Synthesis of methylene- and difluoromethylenephosphonate analogues of uridine-4-phosphate and 3-deazauridine-4-phosphate

Article abstract of DOI:10.1021/jo0617666

(Chemical Equation Presented) Cytidine triphosphate synthetase (CTPS) catalyzes the formation of cytidine triphosphate from glutamine, uridine-5′-triphosphate (UTP), and adenosine-5′-triphosphate. Inhibitors of CTPS are of interest because of their potential as therapeutic agents. One approach to potent enzyme inhibitors is to use analogues of high energy intermediates formed during the reaction. The CTPS reaction proceeds via the high energy intermediate UTP-4-phosphate (UTP-4-P). Four novel analogues of uridine-4-phosphate (U-4-P) and 3-deazauridine-4-phosphate (3-deazaU-4-P) were synthesized in which the labile phosphate ester oxygen was replaced with a methylene and difluoromethylene group. The methylene analogue of U-4-P, compound 1, was prepared by a reaction of the sodium salt of tert-butyl diethylphosphonoacetate with protected, 4-O-activated uridine followed by acetate deprotection and decarboxylation. It was found that this compound undergoes relatively facile dephosphonylation presumably via a metaphosphate intermediate. The difluoromethylene derivative, compound 2, was prepared by electrophilic fluorination of protected 1. This compound was stable and did not undergo dephosphonylation. Synthesis of the methylene analogue of 3-deazaU-4-P, compound 3, was achieved by ribosylation of protected 4-(phosphonomethyl)-2- hydroxypyridine. Electrophilic fluorination was also employed in the preparation of protected 4-(phospho-nodifluoromethyl)-2-hydroxypyridine which was used as the key building block in the synthesis of difluoro derivative 4. These compounds represent the first examples of a nucleoside in which the base has been chemically modified with a methylene or difluormethylenephosphonate group.

Full text of DOI:10.1021/jo0617666

Synthesis of Methylene- and Difluoromethylenephosphonate
Analogues of Uridine-4-phosphate and 3-Deazauridine-4-phosphate
Scott D. Taylor,*,† Farzad Mirzaei,† Ali Sharifi,† and Stephen L. Bearne‡
Department of Chemistry, UniVersity of Waterloo, 200 UniVersity AVenue West,
Waterloo, Ontario N2L 3G1, Canada, and Department of Biochemistry and Molecular Biology,
Dalhousie UniVersity, Halifax, NoVa Scotia, B3H 4H7 Canada
s5taylor@sciborg.uwaterloo.ca
ReceiVed August 27, 2006
Cytidine triphosphate synthetase (CTPS) catalyzes the formation of cytidine triphosphate from glutamine,
uridine-5′-triphosphate (UTP), and adenosine-5′-triphosphate. Inhibitors of CTPS are of interest because
of their potential as therapeutic agents. One approach to potent enzyme inhibitors is to use analogues of
high energy intermediates formed during the reaction. The CTPS reaction proceeds via the high energy
intermediate UTP-4-phosphate (UTP-4-P). Four novel analogues of uridine-4-phosphate (U-4-P) and
3-deazauridine-4-phosphate (3-deazaU-4-P) were synthesized in which the labile phosphate ester oxygen
was replaced with a methylene and difluoromethylene group. The methylene analogue of U-4-P, compound
1, was prepared by a reaction of the sodium salt of tert-butyl diethylphosphonoacetate with protected,
4-O-activated uridine followed by acetate deprotection and decarboxylation. It was found that this
compound undergoes relatively facile dephosphonylation presumably via a metaphosphate intermediate.
The difluoromethylene derivative, compound 2, was prepared by electrophilic fluorination of protected
1. This compound was stable and did not undergo dephosphonylation. Synthesis of the methylene analogue
of 3-deazaU-4-P, compound 3, was achieved by ribosylation of protected 4-(phosphonomethyl)-2-
hydroxypyridine. Electrophilic fluorination was also employed in the preparation of protected 4-(phospho-
nodifluoromethyl)-2-hydroxypyridine which was used as the key building block in the synthesis of difluoro
derivative 4. These compounds represent the first examples of a nucleoside in which the base has been
chemically modified with a methylene or difluormethylenephosphonate group.
Introduction
the formation of cytidine triphosphate (CTP) from glutamine,
uridine-5′-triphosphate (UTP), and adenosine-5′-triphosphate
(ATP; Scheme 1). It has been proposed, on the basis of
numerous mechanistic investigations,2 that CTPS performs this
reaction by transferring the γ-phosphate from ATP to the oxygen
at position 4 of UTP to form UTP-4-phosphate (UTP-4-P) as
an intermediate and adenosine-5′-diphosphate (ADP). Ammonia,
which is generated at a separate site via rate-limiting glutamine
The synthesis of chemically modified nucleosides is a very
active area of chemical research. This is mainly due to their
therapeutic potential as evidenced by the considerable number
of nucleoside analogues that are now on the market as antiviral
and anticancer agents.1 Our interest in nucleoside analogues
stems from our desire to prepare inhibitors of cytidine tri-
phosphate synthetase (CTPS, E. C. 6.4.3.2). CTPS catalyzes
(1) (a) Jordheim, L. P.; Galmarini, C. M.; Dumontet, C. Recent Pat. Anti-
Cancer Drug DiscoVery 2006, 1, 163. (b) Carroll, S. S.; Olen, D. B. Infect.
Disord.: Drug Targets 2006, 6, 17.
† University of Waterloo.
‡ Dalhousie University.
10.1021/jo0617666 CCC: $33.50 © 2006 American Chemical Society
Published on Web 11/14/2006
9420
J. Org. Chem. 2006, 71, 9420-9430

Uridine- and 3-Deazauridine-4-phosphate Analogues
SCHEME 1. Reaction Catalyzed by CTPS
CHART 1. Structures of Target Analogues of U-4-P
hydrolysis, then displaces the phosphate at the O-4 of UTP-
4-P to give CTP. The reaction catalyzed by CTPS is the rate-
limiting step in the biosynthesis of all cytosine nucleotides and
provides one of the crucial building blocks for DNA synthesis.
Therefore, it is hardly surprising that inhibitors of CTPS are
being examined as a means of decreasing the viability of certain
cancers and pathogenic organisms. For example, the nucleoside
analogues 3-deazauridine (3-DaU) and cyclopentenylcytosine
(CPEC) are triphosphorylated intracellularly which inhibits
CTPS and reduces intracellular CTP levels.3,4 Studies with
several cancer cell lines and viruses indicate that 3-DaU and
CPEC potentiate the effects of various anticancer and antiviral
agents which suggests that they could be employed in combina-
tion drug therapy.5 More recently, 3-DaU alone has been shown
to induce apoptosis in myeloid leukemia cells in a dose-
dependent manner.6 One method of generating potent inhibitors
of enzymes is to use analogues either of the transition state of
the reaction or of a high energy intermediate formed during
catalysis.7 UTP-4-P is a high energy intermediate formed during
the CTPS reaction. Consequently, a hydrolytically stable
analogue of this species may exhibit potent inhibition of CTPS.
Analogues of phosphorylated compounds which contain the
isosteric and/or isoelectronic methylene or difluoromethylene
phosphonate moiety in place of the phosphate group often inhibit
enzymes which act upon phosphorylated substrates.8 Here we
report the synthesis of four novel analogues of uridine-4-
phosphate (U-4-P) and 3-deazauridine-4-phosphate (3-DaU-4-
P), compounds 1-4 (Chart 1), in which the labile phosphate
group is replaced with a methylene- or difluoromethylenephos-
honate moiety.
Results and Discussion
There are only a limited number of reports of phosphonopy-
rimidine ribonucleosides. Honjo et al. have reported the synthesis
of 2′,3′,5′-tri-O-benzoyl-4-diethylphosphonouridine by reacting
1-(2′,3′,5′-tri-O-benzoyl-â-D-ribofuranosyl)-4-chloro-2(H)-py-
rimidinone with triethyl phosphite.9 These workers also reported
the synthesis of 5- and 6-phosphonouridine by reaction of
lithiated 2′,3′,5′-tri-O-protected uridine with diethylchlorophos-
phate followed by deprotection.9 Maruyama and co-workers
have reported the synthesis of phosphonopyrimidine ribonucleo-
sides by photolysis of 2′,3′-di-O- or 2′,3′,5′-tri-O-protected
5-bromouridine and triethylphosphite.10 In all of these cases,
the phosphonate group is attached directly to the pyrimidine
ring. To our knowledge, nucleosides where the base bears a
methylene or difluormethylenephosphonic acid group have never
been reported.
We began our studies with the synthesis of compound 1, in
which the O-4 of U-4-P was replaced with a methylene unit.
Synthetic approaches to C-4 substituted pyrimidine nucleosides
include ribosylation of C-4 substituted pyrimidines,11 sulfur
extrusion of S-alkylated 4-thiouridines,12 palladium-catalyzed
cross-coupling of 4-chloropyrimidines with terminal alkynes,13
or Bischofberger’s approach which involves activation of the
(2) (a) Long, C. W.; Levitzki, A.; Koshland, D. E., Jr. J. Biol. Chem.
1979, 245, 80. (b) Koshland, D. E., Jr.; Levitzki, A. Biochemistry 1971,
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(d) von der Saal, W.; Villafranca, J. J.; Anderson, P. M. J. Am. Chem. Soc.
1985, 107, 703. (e) von der Saal, W.; Anderson, P. M.; Villafranca, J. J. J.
Biol. Chem. 1985, 260, 14993. (f) Levitzki, A.; Koshland, D. E., Jr.
Biochemistry 1972, 11, 241. (g) Levitzki, A.; Koshland, D. E., Jr.
Biochemistry 1971, 10, 3365. (h) Lewis, D. A.; Villafranca, J. J. Biochem-
istry 1989, 28, 8454.
(3) McPartland, R. P.; Wang, M. C.; Bloch, A.; Weinfeld, H. Cancer
Res. 1974, 34, 3107.
(8) (a) Blackburn, G. M. Chem. Ind. (London) 1981, 134. (b) Lazarus,
R. A.; Benkovic, P. A.; Benkovic, S. J. Arch. Biochem. Biophys. 1979,
197, 218. (c) Engel, R. Chem. ReV. 1977, 77, 349.
(4) Kang, G. J.; Cooney, D. A.; Moyer, J. D.; Kelley, J. A.; Kim, H.-Y.;
Matquez, V. E.; Johns, D. G. J. Biol. Chem. 1989, 264, 713.
(5) (a) Barlogie, B.; Plunkett, W.; Raber, M.; Latreille, J.; Keating, J.;
McCredie, K. B. Cancer Res. 1981, 41, 1227. (b) Mills-Yamamoto, C.;
Lauzon, G. J.; Peterson, A. R. P. Biochem. Pharmacol. 1978, 27, 181. (c)
Momparler, R. L.; Momparler, L. F. Cancer Chemother. Pharmacol. 1989,
25, 51. (d) Momparler, R. L.; Bouffard, D. Y.; Momparler, L. F.; Marquet,
J.; Zittoun, J.; Marie, J.-P.; Zittoun, R. Int. J. Cancer 1991, 49, 573. (e)
Momparler, R. L.; Vesely, J.; Momparler, L. F.; Rivard, G. E. Cancer Res.
1979, 39, 573. (f) Bouffard, D. Y.; Momparler, L. F.; Momparler, R. L.
Anti-Cancer Drugs 1994, 5, 223.
(9) Honjo, M.; Maruyama, T.; Horikawa, M.; Balzarini, J.; de Clercq,
E. Chem. Pharm. Bull. 1987, 35, 3227.
(10) Maruyama, T.; Kimura, S.; Honjo, M. Org. Prep. Proced. Int. 1988,
20, 485.
(11) (a) Kim, C.-H.; Marquez, V. E.; Mao, D. T.; Haines, D. R.;
McCormack, J. J. J. Med. Chem. 1986, 29, 1374 (b) Schlieper, C. A.;
Wemple, J. Nucleosides Nucleotides 1984, 3, 369.
(12) (a) Vorbruggen, H.; Korlikiewicz, K. Angew. Chem., Int. Ed. Engl.
1976, 15, 689. (b) Yamane, A.; Inoue, H.; Ueda, T. Chem. Pharm. Bull.
1980, 28, 157.
(13) Acevedo, O. L.; Andrews, R. S.; Dunkel, M.; Cook, P. D. J.
Heterocycl. Chem. 1994, 31, 989.
(6) Savickiene, J.; Gineitis, A. Int. J. Biochem. Cell Biol. 2003, 35, 1482.
(7) (a) Wolfenden, R. Acc. Chem. Res. 1972, 5, 10. (b) Wolfenden, R.
Annu. ReV. Biophys. Bioeng. 1976, 5, 271.
J. Org. Chem, Vol. 71, No. 25, 2006 9421

Taylor et al.
SCHEME 2. Synthesis of Compound 1
4-position of protected uridines followed by reaction with
malonyl-type nucleophiles.14 We anticipated that Bischofberger’s
approach would be applicable to the synthesis of 1 by reacting
a protected phosphonoacetate derivative with protected, 4-O-
activated uridine followed by acetate deprotection and decar-
boxylation. Thus, 2′,3′,5-tri-O-benzoyluridine was activated at
the 4-position by reaction with triisopropylbenzenesulfonyl-
chloride (TIPBSCl) in the presence of triethylamine which gave
compound 5 in a 62% yield (Scheme 2). The reaction of 5 with
2.1 equiv of benzyl dimethylphosphonoacetate (6)15 gave
compound 7 in an 88% yield. We anticipated that upon
hydrogenolysis of the benzyl group in 7 the resulting carboxylic
acid would then spontaneously decarboxylate to give phospho-
nate 10.12b However, the yield of this reaction was surprisingly
low, and although some of the desired product was isolated, it
was contaminated with a considerable amount of the tetrahy-
drouridine derivative which resulted from hydrogenation of the
pyrimidine ring at C-5 and C-6; we were unable to obtain 10
in pure form. Therefore, the sodium salt of tert-butyl dimeth-
ylphosphonoacetate 816 was used, which gave compound 9 in
an 83% yield. Treatment of 9 with TFA in CH2Cl2 at -10 °C
resulted in the removal of the tert-butyl group and decarbox-
Spiking the sample with sodium phosphate resulted in an
increase in the size of the minor peak, which suggested that
the minor peak was inorganic phosphate. Attempts to purify
nucleoside 1 by RP-HPLC using a variety of different eluents
at different pH values proved unsuccessful, in that the minor
peak was always present in varying amounts after HPLC and
conversion to the sodium salt. Additional 31P NMR studies in
D2O showed that compound 1 decomposed over time as the
peak at 15.8 ppm decreased and the peak at 1.7 ppm increased.
After 48 h, the peak at 1.7 ppm exhibited 25% of the intensity
of the peak at 15.8 ppm. Decomposition of 10 occurred faster
in a 3:2 DMSO-d6-D2O mixture. A 31P NMR spectrum of the
compound taken within the first 20 min of dissolution showed
a major peak at 13.0 ppm and a small peak at 0.8 ppm. After
just 48 h, the peak at 13.0 ppm had almost disappeared, and
the peak at 0.8 ppm was by far the major peak (Figure 1). The
negative ESI mass spectrum revealed a strong peak with a m/z
of 321 which corresponds to compound 1. Among the peaks in
the positive ESI mass spectrum was a peak with a m/z of 243.
This peak increased in intensity during storage of 1 in water or
1
DMSO-water. The H NMR of this material revealed that
1
compound 1 exists mainly as the CdN tautomer in D2O as
evidenced by a doublet at approximately 3.0 ppm, which
corresponds to that of the methylene protons adjacent to the
phosphorus. This doublet almost completely disappeared after
9 h in D2O indicating that these protons are readily exchanged
with the solvent’s deuterons. A doublet at 6.6 ppm correspond-
ing to the H-5 proton of 1 slowly decreased with time while
another doublet at 6.5 ppm, which initially constituted 5% of
these two doublets, increased over time and, after 48 h, then
constituted 25% of these doublets. In the 3:2 DMSO-d6-D2O
mixture, the doublet at 6.5 ppm constituted approximately 87%
of these two doublets after 48 h. Finally, a small broad singlet
at 2.3 ppm, which was present within the first 15 min after
dissolution in D2O, was barely evident after 48 h.
ylation, which gave compound 10 in an 83% yield. H NMR
revealed that, in CDCl3, compound 10 existed exclusively as
the tautomer in which the double bond is exocyclic to the
pyrimidine ring.
Deprotection of the phosphonate moiety in 10 was achieved
by a reaction with TMSBr in CH2Cl2 for 8 h. After removal of
the solvent and unreacted TMSBr, the residue was subjected to
NH4OH/MeOH for 24 h to remove the benzoyl-protecting
groups, after which the reaction was concentrated and then
lyophilized several times to remove excess NH4OH. A 31P NMR
spectrum of the crude product in D2O showed that, in addition
to a large peak at 15.8 ppm which corresponded to compound
1, a very small peak at 1.7 ppm, which was approximately 2%
of the intensity of the peak at 15.8 ppm, was also present.
These studies reveal that 10 dephosphonylates in solution to
give 1-(â-D-ribofuranosyl)-4-methyl-2-pyrimidinone (11; Scheme
3) and inorganic phosphate. The new doublet at 6.5 ppm and
the singlet at 2.3 ppm correspond to the H-5 proton and the
(14) (a) Bischofberger, N. Tetrahedron Lett. 1987, 28, 2821. (b) Blanalt-
Feidt, S.; Doronina, S. O.; Behr, J.-P. Tetrahedron Lett. 1999, 40, 6229.
(15) Bradley, J. C.; Buechi, G. J. Org. Chem. 1976, 41, 699.
(16) Giese, B.; Linker, T. Synthesis 1992, 1-2, 46.
9422 J. Org. Chem., Vol. 71, No. 25, 2006

Uridine- and 3-Deazauridine-4-phosphate Analogues
FIGURE 1. 31P NMR spectra of compound 1 in 3:2 DMSO-d6:D2O taken at various time intervals over a period of 48 h.
SCHEME 3. Possible Mechanisms for the
Dephosphonylation of Compound 1
phate-like transition state (Scheme 3). We are not aware of any
studies demonstrating this phenomenon with 4-phosphono-
methyl-2-pyrimidinones; however, to our knowledge, this is the
first synthesis of this class of compounds. Nevertheless, Warren
has shown that aminomethylphosphonic acid reacts instanta-
neously at room temperature with ninhydrin in aq ethylene
glycol and triethylamine to give a mixture of inorganic
phosphate and 2-hydroxyethylphosphate.19 It was suggested that
this reaction proceeds by formation of imine 12 followed by
rapid decomposition via formation of metaphosphate (Scheme
4).19 Martell and Langohr have suggested that the facile
dephosphonylation of the metal chelate of the imine resulting
from the reaction of 2-amino-3-phosphonopropionic acid with
pyridoxal proceeds by the unimolecular dissociation of meta-
phosphate.20
To prepare the fluorinated analogue of 1, phosphonate 10
was subjected to electrophilic fluorination21 using potassium
hexamethyldisilazane (KMHDS) and N-fluorobenzenesulfon-
imide (NFSi) which gave compound 13 in a 45% yield (Scheme
5). Complete deprotection of 13, as described above for
compound 10, gave compound 2 in a 51% yield after purifica-
tion by HPLC and conversion to its sodium salt. Unlike
compound 1, compound 2 did not undergo any detectable
dephosphonylation even when stored for days in D2O or DMSO.
Very little has appeared in the literature concerning the influence
of R-fluorines on the rates of decarboxylation of â-keto
carboxylic acid, and, to our knowledge, no studies concerning
their effect on the dephosphonylation of â-ketophosphonates
or â-iminophosphonates have been reported. Bartlett et al. have
noted that difluorooxaloacetic acid is extraordinarily resistant
to decarboxylation in an aqueous solution. This was attributed
to the difficulty in cracking the hydrate which dominates in
aqueous solutions as well as to the destabilizing influence of
methyl protons in compound 11.17 The methyl protons exchange
slowly with the solvent deuterons. The high-resolution positive
ion ESI mass spectrum of the peak with a m/z of 243 was also
consistent with the decomposition product being compound 11.
Carbon-phosphorus bonds are usually highly resistant to
hydrolysis. Nevertheless, some phosphonates are unstable
toward C-P bond cleavage giving inorganic phosphate and/or
polyphosphates. This has been well documented for â-carbonyl
phosphonates, though usually these reactions require elevated
temperatures and prolonged reaction times at a neutral pH.18 It
has been suggested that these reactions proceed by a dissociative
mechanism involving metaphosphate.18 The dephosphonylation
of 10 occurs with considerable ease and probably proceeds via
a similar mechanism involving metaphosphate or a metaphos-
(17) 1-(â-D-Ribofuranosyl)-4-methyl-2-pyrimidinone (11) is a known
(19) Warren, S. G. J. Chem. Soc. C 1966, 1349.
(20) Martell, A.; Langohr, M. F. J. Chem. Soc., Chem. Commun. 1977,
342.
compound. See: Holy, A.; Ludzisa, A.; Votruba, I.; Sediva, K.; Pischel,
1
H. Collect. Czech. Chem. Commun. 1985, 50, 393. Its H NMR spectrum
was not reported.
(21) (a) Differding, E.; Duthaler, R. O.; Kreiger, A.; Ruegg, G. M.;
Schmit, C. Synlett 1991, 395. (b) Taylor, S. D.; Kotoris, C., Dinaut, A. N.;
Chen, M.-J. Tetrahedron 1998, 54, 1691.
(18) Freeman, S.; Irwin, W. J.; Schwalbe, C. H. J. Chem. Soc., Perkin
Trans. 2 1991, 263.
J. Org. Chem, Vol. 71, No. 25, 2006 9423

Taylor et al.
SCHEME 4. Reaction of Aminomethylphosphonic Acid with Ninhydrin
SCHEME 5. Synthesis of Compound 2
SOCl2.23 To this end, compound 18 was prepared in a 29% yield
from 4-carbomethoxypyridine using the method of Boekelheide
and Lehn which involved oxidation of 4-carbomethoxypyridine
to the N-oxide (19) followed by a reaction with refluxing Ac2O
(Scheme 7).24,25 In our hands, Uchida’s procedure for the
synthesis of 17 from 18 proceeded with a yield of only 21%.
Finally, we were unable to obtain phosphonate 16 from 17 in
yields greater than 35% which translates to an overall yield of
16 from 4-carbomethoxy-2-pyridinone of just 2.1%.
CHART 2. Eneamine-Type Compounds 14 and 15
Because of the low yields using the approach outlined in
Scheme 7, alternative routes to 16 were examined (Scheme 8).
A reaction of 2-chloroisonicotinic acid (20) with thionyl chloride
followed by a reaction of the crude acid chloride with benzyl
alcohol gave the corresponding 2-O-benzyl derivative 21.26 The
crude ester was reacted with the sodium salt of benzyl alcohol
which gave the dibenzyl-protected compound 21 in an 80% yield
(three steps).26 Reduction of the ester with LiBH4 in the presence
of a small amount of methanol in refluxing ether gave compound
22 in an 86% yield. Reaction of compound 22 with POBr3 in
DMF/CH2Cl2 gave alkyl bromide 23 in a 77% yield. A
Michaelis-Arbuzov reaction between 23 and trimethylphosphite
performed either neat or in benzene resulted in reactions at both
the methylbromide moiety and the benzylic carbon of the
benzyl-protecting group. This afforded a mixture of the desired
product 24 and dimethyl benzylphosphonate in an overall low
yield, and we were unable to separate 24 from the contaminating
dimethyl benzylphosphonate. Hydrogenolysis of crude 24 gave
16 in a 24% yield from 23 (two steps). The reaction of 23 with
the sodium salt of dimethylphosphite also proceeded very poorly.
Although this route was an improvement over the approach
taken in Scheme 7, the difficulties encountered with the
Michaelis-Arbuzov reaction prompted us to devise an alterna-
tive route to 16 (Scheme 8). Compound 20 was converted to
2-hydroxyisonicotinic acid (25) in an 89% yield by heating it
in concd HCl for 48 h. Reduction of the acid moiety using BH3-
THF gave alcohol 26 in a 54% yield. The reaction of 26 with
SCHEME 6. Retrosynthesis of Compound 3
the fluorines on enol formation.22 It is possible that the presence
of the fluorines may have a destabilizing influence on the
formation of the enamine-type compounds 14 and 15 (Chart
2), and this may also account for the stability of 2 toward
dephosphonylation.
3-DeazaUTP is an inhibitor of CTP synthetase.3,4 CTPS
exhibits the same affinity for 3-deazaUTP as it does for the
substrate UTP which indicates that the enzyme does not require
the nitrogen at the 3-position for binding, at least in the ground
state.2 We anticipated that deaza nucleotides 3 and 4 would
exhibit an affinity for CTPS which is similar to that of aza
compounds 1 and 2, yet neither would undergo dephosphony-
lation because of the lack of a nitrogen at the 3-position. Our
approach to compound 3, as illustrated in Scheme 6, was to
couple phosphonate 16 to an appropriately protected ribofura-
nose derivative. Our initial route to phosphonate 16 was via
alkyl chloride 17 which has been prepared by Uchida et al. by
LiAlH4 reduction of the 4-carbomethoxy-2-hydroxypyridine (18)
followed by a reaction of the crude alcohol product with
(23) Uchida, M.; Tabusa, F.; Komatsu, M.; Morita, S.; Kanbe, T.;
Nakagawa, K. Yakugaku Zasshi 1985, 105, 1040.
(24) Ochai, E. J. Org. Chem. 1953, 18, 534.
(25) Boekelheide, V.; Lehn, W. L. J. Org. Chem. 1961, 26, 428.
(26) Shiozawa, A.; Masahiro, S.; Kazuto, S.; Ichiro, K.; Narita, K.;
Inubushi, A.; Araki, J. Japn. Patent 06080635, 1994.
(22) Alberg, D. G.; Lauhon, C. T.; Nyfeler, R.; Fassler, A.; Bartlett, P.
A. J. Am. Chem. Soc. 1992, 114, 3535.
9424 J. Org. Chem., Vol. 71, No. 25, 2006

Uridine- and 3-Deazauridine-4-phosphate Analogues
SCHEME 7. Synthesis of Phosphonate 16 from 4-Carbomethoxypyridine
SCHEME 8. Formation of Phosphonate 16 from 2-Chloroisonicotinic Acid
SCHEME 9. Synthesis of Compound 3 from Phosphonate 16
methane sulfonylbromide in the presence of 2,6-lutidine resulted
both in bromination of the hydroxylmethyl functionality and in
sulfonation at the 2-position to give compound 27 in a 71%
yield. The Michaelis-Arbuzov reaction between 27 and tri-
methylphosphite proceeded smoothly to give phosphonate 28
in a 93% yield. Finally, the removal of the sulfonate group was
achieved using LiOH in THF-H2O which gave compound 16
in a 76% yield. Using this approach, we found that the overall
yield of 16 improved to 24%.
To obtain compound 3, phosphonate 16 was treated with bis-
(trimethylsilyl)trifluoroacetamide (BTMSTFA) followed by a
reaction with acetyl 2,3,5-tri-O-benzoyl-â-D-ribofuranose in the
presence of trimethylsilyl triflate (TMSOTf; Scheme 9). This
gave nucleoside 29 in a 65% yield. Complete deprotection of
29 was achieved by treatment with 3 equiv of TMSBr for 8 h
followed by a reaction with aq NH4OH in methanol for 24 h.
HPLC purification of the resulting material using TFA (0.1%)
in H2O-CH3CN as the eluent gave 3 as its free acid in a 90%
yield. Nucleoside 3 did not exhibit any decomposition even after
storage for several weeks in water or DMSO indicating that
the ring nitrogen is essential for dephosphonylation.
We initially attempted to construct compound 4 by electro-
philic fluorination of compound 29 followed by deprotection
in a manner analogous to that described for the synthesis of
compound 2 (Scheme 9). Although 19F NMR spectra of the
crude product suggested that electrophilic fluorination of 29 gave
the desired compound 30, a number of byproducts were also
formed, and we were unable to obtain 30 in pure form.
J. Org. Chem, Vol. 71, No. 25, 2006 9425

Taylor et al.
SCHEME 10. Synthesis of Compound 4
Therefore, for the synthesis of compound 4, we envisaged a
route similar to that taken for nucleoside 29 where the
difluoromethylenephosphonate analogue of 16 would be pre-
pared and then coupled to the protected ribose. Unfortunately,
electrophilic fluorination of 16 gave a complex mixture of
products. Nevertheless, we reasoned that a hydroxyl-protected
version of 16 would undergo electrophilic fluorination more
cleanly. Phosphonate 28 was not considered to be a viable
candidate for electrophilic fluorination because of the presence
of the methylsulfonate moiety. Nonetheless, we reasoned that
phosphonate 24, which contained a benzyl-protected hydroxyl
group, would readily undergo electrophilic fluorination. Unfor-
tunately, preparing this compound was problematic because of
the low yield of the Michaelis-Arbuzov reaction. However,
we were surprised to find that when performing the Michaelis-
Arbuzov reaction on 23 with triethylphosphite in benzene at
95-105 °C, phosphonate 31 could be obtained in a 93% yield,
and no diethyl benzylphosphonate was formed (Scheme 10).27
The reason for the difference in yield and product distribution
when using the two different phosphites is not clear. One
possible explanation is that the benzylic carbon may be slightly
more sterically hindered than the alkyl bromide carbon and,
therefore, might be less susceptible to an attack by the more
bulky triethylphosphite. Fluorination of 31 using 2.2 equiv of
NaHMDS and 2.5 equiv of NFSi gave compound 32 in an 87%
yield. The benzyl group in 32 was removed by hydrogenolysis
using catalyst Pd/C to give compound 33 in a 95% yield.
Coupling of phosphonate 33 to acetyl 2,3,5-tri-O-acetyl-â-D-
ribofuranose using the procedure described above for the
synthesis of 29 gave the protected nucleoside 34 in a 78% yield.
Complete deprotection of 34 was achieved in the usual manner,
except removal of the ethyl-protecting groups from 34 required
8 equiv of TMSBr added over a period of 3 days. It has been
reported that anomerization can occur when subjecting 2′-deoxy
nucleotide analogues to TMSBr.28 However, 31P and 1H NMR
analysis of the crude reaction mixture revealed that anomer-
ization had not taken place. This could be due to the presence
of the benzoyl-protecting group at the 2′-position. After HPLC
purification using TFA (0.1%) in H2O-CH3CN as the eluent,
nucleoside 4 was obtained as its free acid in a 45% yield. This
deprotection yield is somewhat lower than that of its nonflu-
orinated analogue 3, and this is probably due to the prolonged
exposure of 34 to TMSBr. As with nucleoside 3, compound 4
was stable and did not exhibit any dephosphonylation even after
prolonged storage in various solvents.
In summary, several unique analogues of U-4-P and 3-de-
azaU-4-P were prepared in which the labile phosphate ester
oxygen was replaced with either a methylene or a difluoro-
methylene group. The methylene analogue of U-4-P, compound
1, was readily prepared by a reaction of the sodium salt of a
tert-butyl diethylphosphonoacetate with protected, 4-O-activated
uridine followed by acetate deprotection and decarboxylation.
However, we were not able to isolate 1 in pure form because
of its tendency to undergo dephosphonylation, possibly via a
metaphosphate intermediate. The difluoromethylene analogue
of 1, nucleoside 2, was prepared by electrophilic fluorination
of protected 1. This compound did not undergo dephosphon-
ylation. The synthesis of the methylene analogue of 3-deazaU-
4-P, compound 3, was achieved by ribosylation of protected
4-(phosphonomethyl)-2-hydroxypyridine followed by deprotec-
tion. Electrophilic fluorination was also employed in the
preparation of protected 4-(phosphonodifluoromethyl)-2-hy-
droxypyridine which was ribosylated and then deprotected to
give difluoro derivative 4. To our knowledge, compounds 1-4
represent the first examples of ribonucleosides in which the base
has been chemically modified with a methylene or difluoro-
methylenephosphonate group. The antiproliferative properties
of these compounds will be examined, and the results will be
reported in due course. Finally, it has not escaped our attention
that the cell permeability of these compounds may be compro-
mised because oftheir highly anionic nature. If necessary, more
cell permeable phosphonate diester or phosphoramidate prodrug
derivatives29 of these compounds will be prepared and examined
as antiproliferative agents.
(27) This was discovered after compound 3 was prepared, and so
phosphonate 31 was not used for the synthesis of compound 3. In any case,
we preferred methyl-protecting groups as opposed to ethyl-protecting groups
for phosphonate protection since methyl groups are removed more readily
by TMSBr than ethyl groups; this reduces the compounds exposure to
TMSBr which can cause side reactions and lower yields. We would have
preferred methyl protection for the synthesis of compound 4; however, this
was not practical.
(28) Brossette, T.; Le Faou, A.; Goujon, L.; Valleix, A.; Creminon, C.;
Grassi, J.; Mioskowski, C.; Lebeau, L. J. Org. Chem. 1999, 64, 5083.
(29) Kurz, T.; Schluter, K.; Kaula, U.; Bergmann, B.; Walter, R. D.;
Geffken, D. Bioorg. Med. Chem. 2006, 14, 5121.
9426 J. Org. Chem., Vol. 71, No. 25, 2006

Uridine- and 3-Deazauridine-4-phosphate Analogues
the organic layer was washed with 10% NH4Cl and saturated brine.
The combined aq layers were back extracted with 200 mL of ether.
The combined organics were dried (MgSO4) and concentrated, and
the resulting foam was purified by flash chromatography (60%
hexane-40% EtOAc to 50% hexane-50% EtOAc) to give pure 9
Experimental Section
2′,3′,5′-Tris-(O-benzoyl)-4-O-[(2,4,6-triisopropylphenyl)sulfon-
yl]uridine (5). To a solution of 2,3,5-tri-O-benzoyluridine (7.00
g, 12.6 mmol) and DMAP (153 mg, 1.26 mmol) in dry CH2Cl2
(70 mL) at 0 °C (ice bath) was added triethylamine (10.5 mL, 75.6
mmol), and the solution was stirred for 30 min. A solution of
TIPBSCl (6.46 g, 21.4 mmol) in dry CH2Cl2 (15 mL) was added
dropwise over 5 min, and the mixture was stirred for 1 h at 4 °C;
then the ice bath was removed, and the mixture was stirred for a
further 6 h. A 1:1 mixture of EtOAc-hexane (400 mL) was added,
and the mixture was filtered. The filtrate was concentrated to about
50 mL, 400 mL of ether was added, and the mixture was filtered
again. The filtrate was concentrated. Purification of the residue by
flash chromatography (70% hexane-30% EtOAc) gave pure 5 as
1
as a white foam (3.0 g, 83% yield). H NMR (300 MHz) δ 12.89
(s, 0.5H), 12.51 (s, 0.5H), 8.03-8.09 (m, 2H), 7.85-7.95 (m, 4H),
7.64 (d, J ) 8.2 Hz, 0.5H), 7.20-7.60 (m, 9.5H), 7.09 (d, J ) 8.5
Hz, 0.5 H), 7.06 (d, J ) 8.6 Hz, 0.5H), 6.28 (d, J ) 5.8 Hz, 0.5H),
6.25 (d, J ) 5.6 Hz, 0.5H), 5.83-5.88 (m, 1H), 5.70-5.75 (m,
1H), 4.60-4.79 (m, 3H), 3.60-3.70 (m, 6H), 1.45 (s, 4.5H), 1.42
(s, 4.5H); 13C NMR (75 MHz) δ 168.8 (d, J ) 7.5 Hz), 166.0,
165.2 (d, J ) 14.2 Hz), 165.2, 165.2, 159.4 (d, J ) 24 Hz), 158.8
(d, J ) 8.6 Hz), 147.5, 147.5 (d, J ) 2.1 Hz), 134.9, 134.7, 133.6,
133.6, 133.5, 129.8, 129.7, 129.6, 129.2, 129.1, 128.6, 128.5, 128.4,
128.3, 102.5, 102.0 (d, J ) 15.4 Hz), 89.0, 88.6, 83.0 (d, J ) 203
Hz), 82.5 (d, J ) 186 Hz), 80.6, 80.4, 80.3, 73.4, 71.1, 71.0, 63.7,
52.4 (d, J ) 5.6 Hz), 51.9 (d, J ) 5.4 Hz), 28.1; 31P NMR (121
MHz) δ 28.0, 24.2; LR+ESIMS m/z (relative intensity) 763 (M +
1, 100), 707 (28). HR+ESIMS m/z: calcd for C38H40N2O13P,
763.2268; found, 763.2262.
