Establishment of a human small-cell lung-cancer subline resistant to okadaic acid.
Okadaic acid (OA), a specific protein phosphatase inhibitor, has various biological functions. To elucidate the mechanism of OA resistance, we have established a small-cell lung-cancer subline (H69/OA100) resistant to the growth-inhibitory effect of OA; this was done by using the parental cell line (H69) and increasing the concentration of OA. H69/OA100 was about 8 times more resistant to OA than H69. Intracellular retention of the fluorescent OA derivative in H69/OA100 was the same as that in H69. The catalytic activity of protein phosphatase from H69/OA100 was significantly reduced compared with that from H69. The protein phosphatase from H69/OA100 was 3.6 times more resistant to OA than that from H69. We examined the effect of OA on the activity of the immunoprecipitated protein phosphatase type I (PPI) and type 2A (PP2A) from the 2 cell lines. The PPI and PP2A from H69/OA100 showed more resistance to OA than those from H69. We next examined the effect of OA on the cell cycle of H69 and H69/OA100. In H69, G2/M block was observed at an OA concentration of 30 ng/ml whereas in H69/OA100, no G2/M block was observed at concentrations up to 100 ng/ml OA. We finally evaluated the amount of p34cdc2 kinase expression and the phosphorylation status of p34cdc2. There was no difference in p34cdc2 expression between H69 and H69/OA100 at several concentrations of OA. However, dephosphorylation of p34cdc2 was observed at 30 ng/ml OA in H69, but not in H69/OA100 up to 100 ng/ml OA. These data suggest that the resistance to OA and the resistance of the cell-cycle block to OA in H69/OA100 might be due to alteration of protein phosphatase activity.
Takeda
Y
Y
Pharmacology Division, National Cancer Center Research Institute, Tokyo, Japan.
Nishio
K
K
Kubota
N
N
Miura
K
K
Morikage
T
T
Ohmori
T
T
Kudoh
S
S
Niitani
H
H
Saijo
N
N
D001483:Base Sequence; D015153:Blotting, Western; D018288:Carcinoma, Small Cell / Q000188:drug therapy* / Q000473:pathology*; D002384:Catalysis; D002455:Cell Division / Q000187:drug effects; D004351:Drug Resistance; D004988:Ethers, Cyclic / Q000493:pharmacokinetics / Q000494:pharmacology*; D005434:Flow Cytometry; D015870:Gene Expression; D006801:Humans; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology*; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D016133:Polymerase Chain Reaction; D014407:Tumor Cells, Cultured / Q000187:drug effects*
D004988:Ethers, Cyclic; D046911:Macromolecular Substances; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases