Deoxycytidine kinase

In addition to possible applications for the development of cancer drugs, deoxycytidine kinase (dCK) inhibitors also hold promise for the treatment of immune disorders. Drug discovery efforts aimed at inhibiting deoxycytidine kinase have been reported by Lexicon Pharmaceuticals. They disclosed an HTS assay for the discovery of dCK inhibitors.30 

Lexicon also disclosed another class of dCK inhibitors. Compound (14) potently inhibited dCK in enzyme and cellular assays, engaged the target in isolated mouse cells and prevented the incorporation of radiolabeled 3H-dC 

---

P-D-Arabinofuranosylcytosine [147-94-4] (ara-C, 16), C H N O, reportedly has had significant therapeutic effects in patients with localized herpes zoster, herpes eye infections, and herpes encephaUtis (33), although several negative results have also been reported (34) (Fig. 2). Ara-C, also known as cytarabine, is quite toxic and is only recommended for very severe viral infections. It is rapidly deaminated in humans to the relatively inactive ara-U Ara-C is converted in the cell to the 5 -monophosphate by deoxycytidine kinase, followed by formation of the corresponding di- and triphosphate. The triphosphate has been shown to inhibit DNA polymerase. 

Ara-A is phosphorylated in mammalian cells to ara-AMP by adenosine kinase and deoxycytidine kinase. Further phosphorylation to the di- and triphosphates, ara-ADP and ara-ATP, also occurs. In HSV-1 infected cells, ara-A also is converted to ara-ATP. Levels of ara-ATP correlate directly with HSV rephcation. It has recently been suggested that ara-A also may exhibit an antiviral effect against adenovims by inhibiting polyadenylation of viral messenger RNA (mRNA), which may then inhibit the proper transport of the viral mRNA from the cell nucleus. 

Cladribine (2-Chlordeoxyadenosine) is a synthetic purine nucleoside that is converted to an active cytotoxic metabolite by the deoxycytidine kinase. The drug is relatively selective for both normal and malignant lymphoid cells. 

The cytotoxic activities of the 2, 2 -difluoro analog (775) of 737 against Chinese hamster ovary and tumor cells, in comparison with those of 1- -d-arabinofuranosylcytosine ara-C, a drug for leukemia), have been studied 775 is transported the faster through membrane into cells, more effectively phosphorylated by the deoxycytidine kinase (to the 5 -mono-phosphate) and, after conversion into the 5 -triphosphate, more highly accumulated in the cells, with longer duration time, than is ara-C, but nevertheless 775 is incorporated into the DNA to a lesser extent than is ara-C. These characteristics of 775 were discussed. 

The 2 -chloro and 2 -bromo congeners of either 748 (FIAC) or 758 (FMAU) are more cytotoxic than FIAC and FMAU, suggesting that these chloro and bromo nucleosides, in contrast to the 2 -fluoro compounds, are comparatively better substrates for deoxycytidine kinase of human lymphocytes than the substrates for viral-specific thymidine kinase. The disposition of the 2 -fluoro group may also be important from the biological viewpoint. It should be noted that the structural difference between RNA and DNA is at the 2 -position. The ribo type of analog (738) of FIAC is 10 times less effective in suppression of HSV replication than is FIAC. Thus Fox, and Watanabe and coworkers concluded that the 2 - up fluorine disposition and the species of the substituent at C-5 are the two important factors influencing antiviral activity. Nevertheless, the mechanism of action of 2 -deoxy-2 -fluorocytidine (737) on certain herpes viruses, including HSV-1... 

The first etCCR application has been reported for a partially C— N-labeled phosphotyrosine peptide derived from interleukin-4 receptor ligated to STAT-6 [107] and subsequent studies involve nucleotide cofactors ligated to human recombinant deoxycytidine kinase [108] and epothilone A bound to tubulin [109]. Since etCCR usually involves isotope-labeling schemes for the ligand, its applicability is limited to specific molecular classes. 

Chattopadhayaya,. An NMR conformational smdy of the complexes of C/ H double-labeled 2 -deoxynucleosides and deoxycytidine kinase (dCK)./. Chem. Soc. Perkin. 

