title: Dose-dependent effects of the caspase inhibitor Q-VD-OPh on different apoptosis-related processes.
abstract: The effects of the pan-caspase inhibitor Q-VD-OPh on caspase activity, DNA fragmentation, PARP cleavage, 7A6 exposition, and cellular adhesivity to fibronectin were analyzed in detail in three different apoptotic systems involving two cell lines (JURL-MK1 and HL60) and two apoptosis inducers (imatinib mesylate and suberoylanilide hydroxamic acid). Q-VD-OPh fully inhibited caspase-3 and -7 activity at 0.05  µM concentration as indicated both by the measurement of the rate of Ac-DEVD-AFC cleavage and anti-caspase immunoblots. Caspase-8 was also inhibited at low Q-VD-OPh concentrations. On the other hand, significantly higher Q-VD-OPh dose (10 µM) was required to fully prevent the cleavage of PARP-1. DNA fragmentation and disruption of the cell membrane functionality (Trypan blue exclusion test) were both prevented at 2 µM Q-VD-OPh while 10 µM inhibitor was needed to inhibit the drug-induced loss of cellular adhesivity to fibronectin which was observed in JURL-MK1 cells. The exposition of the mitochondrial antigen 7A6 occurred independently of Q-VD-OPh addition and may serve to the detection of cumulative incidence of the cells which have initiated the apoptosis. Our results show that Q-VD-OPh efficiency in the inhibition of caspase-3 activity and DNA fragmentation in the whole-cell environment is about two orders of magnitude higher than that of z-VAD-fmk. This difference is not due to a slow permeability of the latter through the cytoplasmic membrane.
authors.affiliation: Department of Cellular Biochemistry, Institute of Hematology and Blood Transfusion, U Nemocnice 2, 128 20 Prague 2, Czech Republic. katerina.kuzelova@uhkt.cz
authors.forename: Kateřina
authors.initials: K
authors.lastname: Kuželová
authors.forename: Dana
authors.initials: D
authors.lastname: Grebeňová
authors.forename: Barbora
authors.initials: B
authors.lastname: Brodská
mesh_terms: D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D061945:Caspase Inhibitors; D002448:Cell Adhesion; D045744:Cell Line, Tumor; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D011804:Quinolines / Q000494:pharmacology*
chemical_list: D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D011804:Quinolines; C468548:quinoline-val-asp(OMe)-CH2-OPH