title: New functional assay of P-glycoprotein activity using Hoechst 33342.
abstract: In this study we describe a simplified, HTS-capable functional assay for the multidrug resistance (MDR) transporter P-glycoprotein (P-gp) based on its substrate Hoechst 33342. The physicochemical properties of Hoechst 33342 and the enormous milieu dependency of its fluorescence intensity allowed performing the assay in a homogeneous manner. This new assay served as an effective tool to estimate the potency of 10 well recognized P-gp substrates and modulators. Further, the potency of these compounds was also estimated in the calcein AM assay. The Hoechst 33342 and calcein AM assays yielded significantly comparable results for all compounds tested. Principal component analysis (PCA) applied to literature data on inhibition of P-gp activity and our results obtained in the Hoechst 33342 and calcein AM assay indicated similarity of compared functional transport assays. However, no correlation could be detected between these functional assays and the ATPase activity assay.
authors: Institute of Pharmacy, University of Bonn, An der Immenburg 4, 53121 Bonn, Germany.
authors: Henrik
authors: H
authors: Müller
authors: Werner
authors: W
authors: Klinkhammer
authors: Christoph
authors: C
authors: Globisch
authors: Matthias U
authors: MU
authors: Kassack
authors: Ilza K
authors: IK
authors: Pajeva
authors: Michael
authors: M
authors: Wiese
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000096:biosynthesis / Q000737:chemistry* / Q000187:drug effects; D000251:Adenosine Triphosphatases / Q000187:drug effects / Q000378:metabolism; D001562:Benzimidazoles / Q000737:chemistry* / Q000493:pharmacokinetics; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D019008:Drug Resistance, Neoplasm; D005452:Fluoresceins / Q000737:chemistry; D006801:Humans; D008856:Microscopy, Fluorescence / Q000379:methods; D015394:Molecular Structure; D025341:Principal Component Analysis; D013997:Time Factors
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D001562:Benzimidazoles; D005452:Fluoresceins; C085925:calcein AM; D000251:Adenosine Triphosphatases; C017807:bisbenzimide ethoxide trihydrochloride