CARNOSINE SYNTHETASE CHAPERONES CHOLINE KINASE CHOLOYL-CoA SYNTHETASE COBALAMIN ADENOSYLTRANSFERASE 4-COUMAROYL-CoA SYNTHETASE CREATINE KINASE CTP SYNTHETASE CYTIDYLATE KINASE 2-DEHYDRO-3-DEOXYGLUCONOKINASE DEHYDROGLUCONOKINASE DEOXYADENOSINE KINASE DEOXYADENYLATE KINASE DEOXYCYTIDINE KINASE (DEOXYjNUCLEOSIDE MONOPHOSPHATE KINASE DEOXYTHYMIDINE KINASE DEPHOSPHO-CoA KINASE DETHIOBIOTIN SYNTHASE DIACYLGLYCEROL KINASE DIHYDROFOLATE SYNTHETASE DNA GYRASES DNA REVERSE GYRASE ETHANOLAMINE KINASE EXONUCLEASE V... 

DEOXYADENOSINE KINASE DEOXYADENYLATE KINASE DEOXYCYTIDINE KINASE DEOXYCYTIDYLATE HYDROXYMETHYL-TRANSFERASE Deoxycytidylate kinase,... 

Deoxycytidine kinase, which phosphorylates bases, is inhibited by deoxycytidine and dCTP, so, as their intracellular levels fall, more gemcitabine can be phosphorylated. 

Bouffard DY, Laliberte J, Momparler RL. Kinetic studies on 2, 2 -difluorodeoxycytidine (Gemcitabine) with purified human deoxycytidine kinase and cytidine deaminase. Biochem Pharmacol 1993 45(9) 1857—1861. 

Hudarabine phosphate is a fluorinated nucleotide analog of the antiviral agent vidarabine. Its cytotoxicity is not well understood. It is rapidly dephospho-rylated at the cell membrane level and then rephos-phorylated intracellularly by deoxycytidine kinase to the active triphosphate derivative. It inhibits DNA polymerase and DNA primase. It is also incorporated into DNA and RNA. Hudarabine is administered intravenously by infusion over 30-120 min. It is eliminated by renal excretion with a terminal half life 10 hours. Adverse effects include myelosuppres-sion, nausea, vomiting, chills and fever. The number of CD4 positive cells is reduced and the incidence of opportunistic infections is increased. 

Cladribine, or 2-chlorodeoxyadenosine, is resistant to adenosine deaminase and after intracellular phosphorylation by deoxycytidine kinase, it is incorporated into DNA. It is considered the drug of choice in hairy cell leukemia because of high activity combined with acceptable toxicity. Cladribine shows variable oral absorbtion and is usually administered intravenously. Its concentration-time course is biphasic with plasma half-lives of 35 minutes and 6.7 hours. Excretion is primarily by the kidneys. Its most prominent dose-limiting toxicity is myelosup-pression. 

A.W. Blackstock, H. Lightfoot, L.D. Case, J.E. Tepper, S.K. Mukhetji, B.S. Mitchell, S. G. Swarts, S.M. Hess, Tumor uptake and elimination of 2, 2 -difluoro-2 -deoxycyti-dine (gemcitabine) after deoxycytidine kinase gene transfer Correlation with in vivo tumor response, Clin. Cancer Res. 7 (2001) 3263-3268. 

More recently fluoromethylene deoxycytidines have been rationally designed by McCarthy et al. as bio-precursors of mechanism-based inhibitors of RDPR, after phosphorylation in vivo by the deoxycytidine kinase. Among these, tezacita-bine (MDL 101731) [92] has a high anti-proliferative activity and is currently in clinical phase III evaluation for the treatment of solid tumours [92],... 

Cytarabine (ara-C) is an S phase-specific antimetabolite that is converted by deoxycytidine kinase to the 5 -mononucleotide (ara-CMP). Ara-CMP is further metabolized to the diphosphate and triphosphate metabolites, and the ara-CTP triphosphate is felt to be the main cytotoxic... 

Gaubert, G. Gosselin, G. Boudou, V. Imbach, J.L. Eriksson, S. Maury, G. Low enantioselectivities of human deoxycytidine kinase and human deoxyguanosine kinase with respect to 2 -deoxyadenosine, 2 -deoxyguanosine and their analogs. Biochimie, 81, 1041-1047 (1999)... 

