Effect of P-Glycoprotein Expression on the Accumulation and Cytotoxicity of Topotecan (SK&F 104864), a New Camptothecin Analogue

Topotecan (TFT, 9-dimethylaminomethyI-lO-hydroxycamptothecin) is the first topoisomerase I-directed cytotoxic agent to enter clinical trials in the United States in two decades. The effect of P-glycoprotein (Pgp) overexpression on TPT cytotoxicity was examined in CH”C5 (colchicinerosâ¡ slant)and AuxBl (parental) Chinese hamster ovary cells. Examina tion of the K.in values observed in colony-forming assays revealed that the CH”C5 cells were 15-fold (SD, ±3;n = 3) resistant to TPT after a 1-h exposure and 3.2-fold (SD, ±1.4;n = 4) resistant in continuous exposure experiments. Band depletion immunoblotting revealed that 4- fold higher concentrations of extracellular TPT were required to induce the formation of topo I-DNA complexes in CH”C5 cells as compared to AuxBl cells. To assess the role of Pgp in this resistance, drug accumu lation and cytotoxicity assays were repeated in the absence and presence of quinidine. Addition of quinidine enhanced TPT accumulation (meas ured by high-performance liquid chromatography) and diminished the IC«, for TPT to a greater extent in CH”C5 cells than in AuxBl cells. To examine whether similar effects could be detected in Pgp-expressing human cells, MCF-7/Adria' breast cancer cells and KGla human acute myelogenous leukemia cells were examined. Quinidine or verapamil enhanced TPT accumulation in both of these cell lines but had no effect in parental MCF-7 cells or a variety of human leukemia cell lines that do not overexpress Pgp. Cytotoxicity measurements performed by counting the number of surviving cells (MCF-7/Adria') or employing a modi- Tied, highly stable tetrazolium dye reduction assay (leukemia cell lines) revealed that quinidine diminished the ICso for TPT in the Pgp-overexpressing cell lines but not in the control lines. These results suggest that Pgp overexpression diminishes TPT accu mulation and TPT cytotoxicity in hamster and human cells. It should be stressed, however, that these effects were substantially smaller than the effects of Pgp overexpression on the accumulation and cytotoxicity of the anthracycline daunorubicin and the epipodophyllotoxin etoposide in the same cell lines.