New mechanism of action of the cancer chemotherapeutic agent 5-fluorouracil in human cells.
5-Fluorouracil (5-FlUra), a cancer chemotherapeutic agent used in the treatment of colon, breast, ovarian and prostate cancer, is incorporated into DNA as a result of its utilization as 5-FldUTP during DNA synthesis. This promutagenic DNA lesion is excised by the base excision repair enzyme uracil DNA glycosylase (UDG). In this report we describe for the first time a mechanism by which 5-FlUra as the free base specifically binds in vivo to the UDG in noncycling human cells, thereby inhibiting its activity. By using 5-FlUra concentrations which did not elicit demonstrable cell toxicity, a dose-dependent decrease in UDG activity was detected which approached 30% of that observed in control cells. In contrast, exposure of cells to equivalent concentrations of uracil, 5-fluorodeoxyuridine or 5-bromouracil had no effect on UDG activity. Subsequent studies demonstrated a reversible binding of 5-FlUra to the glycosylase. Kinetic analysis using nonlinear regression analysis demonstrated a competitive mode of inhibition and indicated a tight binding of 5-FlUra to UDG in vivo, although the 5-FlUra-UDG complex was easily dissociated in vitro. These findings describe a potentially new and novel mechanism of action of 5-FlUra in a nonproliferating human cell population. The potential relevance of these findings to the utility of 5-FlUra as a cancer chemotherapeutic agent is considered.
Wurzer
J C
JC
Fels Institute for Cancer Research and Molecular Biology, Temple University School of Medicine, Philadelphia, Pennsylvania.
Tallarida
R J
RJ
Sirover
M A
MA
D001667:Binding, Competitive; D002452:Cell Count / Q000187:drug effects; D002478:Cells, Cultured; D004247:DNA / Q000096:biosynthesis; D045647:DNA Glycosylases; D005347:Fibroblasts / Q000187:drug effects / Q000201:enzymology; D005467:Floxuridine / Q000494:pharmacology; D005472:Fluorouracil / Q000494:pharmacology*; D006801:Humans; D007700:Kinetics; D009699:N-Glycosyl Hydrolases / Q000037:antagonists & inhibitors*; D014498:Uracil / Q000494:pharmacology; D051981:Uracil-DNA Glycosidase
D005467:Floxuridine; D014498:Uracil; D004247:DNA; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; D051981:Uracil-DNA Glycosidase; D005472:Fluorouracil