Intracellular concentrations of mitoxantrone in leukemic cells in vitro vs in vivo

The aim of this study was to determine the intracellular pharmacokinetics of mitoxantrone in vivo and to use these results to establish how leukemic cells should be incubated to perform clinically relevant in vitro studies of this drug. Blood samples were obtained from 11 patients with acute nonlymphoblastic leukemia at certain intervals up to 20 h after the infusion of mitoxantrone 12 mg/m2. Plasma and leukemic cells were separated and the drug concentrations were determined with HPLC. Before treatment, leukemic cells from 12 patients were incubated with 0.02, 0.05, 0.1, 0.2 and 1.0 microM mitoxantrone for 1-4 h and thereafter cultured in suspension culture for 20 h; during this time cell samples were taken at certain intervals for drug determination. In cells incubated with 0.05 and 0.2 microM mitoxantrone the cytotoxic effect was measured with the DiSC assay after cultivation for 4-5 days. In vivo, the intracellular levels exceeded the plasma concentrations already at the end of infusion and after 2 h the intracellular concentrations were 200-300 times higher than in plasma. In vitro, the intracellular steady state level of mitoxantrone was reached after 1-2 h and there was a pronounced intracellular retention even after 20 h culture in drug-free medium. Incubation with 0.05 microM during 1 h gave intracellular concentrations of mitoxantrone similar to those achieved in vivo. This incubation concentration gave a mean cytotoxic effect of 53% living cells measured with the DiSC assay, which gives good possibilities to discriminate between mitoxantrone-sensitive and unsensitive cells. We believe that exposing leukemic cells in vitro for in vivo mimicking mitoxantrone concentrations could increase the clinical relevance of predictive assays.