Dihydropyrimidine dehydrogenase: a tumoral target for fluorouracil modulation

Dihydropyrimidine dehydrogenase (DPD) is the rate-limiting enzyme of 5-fluorouracil (FU) catabolism. Ethynyluracil (776C) is a very potent, mechanism-based irreversible DPD inhibitor that improves the antitumor efficacy and the therapeutic index of FU in laboratory animals. We tested the cytotoxic effects of the FU-776C combination on a panel of 12 human cancer cell lines (4 breast, 4 head and neck, 3 colon, and 1 duodenum). Basal DPD activity (radioenzymatic assay) and FU sensitivity [FU 50% inhibitory concentration (IC50), 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide test] were determined. The FU potentiation by 776C was calculated from the ratio (F) of FU IC50 without 776C divided by FU IC50 with 776C. 776C was not cytotoxic to any of the cell lines tested. On CAL51 cell line expressing a high basal DPD activity, FU enhancement by 776C was a saturable phenomenon related to the 776C concentration; the inhibition of DPD increased between 10(-12) to 10(-6) M of 776C. For the following studies, 776C was tested at 10(-6) M. FU IC50 varied from 15 to 7770 microM among cell lines (median, 390 microM). Basal DPD activity ranged from not detectable (< pmol/min/mg protein) to 320 pmol/min/mg protein among cell lines (median, 53 pmol/min/mg protein). For the 12 cell lines tested, the mean F ranged from 0.7 (no enhancement of FU cytotoxicity by 776C) up to 5.2 and was significantly related to the basal DPD activity: the greater the DPD activity, the greater the FU enhancement factor (Spearman rank correlation, P = 0.019). Enhancement of FU cytotoxicity by 776C occurred only in the six cell lines expressing the greatest basal DPD activity (>50 pmol/min/mg protein, F ranging between 1.7 and 5. 2), whereas 776C did not modify FU cytotoxicity in the remaining cell lines expressing the lowest DPD activity (<50 pmol/min/mg protein, F ranging between 0.7 and 1.4). F was significantly different between these two groups of cell lines (P = 0.005). These results point out that DPD is an interesting target for FU pharmacomodulation.