ABSTRACT
A series of substituted 5-aminomethyl-2'-deoxyuridines was synthesized as analogues of 5-thymidylyltetrahydrofolic acid, a proposed intermediate in the thymidylate synthetase catalyzed reaction. 1-(3,5-Di-O-p-toluoyl-2-deoxy-beta-D-ribofuranosyl)-5-chloromethyluracil (3) was treated with the appropriate amine to give the ester protected 5-aminomethyl nucleoside. Removal of the ester groups was accomplished with anhydrous potassium carbonate in methanol to afford the free beta-nucleoside. In this way 5-(2-dimethylaminoethylaminomethyl)-2'-deoxyuridine (5a), 5-dimethylaminomethyl-2'-deoxyuridine (5b), 5-N-mehtylpiperazinylmethyl-2'-deoxyuridine (5c), and 5-pyrrolidinylmethyl-2'-deoxyuridine (5d) were prepared. Compounds 5a,b,d were converted to the respective 5'-phosphates 6a,b,d. All three compounds were subtrate competitive inhibitors of thymidylate synthetase purified from Escherichia coli, calf thymus, and Ehrlich ascites tumor cells. The most active compound was 6a with KI's of 6,3.1, and 14 micronM observed for the respective enzymes.
Subject(s)
Deoxyribonucleotides/chemical synthesis , Methyltransferases/antagonists & inhibitors , Thymidylate Synthase/antagonists & inhibitors , Uracil Nucleotides/chemical synthesis , Animals , Carcinoma, Ehrlich Tumor/enzymology , Cattle , Deoxyribonucleotides/pharmacology , Escherichia coli/enzymology , Spectrophotometry, Ultraviolet , Thymus Gland/enzymology , Uracil Nucleotides/pharmacologyABSTRACT
In a study of active site binding the inhibition of thymidylate synthetase derived from Escherichia coli, calf thymus, and Ehrlich ascites tumor was examined using eight inhibitors. 5-Substituted 2'-deoxyuridine 5'-phosphate analogues used in this study are the hydroxymethyl, methoxymethyl, benzyloxymethyl, formyl, acetyl, allyl, and two potential active site alkylating substituents: 2,3-oxypropyl and the azidomethyl analogues. All compounds were competitive with the substrate, 2'-deoxyuridine 5'-phosphate; the most potent inhibitor was 5-formyl-dUMP (Ki = 0.1, 0.09, and 0.08 muM for the respective enzyme). The 5-hydroxymethyl, 5-benzyloxymethyl, and 5-azidomethyl derivatives of dUMP showed some differential inhibition; these compounds were two to three times more active against the ascites tumor enzyme than against the thymus enzyme.
