82 Purine and Pyrimidine Metabolism of Normal and Leukemic Cells

Abstract

Using human leukemia cell lines, w e investigated the biochemical basis for the synergistic interaction between ara-C and hydroxyurea(HU) or methotrexate(MTX1 I n the cells of B cell phenotype, pretreatment with 1mM HU increased ara-CTP formation up to 2-10 fold, whereas it did not have significant effects o n T cell lines. The phosphorylation of ara-A or deoxyadenosine to the corresponding triphosphates was also augmented by HU, indicating the activation o f deoxycytidine(CdR) kinase in intact cells through HU treatment. As a possible mechanism, we propose the important role of d e novo CdR production, since (i) the changes o f pyrimidine dNTP pools by HU ( the decrease in dCTP and increase in TTP ) were equally seen in both cell lineg (ii) the excretion of CdR into the medium was much higher in B cell lines, which was significantly inhibited by HU. On the other hand, MTX(1p) increased ara-CTP generation significantly in both cell lines, albeit it was less than 50%. These results suggest the cell type specific effectiveness of such combination chemotherapyin acute leukemia. We have succeeded in the isolation for a 2.3 kb cDNA for PRPP synthetase from pig.The DNA sequence determined from che coding region shows a high degree of homology with the cDNA sequence recently determined from rat.The enzyme urate oxidase catalyzes the oxidation of uric acid to allantoin in all mammals except humans and certain primates. Recently, mice with complete HPRT deficiency have been identified. These animals display none of the neurological symptoms observed in human patients. This has raised the possibility that the absence of urate oxidase activity in man may contrihute to the neurological symptoms observed in L-N patients. A 2.2 kb cDNA for urate oxidase has been isolated from a porcine liver X cDNA library.Although humans demonstrate no urate oxidase activity, analysis of human genomic DNA has revealed the presence of homologous sequences in human.The loss of urate oxidase activity in human is probably due to a lack of gene transcription since a Northern analysis detects no urate oxidase mRNA in human liver. DEGRADATION OF PURINE NUCLEOSIDES BY MITOCHONDRIAL ENZYMES OF BOVINE LIVER.79 Roger A. Lewis, Laura Link and Wilbert Chen.University of Nevada-Reno. Dept. of Biochemistry, Reno, NV 89557, U.S.A.In recent years, it has been shown that mitochondria actively metabolize purine nucleosides. It is documented that both deoxyguanosine and deoxyadenosine are phosphorylated by mitochondrial enzymes. In the case of deoxyguanosine, intact mitochondria form dGHP, dGDP and dGTP. Herein we report that mitochondria are also involved in the degradation both deoxvauanosine and deoxyadenosine. When intact or fractured . -mitochondria are treated with deoxyadenosine, there is an immediate loss of the deoxynucleoside and a corresponding formation of deoxyinosine. A partially purified preparation of the mitochondrial adenosine deaminase activity was able to deaminate both adenosine and deoxyadenosine and a LineweaverBurk plot showed that an ap...