Effect of alpha-difluoromethyl-ornithine on the expression and function of the epidermal growth factor receptor in human breast epithelial cells in culture.
We have previously shown that ornithine decarboxylase (ODC) overexpression enhances the transforming effects of HER-2neu and epidermal growth factor (EGF) in normal MCF-10A human breast epithelial cells. Our data suggest that such potentiation may be mediated by activation of the mitogen-activated protein kinase (MAPK) pathway and, possibly, STAT signalling. To further explore the interaction between the polyamine pathway and EGF/HER-2neu signalling in this system, we inhibited endogenous ODC activity with alpha-difluoromethylornithine (DFMO) and assessed the effects of this blockade on the expression of EGF receptors (EGFR) and HER-2neu as well as activation of downstream EGF target genes. We found that DFMO administration to MCF-10A cells increased EGF-R mRNA and protein levels in a dose-response fashion, while HER-2neu expression was not affected. The effect of DFMO was mediated through polyamine depletion since it could be reversed by exogenous putrescine administration. Our results also indicated that the increase in EGFR induced by DFMO was not a non-specific consequence of inhibition of cell proliferation. The upregulated EGFRs were functional since they could be phosphorylated by EGF and they were able to promote phosphorylation of downstream signalling molecules including ERK, STAT-3, and STAT-5. We propose that physiologic levels of ODC activity may be critical for regulation of a yet undefined signalling pathway, whose blockade by DFMO leads to a compensatory increase in functional EGFR.
Manni
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A
Department of Medicine, Pennsylvania State University College of Medicine, Hershey, USA. amanni@psu.edu
Trout
D
D
Verderame
M F
MF
Washington
S
S
Mauger
D
D
Demers
L
L
D000970:Antineoplastic Agents / Q000494:pharmacology*; D015152:Blotting, Northern; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000378:metabolism* / Q000473:pathology; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D000518:Eflornithine / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000378:metabolism / Q000473:pathology; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006918:Hydroxyurea / Q000494:pharmacology; D015739:Nocodazole / Q000494:pharmacology; D009955:Ornithine Decarboxylase / Q000378:metabolism; D065108:Ornithine Decarboxylase Inhibitors; D011073:Polyamines / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism
D000970:Antineoplastic Agents; D000518:Eflornithine; D004791:Enzyme Inhibitors; D066246:ErbB Receptors; D006918:Hydroxyurea; D015739:Nocodazole; D009955:Ornithine Decarboxylase; D065108:Ornithine Decarboxylase Inhibitors; D011073:Polyamines; D017346:Protein Serine-Threonine Kinases; D018719:Erb-b2 Receptor Tyrosine Kinases