Adenosine triphosphate-dependent transport of estradiol-17beta(beta-D-glucuronide) in membrane vesicles by MDR1 expressed in insect cells.
MDR1, an ABC transporter that confers multidrug resistance in tumor cells, is constitutively expressed in normal liver canalicular membrane. Human MDR1-expressing multidrug-resistant cells display increased resistance to estradiol-17beta(beta-D-glucuronide) (E217G). MDR1 substrates/modulators inhibit adenosine triphosphate (ATP)-dependent transport of E217G in the rat canalicular membrane and protect against E217G-mediated cholestasis in isolated perfused rat liver. The present studies were designed to determine if E217G is a substrate for MDR1 using a baculovirus expression system and if other estrogen glucuronides interact with MDR1. ATP-dependent transport of E217G (10 micromol/L) was linear for up to 2 minutes and yielded a rate of 45.6 pmol/min/mg protein in membrane vesicles from Sf9 cells infected with MDR1-baculovirus. This transport was saturable (Km = 62 micromol/L) and occurred into an osmotically sensitive space. ATP-dependent transport of E217G (10 micromol/L) was inhibited 63% by 10 micromol/L daunomycin, but not by 100 micromol/L S-(2,4-dinitrophenyl)glutathione (GS-DNP) (a substrate for canalicular multispecific organic anion transporter [cMOAT]). Glucuronide conjugates of the estrogen D-ring (100 micromol/L), estriol-17beta(beta-D-glucuronide) (E317G) and estriol-16(beta-D-glucuronide) (E316G), inhibited MDR1-mediated E217G transport by 58% and 35%, respectively. In contrast, noncholestatic glucuronides, estradiol-3-(beta-D-glucuronide) (E23G) or estradiol-3-sulfate-17beta(beta-D-glucuronide) (E23SO417G), had no effect. E217G neither stimulated MDR1 ATPase activity nor inhibited verapamil-stimulated ATPase activity. Infusion of 1.5 micromol/L doxorubicin or 1 micromol/L taxol protected against cholestasis induced by E316G and E317G in isolated perfused rat liver. These studies identify E217G, and probably E316G and E317G, as endogenous substrates for MDR1.
Huang
L
L
Department of Pharmacology, College of Medicine, Graduate Center for Toxicology, University of Kentucky, Lexington, KY, USA.
Hoffman
T
T
Vore
M
M
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics* / Q000378:metabolism*; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D001693:Biological Transport, Active / Q000187:drug effects; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D002779:Cholestasis / Q000139:chemically induced / Q000517:prevention & control; D004317:Doxorubicin / Q000627:therapeutic use; D004958:Estradiol / Q000031:analogs & derivatives* / Q000378:metabolism / Q000633:toxicity; D015870:Gene Expression; D007700:Kinetics; D008081:Liposomes / Q000378:metabolism; D008297:Male; D017239:Paclitaxel / Q000627:therapeutic use; D051381:Rats; D017207:Rats, Sprague-Dawley; D018411:Spodoptera; D014162:Transfection
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D008081:Liposomes; C025483:estradiol-17 beta-glucuronide; D004958:Estradiol; D004317:Doxorubicin; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D017239:Paclitaxel