Effect of alpha-difluoromethyl-ornithine on the expression and function ofthe 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 p rotein kinase (MAPK) pathway and, possibly, STAT signalling. To further exp lore the interaction between the polyamine pathway and EGF/HER-2neu signall ing in this system, we inhibited endogenous ODC activity with alpha -difluo romethylornithine (DFMO) and assessed the effects of this blockade on the e xpression of EGF receptors (EGFR) and HER-2neu as well as activation of dow nstream EGF target genes. We found that DFMO administration to MCF-10A cell s 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 thro ugh polyamine depletion since it could be reversed by exogenous putrescine administration. Our results also indicated that the increase in EGFR induce d by DFMO was not a non-specific consequence of inhibition of cell prolifer ation. The upregulated EGFRs were functional since they could be phosphoryl ated by EGF and they were able to promote phosphorylation of downstream sig nalling molecules including ERK, STAT-3, and STAT-5. We propose that physio logic levels of ODC activity may be critical for regulation of a yet undefi ned signalling pathway, whose blockade by DFMO leads to a compensatory incr ease in functional EGFR.