Role of deoxycytidine kinase in an in vitro model for AraC- and DAC-resistance: substrate-enzyme interactions with deoxycytidine, 1-beta-D-arabinofuranosylcytosine and 5-aza-2'-deoxycytidine.
Deoxycytidine kinase activity (dCk) was monitored in cell lines from a rat acute myeloid leukemia model of acquired resistance to cytosine arabinoside (AraC) and decitabine (DAC). In both AraC-resistant cell lines (RCL/A and its subclone RA/7), as well as in a DAC-resistant cell line (RCL/D) which we generated from the drug-sensitive RCL/0 cell line, a total deficiency of dCk activity and a cross-resistance for AraC and DAC was demonstrated. Furthermore, the metabolization of deoxycytidine (dC) was severely impaired in all these cell lines. Km values for dC (9.4 microM in RCL/0 cells) had increased 70- to 100-fold in RCL/D (Km = 673.2 microM), in RCL/A (Km = 947.2 microM) and in RA/7 (Km = 817.5 microM). Vmax values were unaltered in RCL/D and RA/7, and twofold increased in RCL/A. Addition of hydroxyurea (HU) to cell cultures stimulated dCk salvage pathway activity in RCL/0 cells for dC, AraC, and DAC by increasing Vmax values approximately 160% leaving Km constants unchanged. In all resistant cell lines, HU pre-incubation did not influence the level of dCk activity, leaving Km and Vmax values unaltered. These data indicate that deficiency of dCk activity is crucial in the mechanism of drug resistance in this model.
Stegmann
A P
AP
Department of Hematology, University Hospital Leiden, The Netherlands.
Honders
M W
MW
Kester
M G
MG
Landegent
J E
JE
Willemze
R
R
D000208:Acute Disease; D000818:Animals; D000970:Antineoplastic Agents / Q000378:metabolism*; D001374:Azacitidine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D001667:Binding, Competitive; D003561:Cytarabine / Q000378:metabolism / Q000494:pharmacology*; D000077209:Decitabine; D003841:Deoxycytidine / Q000378:metabolism; D003842:Deoxycytidine Kinase / Q000378:metabolism*; D004351:Drug Resistance; D066298:In Vitro Techniques; D007700:Kinetics; D007951:Leukemia, Myeloid / Q000188:drug therapy; D051381:Rats; D011914:Rats, Inbred BN; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D003561:Cytarabine; D003841:Deoxycytidine; D000077209:Decitabine; D003842:Deoxycytidine Kinase; D001374:Azacitidine