Isolation and Characterization of a Deoxycytidine Kinase-deficient Human Promyelocyte Leukemic Cell Line Highly Resistant to 1-β-D- Arabinofuranosylcytosine

A deoxycytidine kinase-deficient variant of a human promyelocytic leukemic cell line (HL-60/ara-C) has been isolated and characterized. These cells are capable of proliferating in the presence of 10^-6 M 1-β-D-arabinofuranosylcytosine (ara-C), a level achieved in the plasma of leukemic patients undergoing conventional-dose ara-C therapy. The cells share numerous biological and biochemical features with the parent line, including: morphology; rate of growth; cloning characteristics; karyotype; rates of DNA, RNA, and protein synthesis; and ability to undergo terminal differentiation in the presence of agents such as 12-O-tetradecanoylphorbol acetate and dimethyl sulfoxide. In contrast, these cells display a great reduction in the total intracellular accumulation of ara-C following a 4-hr exposure to 10^-6 M ara-C (2.4 versus 99.0 pmol ara-C/10⁶ cells). Resistant cells exposed to 10^-6 M ara-C for 1 hr also exhibited a reduction in the generation [1.2 versus 31.9 pmol 1-β-D-arabinofuranosylcytosine 5'-triphosphate (ara-CTP)/10⁶ cells] and the 4-hr retention (0.30 versus 3.87 pmol ara-CTP/10⁶ cells) of ara-CTP, the lethal ara-C metabolite, in comparison to parent cells. Incorporation of ara-C into resistant HL-60 cell DNA was also profoundly decreased. These biochemical alterations were associated with a 1000-fold decrease in the sensitivity of clonogenic cells to continuously administered ara-C (ara-C 50% inhibitory concentration: 1.8 x 10^-6 M for HL-60/ara-C; 3.0 x 10^-9 M for HL-60). A variety of antagonists of de novo pyrimidine synthesis inhibited the growth of ara-C-sensitive and -resistant cells to a similar extent. When HL-60 cells were exposed to a lethal concentration of thymidine (5 x 10^-3 M), coadministration of 5 x 10^-6 M deoxycytidine restored 90 ± 4% (S.D.) of colony-forming capacity. Normal human bone marrow progenitor cells were protected to a similar degree by 3 x 10^-3 M deoxycytidine. In contrast, deoxycytidine concentrations as high as 5 x 10^-3 M were unable to confer any protection to HL-60/ara-C cells under identical conditions. These studies suggest that an enzymatic perturbation rendering human leukemic cells highly resistant to ara-C may be exploited to achieve a selective in vitro chemotherapeutic effect.