Impact of 9‐(2‐phosphonylmethoxyethyl)adenine on (deoxy)ribonucleotide metabolism and nucleic acid synthesis in tumor cells

FEBS Letters

Hatse, Sigrid¹  De Clercq, Erik¹  Balzarini, Jan¹ 

[1] Rega Institute for Medical Research, Minderbroedersstraat 10, K.U. Leuven, B-3000 Leuven, Belgium

关键词: (Deoxy)ribonucleotide metabolism;    Acyclic nucleoside phosphonate;    Thymidine kinase;    Cell cycle;    DNA synthesis;    Ara-C;    1-β-d-arabinofuranosylcytosine;    CDK;    cyclin-dependent kinase;    dNTP;    2′-deoxyribonucleoside 5′-triphosphate;    HPLC;    high performance liquid chromatography;    IC50;    50% inhibitory concentration for cell proliferation;    NTP;    ribonucleoside 5′-triphosphate;    PBS;    phosphate-buffered saline;    PMEA;    9-(2-phosphonylmethoxyethyl)adenine;    PMEApp;    the diphosphorylated metabolite of 9-(2-phosphonylmethoxyethyl)adenine;    TCA;    trichloroacetic acid;    TK;    thymidine kinase;    TS;    thymidylate synthase;   

DOI  :  10.1016/S0014-5793(99)00104-0

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Following exposure to 9-(2-phosphonylmethoxyethyl)adenine (an inhibitor of the cellular DNA polymerases α, δ and ϵ), human erythroleukemia K562, human T-lymphoid CEM and murine leukemia L1210 cells markedly accumulated in the S phase of the cell cycle. In contrast to DNA replication, RNA synthesis (transcription) and protein synthesis (mRNA translation) were not affected by 9-(2-phosphonylmethoxyethyl)adenine. The ribonucleoside triphosphate pools were slightly elevated, while the intracellular levels of all four deoxyribonucleoside triphosphates were 1.5–4-fold increased in 9-(2-phosphonylmethoxyethyl)adenine-treated K562, CEM and L1210 cells. The effect of 9-(2-phosphonylmethoxyethyl)adenine on de novo (thymidylate synthase-mediated) and salvage (thymidine kinase-mediated) dTTP synthesis was investigated using radiolabelled nucleoside precursors. The amount of thymidylate synthase-derived dTTP in the acid soluble pool was 2–4-fold higher in PMEA-treated than in untreated K562 cells, which is in accord with the 3–4-fold expansion of the global dTTP level in the presence of 9-(2-phosphonylmethoxyethyl)adenine. Strikingly, 2-derived dTTP accumulated to a much higher extent (i.e. 16–40-fold) in the soluble dTTP pool upon 9-(2-phosphonylmethoxyethyl)adenine treatment. In keeping with this finding, a markedly increased thymidine kinase activity could be demonstrated in extracts of 9-(2-phosphonylmethoxyethyl)adenine-treated K562 cell cultures. Also, in the presence of 200 μM 9-(2-phosphonylmethoxyethyl)adenine, 14-fold less thymidylate synthase-derived but only 3-fold less thymidine kinase-derived dTTP was incorporated into the DNA of the K562 cells. These data show that thymidine incorporation may be inappropriate as a cell proliferation marker in the presence of DNA synthesis inhibitors such as 9-(2-phosphonylmethoxyethyl)adenine. Our findings indicate that 9-(2-phosphonylmethoxyethyl)adenine causes a peculiar pattern of (deoxy)ribonucleotide metabolism deregulation in drug-treated tumor cells, as a result of the metabolic block imposed by the drug on the S phase of the cell cycle.

【 授权许可】

Unknown