Deoxyuridine kinase

This deoxyribonucleoside prodrug (Fig. 42.26) is bioconverted via 2 -deoxyuridine kinase-mediated phosphorylation to the same active 5-fluoro-dUMP structure generated in the multistep biotransformation of... 

Ribonucleotide reductase works on ribo-A, -U, -G, -C diphosphates to give the deoxynucleotide. The deoxyuridine, which is useless for RNA synthesis, is converted to deoxythymidine by the enzyme thymidylate synthase, which uses methylene tetrahydrofolate as a one-carbon donor. The odd thing here is that ribonucleotide reductase uses the UDP as a substrate to give the dUDP. This must then be hydrolyzed to the dUMP before thymidylate synthase will use it to make dTMP. Then the dTMP has to be kinased (phosphorylated) up to dTTP before DNA can be made. 

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... 

Fig. lA. Anabolic and catabolic pathways of 5-FU. DPD dihydropyrimidine dehydrogenase, DP di-hydropyrimidinase, pUP beta-ureidopropionase, UP uridine phosphorylase, OPRT orotate phospho-ribosyl transferase, UK uridine kinase, TP thymidine phosphorylase, TK thymidine kinase, RNR ribonucleotide reductase. The three active metabolites (shown in rectangles) are FdUMP (5-fluoro-2 -deoxyuridine 5 -monophosphate) inhibiting TS (thymidylate synthase), and FUTP (5-fluorouridine 5 -triphosphate) and FdUTP (5-fluoro 2 -deoxyuridine 5 -triphosphate) interfering with RNA and DNA, respectively. 

Cells that have been preincubated with deoxyuridine, then exposed to [ H] thymidine, incorporate little or none of the labeled material into DNA. This is because of both dilution of the labeled material in the larger intracellular pool of newly synthesized TMP and also inhibition of thymidylate kinase by thymidine triphosphate. 

The deoxyuridine analogue 5-azidomethyl-2 -deoxyuridine (159) was one of a large number of compounds investigated as inhibitors of thymidylate synthase and thymidine kinase from L1210 cells, but the compound was... 

A new series of 5-carboranyl-substituted-2 -deoxyuridine (25a) and deoxy-thymidine (25b) derivatives containing a range of alkyl spacers has been prepared to access a more effective use of boron neutron capture therapy. Evaluation of these derivatives as substrates for the human thymidine kinases TKl and TK2 showed that a decrease in the length of the spacer (from 8 methylene units to 4) between the carborane moiety and the pyrimidine base resulted in better substrate characteristics. 

AflBnity Labellii. —Of a large number of 5-substituted-2 -deoxyuridine 5 -mono-phosphates prepared as potential inhibitors of thymidylate kinase, 5-formyl-dUMP was found to be a potent non-competitive inhibitor of the enzyme from calf thymus, and 5-azidomethyl-dUMP irreversibly inactivated both this enzyme and the enzyme from Ehrlich ascites tumour cells. Protection by cofactor addition could only be demonstrated for the latter case, however. 5-Iodoacetamidomethyl-dUMP (68) irreversibly inactivates the tumour enzyme more rapidly than the calf thymus enzyme, and protection could also be demonstrated in this case it has therefore been claimed that (68) is isozyme-specific for the tumour enzyme. ... 

Cristofoli, W. A., Wiebe, L. 1., De Clercq, E., Andrei, G., Snoeck, R., Balzarini, J., Knaus, E. E. 5-Alkynyl analogs of arabinouridine and 2 -deoxyuridine cytostatic activity against herpes simplex virus and varicella-zoster thymidine kinase gene-transfected cells. J. Med. Chem. 2007, 50(12), 2851-2857. 

FO, which is a Sp2 + Sp2 hybrid, may require frequent recloning Y3 is a rat myeloma, AI2 a human myeloma, the others are from mouse. Drug resistance 8-Ag and 6-Tg 8-azaguanine and 6-thioguanine (cells lack hypoxanthine phosphoribosyl transferase) BUdR S-bromo-2-deoxyuridine (cells lack thymidine kinase). 

Takemura M, Yamamoto T, Kitagawa M, Taya Y, Akiyama T, Asahara H, Linn S, Suzuki S, Tamai K, Yoshida S (2001) Stimulation of DNA polymerase a activity by Cdk2-phosphorylated Rb protein. Biochem Biophys Res Commun 282 984-990 Tsukamoto I, Taniguchi Y, Miyoshi M, Kojo S (1991) Purification and characterization of thymidine kinase from regenerating rat liver. Biochim Biophys Acta 1079 348-352 VUpo JA (1983) Mitogen induction of deoxyuridine triphosphatase activity in human T and B lymphocytes. Med Biol 61 54—58... 

Figure 22.17 shows the complicated de novo and salvage pathways for the synthesis of dTTP. The de novo pathways start at the top with either UDP or CDP and lead to dTTP. The salvage pathways begin with deoxycytidine, deoxyuridine, or deoxythymidine nucleosides, which are each converted to nucleoside monophosphates in the first step by appropriate kinases. 

Thymidine kinase (located in mitochondria) - acts on deoxythymidine, deoxycytidine, and deoxyuridine... 

