Acquired resistance to cisplatin and doxorubicin in a small cell lung cancer cell line is correlated to elevated expression of glutathione-linked detoxification enzymes

A human small cell lung cancer cell line, U-1906, developed altered functional properties upon continuous in vitro cultivation. Cells obtained at late (U-1906 L) and early (U-1906 E) passages of cultivation differ in drug resistance to the cytostatic therapeutic agents cisplatin and doxorubicin. The U-1906 L cells are 1.6-fold and 1.3-fold more resistant to cisplatin and doxorubicin respectively, than are the U-1906 E cells. In the more resistant U-1906 L cells, the total glutathione (GSH plus GSSG) level is 40% lower, whereas the activities of GSH-linked enzymes such as GSH peroxidase and GSH transferases are significantly higher. Quantitative analysis with isoenzyme-specific ELISAs demonstrated increased concentrations of all three of the measurable GSTs, M1-1, M3-3 and P1-1, in the more resistant cells. The intracellular protein expression patterns of the U-1906 E and the U-1906 L cells are very similar as revealed by two-dimensional denaturing electrophoresis, but show significant alterations in the concentrations of some components. Two 35 kDa proteins of different pI values, the concentrations of which are increased in the U-1906 L cells, were both identified as glyceraldehyde-3-phosphate dehydrogenase, either by N-terminal or by internal amino acid sequence analysis. The present study demonstrates that the increased resistance of the U-1906 L cells may involve multiple detoxification mechanisms and that the contribution of the GSH-linked detoxification can be ascribed to the elevation of cytosolic GST isoenzymes, GSH peroxidase and glutathione reductase, rather than to the intracellular GSH concentrations.