title: Comparison of the phosphorylation of 4'-ethynyl 2',3'-dihydro-3'-deoxythymidine with that of other anti-human immunodeficiency virus thymidine analogs.
abstract: Thymidine analogs, including 3'-azido-3'-deoxythymidine (AZT) and 2',3'-dideoxy-3'-deoxythymidine (D4T), are important antiretroviral agents. To exert antiretroviral activity, these analogs undergo a stepwise phosphorylation intracellularly to the active triphosphate metabolites. We previously reported that 4'-substituted D4T with an ethynyl group (i.e., 4'-ethynyl D4T) increased the anti-human immunodeficiency virus (HIV) activity and was active against multidrug-resistant HIV strains. 4'-Ethynyl D4T is a better substrate for phosphorylation by human thymidine kinase 1 than D4T is. In this report, we first studied the enzymes involved in the phosphorylation of 4'-ethynyl D4T from monophosphate to triphosphate metabolites. The 4'-ethynyl D4TMP is phosphorylated by recombinant human TMP kinase with a K(m) of 19 +/- 4 microM and a k(cat) of 0.007 +/- 0.001 s(-1); the relative efficiency is about 9 and 15% of those of D4TMP and AZTMP, respectively. Several enzymes from crude cellular extracts, including nucleoside diphosphate kinase, pyruvate kinase, creatine kinase, and 3-phosphoglycerate kinase, could phosphorylate 4'-ethynyl D4T-diphosphate. The relative phosphorylation efficiencies of 4'-ethynyl D4TDP were about 3 to 25% of those of D4TDP and were generally similar to those of AZTDP. In T-lymphoid cell lines, there was a preponderant accumulation of 4'-ethynyl D4TMP, suggesting that TMP kinase could be the rate-limiting enzyme in the metabolism of 4'-ethynyl D4T. Although the same enzymes are involved in the stepwise phosphorylation of thymidine analogs, their behaviors in phosphorylating metabolites of 4'-ethynyl D4T are different from those of D4T and AZT. Qualitatively, the metabolism of 4'-ethynyl D4T is more similar to that of AZT than to that of its progenitor, D4T.
authors.affiliation: Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06520, USA.
authors.forename: Chih-Hung
authors.initials: CH
authors.lastname: Hsu
authors.forename: Rong
authors.initials: R
authors.lastname: Hu
authors.forename: Ginger E
authors.initials: GE
authors.lastname: Dutschman
authors.forename: Guangwei
authors.initials: G
authors.lastname: Yang
authors.forename: Preethi
authors.initials: P
authors.lastname: Krishnan
authors.forename: Hiromichi
authors.initials: H
authors.lastname: Tanaka
authors.forename: Masanori
authors.initials: M
authors.lastname: Baba
authors.forename: Yung-Chi
authors.initials: YC
authors.lastname: Cheng
mesh_terms: D019380:Anti-HIV Agents / Q000494:pharmacology*; D006801:Humans; D009703:Nucleoside-Phosphate Kinase / Q000378:metabolism; D010766:Phosphorylation; D018119:Stavudine / Q000494:pharmacology*; D013329:Structure-Activity Relationship; D015215:Zidovudine / Q000494:pharmacology*
chemical_list: D019380:Anti-HIV Agents; D015215:Zidovudine; D018119:Stavudine; D009703:Nucleoside-Phosphate Kinase; C020093:dTMP kinase