Kinetics of Anthracycline Efflux from Multidrug Resistance Protein-Expressing Cancer Cells Compared with P-Glycoprotein-Expressing Cancer Cells

Abstract

The multidrug resistance protein (MRP) has been shown to mediate ATP-dependent efflux of anticancer agents of diverse structure, such as daunorubicin (DNR), vincristine and etoposide. Thus, this protein does confer a multidrug resistant phenotype to cancer cells, similar to P-glycoprotein (Pgp). The substrate specificity of both transporter proteins is partly overlapping but is otherwise very distinct; because MRP is a multiple organic anion transporter, it transports certain glutathione conjugates and may be partly dependent on intracellular glutathione levels for the transport of anthracyclines. We have studied the transport kinetics of a series of anthracyclines in MRP and Pgp that overexpress tumor cell lines to obtain information on the substrate specificity of these proteins. The anthracyclines have modifications in the sugar moiety. The mean active efflux coefficientk_a, used to characterize the efficiency of the active efflux, was very similar for DNR and one of its 4′-deoxy- derivatives (eso-DNR) for MRP and Pgp [10–20 × 10⁻¹⁰/sec/(cells/ml)]. The permanently neutral derivatives 3′-deamino-3′-hydroxy-doxorubicin (OH-DOX) and 3′-deamino-3′-hydroxy-daunorubicin (OH-DNR) were effluxed by both proteins but had a lower k_a[2 × 10⁻¹⁰ and 6 × 10⁻¹⁰/sec/(cells/ml) (OH-DOX) and 2 × 10⁻¹⁰ and 5 × 10⁻¹⁰/sec/(cells/ml) (OH-DNR)] for MRP and Pgp. Two anthracyclines, the doxorubicin derivative pirarubicin and 2′-bromo-4′-epi-DNR seemed to have a slightly higher k_a value for Pgp than for MRP. The apparent Michaelis-Menten constants (K_m) and maximal efflux rates (V_M) for the active transport were within a narrow range for both transporters, except for OH-DOX and OH-DNR, which had a lower V_M in the case of MRP-mediated transport, suggesting a role of the amino group in the interaction with glutathione. Determination of the Hill coefficient (n_H) of the MRP-mediated efflux gave most values close to 2, which suggests cooperativity of the transport of anthracyclines as reported before for Pgp. In conclusion, the transport kinetics of anthracyclines by MRP and Pgp are very similar.