ALTERATIONS IN GLUTATHIONE AND GLUTATHIONE-RELATED ENZYMES IN A MULTIDRUG-RESISTANT SMALL CELL LUNG-CANCER CELL-LINE

  • 1 February 1990
    • journal article
    • research article
    • Vol. 37  (2) , 192-197
Abstract
H69AR is a multidrug-resistant small cell lung cancer cell line derived from a drug-sensitive cell line, H69, by selection in doxorubicin. It is cross-resistant to a wide variety of natural product-type antineoplastic agents but does not overexpress P-glycoprotein. In the present study, the levels of GSH and GSH-related enzymes in the H69AR cell line were determined and compared with those found in H69 cells. Unlike other drug-resistant cell lines, GSH levels were diminished 6-fold in H69AR cells (0.67 .+-. 0.28 .mu.g/mg of protein), compared with H69 cells (4.23 .+-. 1.17 .mu.g/mg of protein) (P < 0.01). This unusually low level of GSH may explain the pronounced collateral sensitivity of H69AR cells to buthionine sulfoximine (BSO), an inhibitor of the rate-limiting enzyme in GSH biosynthesis (ID50 of 4.4 .mu.M BSO for H69AR cells versus ID50 of 300 .mu.M BSO for H69 cells). BSO did not enhance doxorubicin cytotoxicity in the H69AR cell line, despite further depletion of GSH. GSH-reductase (EC 1.6.4.2) activity was elevated 2-fold in H69AR cells, compared with sensitive H69 cells (75.34 .+-. 14.94 versus 38.62 .+-. 5.06 nmol of NADPH/min/mg of protein) (p < 0.05). Both selenium-dependent and -independent GSH-peroxidase (EC 1.11.1.9) activities were unchanged in the resistant H69AR cell line, compared with its parent cell line. .gamma.-Glutamyl transpeptidase (EC 2.3.2.2) activity was 5-fold elevated in H69 AR cells, compared with H69 cells (2.50 .+-. 0.44 versus 0.46 .+-. 0.21 nmol of p-nitroaniline/min/mg of protein) (p < 0.01), whereas GSH-S-transferase (EC 2.5.1.18) activity was 10-fold higher (201.98 .+-. 43.62 versus 19.77 .+-. 1.72 nmol of 1-chloro-2,4-dinitrobenzene/min/mg of protein in H69AR and H69, cells, respectively) (p < 0.01). The GSH-S-transferases from both cell lines were purified by affinity chromatography and immunoblot analysis identified the GSH-S-transferases as belonging to the anionic .pi. class. GSH-S-transferases from the .mu. or .alpha. classes were not detectable in either cell line. In conclusion, marked differences in GSH levels and the activities of three of four GSH-related enzymes were observed between the multidrug-resistant H69AR cell line and its parent cell line. Further study is required to determine whether these changes are causally related to the development of drug resistance in this model system.

This publication has 31 references indexed in Scilit: