Inhibition of Mek 1/2 kinase activity and stimulation of melanogenesis by 5,7-dimethoxycoumarin treatment of melanoma cells.
In this study, the processes of differentiation and melanogenesis induced by 5,7-dimethoxycoumarin in murine (B16) and human (A375) melanoma cells were investigated. Taking into account the previously demonstrated antiproliferative and differentiation activities of this compound, we examined Ras/Raf/Mek/Erk mitogen-activated protein kinase activity following treatment; inhibition of Mek 1/2 kinase activity and subsequent reduction in Erk 1/2 activation were detected in both cell types. We observed melanogenesis induction associated to an increase in cAMP-response element-binding protein (CREB) and microphthalmia-associated transcription factor (Mitf) expression, both involved in its regulation. Mitf is fundamental for development, survival and differentiation of melanocyte and melanoma, since it regulates transcription of genes encoding for proteins involved in cell cycle progression or in melanogenesis, such as the enzyme tyrosinase. A significant increase of tyrosinase activity was revealed following treatment in B16 but not in A375 cells, although a strong synthesis of melanin was induced by 5,7-dimethoxycoumarin in both cell lines. The treatment induced protoporphyrine IX accumulation involved in melanogenesis since it promotes stability of cAMP. Finally, the Mek 1/2 inhibitor U0126 significantly potentiated growth inhibition of B16 cells triggered by 5,7-dimethoxycoumarin, suggesting that down-regulation of Raf/Mek/Erk pathway sensitizes melanoma cells to 5,7-dimethoxycoumarin treatment.
Department of Biology, University of Rome 'Tor Vergata', I-00133 Rome, Italy.
Daniela
D
Alesiani
Rosella
R
Cicconi
Maurizio
M
Mattei
Roberto
R
Bei
Antonella
A
Canini
D000818:Animals; D015153:Blotting, Western; D002070:Butadienes / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects; D003374:Coumarins / Q000494:pharmacology*; D000242:Cyclic AMP / Q000378:metabolism; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D047468:Immunoprecipitation; D048369:MAP Kinase Kinase 1 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D048370:MAP Kinase Kinase 2 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D008543:Melanins / Q000378:metabolism; D008545:Melanoma / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology*; D051379:Mice; D051739:Microphthalmia-Associated Transcription Factor / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D009570:Nitriles / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects*; D014407:Tumor Cells, Cultured; D048490:raf Kinases / Q000378:metabolism; D018631:ras Proteins / Q000378:metabolism
D002070:Butadienes; D003374:Coumarins; D017362:Cyclic AMP Response Element-Binding Protein; D004791:Enzyme Inhibitors; D008543:Melanins; D051739:Microphthalmia-Associated Transcription Factor; D009570:Nitriles; C113580:U 0126; D000242:Cyclic AMP; D048490:raf Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; D018631:ras Proteins; C037446:5,7-dimethoxycoumarin