Molecular basis for the induction of an angiogenesis inhibitor, thrombospondin-1, by 5-fluorouracil.
5-Fluorouracil (5-FU) is one of the most commonly used anticancer drugs in chemotherapy against various solid tumors. 5-FU dose-dependently increased the expression levels of intrinsic antiangiogenic factor thrombospondin-1 (TSP-1) in human colon carcinoma KM12C cells and human breast cancer MCF7 cells. We investigated the molecular basis for the induction of TSP-1 by 5-FU in KM12C cells. Promoter assays showed that the region with the Egr-1 binding site is critical for the induction of TSP-1 promoter activity by 5-FU. The binding of Egr-1 to the TSP-1 promoter was increased in KM12C cells treated with 5-FU. Immunofluorescence staining revealed that 5-FU significantly increased the level of Egr-1 in the nuclei of KM12C cells. The suppression of Egr-1 expression by small interfering RNA decreased the expression level of TSP-1. Furthermore, 5-FU induced the phosphorylation of p38 mitogen-activated protein kinase (MAPK) and heat shock protein 27 (HSP27). Blockade of the p38 MAPK pathway by SB203580 remarkably inhibited the phosphorylation of HSP27 induced by 5-FU and decreased the induction of Egr-1 and TSP-1 by 5-FU in KM12C cells. These findings suggest that the p38 MAPK pathway plays a crucial role in the induction of Egr-1 by 5-FU and that induced Egr-1 augments TSP-1 promoter activity, with the subsequent production of TSP-1 mRNA and protein.
Zhao
Hong-Ye
HY
Department of Molecular Oncology, Graduate School of Medical and Dental Sciences, Kagoshima University, Sakuragaoka, Kagoshima, Japan.
Ooyama
Akio
A
Yamamoto
Masatatsu
M
Ikeda
Ryuji
R
Haraguchi
Misako
M
Tabata
Sho
S
Furukawa
Tatsuhiko
T
Che
Xiao-Fang
XF
Zhang
Shaoxuan
S
Oka
Toshinori
T
Fukushima
Masakazu
M
Nakagawa
Masayuki
M
Ono
Mayumi
M
Kuwano
Michihiko
M
Akiyama
Shin-ichi
S