Genome Sequence DataBase (GSDB; http://www.ncgr.org/gsdb ) located at the National Center for Genome Resources (NCGR) has been to improve data quality, improve data collections, and provide new methods and tools to access and analyze data. Data quality has been improved by extensive curation of certain data fields necessary for maintaining data collections and for using certain tools. Data quality has also been increased by improvements to the suite of programs that import data from the International Nucleotide Sequence Database Collaboration (IC). The Sequence Tag Alignment and Consensus Knowledgebase (STACK), a database of human expressed gene sequences developed by the South African National Bioinformatics Institute (SANBI), became available within the last year, allowing public access to this valuable resource of expressed sequences. Data access was improved by the addition of the Sequence Viewer, a platform-independent graphical viewer for GSDB sequence data. This tool has also been integrated with other searching and data retrieval tools. A BLAST homology search service was also made available, allowing researchers to search all of the data, including the unique data, that are available from GSDB. These improvements are designed to make GSDB more accessible to users, extend the rich searching capability already present in GSDB, and to facilitate the transition to an integrated system containing many different types of biological data

Resident macrophages influence stem cell activity in the mammary gland

Introduction Macrophages in the mammary gland are essential for morphogenesis of the ductal epithelial tree and have been implicated in promoting breast tumor metastasis. Although it is well established that macrophages influence normal mammopoiesis, the mammary cell types that these accessory cells influence have not been determined. Here we have explored a role for macrophages in regulating mammary stem cell (MaSC) activity, by assessing the ability of MaSCs to reconstitute a mammary gland in a macrophage-depleted fat pad. Methods Two different in vivo models were used to deplete macrophages from the mouse mammary fat pad, allowing us to examine the effect of macrophage deficiency on the mammary repopulating activity of MaSCs. Both the Csf1(op/op) mice and clodronate liposome-mediated ablation models entailed transplantation studies using the MaSC-enriched population. Results We show that mammary repopulating ability is severely compromised when the wild-type MaSC-enriched subpopulation is transplanted into Csf1(op/op) fat pads. In reciprocal experiments, the MaSC-enriched subpopulation from Csf1(op/op) glands had reduced regenerative capacity in a wildtype environment. Utilizing an alternative strategy for selective depletion of macrophages from the mammary gland, we demonstrate that co-implantation of the MaSC-enriched subpopulation with clodronate-liposomes leads to a marked decrease in repopulating frequency and outgrowth potential. Conclusions Our data reveal a key role for mammary gland macrophages in supporting stem/progenitor cell function and suggest that MaSCs require macrophage-derived factors to be fully functional. Macrophages may therefore constitute part of the mammary stem cell nich

Diverse Macrophage Populations Contribute to the Inflammatory Microenvironment in Premalignant Lesions During Localized Invasion.

Myeloid cell heterogeneity remains poorly studied in breast cancer, and particularly in premalignancy. Here, we used single cell RNA sequencing to characterize macrophage diversity in mouse pre-invasive lesions as compared to lesions undergoing localized invasion. Several subpopulations of macrophages with transcriptionally distinct profiles were identified, two of which resembled macrophages in the steady state. While all subpopulations expressed tumor-promoting genes, many of the populations expressed pro-inflammatory genes, differing from reports in tumor-associated macrophages. Gene profiles of the myeloid cells were similar between early and late stages of premalignancy, although expansion of some subpopulations occurred. These results unravel macrophage heterogeneity in early progression and may provide insight into early intervention strategies that target macrophages

Evaluation of the current knowledge limitations in breast cancer research: a gap analysis

