De novo induced mutations in the deoxycytidine kinase (dck) gene in rat leukemic clonal cell lines confer resistance to cytarabine (AraC) and 5-aza-2'-deoxycytidine (DAC).
We have investigated whether cytarabine (AraC) or decitabine (DAC) induce deficiency of deoxycytidine kinase (DCK) through different mutations of the dck gene, related to their distinct interference with DNA replication. Also, it is not known whether mutations of the dck gene are the result of selection of mutants or de novo induction. To address these issues, three subclones of a rat leukemic cell line (RCL/O), sensitive to cytotoxicity mediated by AraC and DAC, were exposed to gradually increasing concentrations (from 0.1 to 10 microM) of either AraC or DAC over a 140 days vs a 180 days period. During the course of resistance induction DCK activity was monitored. We found that all clones acquired irreversible cross-resistance, at marginally cytotoxic AraC or DAC concentrations of 0.1 to 0.4 times the IC50 for the parental clones. Furthermore, all resistant cell lines were DCK deficient and harbored different mutations in the dck gene. AraC induced both rearrangements and point mutations in the dck gene when administered over 140 days and 180 days, respectively. 140 days DAC induction yielded point mutations only. All point mutations detected were nonrandomly distributed within the dck coding region. SSCP analysis showed that in the majority of resistant clones more than one bandshift was present. The data suggest the presence of multiple resistant clones, originating from one sensitive clone and thus arguing against selection of mutants as a mechanism for the development of AraC and DAC resistance.
Stegmann
A P
AP
Department of Hematology, University Hospital Leiden, The Netherlands.
Honders
M W
MW
Willemze
R
R
Landegent
J E
JE
D000818:Animals; D000970:Antineoplastic Agents / Q000633:toxicity*; D001374:Azacitidine / Q000031:analogs & derivatives* / Q000633:toxicity; D015139:Blotting, Southern; D002460:Cell Line; D003561:Cytarabine / Q000633:toxicity*; D000077209:Decitabine; D003842:Deoxycytidine Kinase / Q000096:biosynthesis / Q000235:genetics*; D004351:Drug Resistance / Q000235:genetics; D015870:Gene Expression; D007700:Kinetics; D007942:Leukemia, Experimental / Q000201:enzymology*; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D051381:Rats; D015183:Restriction Mapping; D013997:Time Factors; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D003561:Cytarabine; D000077209:Decitabine; D003842:Deoxycytidine Kinase; D001374:Azacitidine