PKC-independent modulation of multidrug resistance in cells with mutant (V185) but not wild-type (G185) p-glycoprotein by bryostatin 1

• Published in: Biochemical pharmacology Vol. 56; no. 7; pp. 861 - 869
• Main Authors: Spitaler, Martin, Utz, Irene, Hilbe, Wolfgang, Hofmann, Johann, Grunicke, HansH
• Format: Journal Article
• Language: English
• Published: New York, NY Elsevier Inc 01-10-1998; Elsevier Science
• Subjects: Amino Acid Substitution - genetics; Antineoplastic agents; ATP Binding Cassette Transporter, Subfamily B, Member 1 - biosynthesis; ATP Binding Cassette Transporter, Subfamily B, Member 1 - genetics; ATP Binding Cassette Transporter, Subfamily B, Member 1 - metabolism; Binding, Competitive - drug effects; Biological and medical sciences; bryostatin 1; Bryostatins; Drug Resistance, Multiple; Drug Resistance, Neoplasm; Female; General aspects; HeLa Cells; Humans; KB Cells; Lactones - pharmacology; Macrolides; Medical sciences; multidrug resistance; P-glycoprotein; Pharmacology. Drug treatments; Point Mutation; protein kinase C; Protein Kinase C - physiology; reversal; rhodamine 123; P-glycoprotein; multidrug resistance; reversal; rhodamine 123; protein kinase C; bryostatin 1; Antineoplastic agent; Cell culture; Rhodamine; Protein kinase C; Enzyme; Transferases; P Glycoprotein; Enzyme inhibitor; Molecular interaction; Activation; Gene expression; In vitro; Transfection; Multiple resistance; Established cell line; Mutation; Mechanism of action
• ISSN: 0006-2952; 1873-2968
• DOI: 10.1016/S0006-2952(98)00107-5

Bryostatin 1 is a new antitumor agent which modulates the enzyme activity of protein kinase C (PKC, phospholipid-Ca
2+-dependent ATP:protein transferase, EC 2.7.1.37). Several reports have suggested that the pumping activity of the multidrug resistance gene 1 (MDR1)-encoded multidrug transporter P-glycoprotein (PGP) is enhanced by a PKC-mediated phosphorylation. It was shown here that bryostatin 1 was a potent modulator of multidrug resistance in two cell lines over-expressing a mutant MDR1-encoded PGP, namely KB-C1 cells and HeLa cells transfected with an MDR1-V185 construct (HeLa-MDR1-V185) in which glycine at position 185 (G185) was substituted for valine (V185). Bryostatin 1 is not able to reverse the resistance of cells over-expressing the wild-type form (G185) of PGP, namely CCRF-ADR5000 cells and HeLa cells transfected with a MDR1-G185 construct (HeLa-MDR1-G185). Treatment of HeLa-MDR1-V185 cells with bryostatin 1 was accompanied by an increase in the intracellular accumulation of rhodamine 123, whereas no such effect could be observed in HeLa-MDR1-G185 cells. HeLa-MDR1-V185 cells expressed the PKC isoforms α, δ and ζ. Down-modulation of PKC α and δ by 12-
O-tetradecanoylphorbol-13-acetate (TPA) did not affect the drug accumulation by bryostatin 1. Bryostatin 1 depleted PKC α completely and PKC δ partially. In HeLa-MDR1-V185 cells, short-term exposure to bryostatin 1, which led to a PKC activation, was as efficient in modulating the pumping activity of PGP as long-term exposure leading to PKC depletion. Bryostatin 1 competed with azidopine for binding to PGP in cells expressing the MDR1-V185 and MDR1-G185 forms of PGP. It is concluded that bryostatin 1: i) interacts with both the mutated MDR1-V185 and the wild-type MDR1-G185; ii) reverses multidrug resistance and inhibits drug efflux only in PGP-V185 mutants; and iii) that this effect is not due to an interference of PKC with PGP. For gene therapy, it is important to reverse the specific resistance of a mutant in the presence of a wild-type transporter and vice versa. Our results show that it is possible to reverse a specific mutant PGP.