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Title: Transcriptional changes associated with breast cancer occur as normal human mammary epithelial cells overcome senescence barriers and become immortalized

Journal Article · · Molecular Cancer
 [1];  [2];  [1];  [2];  [3];  [3];  [1];  [3];  [3];  [4];  [2]
  1. Wyeth Research, Cambridge, MA (United States). Dept. of Biological Technologies. Section of Bioinformatics
  2. Wyeth Research, Cambridge, MA (United States). Dept. of Biological Technologies. Applied Genomics
  3. Wyeth Research, Cambridge, MA (United States). Dept. of Biological Technologies. Molecular Profiling and Biomarker Discovery
  4. Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Life Sciences Division
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Background: Human mammary epithelial cells (HMEC) overcome two well-characterized genetic and epigenetic barriers as they progress from primary cells to fully immortalized cell lines in vitro. Finite lifespan HMEC overcome an Rb-mediated stress-associated senescence barrier (stasis), and a stringent, telomere-length dependent, barrier (agonescence or crisis, depending on p53 status). HMEC that have overcome the second senescence barrier are immortalized. Methods: We have characterized pre-stasis, post-selection (post-stasis, with p16 silenced), and fully immortalized HMEC by transcription profiling and RT-PCR. Four pre-stasis and seven post-selection HMEC samples, along with 10 representatives of fully immortalized breast epithelial cell lines, were profiled using Affymetrix U133A/B chips and compared using both supervised and unsupervised clustering. Datasets were validated by RT-PCR for a select set of genes. Quantitative immunofluorescence was used to assess changes in transcriptional regulators associated with the gene expression changes. Results: The most dramatic and uniform changes we observed were in a set of about 30 genes that are characterized as a "cancer proliferation cluster," which includes genes expressed during mitosis (CDC2, CDC25, MCM2, PLK1) and following DNA damage. The increased expression of these genes was particularly concordant in the fully immortalized lines. Additional changes were observed in IFN-regulated genes in some post-selection and fully immortalized cultures. Nuclear localization was observed for several transcriptional regulators associated with expression of these genes in post-selection and immortalized HMEC, including Rb, Myc, BRCA1, HDAC3 and SP1. Conclusion: Gene expression profiles and cytological changes in related transcriptional regulators indicate that immortalized HMEC resemble non-invasive breast cancers, such as ductal and lobular carcinomas in situ, and are strikingly distinct from finite-lifespan HMEC, particularly with regard to genes involved in proliferation, cell cycle regulation, chromosome structure and the DNA damage response. The comparison of HMEC profiles with lines harboring oncogenic changes (e.g. overexpression of Her-2neu, loss of p53 expression) identifies genes involved in tissue remodeling as well as proinflamatory cytokines and S100 proteins. Studies on carcinogenesis using immortalized cell lines as starting points or "normal" controls need to account for the significant pre-existing genetic and epigenetic changes inherent in such lines before results can be broadly interpreted.

Research Organization:
Univ. of California, Berkeley, CA (United States)
Sponsoring Organization:
USDOE Office of Science (SC), Biological and Environmental Research (BER). Biological Systems Science Division; US Dept. of Defense (DOD)
Grant/Contract Number:
AC03-76SF00098; W81XWH-04-1-0580
OSTI ID:
1626574
Journal Information:
Molecular Cancer, Vol. 6, Issue 1; ISSN 1476-4598
Publisher:
BioMed CentralCopyright Statement
Country of Publication:
United States
Language:
English

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Synthesis, characterization, crystal structure and cytotoxicity of 2,4-bis(selenomethyl)quinazoline journal June 2011
Analysis of the ways and methods of signaling pathways in regulating cell cycle of NIH3T3 at transcriptional level journal October 2015
Different culture media modulate growth, heterogeneity, and senescence in human mammary epithelial cell cultures journal October 2018
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Single cell cytometry of protein function in RNAi treated cells and in native populations journal August 2008
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