Comparison of 5-Fluorouracil Metabolism in Two Human Gastrointestinal Tumor Cell Lines1

HuTu 80 and HT 29 are two human gastrointestinal tumor cell lines which differ 20-fold in their sensitivity to growth inhibition by 5-fluorouracil (FUra) [concentration of inhibitor necessary to inhibit cell growth 50% as compared with control cells grown under identical conditions (except that inhibitor was omitted): HuTu 80, 6 /¿M; HT 29, 0.3 /UM].Drug response in the two cell lines is further distinguished by the ability of thymidine to reverse the growth inhibition produced by FUra in HT 29 cells but not in HuTu 80 cells. Furthermore, the presence of deoxyinosine po tentiates FUra action in HuTu 80 cells, while having no effect on growth inhibition by FUra in HT 29 cells. The metabolism of [6-3H]FUra was examined in these cell lines using techniques which allow simultaneous measurement of: (a) the amount of 5-fluoro-2'-deoxyuridylate bound to thymidylate synthetase; (b) the level of free thymidylate synthetase; (c) FUra incorporation into RNA; and (d) acid-soluble metabolites of FUra. A comparison of FUra metabolism in the two cell lines, following a 24-hr exposure of the cells to equitoxic concentrations of FUra, demonstrated that such exposure led to thymidylate synthetase inhibition in both HuTu 80 and HT 29 cells. This inhibition, however, was accompanied by a 6-fold greater level of incorpo ration of FUra into RNA in HuTu 80 cells than in HT 29 cells, and suggests that drug incorporation into RNA may be the dominant determinant of growth inhibition by FUra in HuTu 80 cells. Deoxyinosine, which potentiates FUra action in HuTu 80 cells, was found to alter FUra metabolism in the cells so that thymi dylate synthetase inhibition was now achieved under conditions of reduced FUra incorporation into RNA. Thymidine was able to reverse growth inhibition of HuTu 80 cells by the FUra-deoxyinosine combination, suggesting a change in the mechanism of growth inhibition.