Inhibition of intestinal pyrimidine nucleoside phosphorylases.

The activity of 5'-deoxy-5-fluorouridine (dFUR) depends on its activation to 5-fluorouracil (FU) by pyrimidine nucleoside phosphorylases. These enzymes are found in tumors and normal tissues, with the highest activity in the small intestines. The present study examined the inhibition of dFUR phosphorolysis in intestinal tissues. dFUR metabolism in intestinal homogenates was inhibited by uracil (U), uridine (UR), and thymidine (TdR), which are the normal substrates for the phosphorylases. Conversely dFUR reduced the metabolism of these inhibitors. A good agreement was found between the observed data and the computer-fitted data using the equations for competitive inhibition between dFUR and the inhibitors. In the absence of inhibitors, the Vmax of dFUR phosphorolysis was 47.1 +/- 4.9 microM/min and the apparent Km was 910 +/- 167 microM. The Vmax was unaltered by the inhibitors, while the Km was increased with increasing inhibitor concentrations. The maximal inhibition of dFUR metabolism by UR and TdR was about 80%. The Ki's were 372 microM for U, 87.2 microM for UR, and 112 microM for TdR and are orders of magnitude higher than their reported endogenous serum concentrations. The rate of dFUR phosphorolysis to FU in the intact intestinal epithelial crypt cells, indicated by the ratio of FU to dFUR in the intracellular fluid, was reduced by UR in a concentration-dependent fashion. These data indicate that the naturally occurring pyrimidines inhibit competitively the dFUR metabolism by the intestinal phosphorylases, that this inhibition occurs at concentrations much higher than the circulating endogenous levels, and that phosphorolysis is the major route of dFUR metabolism.