title: Epidermal growth factor receptor signaling cascade as target for tyrphostin (RG 50864) in epithelial cells. Paradoxical effects on mitogen-activated protein kinase kinase and mitogen-activated protein kinase activities.
abstract: Tyrphostins are synthetic compounds that have been described as in vitro inhibitors of epidermal growth factor receptor (EGF-R) tyrosine kinase activity. The inhibitory effect of tyrphostins in intact cells has been shown only after prolonged treatment. However, these compounds appear to be readily incorporated, which suggests that tyrphostin acts indirectly on EGF-R. We studied the effects of a tyrphostin derivative, RG 50864, without preincubation in intact epithelial cells. We selected two human cell lines differing in degree of expression of the p185erbB2 protein, which is closely related to EGF-R. We showed that tyrphostin (RG 50864) had no effect on EGF-dependent EGF-R tyrosine phosphorylation in the parental cell line. On the contrary, it prolonged the EGF-dependent EGF-R and p185erbB2(V-E) tyrosine phosphorylation in p185erbB2(V-E)-expressing cells. Because tyrphostin has been shown to be an inhibitor of p185erbB2 and EGF-R in vitro, this finding indicates that the tyrphostin effect on p185erbB2(V-E) and EGF-R was the result of an indirect mechanism in transfected cells. Tyrphostin treatment alone led to the activation of mitogen-activated protein (MAP) kinase kinase or MAP kinase or extracellular signal-regulated kinase kinase (MEK), suggesting that one of the tyrphostin targets was upstream of MEK1. MAP kinase, however, was not activated after tyrphostin treatment. This finding indicates that tyrphostin had another target in intact cells because MEK1 activation by tyrphostin alone did not correlate with MAP kinase activation. In the two cell lines, tyrphostin modified the time course of EGF-dependent MEK and MAP kinase activation. We conclude that whereas tyrphostins were designed to inhibit EGF-R tyrosine kinase activity, under our conditions EGF-R is not a physiological target for tyrphostin, nor is one of its related protein tyrosine kinases, p185erbB2(V-E). On the contrary, our results show that tyrphostin targets are multiple, leading to complex effects on receptor signaling in these epithelial cells.
authors.affiliation: INSERM CJF 93-01, Faculté de Pharmacie, Châtenay Malabry, France.
authors.forename: F
authors.initials: F
authors.lastname: Nowak
authors.forename: A
authors.initials: A
authors.lastname: Jacquemin-Sablon
authors.forename: J
authors.initials: J
authors.lastname: Pierre
mesh_terms: D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000187:drug effects; D002396:Catechols / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004815:Epidermal Growth Factor / Q000494:pharmacology; D066246:ErbB Receptors / Q000037:antagonists & inhibitors*; D006801:Humans; D048369:MAP Kinase Kinase 1; D020929:Mitogen-Activated Protein Kinase Kinases; D009570:Nitriles / Q000494:pharmacology*; D017346:Protein Serine-Threonine Kinases / Q000187:drug effects; D011505:Protein-Tyrosine Kinases / Q000187:drug effects; D014407:Tumor Cells, Cultured; D020032:Tyrphostins
chemical_list: D002396:Catechols; D004791:Enzyme Inhibitors; D009570:Nitriles; D020032:Tyrphostins; C060152:tyrphostin 47; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D048369:MAP Kinase Kinase 1; C482125:MAP2K1 protein, human; D020929:Mitogen-Activated Protein Kinase Kinases