TY - JOUR
T1 - Characterisation of high-level cisplatin-resistant cell lines established from a human hepatoma cell line and human KB adenocarcinoma cells
T2 - Cross-resistance and protein changes
AU - Shen, D. W.
AU - Akiyama, S. I.
AU - Akiyama, S. I.
AU - Schoenlein, Patricia V
AU - Gottesman, M. M.
PY - 1995/4
Y1 - 1995/4
N2 - Human liver carcinoma cells (BEL-7404) and human KB adenocarcinoma cells were selected by stepwise increases in cisplatin. Drug sensitivity assays indicated that the IC50, value for 7404-CP7.5 cells was 49 μg ml-1 cisplatin, 111-fold higher than for the parental hepatoma cells. The IC50 value for KB-CP10 cells was 38 μg ml-1 cisplatin, which is 1152-fold higher than for the parental KB cells. The 7404-CP7.5 cells were cross-resistant to methotrexate (39 x), 5-fluorouracil (23 x) and 6-mercaptopurine (13 x), but were sensitive to drugs which are known substrates for the multidrug transporter (P-glycoprotein), including colchicine, vinblastine and actinomycin D. Similar cross-resistance patterns were observed for KB-CPIO cells. No evidence of DNA amplification or expression of the MDRI gene was found. One-dimensional sodium dodecyl sulphate-polyacrylamide gel electrophoresis showed increases in 52 kDa protein(s) in both the soluble cytosolic and crude membrane fractions in 7404-Cr cells and in KB-CPF cells. The amount of 52 kDa protein was proportional to the degree of resistance of the 7404-CP cells to cisplatin. Two-dimensional gel analysis demonstrated that two polypeptides of molecular mass 52 and 50 kDa were overexpressed in the membrane fractions in both 7404-CP20 and KB-CP20 cells. Using amino acid microsequencing and Western blotting, the major 52 kDa protein was identified as the mitochondrial heat shock protein hsp60. Two-dimensional gels of [35S]methionine-labelled polypeptides showed many other changes, including reduction in soluble proteins of approximately 57 kDa molecular weight in KB-CP20 cells, and of 35 kDa in both 7404-CP20 and KB-CP20 cells. These results suggest that alterations of certain proteins occur commonly in cisplatin-resistant cells, particularly proteins of molecular weight 52 and 50 kDa.
AB - Human liver carcinoma cells (BEL-7404) and human KB adenocarcinoma cells were selected by stepwise increases in cisplatin. Drug sensitivity assays indicated that the IC50, value for 7404-CP7.5 cells was 49 μg ml-1 cisplatin, 111-fold higher than for the parental hepatoma cells. The IC50 value for KB-CP10 cells was 38 μg ml-1 cisplatin, which is 1152-fold higher than for the parental KB cells. The 7404-CP7.5 cells were cross-resistant to methotrexate (39 x), 5-fluorouracil (23 x) and 6-mercaptopurine (13 x), but were sensitive to drugs which are known substrates for the multidrug transporter (P-glycoprotein), including colchicine, vinblastine and actinomycin D. Similar cross-resistance patterns were observed for KB-CPIO cells. No evidence of DNA amplification or expression of the MDRI gene was found. One-dimensional sodium dodecyl sulphate-polyacrylamide gel electrophoresis showed increases in 52 kDa protein(s) in both the soluble cytosolic and crude membrane fractions in 7404-Cr cells and in KB-CPF cells. The amount of 52 kDa protein was proportional to the degree of resistance of the 7404-CP cells to cisplatin. Two-dimensional gel analysis demonstrated that two polypeptides of molecular mass 52 and 50 kDa were overexpressed in the membrane fractions in both 7404-CP20 and KB-CP20 cells. Using amino acid microsequencing and Western blotting, the major 52 kDa protein was identified as the mitochondrial heat shock protein hsp60. Two-dimensional gels of [35S]methionine-labelled polypeptides showed many other changes, including reduction in soluble proteins of approximately 57 kDa molecular weight in KB-CP20 cells, and of 35 kDa in both 7404-CP20 and KB-CP20 cells. These results suggest that alterations of certain proteins occur commonly in cisplatin-resistant cells, particularly proteins of molecular weight 52 and 50 kDa.
KW - Adenocarcinoma cells
KW - Cisplatin
KW - Crossresistance
KW - Hepatoma cells
KW - Two-dimensional gel electrophoresis
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U2 - 10.1038/bjc.1995.134
DO - 10.1038/bjc.1995.134
M3 - Article
C2 - 7710928
AN - SCOPUS:0028920467
SN - 0007-0920
VL - 71
SP - 676
EP - 683
JO - British Journal of Cancer
JF - British Journal of Cancer
IS - 4
ER -