The goal of this application is to establish pharmacologic and biochemical bases for understanding clinical response of human leukemia and for guiding protocol design. The project concentrates on nucleoside analogues that have proven (ara-C) or newly discovered activity (fludarabine, 2-chlorodeoxyadenosine) and one whose activity is under investigation (difluorodeoxycytidine). The rational combination of growth factors (GM-CFS) with chemotherapy requires knowledge of their influence on the pharmacodynamics and pharmacokinetics of nucleotide analogues in human leukemia cells during therapy. The following approaches serve as focal points. 1. Biochemical modulation. A clinical trial has been designed to test the hypothesis that the schedule-specific infusion of fludarabine and ara-C will augment ara-C 5'-triphosphate accumulation in chronic lymphocytic leukemia cells. These investigations will be modeled with in vitro studies to determine the role of fludarabine 5'-triphosphate and dexoynucleoside triphosphates (dNTP) in modulating deoxycytidine kinase. 2. Biological modulation. The action of GM-CSF on ara-C 5'-triphosphate pharmacokinetics and pharmacodynamics will be investigated following ara-C infusions in chronic myelogenous leukemia (CML) blasts serially before and after three days of GM-CSF treatment. Colonogenicity will be evaluated ex vivo in parallel for prognostic significance. 3.Pharmacologic determinants of response. Correlations are being sought between clinical response to a pharmacologically guided phase I trial of difluorodeoxycytidine in relapsed acute leukemia and CML blast crisis and the metabolism and action of this drug in leukemia cells during therapy and ex vivo after treatment. 4. Mechanisms of programmed cell death. The action of fludarabine and 2-chlorodeoxyadenosine on cells of lymphocytic leukemia will be investigated in vitro and during therapy. These studies will focus on DNA damaging mechanisms known to be active in programmed cell death: lowered ATP, dNTP, and NAD+ pools, ADP ribosylation, and DNA fragmentation.
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