L1210 cells cultivated under the selection pressure of doxorubicin or vincristine express common mechanisms of multidrug resistance based on the overexpression of P-glycoprotein

Multidrug resistance of neoplastic tissue is often associated with the overexpression and increased drug transport activity of plasma membrane transporters like P-glycoprotein (P-gp), multidrug resistance associated proteins (MRPs) or breast cancer resistance protein, as well as with the elevation of the glutathione detoxification pathway. We have already described the overexpression of P-gp under the selection pressure of vincristine in L1210 mouse leukemia cells. In the present study, mechanisms of multidrug resistance induced in L 1210 cells cultivated in the presence of doxorubicin were analyzed. The selection pressure of both vincristine (yielding a resistant subline of L1210 cells, R-v) and doxorubicin (yielding a resistant subline of L1210 cells, R-D) induced a dramatic depression of cell sensitivity to both drugs. Both Rv and RD cells demonstrated a lack of ability to accumulate calcein/AM and fluo-3/AM as fluorescent substrates of P-gp and MRP. The retention of dyes could be reached in both cell sublines by the application of inhibitors of P-gp (like verapamil) but not by probenecid-an inhibitor of anion transporters, including MRPs. Massive protein bands, at a M, range of 130-180 kDa that interact with c219 antibody against P-gp, were detected in the crude membrane fraction isolated from both R-V and R-D (but not from L1210) cells by Western blot. The cytosolic activity of glutathione S-transferase was found to be similar in R-V and R-D cells and did not differ significantly from the activity ascertained in parental L1210 cells. Neither the R-V nor R-D cell sublines differed considerably, as measured by cell ultrastructure. In conclusion, based on P-gp overexpression, both doxorubicin and vincristine induce a common multidrug resistance phenotype in L1210 cells. (c) 2006 Elsevier Ltd. All rights reserved.