5'-(Bromoacetamido)-2',5'-dideoxyuridine (3) and derivatives (8, 10, 12, and 14) substituted at the 5-position with bromo, iodo, fluoro, and ethyl groups have been synthesized as potential inhibitors of enzymes that metabolize pyrimidine nucleosides. Also prepared were 2',5'-dideoxyuridine derivatives (4-6) substituted at the 5'-position with 2-bromopropionamido, iodoacetamido, and 4-(fluorosulfonyl)benzamido groups. Compounds 3, 5, 8, 12, and 14 were examined for effect on macromolecular synthesis in L1210 leukemia cells in culture and compared with 5'-(bromoacetamido)-5'-deoxythymidine (1, BAT), a compound with demonstrated cytotoxicity and activity in vivo against P388 murine leukemia. Compounds 3, 8, 12, and 14 inhibited DNA synthesis without significant inhibition of RNA synthesis, and protein synthesis was affected less than DNA synthesis. Compounds 3, 5, 6, 8, 10, 12, and 14 were cytotoxic to H.Ep.-2 and L1210 cells in culture, and 3, 5, 8, and 12 showed activity in the P388 mouse leukemia screen.