title: Effects of the selective bisindolylmaleimide protein kinase C inhibitor GF 109203X on P-glycoprotein-mediated multidrug resistance.
abstract: Inhibition of protein kinase C (PKC) is discussed as a new approach for overcoming multidrug resistance (MDR) in cancer chemotherapy. For evaluation of this concept we applied the bisindolylmaleimide GF 109203X, which shows a highly selective inhibition of PKC isozymes alpha, beta 1, beta 2, gamma, delta and epsilon in vitro. The efficacy of this compound in modulation of MDR was examined using several P-glycoprotein (P-gp)-overexpressing cell lines including a MDR1-transfected HeLa clone, and was compared with the activities of dexniguldipine-HCI (DNIG) and dexverapamil-HC1 (DVER), both of which essentially act via binding to P-gp. As PKC alpha has been suggested to play a major role in P-gp-mediated MDR, cell lines exhibiting different expression levels of this PKC isozyme were chosen. On crude PKC preparations or in a cellular assay using a cfos(-711)CAT-transfected NIH 3T3 clone, the inhibitory qualities of the bisindolylmaleimide at submicromolar concentrations were demonstrated. At up 1 microM final concentrations of the PKC inhibitor GF 109203X, a concentration at which many PKC isozymes should be blocked substantially, no cytotoxic or MDR-reversing effects whatsoever were seen, as monitored by 72 h tetrazolium-based colorimetric MTT assays or a 90 min rhodamine 123 accumulation assay. Moreover, depletion of PKC alpha by phorbol ester in HeLa-MDR1 transfectants had no influence on rhodamine 123 accumulation after 24 or 48 h. MDR reversal activity of GF 109203X was seen at higher final drug concentrations, however. Remarkably, [3H]vinblastine-sulphate binding competition experiments using P-gp-containing crude membrane preparations demonstrated similar dose dependencies as found for MDR reversion by the three modulators, i.e. decreasing efficacy in the series dexniguldipine-HCl > dexverapamil-HCl > GF 109203X. Similar interaction with the P-gp in the micromolar concentration range was revealed by competition of GF 109203X with photoincorporation of [3H]azidopine into P-gp-containing crude membrane preparations. No significant effect of the PKC inhibitor on MDR1 expression was seen, which was examined by cDNA-PCR. Thus, the bisindolylmaleimide GF 109203X probably influences MDR mostly via direct binding to P-gp. Our work identifies the bisindolylmaleimide GF 109203X as a new type of drug interacting with P-gp directly, but does not support the concept of a major contribution of PKC to a P-gp-associated MDR, at least using the particular cellular model systems and the selective, albeit general, PKC inhibitor GF 109203X.
authors.affiliation: Byk Gulden GmbH, Konstanz, Germany.
authors.forename: V
authors.initials: V
authors.lastname: Gekeler
authors.forename: R
authors.initials: R
authors.lastname: Boer
authors.forename: F
authors.initials: F
authors.lastname: Uberall
authors.forename: W
authors.initials: W
authors.lastname: Ise
authors.forename: C
authors.initials: C
authors.lastname: Schubert
authors.forename: I
authors.initials: I
authors.lastname: Utz
authors.forename: J
authors.initials: J
authors.lastname: Hofmann
authors.forename: K H
authors.initials: KH
authors.lastname: Sanders
authors.forename: C
authors.initials: C
authors.lastname: Schächtele
authors.forename: K
authors.initials: K
authors.lastname: Klemm
authors.forename: H
authors.initials: H
authors.lastname: Grunicke
mesh_terms: D016475:3T3 Cells; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics / Q000502:physiology*; D000818:Animals; D001386:Azides / Q000378:metabolism; D004095:Dihydropyridines / Q000378:metabolism / Q000494:pharmacology; D018432:Drug Resistance, Multiple; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D015870:Gene Expression; D006801:Humans; D007211:Indoles / Q000494:pharmacology*; D008301:Maleimides / Q000494:pharmacology*; D051379:Mice; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors*; D014700:Verapamil / Q000494:pharmacology
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D001386:Azides; D004095:Dihydropyridines; D004791:Enzyme Inhibitors; D007211:Indoles; D008301:Maleimides; C041130:azidopine; D014700:Verapamil; D011493:Protein Kinase C; C070515:bisindolylmaleimide I; C054074:niguldipine