Rodriguez, C.O., Jr. Mitchell, B.S. Ayres, M. Eriksson, S. Candhi, V. Ara-binosylguanine is phosphorylated by both cytoplasmic deoxycytidine kinase and mitochondrial deoxyguanosine kinase. Cancer Res., 62, 3100-3105 (2002)... 

Few pyrimidine bases are salvaged in human cells. However, the pyrimidine nucleosides uridine and cytidine can be salvaged by uri-dine-cytidine kinase, deoxycytidine can be salvaged by deoxycytidine kinase, and thymidine can be salvaged by the enzyme thymidine kinase. Each of these enzymes catalyzes the phosphorylation of a nucleoside(s) utilizing ATP, and forming UMP, CMP, dCMP, and TMP. 

Deoxyadenosine is also a substrate, though a relatively poor one. In some mammalian cells deoxycytidine kinase also phosphorylates deoxyadenosine and deoxyguanosine (see next section). In addition, indirect evidence suggests the existence of a specific deoxyadenosine kinase in human cells. Deoxyadenosine has received special attention as a substrate for kinases because congenital inability to catabo-... 

Fludarabine phosphate (2-fluoro-arabinofuranosyladenine monophosphate) is rapidly dephosphorylated to 2-fluoro-arabinofuranosyladenine and then phosphorylated intracellularly by deoxycytidine kinase to the triphosphate. This metabolite interferes with DNA synthesis through inhibition of DNA polymerase- and ribonucleotide reductase, and it also induces apoptosis. Fludarabine phosphate is used chiefly in the treatment of low-grade non-Hodgkin s lymphoma and chronic lymphocytic leukemia (CLL). Fludarabine phosphate is given parentally and is excreted primarily in the urine its dose-limiting toxicity is myelosuppression. 

After intravenous administration (Table 55-3), the drug is cleared rapidly, with most being deaminated to an inactive form. The ratio of the anabolic enzyme deoxycytidine kinase to the inactivating catalyst cytidine deaminase is important in determining the cytotoxicity of cytarabine. 

Gemcitabine is phosphorylated initially by the enzyme deoxycytidine kinase and then by other nucleoside kinases to the di- and triphosphate nucleotide forms, which then inhibit DNA synthesis. Inhibition is considered to result from two actions inhibition of ribonucleotide reductase by gemcitabine diphosphate, which reduces the level of deoxyribonucleoside triphosphates required for the synthesis of DNA and incorporation of gemcitabine triphosphate into DNA. Following incorporation of gemcitabine nucleotide, only one additional nucleotide can be added to the growing DNA strand, resulting in chain termination. 

An important difference between the two compounds is that azacytidine (2) incorporates into RNA, while decitabine (1) acts on DNA. Within cells, decitabine (1) is phosphorylated by deoxycytidine kinase, and after conversion to decitabine triphosphate, it is incorporated into DNA in place of deoxycytidine triphosphate. Azacitidine (2) is phosphorylated by uridine-cytidine kinase and eventually incorporated into RNA, inhibiting the processing of ribosomal RNA and ultimately protein synthesis. Azacitidine (2) can also inhibit DNMTs when azacytidine diphophate is reduced to decitabine diphosphate, which is further phosporylated by kinases to dcitabine triphosphate and incorporated into DNA. Because of the inefficiency of these extra steps, the hypomethylating potency of 2 is believed to be one fifth to one tenth that of l.6-8... 

The 2 -azido group of cytarazid renders the nucleoside more resistant to deamination to the 2 -azidouridine derivative (153) by deoxycytidine deaminase, but was also observed to reduce substrate affinity for the deoxycytidine kinase necessary for anabolic phosphorylation to the active cytarazid 5 -triphosphate [180]. Conversely, cytarazid was a more potent inhibitor of the target DNA polymerases a and K = 0.6 and 0.7 //M, respectively) than the parent ara-C (.A = 10 and 17 fiM, respectively), and the dissimilar spectrum of antitumour activity exhibited by the two compounds was attributed to differences in stability, metabolic activation and inhibitory potency [180, 181]. Interestingly, the instability of cytarazid to thiols present in the assay media, was commented on but not pursued [180]. In view of previous discussions concerning the bioreduction of AZT... 

---