Fagny Christine, Vandevelde Michel, Svoboda Michal, Robberecht Patrick
Department of Biochemistry and Nutrition, Faculty of Medicine, Université Libre de Bruxelles, Brussels, Belgium.
Biochem Pharmacol. 2002 Aug 1;64(3):451-6. doi: 10.1016/s0006-2952(02)01185-1.
Azodicarbonamide tested as an anti-HIV agent was reported to expulse zinc from viral zinc-cysteine factors and to inhibit calcium mobilization machinery. It has structural analogy with hydroxyurea that inhibits ribonucleotide reductase and could also act on this target. Azodicarbonamide was therefore tested for its capacity to modulate deoxyribonucleotides triphosphate pools alone or in combination with other agents in the lymphoblastic SUP-T1 cell line susceptible to HIV infection. The deoxyribonucleotides triphosphate were evaluated by an enzymatic assay using sequenase. Two hours exposure of SUP-T1 cells to 100 microM azodicarbonamide induced a 50% reduction of each deoxyribonucleotide triphosphate. Among other inhibitors of nucleotide metabolism (hydroxyurea, methotrexate and thymidine), hydroxyurea only reproduces the effect of azodicarbonamide. This suggests, but does not demonstrate directly, that azodicarbonamide inhibits ribonucleotide reductase activity. The combination of azodicarbonamide with each of these inhibitors affected particularly the dCTP pool. During this study it was also suggested that azodicarbonamide could interfere with thymidine phosphorylation. Thymidine phosphorylating activity was measured with 3H-thymidine as substrate. In acellular preparations, azodicarbonamide also non-competitively inhibits thymidine phosphorylating activity. This effect was not reproduced by hydroxyurea. Thus, in vitro azodicarbonamide decreases the intracellular pool of deoxyribonucleotide and thymidine phosphorylation.
据报道，作为一种抗HIV药物进行测试的偶氮二甲酰胺可从病毒锌 - 半胱氨酸因子中排出锌，并抑制钙动员机制。它与抑制核糖核苷酸还原酶的羟基脲具有结构相似性，也可能作用于该靶点。因此，在易受HIV感染的淋巴细胞SUP - T1细胞系中，测试了偶氮二甲酰胺单独或与其他药物联合调节三磷酸脱氧核糖核苷酸池的能力。使用测序酶通过酶促测定法评估三磷酸脱氧核糖核苷酸。将SUP - T1细胞暴露于100 microM偶氮二甲酰胺两小时，可使每种三磷酸脱氧核糖核苷酸减少50%。在其他核苷酸代谢抑制剂（羟基脲、甲氨蝶呤和胸苷）中，只有羟基脲能重现偶氮二甲酰胺的作用。这表明，但未直接证明，偶氮二甲酰胺抑制核糖核苷酸还原酶活性。偶氮二甲酰胺与这些抑制剂中的每一种联合使用尤其影响dCTP池。在这项研究中还表明，偶氮二甲酰胺可能干扰胸苷磷酸化。以3H - 胸苷为底物测量胸苷磷酸化活性。在无细胞制剂中，偶氮二甲酰胺也非竞争性抑制胸苷磷酸化活性。羟基脲未重现这种作用。因此，在体外，偶氮二甲酰胺可降低细胞内脱氧核糖核苷酸池和胸苷磷酸化水平。
