Rapid collapse of mitochondrial transmembrane potential in HL-60 cells and isolated mitochondria treated with anti-tumor 1,4-anthracenediones.
Since synthetic analogs of 1,4-anthraquinone (AQ code number), such as AQ8, AQ9 and AQ10, can trigger cytochrome c release without caspase activation and retain their ability to induce apoptosis in multidrug-resistant (MDR) tumor cells, fluorescent probes of transmembrane potential have been used to determine whether these anti-tumor compounds might directly target mitochondria in cell and cell-free systems to cause the collapse of mitochondrial membrane potential (/Deltapsim) that is linked to permeability transition pore (PTP) opening. Using JC-1 dye, the abilities of various AQ analogs to induce the /Deltapsim in wild-type and MDR HL-60 cells are rapid (within 2.5-10 min), irreversible after drug removal, concentration dependent in the 0.256-10 micromol/l range and generally related to their anti-tumor activities in vitro. The /Deltapsim caused by AQ9 and AQ10, which are more potent than mitoxantrone, staurosporine and the reference depolarizing agent carbonyl cyanide m-chlorophenylhydrazone (CCCP) in HL-60 cells, are not prevented by caspase-2 or -8 inhibitors, suggesting that activations of these apical caspases upstream of mitochondria are not involved in this process. Antitumor AQ analogs (0.256-10 micromol/l) also mimic the abilities of the known depolarizing agents CCCP, alamethicin, gramicidin A and 100 micromol/l CaCl2 to directly induce within 15 min the /Deltapsim in isolated mitochondria prepared from mouse liver and loaded with rhodamine 123 dye. The fact that 20 micromol/l Ca2+, which is insufficient to trigger depolarization on its own, is required to reveal the depolarizing effect of AQ9 in isolated mitochondria suggests that anti-tumor AQ analogs might interact with the PTP to alter its conformation and increase its Ca2+ sensitivity. Indeed, such Ca2+-dependent /Deltapsim of isolated mitochondria treated with 1.6 micromol/l AQ9 or 100 micromol/l Ca2+ are blocked by ruthenium red. Daunorubicin (DAU) is unable to mimic the rapid /Deltapsim caused by anti-tumor AQ analogs within 2.5-40 min of treatment in HL-60 cells or isolated mitochondria. Moreover, the /Deltapsim caused by 1.6 micromol/l AQ9 or 100 micromol/l Ca2+ in isolated mitochondria are similarly blocked by cyclosporin A (CsA), bongkrekic acid and decylubiquinone, which prevent PTP opening, suggesting that, in contrast to DAU, anti-tumor AQ analogs that directly target mitochondria to trigger the Ca2+-dependent and CsA-sensitive /Deltapsim, might induce PTP opening and the mitochondrial pathway of apoptosis even in the absence of nuclear signals.
Wang
Yang
Y
Anti-Cancer Drug Laboratory, Division of Biology, Ackert Hall, Kansas State University, Manhattan, Kansas 66506-4901, USA.
Perchellet
Elisabeth M
EM
Ward
Mary M
MM
Lou
Kaiyan
K
Hua
Duy H
DH
Perchellet
Jean-Pierre H
JP
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics; D000408:Alamethicin / Q000494:pharmacology; D000818:Animals; D000880:Anthraquinones / Q000737:chemistry / Q000494:pharmacology*; D001865:Bongkrekic Acid / Q000494:pharmacology; D002122:Calcium Chloride / Q000494:pharmacology; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D016572:Cyclosporine / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D003630:Daunorubicin / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D018432:Drug Resistance, Multiple / Q000235:genetics; D005260:Female; D006096:Gramicidin / Q000494:pharmacology; D018922:HL-60 Cells; D006801:Humans; D007425:Intracellular Membranes / Q000187:drug effects* / Q000502:physiology; D007473:Ion Channels / Q000037:antagonists & inhibitors / Q000187:drug effects; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D008928:Mitochondria / Q000187:drug effects* / Q000502:physiology; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D008942:Mitoxantrone / Q000494:pharmacology; D012430:Ruthenium Red / Q000494:pharmacology; D019311:Staurosporine / Q000494:pharmacology; D014451:Ubiquinone / Q000031:analogs & derivatives / Q000494:pharmacology
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000880:Anthraquinones; D015853:Cysteine Proteinase Inhibitors; D007473:Ion Channels; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D001865:Bongkrekic Acid; D012430:Ruthenium Red; D014451:Ubiquinone; D006096:Gramicidin; D000408:Alamethicin; C060262:2,3-dimethoxy-5-methyl-6-decyl-1,4-benzoquinone; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; C416442:1,4-anthraquinone; D016572:Cyclosporine; D008942:Mitoxantrone; D019311:Staurosporine; D002122:Calcium Chloride; D003630:Daunorubicin