SEMIAUTOMATED RADIOASSAY FOR DETERMINATION OF DIHYDROPYRIMIDINE DEHYDROGENASE (DPD) ACTIVITY - SCREENING CANCER-PATIENTS FOR DPD DEFICIENCY, A CONDITION ASSOCIATED WITH 5-FLUOROURACIL TOXICITY

Dihydropyrimidine dehydrogenase (DPD) catalyzes the reduction of the n aturally occurring pyrimidines, uracil and thymine, and the fluoropyri midine anticancer drug, 5-fluorouracil (FUra) to 5,6-dihydropyrimidine s. Previous studies have demonstrated that cancer patients who are DPD deficient exhibit severe toxicity (including death) following treatme nt with FUra. To date, the direct measurement of DPD enzyme activity h as been the only reliable method to identify DPD deficient cancer pati ents. We now report a semi-automated radioassay for measuring DPD acti vity in human peripheral lymphocytes. Following incubation of lymphocy te cytosol (at a fixed protein concentration of 200 mu g) with [6-C-14 ]FUra at timepoints ranging from 0 to 30 min, samples are ethanol prec ipitated, filtered and analyzed by HPLC. Determination of radioactivit y is accomplished using an in-line flow scintillation analyzer with au tomatic quantitation of peaks. This method provides the first specific assay for DPD enzyme activity which is rapid, reproducible and of can cer patients for DPD deficiency prior to treatment with FUra. sensitiv e enough to be used in the routine screening (C) 1997 Elsevier Science B.V.