TRANSCRIPTION AND ACTIVITY OF 5-FLUOROURACIL CONVERTING ENZYMES IN FLUOROPYRIMIDINE RESISTANCE IN COLON-CANCER IN-VITRO

Cellular resistance to 5-fluorouracil (5-FU) is not completely underst ood. Since 5-FU shares the pyrimidine pathway with the physiological p yrimidines, we investigated the relationship between fluoropyrimidine metabolism, nucleic acid uptake and cytotoxicity of 5-FU in eight colo n tumour cell lines including 5-FU-resistant subclones. The cytotoxici ty of 5-FU was increased up to 423-fold when the anabolites 5-fluorour idine (FUrd), 5-fluorodeoxyuridine (FdUrd), and 5-fluorodeoxyuridine m onophosphate (FdUMP) were compared with the parent drug in vitro. The enzymes uridine phosphorylase and thymidine phosphorylase were predict ive for the cytotoxicity of 5-FU in 5/7 cell lines. Inhibition of urid ine phosphorylase and thymidine phosphorylase by antisense strategies effectively antagonised 5-FU, abolishing 84% and 79% of its toxicity. The importance of thyimidine phosphorylase was supported by a highly r estricted enzyme activity in 5-FU-resistant cells. In 5-FU naive cells , a stimulating effect of 5-FU on thymidylate synthase mRNA and ribonu cleotide reductase mRNA expression was observed. In these cells, antis ense oligonucleotides to ribonucleotide reductase significantly reduce d cell growth. Downregulation of ribonucleotide reductase mRNA in 5-FU -resistant subclones suggests different mechanisms in primary and seco ndary resistance to 5-FU. Most of the intracellular 5-FU was selective ly incorporated into RNA (range: 45-91%) and generally spared DNA (ran ge: 0.2-11%). In synthesising our data, we conclude that drug resistan ce could be overwhelmed through bypassing limiting steps in the activa tion of 5-FU. In the majority of colonic tumours, the activity of urid ine phosphorylase and thymidine phosphorylase may have prognostic rele vance for the cytotoxicity of 5-FU in vitro. (C) 1997 Elsevier Science Inc.