Selective activation of deoxycytidine kinase by thymidine-5'-thiosulphate and release by deoxycytidine in human lymphocytes.
Deoxycytidine kinase (dCK) catalyses the rate-limiting step of the salvage of three natural deoxyribonucleosides as well as several therapeutic nucleoside analogues, which in turn can enhance its enzymatic activity [Biochem Pharmacol 56 (1998) 1175], improving the efficacy of the cytostatic therapy. Here, we measured the effect of the 5'-thiosulphate (5'-TS) derivatives of four deoxyribonucleosides (deoxyadenosine, deoxycytidine (dCyd), azidothymidine, thymidine) and two ribonucleosides (ribopurine, ribouridine (Urd)) on the activity of the two main salvage deoxynucleoside kinases, and on the salvage of dCyd and deoxythymidine (dThd). It turned out that only 2'-deoxythymidine-5'-thiosulphate (dThd-5'-TS) can potentiate the dCK activity, without influencing the thymidine kinase isoenzymes during short-time treatments of human peripheral blood and tonsillar lymphocytes. The enhancement of dCK activity by dThd-5'-TS can be reversed by dCyd, but dThd had no effect on the enzyme activation in cells. Neither dThd-5'-TS nor Urd-5'-TS had any effect on the dCK and thymidine kinase activities tested in cell-free extracts. The stimulation of dCK activity in cells was accompanied by an imbalance in the dThd and dCyd metabolism. The incorporation of 3H-dThd into DNA was suppressed by 90% in cells by dThd-5'-TS, while Urd-5'-TS only slightly influenced the same process. The 3H-dCyd incorporation into DNA was inhibited only to 50% of the control, while the 3H-dCyd labelling of the nucleotide fraction was enlarged in dThd-5'-TS-treated cells, as a consequence of the increased dCK activity. We suggest that the enhancement of dCK activity is a compensatory mechanism in cells that might be induced by different "inhibitors" of DNA synthesis leading to damage of DNA. The increased dCK activity is able to supply the repair of DNA with dNTPs in quiescent cells; this suggestion seems to be supported by the counteracting effect of extracellular dCyd, too.
Keszler
Gergely
G
Department of Medical Chemistry, Molecular Biology and Pathobiochemistry, Faculty of Medicine, Semmelweis University, Budapest, Hungary.
Szikla
Karoly
K
Kazimierczuk
Zygmunt
Z
Spasokoukotskaja
Tatjana
T
Sasvari-Szekely
Maria
M
Staub
Maria
M
D002474:Cell-Free System; D003841:Deoxycytidine / Q000494:pharmacology*; D003842:Deoxycytidine Kinase / Q000378:metabolism*; D003845:Deoxycytosine Nucleotides / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D066298:In Vitro Techniques; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000378:metabolism; D008198:Lymph Nodes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D008214:Lymphocytes / Q000187:drug effects* / Q000201:enzymology; D014066:Palatine Tonsil / Q000166:cytology; D013885:Thiosulfates / Q000494:pharmacology*; D013936:Thymidine / Q000031:analogs & derivatives / Q000494:pharmacology*; D013937:Thymidine Kinase / Q000378:metabolism
D003845:Deoxycytosine Nucleotides; D013885:Thiosulfates; C471848:thymidine-5'-thiosulfate; D003841:Deoxycytidine; C045813:2'-deoxycytidine diphosphate; D013937:Thymidine Kinase; D003842:Deoxycytidine Kinase; D013936:Thymidine