MECHANISM OF TOXICITY OF FLUOROCYTOSINE AND FLUOROURACIL

The long term objective of this proposal is to improve the clinical effectiveness of fluoropyrimidine (FP) drugs through a better understanding of the biochemical and pharmacologic factors that determine toxicity with a major focus on FP catabolism in humans. Our knowledge of FP catabolism is still quite limited, despite the obvious importance in determining availability of active drug. Initially, the three enzymes in FP catabolism: uracil reductase (UR), dihydropyrimidinase and ureidopropionase will be purified from human liver and characterized. FP metabolism will also be examined in hepatocytes and non-parenchymal cells isolated from human liver. A study will be made of partial and complete deficiencies of UR, initially in affected families (to clarify the genetics of this syndrome) and later in FP-treated patients to determine whether partial deficiency of enzyme is a factor in some cases of FP-induced toxicity. The second major area of this proposal will be to identify and characterize novel FUra metabolite(s) recently isolated from human bile. Preliminary studies suggested these to be conjugates of fluoro- beta-alanine (FBAL) and bile acids (e.g. cholic acid). These metabolites may have a role in the pathogenesis of FP-induced cholestasis, seen frequently with hepatic arterial infusion (HAI) of fluorodeoxyuridine (FdUrd). An attempt will be made to synthesize FBAL-bile acid conjugates for use in in vivo studies. The enzyme cholyl-CoA-amino acid N-acyltransferase will be purified from human liver to determine affinity of FBAL compared with beta-alanine, taurine and glycine. In human hepatocytes, FP catabolism will be assessed in absence or presence of biochemical modulators known to inhibit FP catabolism in an attempt to decrease the formation of these conjugates. Utilizing the isolated perfused rat liver, bile flow and bile acid content will be assessed after HAI of FdUrd or FBAL- conjugate. Further metabolism of FBAL-conjugates by intestinal microflora will be evaluated in a semicontinuous culture system. In clinical studies (3H)-FdUrd will be administered by HAI with bile analysis to determine presence of metabolites. Late the presence of the conjugates will be quantitated in serum from patients treated with HAI FdUrd (non-labelled), with simultaneous assessment of liver dysfunction. It is hoped by understanding FP catabolism and the mechanism of FP-induced cholestasis improved FP regimens can be designed with decreased host toxicity + increased antitumor effect.