title: The potent peptide antagonist to angiogenesis, C16Y, and cisplatin act synergistically in the down-regulation of the Bcl-2/Bax ratio and the induction of apoptosis in human ovarian cancer cells.
abstract: Cisplatin is one of the most potent antitumor agents for ovarian cancer, but has also been implicated in normal tissue cytotoxicity. We examined the effect of cisplatin alone and in combination with C16Y, a newly-identified anti-angiogenic peptide from the NH2-terminal domains of the γ-chain of laminin-1, on the modulation of Bcl-2/Bax expression and induction of apoptosis in ovarian cancer cells (OVACAR3). C16Y did not elicit cell death of human umbilical vein endothelial cells (HUVECs). Cisplatin exerted a lethal effect with an EC50 of 10 µM in OVACAR3s. In the presence of 25 or 50 µg/ml of C16Y (a range which has no effect against HUVECs), the EC50 for cisplatin in OVACAR3s decreased to 3.5 and 2.0 µM, respectively. Using fluorescence-activated cell sorting (FACS) analysis of DNA stained OVACAR3s and terminal deoxynucleotide tranferase-mediated dUTP nick end-labeling (TUNEL), we found that even at concentrations of 1 and 3 µM cisplatin, C16Y at 10 and 25 µg/ml increased the incidence of apoptosis in OVACAR3s by 3-5-fold. Each drug had some measurable effect on Bax protein expression. Furthermore, Bcl-2 protein expression levels were markedly reduced by C16Y alone and cisplatin alone in a dose-dependent manner. The combination of C16Y and cisplatin resulted in a further dramatic reduction in Bcl-2, underscoring the pronounced synergy produced by cisplatin and C16Y together. On the other hand, C16Y did not activate any other signal transduction pathways that usually culminate in the activation of apoptosis, such as the p53, p21waf1, p73, ERK1/2 or PI3-AKT pathways. These observations suggest that the suppression of the Bcl-2/Bax ratio may play an important role in mediating the synergistic effect of cisplatin and C16Y on the induction of apoptosis in OVACAR3 cells.
authors.affiliation: Department of Gynecology, Faculty of Medical Sciences, University of Fukui, Fukui, Japan.
authors.forename: Akiko
authors.initials: A
authors.lastname: Shinagawa
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Yoshida
authors.forename: Tetsuji
authors.initials: T
authors.lastname: Kurokawa
authors.forename: Yuko
authors.initials: Y
authors.lastname: Horiuchi
authors.forename: Hideaki
authors.initials: H
authors.lastname: Tsuyoshi
authors.forename: Makoto
authors.initials: M
authors.lastname: Orisaka
authors.forename: Yoko
authors.initials: Y
authors.lastname: Sawamura
authors.forename: Hynda K
authors.initials: HK
authors.lastname: Kleinman
authors.forename: Fumikazu
authors.initials: F
authors.lastname: Kotsuji
mesh_terms: D020533:Angiogenesis Inhibitors / Q000494:pharmacology*; D000818:Animals; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects* / Q000235:genetics; D049109:Cell Proliferation / Q000187:drug effects; D002945:Cisplatin / Q000494:pharmacology*; D015536:Down-Regulation / Q000187:drug effects; D004357:Drug Synergism; D042783:Endothelial Cells / Q000187:drug effects; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D051379:Mice; D041681:NIH 3T3 Cells; D009842:Oligopeptides / Q000494:pharmacology*; D010051:Ovarian Neoplasms / Q000235:genetics / Q000378:metabolism*; D010455:Peptides / Q000494:pharmacology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D051028:bcl-2-Associated X Protein / Q000378:metabolism*
chemical_list: D020533:Angiogenesis Inhibitors; D009842:Oligopeptides; D010455:Peptides; D019253:Proto-Oncogene Proteins c-bcl-2; C569383:aspartyl-phenylalanyl-lysyl-leucyl-phenylalanyl-alanyl-valyl-tyrosyl-isoleucyl-lysyl-tyrosyl-arginine; D051028:bcl-2-Associated X Protein; D002945:Cisplatin