title: Multidrug-resistant human KB carcinoma cells are highly resistant to the protein phosphatase inhibitors okadaic acid and calyculin A. Analysis of potential mechanisms involved in toxin resistance.
abstract: In this study we show that multidrug-resistant (MDR) human KB-V1 cells are highly resistant to the cytotoxicity of okadaic acid and calyculin A, 2 toxins from marine sponges that are potent inhibitors of type-1 and type-2A protein phosphatases (PP1 and PP2A). Cytotoxicity and colony-forming assays indicated that, relative to parental drug-sensitive KB-3 cells, KB-V1 cells are 35-fold more resistant to okadaic acid and 70-fold more resistant to calyculin A. Cytotoxicity of the toxins was associated with mitotic arrest characterized by chromosome scattering and over-condensation, with KB-3 cells being more sensitive than KB-V1 cells and calyculin A being more potent than okadaic acid. The resistance of KB-V1 cells to both okadaic acid and calyculin A was completely reversed by verapamil, suggesting that the toxins may be transported by P-glycoprotein (P-gp). To further assess the possibility of an interaction with P-gp, the toxins were employed as potential modulators of the photoaffinity labeling of P-gp by [3H]azidopine. Relative to vinblastine, which effectively competed with [3H]azidopine for P-gp photolabeling, calyculin A was 100-fold less potent and okadaic acid did not inhibit photolabeling at concentrations up to 50 microM. To determine whether the resistance mechanism involved differences in toxin-sensitive phosphatase activity, the activity was assayed in extracts from both cell lines and found to be slightly higher (1.6-fold) in KB-V1 than in KB-3 cells. Our results demonstrate a novel, marked resistance of MDR KB-V1 cells to these phosphatase inhibitors and suggest that a major mechanism of resistance may involve toxin transport by P-gp at sites apparently different from those which bind azidopine.
authors.affiliation: Department of Pharmacology, Emory University School of Medicine, Atlanta, GA 30322.
authors.forename: T C
authors.initials: TC
authors.lastname: Chambers
authors.forename: R L
authors.initials: RL
authors.lastname: Raynor
authors.forename: J F
authors.initials: JF
authors.lastname: Kuo
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000345:Affinity Labels; D002470:Cell Survival / Q000187:drug effects; D004351:Drug Resistance / Q000502:physiology*; D004988:Ethers, Cyclic / Q000494:pharmacology*; D006801:Humans; D007624:KB Cells / Q000187:drug effects; D008387:Marine Toxins; D008562:Membrane Glycoproteins / Q000187:drug effects; D009363:Neoplasm Proteins / Q000187:drug effects; D019319:Okadaic Acid; D010080:Oxazoles / Q000494:pharmacology*; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors*; D010777:Photochemistry; D014700:Verapamil / Q000494:pharmacology
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000345:Affinity Labels; D004988:Ethers, Cyclic; D008387:Marine Toxins; D008562:Membrane Glycoproteins; D009363:Neoplasm Proteins; D010080:Oxazoles; D019319:Okadaic Acid; C059041:calyculin A; D014700:Verapamil; D010749:Phosphoprotein Phosphatases