Co-expression of ABCB1 and ABCG2 in a Cell Line Model Reveals Both Independent and Additive Transporter Function

Although overexpression of multiple ATP-binding cassette transporters has been reported in clinical samples, few studies have examined how co-expression of multiple transporters affected resistance to chemotherapeutic drugs. We therefore examined how co-expression of ABCB1 (P-glycoprotein) and ABCG2 contribute to drug resistance in a cell line model. HEK-293 cells were transfected with vector encoding full-length ABCB1, ABCG2, or a bicistronic vector containing both genes, each under the control of a separate promoter. Cells transfected with both transporters (B1/G2 cells) demonstrated high levels of both transporters and uptake of both the ABCB1-specific substrate rhodamine 123 and the ABCG2-specific substrate pheophorbide a was reduced when examined by flow cytometry. B1/G2 cells were also cross-resistant to the ABCB1 substrate doxorubicin, the ABCG2 substrate topotecan, as well as mitoxantrone and the cell cycle checkpoint kinase 1 inhibitor prexasertib, both of which were found to be substrates of both ABCB1 and ABCG2. When B1/G2 cells were incubated with both rhodamine 123 and pheophorbide a, transport of both compounds was observed, suggesting that ABCB1 and ABCG2, when co-expressed, can function independently to transport substrates. ABCB1 and ABCG2 also functioned additively to transport the common fluorescent substrates mitoxantrone and BODIPY®-prazosin, as it was necessary to inhibit both transporters to prevent efflux from B1/G2 cells. ABCG2 expression was also found to decrease the efficacy of the ABCB1 inhibitor tariquidar in B1/G2 cells. Thus, ABCB1 and ABCG2 can independently and additively confer resistance to substrates, underscoring the need to inhibit multiple transporters when they are co-expressed. SIGNIFICANCE STATEMENT: N/A. The American Society for Pharmacology and Experimental Therapeutics.

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