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Inhibition of 1-ß-D-Arabinofuranosylcytosine Transport and Net Accumulation by Teniposide and Etoposide in Ehrlich Ascites Cells and Human Leukemic Blasts¹

Department of Biochemistry, Bowman Gray School of Medicine of Wake Forest University, Winston-Salem, North Carolina 27103

The interactions of the epipodophyllotoxins, teniposide (VM-26) and etoposide (VP-16), with the nucleoside carrier were examined with emphasis on their effects on 1-ß-D-arabinofuranosylcytosine (ara-C) transport and net accumulation. VM-26 inhibited ara-C transport by Ehrlich ascites cells within 1 min of exposure, and inhibition was only partially reversed after 45 min in VM-26-free medium. ara-C transport was slowed by 50% by 7 µM VM-26 or by 35 µM VP-16. Since epipodophyllotoxins were noncompetitive inhibitors, fractional inhibition was independent of the ara-C concentration. Analysis of ara-C transport kinetics revealed only a single saturable transport route, and there was no indication of VM-26-insensitive transport. VM-26, VP-16, and ara-C were competitive inhibitors of the specific binding of nitrobenzylthioinosine to the nucleoside carrier with K_i values of 7.4 µM, 23 µM, and 2.2 mM, respectively. The rate of dissociation of nitrobenzylthioinosine (t = 20.6 min) was accelerated by 5 mM ara-C (t = 18.5 min) but slowed by 100 µM VM-26 (t = 34.6). By these criteria, the interaction of VM-26 with the nucleoside carrier was qualitatively similar to that of dipyridamole. Although VM-26 inhibited ara-C transport, it did not significantly slow the rate of net intracellular accumulation of ara-C by Ehrlich cells, presumably because transport capacity far exceeds the capacity for phosphorylation in these cells. In freshly isolated human leukemic blasts, which have far less nucleoside transport activity, inhibition of ara-C accumulation by VM-26 was dependent on the ara-C concentration. At 1 µM ara-C, a concentration where transport was rate limiting for net uptake, VM-26 inhibited accumulation of ara-C over a 60-min time course. At 50 µM ara-C, transport was in excess, and VM-26 did not slow ara-C metabolism.

¹ Supported by Grant CH-289 from the American Cancer Society.

² To whom requests for reprints should be addressed.

Received 12/26/84. Revised 4/ 2/85. Accepted 4/ 5/85.