Differential modulation of P-glycoprotein transport by protein kinase inhibition.
Previous studies of P-glycoprotein have demonstrated that its function can be modulated by phosphorylation. In the present study, inhibition of protein kinase C with calphostin C or stauroporine or prolonged treatment with the phorbol ester TPA decreased phosphorylation of P-glycoprotein, and impaired transport of vinblastine. Calphostin C also inhibited transport of actinomycin D, vincristine, rhodamine, and azidopine in SW620 Ad300 multidrug-resistant human colon carcinoma cells. Photoaffinity labeling of P-glycoprotein with azidopine was decreased by calphostin C, suggesting that dephosphorylation alters the affinity of P-glycoprotein for its substrates. Impaired transport of rhodamine in normal T lymphocytes treated with staurosporine demonstrates that modulation of P-glycoprotein function is not limited to cells selected for drug resistance in vitro. Transport of P-glycoprotein antagonists in SW620 Ad300 cells was also affected by calphostin C. Cyclosporin A transport decreased, while verapamil transport increased. Cyclosporin A in calphostin C-treated cells resulted in additive P-glycoprotein antagonism, while no additive effect could be demonstrated with verapamil, suggesting that the increase in verapamil transport makes it a poorer P-glycoprotein antagonist. These studies suggest that transport by P-glycoprotein is a dynamic process which can be modulated by phosphorylation, and that antagonists may block P-glycoprotein differently in different phosphorylation states.
Bates
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SE
Medicine Branch, National Cancer Institute, Bethesda, Maryland 20892.
Lee
J S
JS
Dickstein
B
B
Spolyar
M
M
Fojo
A T
AT
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000345:Affinity Labels / Q000494:pharmacology; D000470:Alkaloids / Q000494:pharmacology; D001386:Azides / Q000494:pharmacology; D001692:Biological Transport / Q000187:drug effects; D002352:Carrier Proteins / Q000378:metabolism*; D016572:Cyclosporine / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D004095:Dihydropyridines / Q000494:pharmacology; D004351:Drug Resistance / Q000502:physiology; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism*; D009281:Naphthalenes; D011083:Polycyclic Compounds / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors*; D020112:Rhodamine 123; D012235:Rhodamines / Q000494:pharmacology; D019311:Staurosporine; D013601:T-Lymphocytes / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D014700:Verapamil / Q000494:pharmacology; D014747:Vinblastine / Q000494:pharmacology; D014750:Vincristine / Q000494:pharmacology
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000345:Affinity Labels; D000470:Alkaloids; D001386:Azides; D002352:Carrier Proteins; D004095:Dihydropyridines; D007527:Isoenzymes; D008562:Membrane Glycoproteins; D009281:Naphthalenes; D011083:Polycyclic Compounds; D012235:Rhodamines; C061465:calphostin complex; D003609:Dactinomycin; D020112:Rhodamine 123; D014750:Vincristine; D014747:Vinblastine; C041130:azidopine; D016572:Cyclosporine; D014700:Verapamil; D011493:Protein Kinase C; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate