Synthesis and antitumor evaluation of bis[(pivaloyloxy)methyl] 2′-deoxy-5-fluorouridine 5′-monophosphate (FdUMP): A strategy to introduce nucleotides into cells

David Farquhar, Saeed Khan, Devendra N. Srivastva, Priscilla P. Saunders

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100 Scopus citations


The bis[(pivaloyloxy)methyl] [PIV2] derivative of 2′-deoxy-5-fluorouridine 5′-monophosphate (FdUMP) was synthesized as a potential membrane-permeable prodrug of FdUMP. The compound was designed to enter cells by passive diffusion and to revert to FdUMP after removal of the PIV groups by hydrolytic enzymes. The most convenient preparation of PIV2FdUMP was by condensation of 2′-deoxy-5-fluorouridine (FUdR) with PIV2 phosphate in the presence of triphenylphosphine and diethyl azodicarboxylate (the Mitsunobo reagent). PIV2FdUMP was stable in the pH range 1.0-4.0 (t1/2 > 100 h). It was also fairly stable at pH 7.4 (t1/2 = 40.2 h). In 0.05 M NaOH solution, however, it was rapidly degraded (t1/2 <2 min). In the presence of hog liver carboxylate esterases, PIV2FdUMP was converted quantitatively to the mono-[(pivaloyloxy)methyl] [PIV1] analogue PIV1FdUMP. After a 24 h incubation, only trace amounts of FdUMP (1-3%) were observed, indicating that PIV1FdUMP is a poor substrate for carboxylate esterases. In mouse plasma, PIV2FdUMP was rapidly metabolized, first to PIV1FdUMP and then to FdUMP. With continued incubation, FUdR was formed, presumably due to further catabolism of FdUMP by plasma phosphatases or 5′-nucleotidases. Since PIV1FdUMP is a poor substrate for carboxylate esterase, the cleavage of the second PIV group is most likely mediated by plasma phosphodiesterases. The rate of degradation of PIV2FdUMP in the presence of acid and alkaline phosphatase, 5′-nucleotidase, or spleen phosphodiesterase was the same as that in buffer controls, indicating that the compound is not a substrate for these nucleotide catabolizing enzymes. The concentration of PIV2FdUMP and its 3′-O-acetyl ester (PIV2 3′-O-Ac-FdUMP) required to inhibit the growth of Chinese hamster ovary (CHO) cells in vitro to less than 50 cells per colony was 5 × 10-6 M, the same as that required for 5-fluorouracil (FU). Both nucleotide prodrugs showed the same growth-inhibitory potency against a mutant CHO cell line that was 20-fold resistant to FU (CHO/FU). Administered intraperitoneally at optimal dosage for 5 consecutive days, PIV2FdUMP and PIV2 3′-O-Ac-FdUMP were as effective as FU at prolonging the life spans of mice bearing intraperitoneally implanted P388 leukemia. Both prodrugs retained full therapeutic activity against a P388 subline resistant to FU. Collectively, these data indicate that PIV2FdUMP and PIV2 3′-O-Ac-FdUMP are effective membrane-permeable prodrugs of FdUMP.

Original languageEnglish (US)
Pages (from-to)3902-3909
Number of pages8
JournalJournal of Medicinal Chemistry
Issue number23
StatePublished - 1994
Externally publishedYes

ASJC Scopus subject areas

  • Organic Chemistry


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