BACKGROUND A gap analysis was conducted to determine which areas of breast cancer research, if targeted by researchers and funding bodies, could produce the greatest impact on patients. METHODS Fifty-six Breast Cancer Campaign grant holders and prominent UK breast cancer researchers participated in a gap analysis of current breast cancer research. Before, during and following the meeting, groups in seven key research areas participated in cycles of presentation, literature review and discussion. Summary papers were prepared by each group and collated into this position paper highlighting the research gaps, with recommendations for action. RESULTS Gaps were identified in all seven themes. General barriers to progress were lack of financial and practical resources, and poor collaboration between disciplines. Critical gaps in each theme included: (1) genetics (knowledge of genetic changes, their effects and interactions); (2) initiation of breast cancer (how developmental signalling pathways cause ductal elongation and branching at the cellular level and influence stem cell dynamics, and how their disruption initiates tumour formation); (3) progression of breast cancer (deciphering the intracellular and extracellular regulators of early progression, tumour growth, angiogenesis and metastasis); (4) therapies and targets (understanding who develops advanced disease); (5) disease markers (incorporating intelligent trial design into all studies to ensure new treatments are tested in patient groups stratified using biomarkers); (6) prevention (strategies to prevent oestrogen-receptor negative tumours and the long-term effects of chemoprevention for oestrogen-receptor positive tumours); (7) psychosocial aspects of cancer (the use of appropriate psychosocial interventions, and the personal impact of all stages of the disease among patients from a range of ethnic and demographic backgrounds). CONCLUSION Through recommendations to address these gaps with future research, the long-term benefits to patients will include: better estimation of risk in families with breast cancer and strategies to reduce risk; better prediction of drug response and patient prognosis; improved tailoring of treatments to patient subgroups and development of new therapeutic approaches; earlier initiation of treatment; more effective use of resources for screening populations; and an enhanced experience for people with or at risk of breast cancer and their families. The challenge to funding bodies and researchers in all disciplines is to focus on these gaps and to drive advances in knowledge into improvements in patient care

Key signalling nodes in mammary gland development and cancer. Signalling downstream of PI3 kinase in mammary epithelium: a play in 3 Akts

The protein serine/threonine kinase Akt, also known as protein kinase B (PKB), is arguably the most important signalling nexus in the cell. Akt integrates a plethora of extracellular signals to generate diverse outcomes, including proliferation, motility, growth, glucose homeostasis, survival, and cell death. The phosphatidylinositol 3-kinase (PI3K)/Akt pathway is the second most frequently mutated pathway in cancer, after p53, and mutations in components of this pathway are found in around 70% of breast cancers. Thus, understanding how Akt relays input signals to downstream effectors is critically important for the design of therapeutic strategies to combat breast cancer. In this review, we will discuss the various signals upstream of Akt that impact on its activity, how Akt integrates these signals and modulates the activity of downstream targets to control mammary gland development, and how mutations in components of the pathway result in breast cancer

Fibroblast Growth Factor Receptor Signaling Dramatically Accelerates Tumorigenesis and Enhances Oncoprotein Translation in the Mouse Mammary Tumor Virus-Wnt-1 Mouse Model of Breast Cancer

Fibroblast growth factor (FGF) cooperates with the Wnt/β-catenin pathway to promote mammary tumorigenesis. To investigate the mechanisms involved in FGF/Wnt cooperation, we genetically engineered a model of inducible FGF receptor (iFGFR) signaling in the context of the well-established mouse mammary tumor virus–Wnt-1 transgenic mouse. In the bigenic mice, iFGFR1 activation dramatically enhanced mammary tumorigenesis. Expression microarray analysis did not show transcriptional enhancement of Wnt/β-catenin target genes but instead showed a translational gene signature that also correlated with elevated FGFR1 and FGFR2 in human breast cancer data sets. Additionally, iFGFR1 activation enhanced recruitment of RNA to polysomes, resulting in a marked increase in protein expression of several different Wnt/β-catenin target genes. FGF pathway activation stimulated extracellular signal-regulated kinase and the phosphorylation of key translation regulators both in vivo in the mouse model and in vitro in a human breast cancer cell line. Our results suggest that cooperation of the FGF and Wnt pathways in mammary tumorigenesis is based on the activation of protein translational pathways that result in, but are not limited to, increased expression of Wnt/β-catenin target genes (at the level of protein translation). Further, they reveal protein translation initiation factors as potential therapeutic targets for human breast cancers with alterations in FGF signaling