Synthesis and interactions with thymidylate synthase of 2,4-dithio analogues of dUMP and 5-fluoro-dUMP.
The 2,4-dithio analogues of 2'-deoxyuridine and 2'-deoxy-5-fluorouridine have been synthesized by thiation of the previously described 2-thio analogues, and then phosphorylated enzymatically or chemically to yield 2,4-dithio-dUMP and 2,4-dithio-5-fluoro-dUMP. In striking contrast to the 2-thio and 4-thio analogues of dUMP, which are good substrates of thymidylate synthase, 2,4-dithio-dUMP is not a substrate. But, surprisingly, it is a competitive inhibitor, relative to dUMP, of the purified enzymes from both parental and FdUrd-resistant L1210 cells, with K(i) values of 32 microM and 55 microM, respectively. Although 2,4-dithio-5-fluoro-dUMP behaved as a typical slow-binding inhibitor of the enzyme, its K(i) value was 10(3)-10(4)-fold higher than those for the corresponding 2-thio and 4-thio congeners. Similarly, 2,4-dithio-FdUrd was a much weaker inhibitor of tumour cell growth (IC50 approximately 10(-5)M) than FdUrd (IC50 approximately 10(-9)M), 2-thio-FdUrd(IC50 approximately 10(-7)M) or 4-thio-FdUrd (IC50 approximately 5x10(-8)M), while with 2,4-dithio-dUrd no influence on cell growth could be observed. Theoretical considerations, based on calculated aromaticities of the uracil and thiouracil rings, suggest that lack of substrate activity of 2,4-dithio-dUMP may result from increased pyrimidine ring aromaticity of the latter, leading to resistance of C(6) to nucleophilic attack by the enzyme active center cysteine.
Dzik
J M
JM
Nencki Institute of Experimental Biology, Polish Academy of Sciences, Warszawa, Poland.
Bretner
M
M
Kulikowski
T
T
Gołos
B
B
Jarmuła
A
A
Poznański
J
J
Rode
W
W
Shugar
D
D
D000818:Animals; D001665:Binding Sites; D002455:Cell Division / Q000187:drug effects; D003856:Deoxyuracil Nucleotides / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D019008:Drug Resistance, Neoplasm; D004791:Enzyme Inhibitors / Q000378:metabolism / Q000494:pharmacology; D005467:Floxuridine / Q000494:pharmacology; D005468:Fluorodeoxyuridylate / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D007700:Kinetics; D007939:Leukemia L1210; D009682:Magnetic Resonance Spectroscopy; D008968:Molecular Conformation; D015394:Molecular Structure; D010766:Phosphorylation; D010770:Phosphotransferases / Q000378:metabolism; D011485:Protein Binding; D013873:Thionucleotides / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D013940:Thymidylate Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D014407:Tumor Cells, Cultured
C099748:1-(2-deoxyribofuranosyl)-2,4-dithio-5-fluorouracil-5'-phosphate; C099747:1-(2-deoxyribofuranosyl)-2,4-dithiouracil-5'-phosphate; D003856:Deoxyuracil Nucleotides; D004791:Enzyme Inhibitors; D013873:Thionucleotides; D005467:Floxuridine; D005468:Fluorodeoxyuridylate; C007267:2'-deoxyuridylic acid; D013940:Thymidylate Synthase; D010770:Phosphotransferases; C021892:nucleoside phosphotransferase