IN VITRO-INDUCED RESISTANCE TO THE DEOXYCYTIDINE ANALOGS CYTARABINE (ARAC) AND 5-AZA-2'-DEOXYCYTIDINE (DAC) IN A RAT MODEL FOR ACUTE MYELOID-LEUKEMIA IS MEDIATED BY MUTATIONS IN THE DEOXYCYTIDINE KINASE (DCK) GENE

The deoxycytidine kinase (dck) gene encodes the enzyme responsible for the metabolic activation of the antileukemic drugs cytosine arabinosi de (AraC) and 5-aza-2'-deoxycytidine (decitabine, DAC). The dck locus was analyzed at the chromosomal and the molecular level in a model of rat leukemic cell lines, in which AraC and DAC resistance was induced, that was marked by dck deficiency. At the chromosomal level, karyotyp e analysis of metaphase spreads revealed the presence of an aberrant 2 q + chromosome in the AraC-resistant cell line and a (Xq;11q) transloc ation in its subclone RA/7. The DAC-resistant lines were identical to the parental RCL/O. Fluorescence in situ hybridization on normal rat f ibroblast metaphase spreads localized the rat dck gene to chromosome 1 4q21-q22, a region that was not involved in any of the observed karyot ypic aberrations. Analysis at the molecular level revealed an identica l rearrangement of the dck gene in the AraC-resistant cell line RCL/A and its subclone RA/7 that resulted in the absence of dck expression, as assessed by RT-PCR. No genomic rearrangements were observed in a DA C-resistant cell line RCL/D or in its subclone RD/1. However, detectio n of a single-stranded conformation polymorphism (SSCP) allowed the id entification of a single C to G substitution (His to Gln) in the dck c DNA of the DAC-resistant RD/1 clone. The data demonstrate that exposur e to AraC and DAC induces a resistant phenotype marked by functional d ck deficiency that may be the consequence of mutations occurring in th e dck gene.