title: Resistance of leukemic cells to 2-chlorodeoxyadenosine is due to a lack of calcium-dependent cytochrome c release.
abstract: The purine nucleoside 2-chlorodeoxyadenosine (CdA) is often used in leukemia therapy. Its efficacy, however, is compromised by the emergence of resistant cells. In the present study, 3 CdA-resistant cell lines were generated and characterized. Their ability to accumulate 2-chloroadenosine triphosphate (CdATP) varied, reflecting differences in activities of deoxycytidine kinase (dCK) and deoxyguanosine kinase (dGK). Nonetheless, the selected lines were uniformly resistant to CdA-induced apoptosis, as assessed by caspase activation and DNA fragmentation. In contrast, cytosols from resistant cells were capable of robust caspase activation when incubated in the presence of cytochrome c and dATP. Moreover, replacement of dATP with CdATP also resulted in caspase activation in the parental and some of the resistant cell lines. Strikingly, CdA-induced decreases in mitochondrial transmembrane potential and release of cytochrome c from mitochondria were observed in the parental cells but not in any resistant lines. The lack of cytochrome c release correlated with an increased ability of mitochondria from resistant cells to sequester free Ca2+. Consistent with this enhanced Ca2+ buffering capacity, an early increase in cytosolic Ca2+ after CdA treatment of parental cells but not resistant cells was detected. Furthermore, CdA-resistant cells were selectively cross-resistant to thapsigargin but not to staurosporine- or Fas-induced apoptosis. In addition, CdA-induced caspase-3 activation and DNA fragmentation were inhibited by the Ca2+ chelator BAPTA-AM in sensitive cells. Taken together, the data indicate that the mechanism of resistance to CdA may be dictated by changes in Ca2+-sensitive mitochondrial events.
authors.affiliation: Institute for Environmental Medicine, Division of Toxicology, and Department of Medicine, Karolinska Institutet, Stockholm, Sweden. chandra.joya@mayo.edu
authors.forename: Joya
authors.initials: J
authors.lastname: Chandra
authors.forename: Emma
authors.initials: E
authors.lastname: Mansson
authors.forename: Vladimir
authors.initials: V
authors.lastname: Gogvadze
authors.forename: Scott H
authors.initials: SH
authors.lastname: Kaufmann
authors.forename: Freidoun
authors.initials: F
authors.lastname: Albertioni
authors.forename: Sten
authors.initials: S
authors.lastname: Orrenius
mesh_terms: D000255:Adenosine Triphosphate / Q000031:analogs & derivatives* / Q000096:biosynthesis; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D020169:Caspases / Q000378:metabolism; D002614:Chelating Agents / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D017338:Cladribine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D003574:Cytochrome c Group / Q000378:metabolism* / Q000494:pharmacology; D053938:DNA Fragmentation / Q000187:drug effects; D003838:Deoxyadenine Nucleotides / Q000494:pharmacology; D003842:Deoxycytidine Kinase / Q000378:metabolism; D019008:Drug Resistance, Neoplasm; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D007425:Intracellular Membranes / Q000502:physiology; D008564:Membrane Potentials / Q000187:drug effects; D008928:Mitochondria / Q000201:enzymology*; D009363:Neoplasm Proteins / Q000378:metabolism*; D014411:Neoplastic Stem Cells / Q000187:drug effects* / Q000201:enzymology; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000032:analysis; D019311:Staurosporine / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology
chemical_list: D000964:Antimetabolites, Antineoplastic; D002614:Chelating Agents; D003574:Cytochrome c Group; D003838:Deoxyadenine Nucleotides; D009363:Neoplasm Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; C097601:cladribine triphosphate; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D017338:Cladribine; D004533:Egtazic Acid; D019284:Thapsigargin; D000255:Adenosine Triphosphate; D017853:Phosphotransferases (Alcohol Group Acceptor); C019165:deoxyguanosine kinase; D003842:Deoxycytidine Kinase; D020169:Caspases; D019311:Staurosporine; C026600:2'-deoxyadenosine triphosphate