BacMam recombinant baculovirus in transporter expression: a study of BCRP and OATP1B1.
Human BCRP and OATP1B1 have recently been identified as important transporters in the absorption, distribution, and elimination of clinically significant drugs. In this report, we illustrate the use of modified baculoviruses, termed BacMam viruses for the expression of functional BCRP and OATP1B1 in mammalian cells. We show a variety of host cells efficiently transduced to express BCRP including HEK 293, LLC-PK, and U-2 OS, where protein levels on the cell-surface were modulated by titrating different amounts of viral inoculum. In addition, using the BODIPY-prazosin efflux assay and the BacMam reagent we illustrate inhibition of BCRP activity with GF120918 or Fumitremorgin C. Furthermore, we present data demonstrating simultaneous expression of BCRP and OATP1B1 in BacMam transduced mammalian cells by simply adding viral inoculum of each transporter. Thus these results indicate that BacMam mediated gene delivery provides a novel and efficient research tool for the investigation of single or multiple transporters in vitro.
Hassan
Namir J
NJ
Department of Gene Expression and Protein Biochemistry, GlaxoSmithKline, Harlow, UK. namir.j.hassan@gsk.com
Pountney
David J
DJ
Ellis
Catherine
C
Mossakowska
Danuta E
DE
D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000235:genetics; D000166:Acridines / Q000494:pharmacology; D000818:Animals; D016367:Baculoviridae; D019556:COS Cells; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000235:genetics / Q000378:metabolism*; D002522:Chlorocebus aethiops; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D027381:Liver-Specific Organic Anion Transporter 1; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000235:genetics; D027361:Organic Anion Transporters / Q000096:biosynthesis* / Q000235:genetics; D018411:Spodoptera / Q000166:cytology; D013552:Swine; D044005:Tetrahydroisoquinolines / Q000494:pharmacology; D014161:Transduction, Genetic
C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D000166:Acridines; D007211:Indoles; D027381:Liver-Specific Organic Anion Transporter 1; D009363:Neoplasm Proteins; D027361:Organic Anion Transporters; C503999:SLCO1B1 protein, human; D044005:Tetrahydroisoquinolines; C010522:tryptoquivaline; C083501:Elacridar