Growth inhibition of colon cancer cells by transfection of dominant-negative apoptosis signal-regulating kinase-1.
Apoptosis signal-regulating kinase-1 (ASK-1) is an important molecule for the pro-apoptotic signaling. ASK-1 also contributes to the cellular survival for many types of cells. Thus, ASK-1 has a broad range of biological activities depending on the cell type. The present study assessed the role(s) of ASK-1 in colorectal cancer cells (HT-29) by using adenovirus vectors expressing wild-type (WT)-ASK-1 or dominant-negative (DN) mutant of ASK-1 and recombinant adenovirus containing the bacterial beta-galactosidase gene (Ad-LacZ), a negative control for Ad-DN-ASK-1. Selective phosphorylation of ASK-1 at Thr 845, a kinase domain site, but not Ser 83 nor 967 sites was induced by serum stimulation in a time-dependent manner. Transfection with Ad-DN-ASK-1 inhibited the serum-induced phosphorylation of p38 mitogen-activated protein kinase, a downstream molecule of ASK-1. Transfection with Ad-DN-ASK-1 diminished the serum-induced cell proliferation in a dose-dependent manner, whereas WT-ASK-1 increased it. Apoptosis assessed by Hoechst staining was induced in the Ad-DN-ASK-1 treated cells. In vivo transfection of Ad-DN-ASK-1 into tumor xenografts of HT-29 cells in nude mice significantly decreased the tumor volume on day 29. Cleaved caspase-3 was found in the tumors of DN-ASK-1 treated mice. We obtained the first evidence that DN-ASK-1 transfection exerted significant antitumor effects on colon cancer mediated by apoptosis.
Kuwamura
Hikaru
H
Department of Gastroenterology, Osaka City University, Graduate School of Medicine, Osaka 545-8585, Japan.
Tominaga
Kazunari
K
Shiota
Masayuki
M
Ashida
Reiko
R
Nakao
Takafumi
T
Sasaki
Eiji
E
Watanabe
Toshio
T
Fujiwara
Yasuhiro
Y
Oshitani
Nobuhide
N
Higuchi
Kazuhide
K
Ichijo
Hidenori
H
Arakawa
Tetsuo
T
Iwao
Hiroshi
H
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D053148:Caspase 3 / Q000378:metabolism; D003110:Colonic Neoplasms / Q000628:therapy*; D005799:Genes, Dominant; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D048848:MAP Kinase Kinase Kinase 5 / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D008815:Mice, Inbred Strains; D009154:Mutation; D010766:Phosphorylation; D014162:Transfection; D023041:Xenograft Model Antitumor Assays; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D048051:p38 Mitogen-Activated Protein Kinases; D048848:MAP Kinase Kinase Kinase 5; D053148:Caspase 3