Pharmacokinetic and pharmacodynamic studies of fludarabine and cytosine arabinoside administered as loading boluses followed by continuous infusions after a phase I/II study in pediatric patients with relapsed leukemias. The Children's Cancer Group.

The sequential administration of fludarabine followed by cytosine arabinoside (ara-C) has demonstrated significant synergistic effects against the CEM human leukemic cell line. This in vitro synergism was investigated in a Phase I trial in pediatric patients with relapsed acute leukemia. The optimum concentrations of 9-beta-D-arabinofuranosyl 2-fluoroadenine and ara-C necessary to achieve significant drug synergism from in vitro studies were between 10 and 20 microM. Fludarabine was infused at a dose to attain a target plasma concentration of 10 microM for 48 h, followed by a continuous infusion of escalated ara-C doses to maintain plasma ara-C concentrations of 10, 12.5, 15, or 17.5 microM for 72 h. Thirteen patients with acute lymphocytic leukemia and 18 with acute myelocytic leukemia were entered into the study, 30 of whom were clinically evaluable for toxicity. Pharmacokinetic and pharmacodynamic studies were performed on specimens from 20 patients. The optimal 9-beta-D-arabinofuranosyl 2-fluoroadenine and ara-C concentrations in plasma were easily achieved after continuous infusion regimens of both drugs. Cellular ara-CTP is augmented 5-8-fold in leukemic cells from patients receiving fludarabine phosphate treatment followed by ara-C. The maximum tolerated plasma concentrations for this combination regimen was 10 microM fludarabine for 48 h followed by 72 h of 15 microM ara-C, which were achieved at dose level 3. A significant number of responses were also seen. Nine of 18 evaluable patients (50%) with acute myelocytic leukemia achieved complete or partial responses, and 3 of 9 evaluable patients with acute lymphocytic leukemia achieved complete or partial responses. Fludarabine and ara-C successfully eradicated bone marrow disease in 16 of 27 patients (59%), 23 patients of which had been treated previously with high-dose ara-C. These results verified the synergistic effect fludarabine exhibited in augmenting ara-CTP concentrations in patients' leukemic blasts, thus improving the clinical response in relapsed pediatric leukemias.

Intracellular pharmacodynamic studies of the synergistic combination of 6-mercaptopurine and cytosine arabinoside in human leukemia cell lines.

Selective combinations of purine and pyrimidine analogs increase remission rates in pediatric patients with relapsed leukemias. The combination of 6-mercaptopurine (6-MP) and cytosine arabinoside (ara-C) may exhibit synergism similar to that observed for fludarabine and ara-C and may diminish the potential for development of resistance since the two drugs are activated by separate enzymatic pathways. To determine the efficacy of the combination against human leukemia cells, we investigated the time-concentration relationships of the drugs given alone or in combination to the resultant cytotoxicity. To determine whether the combination leads to enhanced activity of deoxycytidine kinase (dCk), the rate-limiting enzyme in ara-C activation, we characterized the cellular dCk in CCRF/CEM/0, CCRF/CEM/ara-C/7A, and CCRF/CEM/ara-C/3A monoclonal cells before and after treatment with 6-MP. CCRF/CEM/0 (wild type), CCRF/CEM/ara-C/7A (approximately 50% ara-C-resistant as determined by ara-C sensitivity assay and dCk characterization), and CCRF/CEM/ara-C/3A (approximately 90% resistant to ara-C) human leukemia cells were incubated with various concentrations of 6-MP and ara-C given alone or in combination. Cell survival, inhibition of DNA synthetic capacity (DSC), ara-CTP anabolism, and dCk enzymatic characteristics were studied. Incubation of CEM/0 cells with 6-MP for 24 h, followed by ara-C for 48 h, increased cell-growth inhibition by approximately 0.5-1 log10, corresponding to 5- to 10-fold synergism, as compared with ara-C alone after identical drug incubation in all cell lines. Simultaneous administration showed no synergism, whereas reversal of the sequence produced an antagonistic effect. The ara-CTP levels were 2- to 3.5-fold and 3- to 5-fold higher in CEM/0 and CEM/ara-C/7A cells, respectively, in cells exposed to 6-MP followed by ara-C than in those exposed to ara-C alone at the same concentrations. Furthermore, a progressive increase in ara-CTP levels was noted in CEM/0 cells exposed to increasing concentrations of 6-MP followed by 10 or 20 microM ara-C. A significant decrease in DSC was observed upon treatment of wild-type and ara-C-resistant cells with 6-MP and ara-C. The combination of 6-MP and ara-C exhibits significant sequence-specific synergism in both wild-type and partially ara-C-resistant leukemia cell lines. The combination also exerts collateral sensitivity in the ara-C-resistant cell lines. 6-MP pretreatment may play a role in enhancing ara-C activation, thus producing drug synergism in sensitive and resistant leukemia cell lines.