Abstract
Breast cancer resistance protein (BCRP) is an ATP-driven efflux pump at the blood-brain barrier that limits central nervous system pharmacotherapy. Our previous studies showed rapid loss of BCRP transport activity in rat brain capillaries exposed to low concentrations of 17-β-estradiol (E2); this occurred without acute change in BCRP protein expression. Here, we describe a pathway through which sustained, extended exposure to E2 signals down-regulation of BCRP at the blood-brain barrier. Six-hour exposure of isolated rat and mouse brain capillaries to E2 reduced BCRP transport activity and BCRP monomer and dimer expression. Experiments with brain capillaries from estrogen receptor (ER)α and ERβ knockout mice and with ER agonists and antagonists showed that E2 signaled through ERβ to down-regulate BCRP expression. In rat brain capillaries, E2 increased unphosphorylated, active phosphatase and tensin homolog (PTEN); decreased phosphorylated, active Akt; and increased phosphorylated, active glycogen synthase kinase (GSK)3. Consistent with this, inhibition of phosphoinositide 3-kinase (PI3K) or Akt decreased BCRP activity and protein expression, and inhibition of PTEN or GSK3 reversed the E2 effect on BCRP. Lactacystin, a proteasome inhibitor, abolished E2-mediated BCRP down-regulation, suggesting internalization followed by transporter degradation. Dosing mice with E2 reduced BCRP activity in brain capillaries within 1 h; this reduction persisted for 24 h. BCRP protein expression in brain capillaries was unchanged 1 h after E2 dosing but was substantially reduced 6 and 24 h after dosing. Thus, E2 signals through ERβ, PTEN/PI3K/Akt/GSK3 to stimulate proteasomal degradation of BCRP. These in vitro and in vivo findings imply that E2-mediated down-regulation of blood-brain barrier BCRP has the potential to increase brain uptake of chemotherapeutics that are BCRP substrates.
Footnotes
This work was supported by the Intramural Research Program at the National Institutes of Health National Institute of Environmental Health Sciences (to D.S.M.); a University of Minnesota College of Pharmacy Melendy Research Scholarship (to E.K.M.); a research grant from the Whiteside Institute for Clinical Research (to A.M.S.H. and B.B.); and University of Minnesota College of Pharmacy startup funds (to B.B.).
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
doi:10.1124/jpet.110.168930.
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ABBREVIATIONS:
- BCRP
- breast cancer resistance protein
- CNS
- central nervous system
- ABC
- ATP-binding cassette
- E2
- 17-β-estradiol
- KO
- knockout
- ER
- estrogen receptor
- PTEN
- phosphatase and tensin homolog
- PI3K
- phosphoinositide 3-kinase
- GSK
- glycogen synthase kinase
- PPT
- propylpyrazoletriol
- DPN
- diarylpropionitrile
- MPP
- methylpiperidinopyrazole
- ICI
- fulvestrant
- LY294002
- 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
- XIII
- 5-methyl-1H-pyrazol-3-yl)-(2-phenylquinazolin-4-yl)amine
- PBS
- phosphate-buffered saline
- BSA
- bovine serum albumin
- PCR
- polymerase chain reaction
- RT
- reverse transcription
- FTC
- fumitremorgin C
- ELISA
- enzyme-linked immunosorbent assay
- bp
- base pair(s).
- Received April 1, 2010.
- Accepted May 10, 2010.
- U.S. Government work not protected by U.S. copyright
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