Chromosomal instability and lack of cyclin E regulation in hCdc4 mutant human breast cancer cells

INTRODUCTION: Cyclin E, a G(1 )cyclin essential for G(1)–S phase transition, is known to have a profound effect on tumorigenesis. Elevated levels of cyclin E have been associated with breast cancer, and chromosomal instability observed in breast cancer is suggested to be associated with constitutive expression of cyclin E. It was previously demonstrated that SUM149PT human breast cancer cells show very high levels of cyclin E expression by western analysis and that they express a nonfunctional cyclin E ubiquitin ligase due to a mutation in the F-box protein hCdc4. METHODS: We examined cyclin E expression in both MCF10A and SUM149PT cells using western blot analysis and flow cytometry. Immunofluorescence was utilized for the localization of cyclin E in both normal and breast cancer cells. In addition, array comparative genomic hybridization analysis was performed to compare chromosome copy number alterations with levels of cyclin E expression among a panel of breast cancer cell lines. RESULTS: SUM149PT cells overexpress cyclin E on a cell per cell basis for the duration of the cell cycle. High cyclin E levels are maintained throughout the S phase, and SUM149PT cells exhibit an S phase delay or arrest probably due to cyclin E overexpression. In addition, comparative genomic hybridization indicated that SUM149PT cells exhibit many chromosome copy number alterations, which may reflect prior or ongoing genomic instability. However, no direct correlation was observed between cyclin E levels and genomic copy number alteration in a panel of human breast cancer cell lines. CONCLUSIONS: Cyclin E is overexpressed at high levels throughout the cell cycle in SUM149PT cells, which is in stark contrast to cyclin E degradation observed in the mid to late S phase of normal cells. SUM149PT cells are unable to regulate cyclin E and also exhibit many copy number alterations. However, there was a lack of direct correlation between cyclin E overexpression and chromosomal instability across a panel of other breast cancer cell lines examined

The influence of growth factors on the proliferative potential of normal and primary breast cancer-derived human breast epithelial cells

In previous studies, we developed serum-free, bovine pituitary extract (BPE)-free culture conditions for the growth of normal and neoplastic rat mammary epithelial cells. The present studies were aimed at determining if these culture methods could be used to study the influence of specific growth factors on the proliferative potential of normal human mammary epithelial (HME) cells and cells derived from human breast cancer (HBC) specimens. Our results indicate that normal HME cells in primary culture express stringent requirements for insulin (IN), epidermal growth factor (EGF), and cholera toxin (CT). Of these factors, EGF is most important, with essentially no proliferation taking place in the absence of this factor. By contrast, when cells are grown in serum-free primary culture in the presence of a full complement of growth factors and then subcultured, growth in secondary culture is not influenced by the removal of individual growth factors. Growth in secondary culture in the absence of EGF is mediated by autocrine factors secreted by the cells. However, there is no evidence for autocrine activity that mediates growth in the absence of IN in secondary cultures. Primary culture of HBC cells in serum-free, BPE-free medium revealed two patterns of growth factor requirements. One set of HBC cells expressed identical requirements for IN and EGF in primary culture as normal cells. Likewise, these cells grew in secondary culture in the absence of either factor. The second set of tumors expressed independence of IN for growth in primary culture. These cells grew to confluence in primary culture in the absence of IN and could be subcultured in this medium. All tumor cells examined expressed a requirement for EGF for primary culture growth, whereas none of the HBC cells examined expressed a significant CT requirement. In many cases, growth in the absence of CT exceeded that observed in its presence. Thus, our culture system allows analysis of the growth factor requirements of HME and HBC cells in primary culture. Our results indicate significant differences between HME and HBC cells in this regard. However, the results of secondary culture experiments indicate that the growth factor milieu from which cells are taken can have a profound effect on the requirements for growth factors in culture.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/44196/1/10549_2005_Article_BF01806371.pd

Selecting Indicator Portfolios for Marine Species and Food Webs: A Puget Sound Case Study

Ecosystem-based management (EBM) has emerged as a promising approach for maintaining the benefits humans want and need from the ocean, yet concrete approaches for implementing EBM remain scarce. A key challenge lies in the development of indicators that can provide useful information on ecosystem status and trends, and assess progress towards management goals. In this paper, we describe a generalized framework for the methodical and transparent selection of ecosystem indicators. We apply the framework to the second largest estuary in the United States – Puget Sound, Washington – where one of the most advanced EBM processes is currently underway. Rather than introduce a new method, this paper integrates a variety of familiar approaches into one step-by-step approach that will lead to more consistent and reliable reporting on ecosystem condition. Importantly, we demonstrate how a framework linking indicators to policy goals, as well as a clearly defined indicator evaluation and scoring process, can result in a portfolio of useful and complementary indicators based on the needs of different users (e.g., policy makers and scientists). Although the set of indicators described in this paper is specific to marine species and food webs, we provide a general approach that could be applied to any set of management objectives or ecological system

Choosing the right cell line for breast cancer research

Breast cancer is a complex and heterogeneous disease. Gene expression profiling has contributed significantly to our understanding of this heterogeneity at a molecular level, refining taxonomy based on simple measures such as histological type, tumour grade, lymph node status and the presence of predictive markers like oestrogen receptor and human epidermal growth factor receptor 2 (HER2) to a more sophisticated classification comprising luminal A, luminal B, basal-like, HER2-positive and normal subgroups. In the laboratory, breast cancer is often modelled using established cell lines. In the present review we discuss some of the issues surrounding the use of breast cancer cell lines as experimental models, in light of these revised clinical classifications, and put forward suggestions for improving their use in translational breast cancer research

The impact of surgical delay on resectability of colorectal cancer: An international prospective cohort study

AIM: The SARS-CoV-2 pandemic has provided a unique opportunity to explore the impact of surgical delays on cancer resectability. This study aimed to compare resectability for colorectal cancer patients undergoing delayed versus non-delayed surgery. METHODS: This was an international prospective cohort study of consecutive colorectal cancer patients with a decision for curative surgery (January-April 2020). Surgical delay was defined as an operation taking place more than 4 weeks after treatment decision, in a patient who did not receive neoadjuvant therapy. A subgroup analysis explored the effects of delay in elective patients only. The impact of longer delays was explored in a sensitivity analysis. The primary outcome was complete resection, defined as curative resection with an R0 margin. RESULTS: Overall, 5453 patients from 304 hospitals in 47 countries were included, of whom 6.6% (358/5453) did not receive their planned operation. Of the 4304 operated patients without neoadjuvant therapy, 40.5% (1744/4304) were delayed beyond 4 weeks. Delayed patients were more likely to be older, men, more comorbid, have higher body mass index and have rectal cancer and early stage disease. Delayed patients had higher unadjusted rates of complete resection (93.7% vs. 91.9%, P = 0.032) and lower rates of emergency surgery (4.5% vs. 22.5%, P < 0.001). After adjustment, delay was not associated with a lower rate of complete resection (OR 1.18, 95% CI 0.90-1.55, P = 0.224), which was consistent in elective patients only (OR 0.94, 95% CI 0.69-1.27, P = 0.672). Longer delays were not associated with poorer outcomes. CONCLUSION: One in 15 colorectal cancer patients did not receive their planned operation during the first wave of COVID-19. Surgical delay did not appear to compromise resectability, raising the hypothesis that any reduction in long-term survival attributable to delays is likely to be due to micro-metastatic disease