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PMID: 23470221 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't

Fluorescent substrates for flow cytometric evaluation of efflux inhibition in ABCB1, ABCC1, and ABCG2 transporters.

Analytical biochemistry ·Vol. 437 ·No. 1 ·2013-06-01 ·Pages 77-87

Strouse JJ, Ivnitski-Steele I, Waller A, Young SM, Perez D, Evangelisti AM, Ursu O, Bologa CG, Carter MB, Salas VM, Tegos G, Larson RS, Oprea TI, Edwards BS, Sklar LA

Abstract

ATP binding cassette (ABC) transmembrane efflux pumps such as P-glycoprotein (ABCB1), multidrug resistance protein 1 (ABCC1), and breast cancer resistance protein (ABCG2) play an important role in anticancer drug resistance. A large number of structurally and functionally diverse compounds act as substrates or modulators of these pumps. In vitro assessment of the affinity of drug candidates for multidrug resistance proteins is central to predict in vivo pharmacokinetics and drug-drug interactions. The objective of this study was to identify and characterize new substrates for these transporters. As part of a collaborative project with Life Technologies, 102 fluorescent probes were investigated in a flow cytometric screen of ABC transporters. The primary screen compared substrate efflux activity in parental cell lines with their corresponding highly expressing resistant counterparts. The fluorescent compound library included a range of excitation/emission profiles and required dual laser excitation as well as multiple fluorescence detection channels. A total of 31 substrates with active efflux in one or more pumps and practical fluorescence response ranges were identified and tested for interaction with eight known inhibitors. This screening approach provides an efficient tool for identification and characterization of new fluorescent substrates for ABCB1, ABCC1, and ABCG2.

MeSH Terms
ATP Binding Cassette Transporter, Subfamily B ATP Binding Cassette Transporter, Subfamily B, Member 1/antagonists & inhibitors,genetics,metabolism ATP Binding Cassette Transporter, Subfamily G, Member 2 ATP-Binding Cassette Transporters/antagonists & inhibitors,genetics,metabolism Cell Line Flow Cytometry/methods Fluorescent Dyes/metabolism Humans Multidrug Resistance-Associated Proteins/antagonists & inhibitors,genetics,metabolism Neoplasm Proteins/antagonists & inhibitors,genetics,metabolism Protein Binding
Chemicals
ABCB1 protein, human ABCG2 protein, human ATP Binding Cassette Transporter, Subfamily B ATP Binding Cassette Transporter, Subfamily B, Member 1 ATP Binding Cassette Transporter, Subfamily G, Member 2 ATP-Binding Cassette Transporters Fluorescent Dyes Multidrug Resistance-Associated Proteins Neoplasm Proteins multidrug resistance-associated protein 1
Authors & Affiliations
15 authors, click to expand affiliations / ORCID
Strouse J Jacob
Cytometry, Cancer Research and Treatment Center, University of New Mexico Health Sciences Center, Albuquerque, NM 87131, USA.
Ivnitski-Steele Irena
Waller Anna
Young Susan M
Perez Dominique
Evangelisti Annette M
Ursu Oleg
Bologa Cristian G
Carter Mark B
Salas Virginia M
Tegos George
Larson Richard S
Oprea Tudor I
Edwards Bruce S
Sklar Larry A
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Article Info
Journal
Analytical biochemistry
Abbr.
Anal Biochem
ISSN
1096-0309
DOI
10.1016/j.ab.2013.02.018
Published
2013-06-01
Epub
2013-00-05
Pages
77-87
Language
English
Region
United States
NLM ID
0370535
PMCID
PMC3785545
Subset
IM
Grants
NIMH NIH HHS · U54 MH084690 · United States
NCATS NIH HHS · UL1 TR000041 · United States
NCATS NIH HHS · UL1 TR001449 · United States
NIMH NIH HHS · U54MH084690 · United States
External Links
PubMed source DOI: 10.1016/j.ab.2013.02.018 PMC Full Text View journal details
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