In-vitro studies on phosphorylation and dephosphorylation of cytosine arabinoside in human leukemic cells

Abstract

The cytotoxic effect of cytosine arabinoside (ara-C) depends on the capacity of cells to form and retain intracellularly the phosphorylated metabolite cytosine arabinoside triphosphate (ara-CTP). In this study accumulation and cellular retention of ara-CTP have been measured in vitro in the bone marrow cells of 69 patients with acute leukemia. Cells were incubated with ³H-ara-C and the amount of ara-CTP formed was determined after separation of the nucleotides by thin-layer chromatography.

Phosphorylation of ara-C to ara-CTP appeared to be a saturable process. The K_m-equivalents varied between 1.1 and 16.2 μM ara-C. Maximal ara-CTP formation ranged from 12 to 125 pmol ara-CTP/10⁶ cells in 30 min. The phosphorylation activity did not correlate with the percentage of S-phase cells. The intracellular half-life time of ara-CTP measured in vitro ranged from 53 to 210 min.

Phosphorylation of ara-C was comparable in patients with acute myeloid leukemia (n = 51) and in patients with acute lymphoblastic leukemia (n = 18). Ara-CTP elimination appeared slower in lymphoblasts than in myeloblasts.

The average intracellular ara-CTP level in relapsed patients (n = 34) appeared higher than in previously untreated patients (n = 52). The less favourable outcome of second remission induction therapy with conventional doses of ara-C compared to the first remission induction treatment is not explained by an alteration in the intracellular metabolism of ara-C.