The form of the enzyme found in the cytosol acts only on deoxythymidine. The thymidine kinase found in the mitochondria acts on deoxythymidine, deoxycytidine, and deoxyuridine. The mitochondrial thymidine kinase s activity is sufficiently broad that it will also act on the anti-HIV drug, 3 -azido-2 3 -dideoxythymidine (AZT). Evidence suggest that deoxyribonucleotides of AZT interfere with mitochondrial function, possibly by inhibiting mitochondrial DNA replication or transcription, which may explain some of the side effects of cardiotoxicity observed with its use. 

FIG. 6.13 Mammalian pyrimidine salvage and interconversion pathways. Enzymes listed in Figs 6.13-6.17 are as follows (1) deoxyCMP deaminase (2) thymidylate synthase (3) ribonucleotide reductase (4) deoxyuridine triphosphatase (5) CTP synthetase (6) nucleotide kinase (7) deoxyTMP kinase (8) nucleotide diphosphokinase (9) non-specific phosphatase or nucleotidase (10) cytidine kinase (11) pyrimidine phos-phorylase or hydrolase (12) uracil PRTase (13) cytidine deaminase (14) thymidine kinase (15) cytidine phosphotransferase (16) uridine phosphotransferase (17) thymidine phosphotransferase (18) deoxyribo-nucleotide phosphotransferase (19) cytosine PRTase. 

Other halogenated uridine derivatives have been reported to exhibit antiviral activity. Fluorodeoxyuridine has in vitro antiviral activity but is not used in clinical practice. Bromodeoxyuridine is used in subacute sclerosing panencephalitis, a deadly, virus-induced CNS disease. This agent appears to interfere with DNA synthesis in the same way as idoxuridine. The 5 -amino analogue of idoxuridine (5-iodo-5 -amino-2, 5 -dideoxyuridine) is a better antiviral agent than idoxuridine, and it is less toxic. It is metabolized in herpesvirus-infected cells only by thymidine kinase to di- and triphosphoramidates. These metabolites inhibit HSV-specific late RNA transcription, causing reduction of less infective abnormal viral proteins. 5-Bromo-2 -deoxyuridine has an action similar to that of other iodinated compounds. None of these compounds are commercially available in the United States. 

The synthetic pirimidine, 5-fluouracil (5-FU), is used in the treatment of a variety of epithelial tumors including, colorectal, breast cancer, ovarian, head, and neck cancers. 5-FU enters tumor cells by a facilitated nucleobase transporter and is converted into active metabolites (5 -fluoro-2 -deoxyuridine) by a complex metabolic pathway that interferes with both DNA and RNA syntheses. Of particular interest is the conversion of 5-FU to 5 -fluoro-2 -deoxyur-idine (5-FUdR) and subsequent phosphorylation by thymidine kinase to the active metabolite 5 -fluoro-2 -DUMP (5-FdUMP). Unfortunately, the response rate of patients receiving 5-FU-based chemotherapy is relatively poor, varying between 10 and 30%. To further evaluate, 5-FU labeled with F (5- FU) has been used in PET studies to understand the factors behind this strong interindividual variability. As 5- FU is biochemically identical to 5-FU, PET has been used for quantitative pharmacokinetic measurements of 5- FU in human and animal models. 

Carvone was used as a precursor for the synthesis of the cyclohexene 159, and this was used to make the cyclohexenyl nucleoside analogue 160. The same chiral precursor was also manipulated to give the enantiomers of 159 and 160. Both enantiomers of 160 showed potent antiherpetic activity. Molecular modelling of the binding of both compounds to the active site of HSV-1 thymidine kinase was carried out, and a model for the binding of both enantiomers was proposed.An analogue of carbocyclic 2 -deoxyuridine, conforma-tionally-restricted due to a 6,6 -oxido-link (Vol. 32, p. 278), has now been reported in optically-active form, in the L-series. ... 

As seen in the diagram above, both deoxycytidine and deoxyuridine may be converted to deox3ruridylate. Deamination of the former by cyti-dine deaminase yields deoxyuridine, which is phosphorylated by thymidine kinase (see Chapter 14). Cytidine deaminase is responsible for the deamination of both cytidine and deoxycytidine (see Chapter 12) the enzyme may be under regulatory control in that it is inhibited by dTTP 18). 

While it is known that the TK induced by HSV-1 has the unusual property of bifunctionality, little has been reported about the specificity or importance of the second enzyme activity, the deoxythymidylate (dTMP) kinase. The following report compares its role in the activation of the nucleoside analogs, acyclovir (9-(2-hydroxyethoxymethyl)guanine and BrVdUrd ((E)-5-(2-bromovinyl)-2 -deoxyuridine) in both HSV-1 and HSV-2 infected cells. 

These results are consistent with anabolism studies with cell cultures. A close analog of BrVdUrd, (E)-5-(2-iodovinyl)-2 -deoxyuridine, was efficiently converted to its monophosphate derivative in cells infected with both HSV-1 and HSV-2. However, a diphosphate (or triphosphate) could only be detected with extracts of cells infected with HSV-1 . The inefficient phosphorylation of BrVdUMP by the dThd-dTMP kinase from HSV-2 correlates well with the observation that replication of HSV-2 is at least 100-fold less sensitive to BrVdUrd than is that of HSV-1 and suggests a cause-and-effect relationship. The sensitivities of the two viruses to acyclovir are about equal. ... 

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