Intracellular glutathione level and efflux in human melanoma and cervical cancer cells differing in doxorubicin resistance

Drug resistance continues to be a major problem in cancer treatment. Occurrence of this phenomenon is often associated with altered levels of glutathione (GSH) and GSH-related enzymes. The aim of the study was to evaluate the possible involvement of GSH and GSH-related enzymes in doxorubicin (DOX) resistance in two types of cancer cells of different etiology, from both parental and DOX-resistant sublines. The human melanoma (ME18 and ME18/R) and cervical cancer cells (HeLa and KB-V1) were tested in terms of their DOX sensitivity (EZ4U test), GSH level (HPLC) and its efflux (spectrofluorometrically). The effects of inhibition of the GSH-related enzymes. gamma-glutamylcysteine synthetase (gamma-GCS) and glutathione S-transferase (GST) were also evaluated. Exposure to DOX caused an increase of GSH levels in all tested cells except for HeLa cells. However, depletion of GSH did not have a significant influence on the sensitivity of the cells to DOX. Inhibition of the activity of GST also did not have a major effect on DOX sensitivity, although it caused changes of the GSH content. Our attempts to use the spectrofluorometric method for measurements of GSH efflux were not successful. It could be suggested that in ME18 and HeLa cells treated with DOX, GSH efflux does occur. The obtained results seem to refute the hypothesis of a central role of GSH in DOX resistance of the tested cells. Despite observations of different effects related to GSH, they do not seem to be essential in terms of DOX resistance. The mechanisms underlying DOX resistance are highly cell-specific.