Thymidine and 3'-azido-3'-deoxythymidine metabolism in human peripheral blood lymphocytes and monocyte-derived macrophages. A study of both anabolic and catabolic pathways.
3'-Azido-3'-deoxythymidine (AZT) is HIV-inhibitory in human macrophages, which is surprising in view of the low AZT phosphorylation reported in macrophage extracts. To elucidate the mechanism of AZT activation, we studied AZT anabolism as well as catabolism in human lymphocytes and macrophages, and compared it to that of thymidine. Thymidine kinase (TK)-specific activity in mitogen-stimulated lymphocytes was 15 times higher than in macrophages. However, the TK activity per cell was only 1.3 times higher, because of the large macrophage cell volume. Total cellular TK activity, but not specific activity, matched the level of intracellular AZT anabolism. The substrate specificity of TK in macrophages strongly suggests that mitochondrial TK2 was the enzyme phosphorylating thymidine and AZT in these cells, whereas it was cytosolic TK1 in stimulated lymphocytes. In vivo thymidine catabolism was extensive, forming thymine and dihydrothymine. In macrophages more than 95% of the added thymidine (0.5 microM) was degraded within 60 min. AZT, in contrast, was not catabolized, which explains the high AZT nucleotide accumulation, a process opposed only by AZTMP excretion. The lack of catabolism together with phosphorylation by TK2 clarifies how AZT can inhibit human immunodeficiency virus in macrophages. The fact that TK2 and not TK1 phosphorylates AZT in macrophages should have important implications for combination chemotherapy.
Arnér
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ES
Medical Nobel Institute, Department of Biochemistry I, Karolinska Institute, Stockholm, Sweden.
Valentin
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A
Eriksson
S
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D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D004247:DNA / Q000378:metabolism; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D008214:Lymphocytes / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D008264:Macrophages / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D010766:Phosphorylation; D010835:Phytohemagglutinins; D013379:Substrate Specificity; D013936:Thymidine / Q000378:metabolism*; D013937:Thymidine Kinase / Q000378:metabolism; D015215:Zidovudine / Q000378:metabolism*
D007527:Isoenzymes; D010835:Phytohemagglutinins; D015215:Zidovudine; D004247:DNA; D013937:Thymidine Kinase; D013936:Thymidine