1
a white foam (6.6 g, 64% yield). H NMR (300 MHz) δ 8.04 (d,
J ) 7.1 Hz, 2H), 7.92 (d, J ) 5.7 Hz, 3H), 7.87 (d, J ) 7.2 Hz,
2H), 7.30-7.62 (m, 11H), 7.19 (s, 2H), 6.28 (bs, 1H), 5.91 (d, J )
6.9 Hz, 1H), 5.82 (d, J ) 5.4 Hz, 1H), 5.75 (bs, 1H), 4.62-4.84
(m, 3H), 4.20-4.25 (m, 2H), 2.81-2.93 (m, 1H), 1.25 (d, J ) 7.6
Hz, 18H);13C NMR (75 MHz) δ 167.2, 165.9, 165.1, 165.0, 154.6,
153.3, 151.2, 145.3, 133.6, 130.3, 129.9, 129.7, 129.5, 129.1, 128.7,
128.4, 124.0, 95.7, 89.9, 80.4, 74.6, 70.4, 63.2, 34.2, 29.6, 24.5,
23.3; LR+ESIMS m/z (relative intensity) 823.5 (M + 1, 100), 445
(57). HR+ESIMS m/z: calcd for C45H47N2O11S, 823.2901; found,
823.2920.
{1-[3,4-Bis-(benzoyloxy)-5-benzoyloxymethyltetrahydrofur-
an-2-yl]-2-oxo-2,3-dihydro-1H-pyrimidin-4-ylidenemethyl}-
phosphonic Acid Dimethyl Ester (10). To a solution of nucleoside
9 (2.90 g, 3.81 mmol) in dry CH2Cl2 (60 mL) cooled in a KCl ice
bath (-10 °C) under Ar was added a solution of TFA (3.5 mL, 46
mmol, 12 equiv) in CH2Cl2 (60 mL) over a period of 30 min. The
bath was removed, and the reaction was stirred for 4 h. The mixture
was diluted with 800 mL of ether and washed with saturated
NaHCO3, water, and saturated brine. The combined aq layers were
back extracted with ether (2 × 200 mL). The combined organics
were dried (MgSO4) and concentrated, and the resulting oil was
purified by flash chromatography (30% hexane-70% EtOAc) to
[2-Oxo-2-{1-[3,4-bis-(benzoyloxy)-5-(benzoyloxymethyl)tet-
rahydrofuran-2-yl]-2-oxo-2,3-dihydro-1H-pyrimidin-4-ylidene}-
2-(bis-(benzyloxy)phosphoryl)acetic Acid Benzyl Ester (7). NaH
(75 mg, 1.86 mmol, 2.1 equiv, 60% dispersion in oil) was added
to a solution of phosphonate 615 (0.50 g, 1.93 mmol, 2.18 equiv)
in dry THF (10 mL) at 0 °C (ice bath) under Ar. The ice bath was
removed, and the mixture was stirred for 1 h. The mixture was
cooled using a KCl ice bath (-10 °C), and a solution of nucleoside
5 (0.728 g, 0.886 mmol, 1 equiv) in dry THF (10 mL) was added.
The mixture was stirred for 30 min; the ice bath was removed, and
stirring was continued for an additional 4 h. The reaction was diluted
with 200 mL of Et2O and was washed with 10% NH4Cl and
saturated brine. The organic layer was dried (MgSO4) and concen-
trated. Purification of the residue by flash chromatography (60%
hexane-40% EtOAc) gave pure 7 as a white foam (0.619 g, 88%
1
give pure 10 as a colorless oil (2.1 g, 83% yield). H NMR (300
MHz) δ 10.39 (s, 1H), 8.10 (d, J ) 7.8 Hz, 2H), 7.93 (d, J ) 7.8
Hz, 4H), 7.45-7.63 (m, 5H), 7.30-7.40 (m, 4H), 6.67 (d, J ) 8.0
Hz, 1H), 6.34 (d, J ) 6.5 Hz, 1H), 5.80-5.87 (m, 1H), 5.65
(overlapping dd, J ) 6.4 Hz, 1H), 5.43 (d, J ) 8.3 Hz, 1H), 4.70-
4.84 (m, 1H), 4.50-4.65 (m, 2H), 3.60-3.69 (m, 6H); 13C NMR
(75 MHz) δ 165.9, 165.2, 151.8 (d, J ) 4.3 Hz), 148.1, 133.6,
133.6, 133.4, 130.7, 129.8, 129.7, 129.5, 129.2, 128.6, 128.4, 104.3
(d, J ) 12.9 Hz), 86.9, 79.8, 74.6 (d, J ) 196 Hz), 72.9, 71.1,
63.9, 51.0 (d, J ) 4.3 Hz); 31P NMR (121 MHz) δ 27.6; LREIMS
m/z (relative intensity) 662 (M, 5), 445 (100), 201 (45), 105 (73).
HREIMS m/z: calcd for C33H31O11N2P, 662.1665; found, 662.1668.
General Procedure for the Deprotection of Nucleosides 1-4.
To a solution of the protected nucleoside in dry CH2Cl2 (≈0.8
mmols of nucleoside/mL of solvent) was added TMSBr (3 equiv
for compounds 1-3, 8 equiv added over 3 days for compound 4),
and the reaction was stirred for 20 h for compounds 1-3 or for 3
days for compound 4. The reaction was concentrated. Dry CH2Cl2
(5 mL) was added, and the solution was concentrated by rotary
evaporation. This was repeated. The residue was subjected to high
vacuum for 2 h. A solution of aq concd ammonium hydroxide-
methanol (3:2, 5 mL) was added, and the mixture was stirred for
24 h. The reaction was concentrated, and the residue was dissolved
in water and lyophilized. Lyophilization was repeated to give a
white powder. Purification was achieved using preparative RP-
HPLC (C-18 column). For compounds 1 and 2, triethylammonium
acetate (pH 8.8)-acetonitrile was used as the eluent. After removal
of the solvent and after lyophilization from water (five times),
compounds 1 and 2 were obtained as their triethylammonium salts.
