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Adenosine triphosphate–dependent transport of estradiol‐17β(β‐ d ‐glucuronide) in membrane vesicles by MDR1 expressed in insect cells
Author(s) -
Huang Liyue,
Hoffman Tim,
Vore Mary
Publication year - 1998
Publication title -
hepatology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.488
H-Index - 361
eISSN - 1527-3350
pISSN - 0270-9139
DOI - 10.1002/hep.510280528
Subject(s) - multidrug resistance associated protein 2 , glucuronide , estriol , atp binding cassette transporter , biochemistry , glutathione , chemistry , adenosine , biology , estrogen , metabolism , endocrinology , transporter , enzyme , gene
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‐17β(β‐ d ‐glucuronide) (E 2 17G). MDR1 substrates/modulators inhibit adenosine triphosphate (ATP)‐dependent transport of E 2 17G in the rat canalicular membrane and protect against E 2 17G‐mediated cholestasis in isolated perfused rat liver. The present studies were designed to determine if E 2 17G is a substrate for MDR1 using a baculovirus expression system and if other estrogen glucuronides interact with MDR1. ATP‐dependent transport of E 2 17G (10 μmol/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 ( K m = 62 μmol/L) and occurred into an osmotically sensitive space. ATP‐dependent transport of E 2 17G (10 μmol/L) was inhibited 63% by 10 μmol/L daunomycin, but not by 100 μmol/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 μmol/L), estriol‐17β(β‐ d ‐glucuronide) (E 3 17G) and estriol‐16α(β‐ d ‐glucuronide) (E 3 16G), inhibited MDR1‐mediated E 2 17G transport by 58% and 35%, respectively. In contrast, noncholestatic glucuronides, estradiol‐3‐(β‐ d ‐glucuronide) (E 2 3G) or estradiol‐3‐sulfate‐17β(β‐ d ‐glucuronide) (E 2 3SO 4 17G), had no effect. E 2 17G neither stimulated MDR1 ATPase activity nor inhibited verapamil‐stimulated ATPase activity. Infusion of 1.5 μmol/L doxorubicin or 1 μmol/L taxol protected against cholestasis induced by E 3 16G and E 3 17G in isolated perfused rat liver. These studies identify E 2 17G, and probably E 3 16G and E 3 17G, as endogenous substrates for MDR1.