Modulation of daunorubicin cellular resistance by combination of P-glycoprotein blockers acting on drug efflux and intracellular drug sequestration in golgi vesicles

Background: S9788 and PSC833 were developped as P-glycoprotein (Pgp) blocke rs and found to act additionally on daunorubicin subcellular distribution, involving different putative targets. On this basis, combinations of S9788 and PSC833 were evaluated in Pgp-expressing MCF7(DXR) cells in which we rec ently demonstrated that daunorubicin was sequestered in Golgi vesicles (Bou r-Dill et al.: Cytometry, 39: 16-25, 200). Methods: Combinations of S9788 and PSC833 consisted in complementary fracti ons of iso-effective concentrations (IEC) leading to 90% (IEC90) and median (IEC50) reversion of daunorubicin resistance. Resistance modulation was as sessed using cytotoxicity assays, flow cytometry determination of intracell ular daunorubicin, and fluorescence microscopy analysis of daunorubicin sub cellular distribution. Results: Individually, both S9788 and PSC833 were found to be very potent w ith IEC90 of 5 and 15 mu mol/l, and IEC50 of 0.1 and 0.2 mu mol/l, respecti vely, for S9788 and PSC853. When combined, synergistic cytotoxicity was obs erved for both IEC90 and IEC50 combinations while intracellular daunorubici n fluorescence was only synergistically increased for IEC90 combinations. F or IEC50 combinations, no increase in intracellular fluorescence was observ ed, and fluorescence microscopy examination of the cells suggested that dau norubicin sequestration in Golgi vesicles could be modulated at concentrati ons that do not significantly increase daunorubicin cellular concentration. Using immunofluorescence and reverse transcription-polymerase chain reacti on analyses, multidrug resistance-associated protein, major vault lung-resi stance protein, and anthracycline-resistance associated protein were not fo und to be implicated. Conclusions: Synergistic combinations of S9788 and PSC833 might offer alter native ways to decrease the toxicity generated by high-dose Pgp-blockers wi thout altering the efficacy of the resistance modulation. (C) 2000 Wiley-Li ss, Inc.