The Role of Human Nucleoside Transporters in Cellular Uptake of 4′-Thio-β-d-arabinofuranosylcytosine and β-d-Arabinosylcytosine

4′-Thio-β- d -arabinofuranosyl cytosine (TaraC) is in phase I development for treatment of cancer. In human equilibrative nucleoside transporter (hENT) 1-containing CEM cells, initial rates of uptake (10 μM; picomoles per microliter of cell water per second) of [ 3 H]TaraC and [ 3 H]1-β- d -arabinofuranosyl cytosine (araC) were low (0.007 ± 003 and 0.034 ± 0.003, respectively) compared with that of [ 3 H]uridine (0.317 ± 0.048), a highactivity hENT1 permeant. In hENT1- and hENT2-containing HeLa cells, initial rates of uptake (10 μM; picomoles per cell per second) of [ 3 H]TaraC, [ 3 H]araC, and [ 3 H]deoxycytidine were low (0.30 ± 0.003, 0.42 ± 0.03, and 0.51 ± 0.11, respectively) and mediated primarily by hENT1 (∼74, ∼65, and ∼61%, respectively). In HeLa cells with recombinant human concentrative nucleoside transporter (hCNT) 1 or hCNT3 and pharmacologically blocked hENT1 and hENT2, transport of 10 μM[ 3 H]TaraC and [ 3 H]araC was not detected. The apparent affinities of recombinant transporters (produced in yeast) for a panel of cytosine-containing nucleosides yielded results that were consistent with the observed low-permeant activities of TaraC and araC for hENT1/2 and negligible permeant activities for hCNT1/2/3. During prolonged drug exposures of CEM cells with hENT1 activity, araC was more cytotoxic than TaraC, whereas coexposures with nitrobenzylthioinosine (to pharmacologically block hENT1) yielded identical cytotoxicities for araC and TaraC. The introduction by gene transfer of hENT2 and hCNT1 activities, respectively, into nucleoside transport-defective CEM cells increased sensitivity to both drugs moderately and slightly. These results demonstrated that nucleoside transport capacity (primarily via hENT1, to a lesser extent by hENT2 and possibly by hCNT1) is a determinant of pharmacological activity of both drugs.