Effects of epidermal growth factor on MDA-MB-468 breast cancer cells: alterations in polyamine biosynthesis and the expression of p21/CIP1/WAF1.
We examined the effects of epidermal growth factor (EGF) on MDA-MB-468 cells to understand its mechanism of action in an EGF receptor-rich breast cancer cell line. EGF inhibited the growth of MDA-MB-468 cells with an IC50 of 1.5 +/- 0.5 nM, as determined by measurements of DNA content of cells in culture over a period of 4 to 6 days. This growth inhibition included apoptosis 24 h after EGF addition, as detected by an enzyme-linked immunosorbent assay (ELISA) and Hoechst 33342 staining. In EGF-treated cells, peak activities of two key enzymes of polyamine biosynthesis, ornithine decarboxylase (ODC) and S-adenosylmethionine decarboxylase (SAMDC), were reduced by 57% and 83%, respectively. EGF treatment also caused a 30 to 50% decrease in cellular putrescine at all time points tested (12 to 48 h). EGF-induced inhibition of DNA synthesis was also partially reversed by the addition of putrescine or spermidine, but not by spermine. Western blot analysis of cell cycle regulatory proteins showed that EGF-mediated growth inhibition was associated with the induction of p21, an inhibitor of cyclin-dependent kinases. However, EGF had no significant effect on the expression of cyclin D1 or cyclin E. Furthermore, putrescine reversal of EGF effects was associated with the down-regulation of EGF-induced p21. These results suggest that the mechanism of growth inhibition by EGF in MDA-MB-468 cells include a down-regulation of polyamine biosynthesis and the induction of p21. Identification of growth regulatory pathways in breast cancer cells might be useful in the development of novel targets for therapeutic intervention.
Thomas
T
T
Department of Environmental and Community Medicine, the Environmental and Occupational Health Sciences Institute and The Cancer Institute of New Jersey, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medica.
Balabhadrapathruni
S
S
Gardner
C R
CR
Hong
J
J
Faaland
C A
CA
Thomas
T J
TJ
D001562:Benzimidazoles; D001943:Breast Neoplasms; D002455:Cell Division / Q000187:drug effects; D019938:Cyclin D1 / Q000378:metabolism; D019927:Cyclin E / Q000378:metabolism; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000235:genetics* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000378:metabolism*; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D005456:Fluorescent Dyes; D015972:Gene Expression Regulation, Neoplastic; D006131:Growth Inhibitors / Q000494:pharmacology; D006801:Humans; D011073:Polyamines / Q000378:metabolism*; D014407:Tumor Cells, Cultured / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
D001562:Benzimidazoles; C493956:CDKN1A protein, human; D019927:Cyclin E; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D006131:Growth Inhibitors; D011073:Polyamines; D016159:Tumor Suppressor Protein p53; D019938:Cyclin D1; D004815:Epidermal Growth Factor; C017807:bisbenzimide ethoxide trihydrochloride