Acyclic analogues of 5-fluoro-dUMP and 5-fluoro-2'-deoxyuridine: synthesis and inhibition of thymidylate synthase and tumour cell growth.
1-[(2-Hydroxyethoxy)methyl]-5-fluorouracil (HEMFU) and 1-[(1,3-dihydroxy-2-propoxy)methyl]-5-fluorouracil (DHPFU) were prepared by alkylation of the di-O-TMS derivative of 5-fluorouracil and phosphorylated with the use of the wheat shoot phosphotransferase system to their monophosphates, HEMFUMP and DHPFUMP. 1-(2-Phosphonylmethoxyethyl)-5-fluorouracil (PMEFU) was obtained by condensation of diethyl-2-chloroethoxymethanephosphonate with 5-fluorouracil and cleavage of the alkylphosphoester with trimethylbromosilane. Inhibition of highly purified thymidylate synthase from mouse tumour Ehrlich carcinoma and leukemia L1210 cells by each of the nucleotide analogues, DHPFUMP, PMEFU and HEMFUMP, and of L5178Y mouse leukemia cell growth by the nucleoside (HEMFU) analogue, were studied. DHPFUMP proved to be the strongest inhibitor, non-competitive vs dUMP, with K(i)app 2.8 microM for time-independent interaction with the enzyme and N5,N10-methylenetetrahydrofolate (CH2H4PteGlu). In the presence of CH2H4PteGlu, DHPFUMP exhibited time-dependent inactivation of the enzyme, the inactivation rate plots being biphasic and pointing to Ki values in the microM range (10(3)-fold higher than for 5-fluoro-dUMP). HEMFUMP and PMEFU were much weaker inhibitors of the enzyme, with K(i)app values of 0.26 mM (non-competitive vs dUMP) and 30 mM (non-competitive vs dUMP), respectively. HEMFU, despite the weak interaction of its nucleotide analogue with the enzyme, proved to be a strong cell (L5178Y) growth inhibitor, with IC50 in the range 10(-5) M.
Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warsaw.
K
K
Felczak
B
B
Gołos
J M
JM
Dzik
W
W
Rode
M
M
Bretner
D
D
Shugar
T
T
Kulikowski
D000818:Animals; D000970:Antineoplastic Agents / Q000138:chemical synthesis* / Q000494:pharmacology; D002455:Cell Division / Q000187:drug effects; D004354:Drug Screening Assays, Antitumor; D004791:Enzyme Inhibitors / Q000138:chemical synthesis* / Q000494:pharmacology; D005468:Fluorodeoxyuridylate / Q000031:analogs & derivatives*; D005472:Fluorouracil / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000494:pharmacology; D007942:Leukemia, Experimental / Q000188:drug therapy / Q000473:pathology; D051379:Mice; D013940:Thymidylate Synthase / Q000037:antagonists & inhibitors*; D014407:Tumor Cells, Cultured
C115026:1-((1,3-dihydroxy-2-propoxy)methyl)-5-fluorouracil; C115025:1-((2-hydroxyethoxy)methyl)-5-fluorouracil; D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; D005468:Fluorodeoxyuridylate; D013940:Thymidylate Synthase; D005472:Fluorouracil