Cyclooxygenase-2 expression induces genomic instability in MCF10A breast epithelial cells

J Surg Res. 2007 Jun 15;140(2):220-6. doi: 10.1016/j.jss.2007.01.039. Epub 2007 Apr 9.

Abstract

Background: Cyclooxygenase-2 (COX-2) is induced in many breast cancers and COX-2 expression correlates with a worse outcome in the clinic. We hypothesized that the induction of genomic instability is a major mechanism through which COX-2 contributes to breast cancer progression.

Methods: We transfected a normal immortalized breast epithelial cell line of Basal B subtype, MCF10A, with the pSG5-COX-2 vector and established the stably transfected cell line MCF10A/COX-2. We analyzed the genomic instability phenotype by chromosomal analysis of metaphase-arrested MCF10A and MCF10A/COX-2 cells after Giemsa staining. Groups were compared using chi(2) tests. To investigate the DNA damage checkpoint signaling, we analyzed the phosphorylation status of CHK1 protein with a phospho-specific antibody.

Results: Cytogenetic analysis of early passage transfected cells showed that COX-2 expression increased genomic instability compared with the MCF10A cells transfected with a luciferase vector alone. COX-2 overexpression was associated with a significant increase in chromosomal aberrations (fusions, breaks, and tetraploidy). There was a statistically significant increase in the number of polyploid cells in the COX-2 transfected cells versus the control (P=0.004). We also found that an inhibitory CHK1 phosphorylation at Ser-280 was dramatically increased upon COX-2 overexpression in MCF10A cells, thus explaining the mechanism of inactivation of an important cell cycle checkpoint. Further analysis of the MCF10A/COX-2 cells showed that these cells have acquired a premalignant phenotype characterized by a morphological transformation, a resistance to anoikis, a reduced requirement of epidermal growth factor for growth in culture, but their inability to establish tumors in a nude mouse model of malignancy.

Conclusion: We found that COX-2 expression in MCF10A breast epithelial cells confers a premalignant phenotype that includes enhanced genomic instability and altered cell-cycle regulation.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Anoikis / genetics
  • Breast / cytology
  • Breast / enzymology*
  • Breast Neoplasms / enzymology
  • Breast Neoplasms / genetics
  • Breast Neoplasms / pathology
  • Cell Cycle
  • Cell Line
  • Checkpoint Kinase 1
  • Cyclooxygenase 2 / genetics
  • Cyclooxygenase 2 / metabolism*
  • DNA Damage
  • Disease Progression
  • Epidermal Growth Factor / physiology
  • Epithelial Cells / cytology
  • Epithelial Cells / enzymology*
  • Female
  • Gene Expression Regulation, Enzymologic
  • Genomic Instability / genetics*
  • Humans
  • Mice
  • Mice, Nude
  • Phenotype
  • Precancerous Conditions / enzymology
  • Precancerous Conditions / genetics
  • Precancerous Conditions / pathology
  • Protein Kinases / genetics
  • Protein Kinases / metabolism
  • Transfection
  • Transplantation, Heterologous

Substances

  • Epidermal Growth Factor
  • Cyclooxygenase 2
  • Protein Kinases
  • CHEK1 protein, human
  • Checkpoint Kinase 1
  • Chek1 protein, mouse