Arabinosylguanine: Metabolism, action, and lineage-specific cytotoxicity in human leukemia cells
9-β-D-arabinosylguanine (ara-G), an analogue of deoxyguanosine, has demonstrated T-lymphoblast selective anti-leukemia activity both in vitro and in vivo in cell lines and primary cells and in phase I investigations. The present work was initiated to identify factors that result in this selectivity. The cytotoxicity of ara-G is manifest only after its phosphorylation. Experiments using cell lines transfected to overexpress specific nucleoside kinases demonstrated that the phosphorylation of ara-G to its monophosphate is by both cytoplasmic deoxycytidine kinase and mitochondria) deoxyguanosine kinase. Ara-G monophosphate is converted to its 5′-triphosphate (ara-GTP) in cells by these kinases and then incorporated into DNA. Mechanistic studies demonstrated that incorporation of ara-GTP into DNA was a necessary event for the induction of cell death. Pharmacokinetic and pharmacodynamic studies utilizing three human acute leukemia cell lines, CEM (T-lymphoblastic), Raji (B-lymphoblastic), and ML-1 (myeloid) were performed. CEM cells were most sensitive to ara-G-induced inhibition of colony formation, accumulated ara-GTP at a faster rate and to a greater degree than either Raji or ML-1, but incorporated the lowest number of ara-G molecules into DNA. The position of incorporation was internal and similar in all cell lines. The terminal elimination phase of ara-GTP was >24 h and similar in these cells. Comparisons between inhibition of colony formation and ara-GTP incorporation into DNA demonstrated that while within a cell line there was correlation among these parameters, between cell lines there was no relationship between number of incorporated ara-G molecules and ara-G(TP)-mediated toxicity suggesting that there were additional factors. The expression of membrane bound Fas and Fast was unchanged in all cell lines. In contrast, there was a 2-fold increase in soluble Fast, which was found exclusively in CEM cells. Ara-G-mediated apoptosis in CEM occurred from all phases of the cell cycle and was abrogated partially by Fas antagonist antibodies. These data suggest that Fas-mediated cell death due to the liberation of sFasL may be responsible for the hypersensitivity to ara-G manifested by immature T-cells such as CEM. The role of Fas in ara-G induced death of acute T-lymphoblastic leukemia cells during therapy needs to be tested.
Rodriguez, Carlos O., "Arabinosylguanine: Metabolism, action, and lineage-specific cytotoxicity in human leukemia cells" (2001). Texas Medical Center Dissertations (via ProQuest). AAI3004456.