Functional intracellular P-glycoprotein.
Efflux of chemotherapy drugs by P-glycoprotein (P-gp) at the plasma membrane is thought to be a major cause of cancer multidrug resistance. In this report, we show by flow cytometry that P-gp also concentrates large amounts of 2 different drugs, Hoechst 33342 and daunorubicin, within a cytoplasmic compartment of multidrug resistant CHRC5 cells. A quantitative assay of Hoechst 33342 revealed that cytoplasmic sequestration by P-gp in CHRC5 cells accounted for about half of the amount of Hoechst 33342 accumulated by the drug-sensitive parental Aux BI cells. Daunorubicin sequestered in the cytoplasm of CHRC5 cells could be released by inhibiting P-gp function with cyclosporin A, resulting in cell death. A likely site of drug sequestration is P-gp-containing cytoplasmic vesicles, in which the P-gp is oriented so that drugs are transported and concentrated in the interior of the vesicles. P-gp was detected in the membranes of cytoplasmic vesicles of CHRC5 cells by confocal immunofluorescence microscopy and immunoelectron microscopy with anti-P-gp monoclonal antibodies (MAbs). Vesicular localization of daunorubicin was observed by epifluorescence microscopy. The origin and nature of the P-gp-containing vesicles are unknown, but they do not correspond to endocytic vesicles. Our results directly demonstrate that chemosensitizer-induced release of drugs sequestered in cytoplasmic vesicles by P-gp can be used to overcome multidrug resistance.
Shapiro
A B
AB
British Columbia Cancer Research Centre, Vancouver, Canada.
Fox
K
K
Lee
P
P
Yang
Y D
YD
Ling
V
V
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000032:analysis / Q000502:physiology*; D001562:Benzimidazoles / Q000493:pharmacokinetics; D002460:Cell Line; D016572:Cyclosporine / Q000494:pharmacology; D003630:Daunorubicin / Q000493:pharmacokinetics; D004351:Drug Resistance
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D001562:Benzimidazoles; D016572:Cyclosporine; C017807:bisbenzimide ethoxide trihydrochloride; D003630:Daunorubicin