Back DJ, Phiboonbanakit D; International Conference on AIDS.
Int Conf AIDS. 1998; 12: 826 (abstract no. 42264).
Pharmacology University of Liverpool, UK.
AIMS: The cellular interaction between the phosphate intermediates of thymidine and zidovudine (ZDV) is not clearly understood. For example, methotrexate (MTX), reduces intracellular thymidine triphosphate (dTTP) by interruption of the thymidine de novo pathway prior to thymidine monophosphate (dTMP) formation, and causes an upregulation of thymidine kinase (TK) activity. When MTX and ZDV are combined an increase in ZDV triphosphate (ZDVTP) is seen. Competitive inhibition of thymidylate kinase (TMPK) by ZDV monophosphate (ZDVMP) inhibits dTMP phosphorylation in vitro (Furman et al., Proc Natl Acad Sci USA 1986, 83: 8333-7). The aim was to study if the level of dTMP, rather than TK activity, controls the extent of ZDV phosphorylation. METHODS: ZDV phosphorylation in the presence and absence of MTX (1 microM) was compared in U937 cells under two physiological conditions. U937 cells were cultured in i) medium replenished daily and ii) medium exhausted of nutrients. Exhausted medium causes a depletion of cellular kinases, therefore ZDV phosphorylation is not regulated by increased TK activity secondary to the depletion of dTTP. RESULTS: In replenished culture medium, MTX significantly suppressed intracellular dTTP levels and significantly increased the intracellular concentrations of all ZDV metabolites (p < 0.0005) compared to controls. In exhausted medium, a significant reduction is dTTP and ZDVMP (p < 0.005) was evident compared to replenished medium. MTX further reduced dTTP concentrations (p < 0.05) compared to exhausted medium alone. In the absence of MTX, ZDVDP and ZDVTP levels were similar in replenished and exhausted medium. Levels were significantly increased (p < 0.05) upon addition of MTX to exhausted medium when compared to both media alone. CONCLUSIONS: Increased TK activity may be partly responsible for the increase in ZDV phosphorylation, in particular ZDVMP levels. However, ZDVDP and ZDVTP are not directly controlled by TK and the data suggest the increases in ZDVDP and ZDVTP are due to a direct effect of reduced dTMP formation rather than increase TK activity brought about by reduced dTTP. The results support the hypothesis that dTMP plays an important role in the control of ZDV phosphorylation.
Publication Types:
Keywords:
- 3'-azido-3'-deoxythymidine 5'phosphate
- In Vitro
- Methotrexate
- Nucleoside-Phosphate Kinase
- Oxidation-Reduction
- Phosphorylation
- Thymidine
- Thymidine Kinase
- Thymidine Monophosphate
- Thymine Nucleotides
- Zidovudine
- dTMP kinase
- enzymology
- prevention & control
- surgery
- thymidine 5'-triphosphate
Other ID:
UI: 102231063
From Meeting Abstracts