title: Breast cancer resistance protein exports sulfated estrogens but not free estrogens.
abstract: Breast cancer resistance protein (BCRP), an ATP-binding cassette transporter, confers resistance to a series of anticancer reagents such as mitoxantrone, 7-ethyl-10-hydroxycamptothecin, and topotecan. We reported previously that estrone and 17beta-estradiol reverse BCRP-mediated multidrug resistance. In the present study, we demonstrate that BCRP exports estrogen metabolites. First, we generated BCRP-transduced LLC-PK1 (LLC/BCRP) cells, in which exogenous BCRP is expressed in the apical membrane, and investigated transcellular transport of 3H-labeled compounds using cells plated on microporous filter membranes. The basal-to-apical transport (excretion) of mitoxantrone, estrone, and 17beta-estradiol was greater in LLC/BCRP cells than in LLC-PK1 cells. Thin-layer chromatography of transported steroids revealed that the transport of estrone and 17beta-estradiol was independent of BCRP expression. Alternatively, increased excretion of estrone sulfate and 17beta-estradiol sulfate was observed in LLC/BCRP cells. BCRP inhibitors completely inhibited the increased excretion of sulfated estrogens across the apical membrane. Conversion of estrogens into their sulfate conjugates was similar between LLC/BCRP and LLC-PK1 cells, suggesting that the increased excretion of estrogen sulfates was attributable to BCRP-mediated transport. Next, the uptake of 3H-labeled compounds in membrane vesicles from BCRP-transduced K562 (K562/BCRP) cells was investigated. 3H-labeled estrone sulfate, but not 3H-labeled estrone or 17beta-estradiol, was taken up by membrane vesicles from K562/BCRP cells, and this was ATP-dependent. Additionally, BCRP inhibitors suppressed the transport of estrone sulfate in membrane vesicles from K562/BCRP cells. These results suggest that BCRP does not transport either free estrone or 17beta-estradiol but exports sulfate conjugates of these estrogens.
authors.affiliation: Division of Molecular Biotherapy, Cancer Chemotherapy Center, Japanese Foundation for Cancer Research, 1-37-1 Kami-Ikebukuro, Toshima-ku, Tokyo 170-8455, Japan.
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Imai
authors.forename: Sakiyo
authors.initials: S
authors.lastname: Asada
authors.forename: Satomi
authors.initials: S
authors.lastname: Tsukahara
authors.forename: Etsuko
authors.initials: E
authors.lastname: Ishikawa
authors.forename: Takashi
authors.initials: T
authors.lastname: Tsuruo
authors.forename: Yoshikazu
authors.initials: Y
authors.lastname: Sugimoto
mesh_terms: D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D001943:Breast Neoplasms / Q000378:metabolism*; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D019008:Drug Resistance, Neoplasm; D004967:Estrogens / Q000378:metabolism*; D004966:Estrogens, Conjugated (USP) / Q000378:metabolism; D004970:Estrone / Q000031:analogs & derivatives* / Q000378:metabolism; D018374:LLC-PK1 Cells; D008958:Models, Molecular; D009363:Neoplasm Proteins / Q000378:metabolism*; D013431:Sulfates / Q000378:metabolism*; D013552:Swine
chemical_list: C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D004967:Estrogens; D004966:Estrogens, Conjugated (USP); D009363:Neoplasm Proteins; D013431:Sulfates; D004970:Estrone; C017296:estrone sulfate