Nucleoside monophosphate kinases metabolism

Entecavir, telbivudine, clevudine, and the other nucleoside analogues (Fig. 4aa) need to be phosphorylated to their 5 -triphosphate form to be antivirally active (Fig. 8). This again implies three phosphorylation steps based successively on a nucleoside kinase, nucleoside 5 -monophosphate kinase, and nucleoside 5 -diphosphate kinase. These reactions have been characterized only in a few cases, that is, thymidylate kinase in the metabolism of clevudine (Hu et al. 2005). 

This question was resolved by demonstrations of the intermediate formation of dTDP by Grav and Smellie (38) and by Ives 39). Ives studied the phosphorylation of thymidylate in extracts of the Novikoff rat hepatoma, in which nucleoside diphosphate kinase activity was about three orders of magnitude higher than that of the other enzymes of thymine metabolism present (thymidine and thymidylate kinases, thymidylate synthetase, and thymidylate phosphatase). Ives showed that dTDP was an intermediate in dTTP formation by a method which was independent of the monophosphate and diphosphate kinase reactions. When ATP[7 P] was employed as the phosphorylating agent in the tumor extracts, the dTTP product was labeled in both y- and /3-phosphates when ATP[/3- P] was employed, dTTP was essentially unlabeled ... 

All NRTIs, as exemplified for AZT (Fig. 7), act in a similar fashion following their uptake by the cells, they are phosphorylated successively to their 5 -monophosphate, 5 -diphosphate, and 5 -triphosphate form (De Clercq 2002). Unlike the first phosphorylation step in the metabolic pathway of the acyclic guanosine analogues (see above), which is carried out by a virus-encoded enzyme (thymidine kinase), the first as well as the subsequent phosphorylations of the 2, 3 -dideoxynucleosides are carried out by cellular enzymes, that is, a 2 -deoxynucleoside (e.g., dThd) kinase, a 2 -deoxynucleotide (e.g., dTMP) kinase, and a (2 -deoxy)nucleoside 5 -diphosphate (NDP) kinase. 

Dideoxyuridine (ddU) is an antiviral agent that proved ineffective at controlling human immunodeficiency virus type 1 (HIV-1) infection in human T-cells. This ineffectiveness was ascribed to a lack of substrate affinity of ddU for cellular nucleoside kinases, which prevent it from being metabolized to the active 5 -triphosphate. To overcome this problem, bis[(pivaloyloxy)methyl] 2, 3 -dideoxyuridine 5 -monophosphate (9.41) was prepared and shown to be a membrane-permeable prodrug of 2, 3 -di-deoxyuridine 5 -monophosphate (ddUMP, 9.42) [93]. Indeed, human T-cell lines exposed to 9.41 rapidly formed the mono-, di-, and triphosphate of ddU, and antiviral activity was observed. This example again documents... 

This approach has been applied to the delivery of the 5 -monophosphate of anticancer nucleoside, 2 -deoxy-5-fluorouridine (FdU) [78, 79], in which prodrug (59, R = 5 -FdU) cleaves to FdU 5 -monophosphate and acrolein in the presence of carboxyesterase. The prodrug inhibited DNA synthesis in both Chinese hamster ovary cells and thymidine kinase-deficient mouse fibroblasts this effect has been attributed to intracellular metabolism to FdU 5 -monophosphate and subsequent inhibition of thymidine monophosphate synthase. The prodrug was also as effective as 5-ffuorouracil in prolonging the life-span of mice with P-388 leukaemia. The prodrug was tolerated better by mice when administered with 2-mercaptoethanesulphonic acid, a scavenger for the toxic by-product, acrolein. 

GCV (9- [ 1,3 dihydroxy-2- propo>Q )melhyl]-guanine) is an acyclic nucleoside that is poorly metabolized by mammalian cells and is therefore generally nontoxic. However, after enzymatic conversion to GCV monophosphate (GCV-MP) by HSV/k, it is rapidly metabolized to GCV triphosphate (GCV-TP) by endogenous mammalian kinases. GCV-TP is a poterrt inhibitor of viral DNA polymerase and competes with normal mammalian nucleosides for DNA replication (12). In addition, incorporation of GCV-MP into the DNA template can... 

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