Breast cancer resistance
protein (BCRP/ABCG2) of an
ATP-binding cassette half-transporter confers resistance against
mitoxantrone and
camptothecin derivatives of
topotecan and
irinotecan.
Novobiocin, a
coumermycin antibiotic, is known to enhance anticancer drug sensitivity of
cancer cells in vitro and in vivo, the mechanism of which remains undetermined. Here we focused on drug efflux pump and examined whether
novobiocin reversed drug resistance in multidrug-resistant cells highly expressing BCRP. To explore the reversal mechanisms, intracellular drug accumulation was measured by flow cytometry, and a
topotecan transport study using plasma membrane vesicles was performed. We used PC-6/SN2-5H2
small cell lung cancer and MCF-7/MX
breast cancer cells selected with
SN-38 of the active
irinotecan metabolite and
mitoxantrone, respectively, and the BCRP
cDNA transfectant MCF-7/clone 8 cells. These cells expressed high levels of BCRP
mRNA but not other known transporters. Compared to the parental PC-6 cells, PC-6/SN2-5H2 cells were 141-, 173- and 57.2-fold resistant to
topotecan,
SN-38 and
mitoxantrone, respectively.
Novobiocin at 60 microM decreased the degree of the above resistance by approximately 26-fold in PC-6/SN2-5H2 cells, and similarly reversed resistance in MCF-7/MX, MCF-7/clone 8 and un-selected NCI-H460 cells highly expressing BCRP. Furthermore,
novobiocin increased the intracellular
topotecan accumulation in these cells and inhibited the
topotecan transport into the membrane vesicles of PC-6/SN2-5H2 cells. No effects of
novobiocin in these assay were observed in the parental PC-6 and MCF-7 cells. The kinetic parameters in the transport study indicated that
novobiocin was a inhibitor for BCRP, resulting in competitive inhibition of BCRP-mediated
topotecan transport. These findings suggest that
novobiocin effectively overcomes BCRP-mediated drug resistance at acceptable concentrations.