The triethylammonium salts are hygroscopic. Therefore, they were
passed through a small Dowex 50 Na+ ion exchange column which
gave compounds 1 and 2 as their sodium salts and as easy-to-handle
white powders after lyophilization. For compounds 3 and 4, 0.1%
TFA in water-acetonitrile was used as the eluent which gave
compounds 3 and 4 as white powders after removal of the eluent
and after lyophilization.
1
yield). H NMR (300 MHz) δ 8.08 (d, J ) 7.3 Hz, 2H), 7.90-
7.96 (m, 4H), 7.70 (d, J ) 8.7 Hz, 0.5H), 7.22-7.59 (m, 14.5H),
7.15 (d, J ) 8.3 Hz, 0.5H), 7.12 (d, J ) 9.3 Hz, 0.5H), 6.29 (d, J
) 5.8 Hz, 0.5H), 6.26 (d, J ) 5.4 Hz, 0.5H), 5.84-5.88 (m, 1H),
5.69-5.72 (m, 1H), 5.22 (s, 1H), 5.17 (s, 1H), 4.64-4.70 (m, 3H),
3.59-3.66 (m, 6H); 13C NMR (75 MHz) δ 169.2 (d, J ) 8.8 Hz),
165.9 (CdO), 165.8 (d, J ) 7.6 Hz), 165.2, 165.1, 160.1 (d, J )
8.7 Hz), 160.2 (d, J ) 23.0 Hz), 147.4, 147.3 (d, J ) 1.9 Hz),
136.3, 135.8, 135.8, 135.76, 133.7, 133.6, 133.5, 129.8, 129.7,
129.6, 129.2, 129.1, 128.6, 128.5, 128.4, 128.3, 127.9, 127.8, 127.7,
127.6, 102.4, 102.0 (d, J ) 15.2 Hz), 88.4, 88.1, 81.3 (d, J ) 205
Hz), 80.1 (d, J ) 188 Hz), 73.6, 71.1, 66.2, 65.8, 63.7, 63.7, 52.7
(d, J ) 5.4 Hz), 52.1 (d, J ) 5.4 Hz); 31P NMR (121 MHz) δ 27.3,
23.3; LR+ESIMS m/z (relative intensity) 797 (M + 1, 100).
HREIMS m/z: calcd for C41H38N2O13P, 797.2112; found, 797.2094.
2-{1-[3,4-Bis-(benzoyloxy)-5-(benzoyloxymethyl)tetrahydro-
furan-2-yl]-2-oxo-2,3-dihydro-1H-pyrimidin-4-ylidene}-2-(bis-
(tert-butyloxy)phosphoryl)acetic Acid Benzyl Ester (9). NaH (398
mg, 9.94 mmol, 2.1 equiv, 60% dispersion in oil) was added to a
solution of phosphonate 816 (2.32 g, 4.73 mmol, 2.2 equiv) in dry
THF (60 mL) at 0 °C (ice bath) under Ar. The ice bath was
removed, and the mixture was stirred for 1 h. The mixture was
cooled using a KCl ice bath (-10 °C), and a solution of nucleoside
5 (3.9 g, 4.73 mmol, 1 equiv) in dry THF (60 mL) was added. The
mixture was stirred for 30 min; the ice bath was removed, and
stirring was continued for an additional 7 h. The reaction was diluted
with 200 mL of saturated NH4Cl and transferred to a separatory
funnel containing 600 mL of ether. The layers were separated, and
J. Org. Chem, Vol. 71, No. 25, 2006 9427

Taylor et al.
{1-[3,4-Bis-(hydroxy)-5-hydroxymethyltetrahydrofuran-2-yl]-
2-oxo-1,2-dihydropyrimidin-4-ylmethyl)phosphonic Acid Bis-
sodium Salt (1). Compound 1 was prepared from 100 mg (0.150
mmol) of 10 using the general procedure described above. After
RP-HPLC (99% triethylammonium acetate, 1% CH3CN (pH 8.8),
isocratic, tR ) 7.7 min) and Na+ ion exchange chromatography,
2-Benzyloxyisonicotinic Acid Benzyl Ester (21). A suspension
of 2-chloroisonicotinic acid (9.00 g, 57.0 mmol), pyridine (0.227
g, 2.87 mmol), and thionyl chloride (13.6 g, 114 mmol, 2 equiv)
in dry toluene (60 mL) was heated to reflux for 2 h during which
the solution became homogeneous. The reaction was concentrated
by high vacuum rotary evaporation. The residue was dissolved in
dry toluene (30 mL) and cooled to 0 °C (ice bath). Dry pyridine
(13.8 mL) was added, which was followed by benzyl alcohol (9.3
g, 86 mmol), and the mixture was stirred at room temperature for
3 h. Water was added (34 mL), which was followed by concentrated
HCl (13.5 g). The layers were separated, and the aq layer was
extracted with toluene (60 mL). The organic layers were combined,
washed with H2O and saturated brine, then dried (MgSO4), and
concentrated to give a yellow oil; this was dried under high vacuum
for several hours. 1H NMR of the residue indicated that this
consisted of the benzyl ester of 2-chloroisonicotinic acid and a small
amount of benzyl alcohol (10%). To a solution of benzyl alcohol
(6.46 mL, 59.7 mmol) in dry DMF (24 mL) at 0 °C (ice bath) was
added NaH (2.39 g, 59.7 mmol). The ice bath was removed; the
mixture was stirred at room temperature for 1 h and then cooled to
0 °C. To this was added a solution of the crude benzyl ester in dry
DMF (24 mL), and the mixture was stirred for 2 h. Glacial acetic
acid (710 mg) was added, and the mixture was stirred for 30 min.
Toluene (150 mL) was added and was followed by a solution of
saturated NaHCO3 (75 mL). The layers were separated, and the
organic layer was washed with water and saturated brine, dried
(MgSO4), and concentrated. The residue was subjected to flash
chromatography (80% hexane-20% EtOAc) to give compound 21
1
41.0 mg of a mixture of compound 1 (≈95% of the total by H
NMR integration of H-5) and dephosphonylated compound 11
1
1
(≈5% of the total by H NMR of H-5) was obtained. H NMR
(D2O, 300 MHz) δ 8.18 (d, J ) 6.9 Hz, 1H), 6.60 (d, J ) 6.7 Hz,
1H), 6.50 (d, J ) 7.5 Hz), 5.76 (s, 1H), 4.16 (s), 4.03 (m), 4.02 (d,
J ) 4.4 Hz), 3.70 (dd, J ) 12.9, 3.9 Hz), 2.92 (m, exchange with
solvent deuterons), 2.62 (bs, exchange with solvent deuterons); 31
P
NMR (D2O, 121 MHz) δ 15.8, 1.7; LR-ESIMS m/z (relative
intensity) 321 (M - 2Na + 1, 100); LR+ESIMS m/z (relative
intensity) 323 (M - 2Na + 3, 100), 243 (66), 191 (65). HR-ESIMS
m/z: calcd for C10H14N2O8P (compound 1 - 2Na + 1), 321.0488;
found, 321.0497. HR+ESIMS m/z: calcd for C10H15N2O5 (com-
pound 11 + 1), 243.0981; found, 243.0986.
