5-Fluorouracil induces apoptosis in human colon cancer cell lines with modulation of Bcl-2 family proteins.
Recently, apoptosis has been implicated as one of the end points of cells exposed to chemotherapeutic agents. The p53 and Bcl-2 family of proteins are involved in chemotherapy-induced apoptosis, but in a cell type-dependent manner. We sought to determine the roles played by the p53 and Bcl-2 family of proteins in 5-fluorouracil (5-FU)-induced apoptosis of human colon cancer cell lines. We first studied the p53 genetic and functional status, and then 5-FU, at inhibitory concentration of 50% (IC50) doses, was used to induce apoptosis, which was confirmed by morphological analysis and enzyme-linked immunosorbent assay (ELISA). Bcl-2, Bcl-X(L), Bax, Bad, Bak and p53 protein expression was analysed by Western blotting. Using five human colon cancer cell lines, we found that equitoxic (IC50) doses of 5-FU induced apoptosis in both wild-type p53 and mutant p53 cells. Analysis of the steady-state levels of Bcl-2 family proteins showed high expression of Bcl-X(L) in all of the cell lines except Colo320. Bcl-2 was expressed in two of them. Bax presented with the lowest basal expression and Bad showed homogeneous expression. On the other hand, Bak expression varied more than fivefold among these cells. In cells containing wild-type p53 (e.g. LoVo), 5-FU-induced apoptosis was accompanied by increased expression of Bax and Bak without consistent modulation of other bcl-2 family proteins. In contrast in cells containing mutant p53 (e.g. DLD1), Bak expression was remarkably increased. There was a significant correlation between chemosensitivity and Bcl-X(L) to Bax ratio, rather than Bcl-2 to Bax. In conclusion, these results suggest that some members of the Bcl-2 family of proteins, in human colon cancer cell lines, are modulated by 5-FU and that the ratio of Bcl-X(L) to Bax may be related to chemosensitivity to 5-FU.
Nita
M E
ME
Department of Surgical Oncology, Graduate School of Medical Sciences, Faculty of Medicine, The University of Tokyo, Japan.
Nagawa
H
H
Tominaga
O
O
Tsuno
N
N
Fujii
S
S
Sasaki
S
S
Fu
C G
CG
Takenoue
T
T
Tsuruo
T
T
Muto
T
T
D017209:Apoptosis / Q000187:drug effects; D001483:Base Sequence; D003110:Colonic Neoplasms / Q000378:metabolism* / Q000473:pathology*; D019008:Drug Resistance, Neoplasm; D005472:Fluorouracil / Q000494:pharmacology*; D006801:Humans; D016133:Polymerase Chain Reaction; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein
C494826:BAX protein, human; C494811:BCL2L1 protein, human; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D016159:Tumor Suppressor Protein p53; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; D005472:Fluorouracil