INHIBITION OF APOPTOSIS IS THE CAUSE OF RESISTANCE TO DOXORUBICIN IN HUMAN BREAST ADENOCARCINOMA CELLS

In our previous paper we have described the isolation and characteriza tion of a doxorubicin (DOX) resistant subline of breast adenocarcinoma SC6 cells. These cells were obtained after the treatment with low, cl inically relevant doses of doxorubicin. They became cross-resistant to different wide used cytostatics. The expression of several genes invo lved in mitotic signal transduction, as well as cathepsins D and L, wa s similar in both parental and doxorubicin treated cells. The aim of t his study was to examine the molecular mechanisms involved in resistan ce of these cells to doxorubicin. Activity of plasma membrane Pgp was examined in parental and resistant cells due to rhodamine-accumulation assay. The involvement of glutathione (GSH) and glutathione S-transfe rase (GST) in resistance to doxorubicin was determined in MTT modified assay due to the addition of specific inhibitors: buthionine sulfoxim ine (for GSH) or ethacrynic acid (for GST). The kinetic of apoptosis w as followed after the treatment with DOX in control and SC6 cells by f luorescent microscope. The occurrence of apoptosis was confirmed by an alysing DNA fragmentation in agarose gel. Our results indicate that P- glycoprotein, glutathione or glutathione transferases were not involve d in resistance of SC6 cells to doxorubicin. However, the apoptosis wa s inhibited in doxorubicin-resistant cells. Therefore, even low doses of doxorubicin can induce the resistance to this drug due to inhibitio n of apoptosis.