A fluorometric screening assay for drug efflux transporter activity in the blood-brain barrier.
OBJECTIVE
To examine the capability of a fluorometric assay to identify and characterize the drug efflux interactions of a broad spectrum of drug agents in an in vitro model of the blood-brain barrier (BBB).


METHODS
Various concentrations of drug agent (1, 10, and 100 microM) were evaluated for their effect on the cellular accumulation of the P-glycoprotein (P-gp) probe R123 (3.2 microM), and the mixed P-gp and multidrug resistance-associated protein (MRP) probe, BCECF (1 microM), in bovine brain microvessel endothelial cell (BBMEC) monolayers. Drugs demonstrating a significant effect were further quantitated using an expanded concentration range and a nonlinear regression curve fit to determine the potency (IC50) and efficacy (Imax) of the drug for P-gp and/or MRP.


RESULTS
Several of the 36 therapeutic agents examined showed drug efflux transporter interactions in BBMEC monlayers. Melphalan and risperidone significantly enhanced the accumulation of R123 over control (1.47- and 1.82-fold, respectively) with resulting IC50s of 1.4 and 14.6 microM, respectively. Chlorambucil and valproic acid significantly enhanced the accumulation of BCECF compared to control monolayers (2.02- and 4.01-fold, respectively) with resulting IC50s of 146.1 and 768.5 microM, respectively.


CONCLUSIONS
The current study demonstrates the feasibility of a fluorometric assay consisting of R123 and BCECF in assessing the drug efflux interactions of a variety of drugs in the BBB.
Bachmeier
Corbin J
CJ
Department of Pharmaceutical Sciences, University of Nebraska Medical Center, Omaha, Nebraska 68198, USA.
Miller
Donald W
DW
D018435:ATP Binding Cassette Transporter, Subfamily B / Q000032:analysis / Q000378:metabolism*; D000818:Animals; D001812:Blood-Brain Barrier / Q000737:chemistry / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D004353:Drug Evaluation, Preclinical / Q000379:methods; D005470:Fluorometry / Q000379:methods; D004364:Pharmaceutical Preparations / Q000032:analysis / Q000378:metabolism*
D018435:ATP Binding Cassette Transporter, Subfamily B; D004364:Pharmaceutical Preparations