[Difluoro-{1-[3,4-bis-(benzoyloxy)-5-benzoyloxymethyl-tet-
rahydrofuran-2-yl]-2-oxo-1,2-dihydro-pyrimidin-4-yl}-methyl]-
phosphonic Acid Dimethyl Ester (13). To a solution of compound
10 (1.00 g, 1.51 mmol, 1 equiv) and NFSi (1.18 g, 3.78 mmol, 2.5
equiv) in dry THF (50 mL) cooled to 0 °C (ice bath) and under Ar
was added dropwise a 0.5 M solution of KHMDS in toluene (6.64
mL, 3.32 mmol, 2.2 equiv) over a period of 70 min. The ice bath
was removed, and stirring was continued further for 2 h at room
temperature. NFSi (0.476 g, 1 equiv) was added, and the reaction
was cooled to 0 °C. A 0.5 M solution of KHMDS in toluene (2.0
mL, 1 mmol, 0.66 equiv) was added over a period of 20 min. The
reaction was stirred at 0 °C for 30 min; the ice bath was removed,
and the reaction was stirred for an additional 1 h. NH4Cl (10%, 2
mL) was added, the reaction was filtered through Celite, and the
Celite was washed with 15 mL of THF. The filtrate was concen-
trated to ≈15 mL, diluted with CH2Cl2 (50 mL), and filtered again
though filter paper. The filtrate was concentrated to ≈15 mL, diluted
with toluene (50 mL), filtered through filter paper, and concentrated.
The residue was subjected to flash chromatography (40% hexane-
60% EtOAc) which gave pure 13 as a colorless oil (0.478 g, 45%
1
as a pale yellow solid (14.7 g, 81%). Mp 42-43 °C; H NMR
(300 MHz) δ 8.26 (d, J ) 5.5 Hz, 1H), 7.22-7.45 (m, 12H), 5.39
(s, 2H), 5.34 (s, 2H); 13C NMR (75 MHz) δ 164.7, 164.2, 147.66,
140.2, 136.9, 135.3, 128.6, 128.4, 128.3, 127.9, 116.0, 111.6, 68.0,
67.3; LREIMS m/z (relative intensity) 319 (M, 100), 213 (35), 91
(65). HREIMS m/z: calcd for C20H17NO3, 319.1208; found,
319.1203.
(2-Benzyloxy-pyridin-4-yl)-methanol (22). A mixture of com-
pound 21 (3.45 g, 10.8 mmol), LiBH4 (0.352 g, 16.2 mmol), and
dry MeOH (0.519 g, 16.2 mmol) in dry ether (40 mL) was refluxed
for 1 h. A 0.3 N HCl solution was added until the reaction was
slightly acidic. The mixture was extracted with CH2Cl2 (3 × 60
mL), and the combined organics were dried (MgSO4) and concen-
trated. The residue was subjected to flash chromatography (80%
hexane-20% EtOAc to 70% hexane-30% EtOAc) to give pure
22 as a colorless liquid (2.0 g, 86%). 1H NMR (300 MHz) δ 8.10
(d, J ) 4.8 Hz, 1H), 7.20-7.45 (m, 5H), 6.84 (d, J ) 4.8 Hz, 1H),
6.80 (s), 5.35 (s, 2H), 4.65 (s, 2H), 2.16 (s, 1H); 13C NMR (75
MHz) δ 163.9, 153.6, 146.5, 137.0, 128.3, 127.8, 114.8, 108.0,
67.7, 63.0; LREIMS m/z (relative intensity) 215 (M, 100), 138 (14),
109 (47), 91 (89). HREIMS m/z: calcd for C13H13NO2, 215.0946;
found, 215.0944.
1
yield). H NMR (300 MHz) δ 8.18 (d, J ) 6.9 Hz, 1H), 8.02 (d,
J ) 7.2 Hz, 2H), 7.92 (d, J ) 7.4 Hz, 1H), 7.87 (d, J ) 7.4 Hz,
2H), 7.20-7.58 (m, 9H), 6.53 (d, J ) 7.1 Hz, 1H), 6.32 (s, 1H),
5.80-5.60 (m, 2H), 4.65-4.87 (m, 3H), 3.92 (d, J ) 4.7 Hz, 6H);
13C NMR (75 MHz) δ 168.4 (dt, J ) 25.0, 15.3 Hz), 165.9, 165.1,
165.0, 153.7, 145.8, 133.6, 133.6, 129.8, 129.7, 129.5, 129.1, 128.6,
128.4, 114.7 (dt, J ) 269, 227 Hz), 100.8, 90.8, 80.8, 74.8, 70.5,
63.2, 55.5 (d, J ) 6.5 Hz); 31P NMR (121 MHz) δ 7.5 (t, J ) 100
Hz); 19F NMR (282 MHz) δ -112.3 (dd, J ) 312, 100 Hz), -113.5
(dd, J ) 312, 100 Hz); LR+ESIMS m/z (relative intensity) 699 (M
+ 1, 100), 445 (37), 371 (42). HR+ESIMS m/z: calcd for
C33H30N2O11F2P, 699.1555; found, 699.1572.
2-Benzyloxy-4-bromomethylpyridine (23). To a solution of
POBr3 (4.89 g, 17.1 mmol, 1.2 equiv) in dry CH2Cl2 (40 mL) at 0
°C was added dropwise dry DMF (19 mL) over a period of 30
min. A solution of compound 22 (3.0 g, 14 mmol) in dry CH2Cl2
(20 mL) was added dropwise over a period of 30 min and stirred
for 1 h. The mixture was transferred via canula to a cold solution
of saturated NaHCO3 (100 mL) keeping the temperature below 10
°C. The layers were separated, and the aq layer was extracted with
CH2Cl2 (3 × 60 mL). The combined organics were washed with
water and saturated brine, dried (MgSO4), and concentrated.
Purification of the residue by flash chromatography (95% hexane-
5% EtOAc) gave pure 23 as a white solid (2.97 g, 77%). Mp 29-
[Difluoro-{1-[3,4-bis-(hydroxy)-5-hydroxymethyltetrahydro-
furan-2-yl]-2-oxo-1,2-dihydro-pyrimidin-4-yl}-methyl]phospho-
nic Acid Bis-sodium Salt (2). Compound 2 was prepared from 52
mg (0.0745 mmol) of 13 using the general procedure described
above. After RP-HPLC (98% triethylammonium acetate, 2% CH3-
CN (pH 8.8), isocratic, tR ) 9.2 min) and Dowex Na+ ion exchange
chromatography, compound 2 was obtained as a white powder (15.1
mg, 51% yield). 1H NMR (D2O, 500 MHz) δ 8.49 (d, J ) 6.9 Hz,
1H), 6.91 (d, J ) 6.9 Hz, 1H), 5.86 (s, 1H), 4.27 (d, J ) 4.7 Hz,
1H), 4.05-4.17 (m, 2H), 3.96 (d, J ) 12.8 Hz, 1H), 3.79 (dd, J )
12.8, 4.1 Hz, 1H); 13C NMR (D2O, 125 MHz) δ 171.9 (dt, J )
22.9, 12.6 Hz), 156.3, 145.5, 118.4 (dt, J ) 265.7, 166.7 Hz), 104.1,
92.2, 83.6, 74.5, 68.2, 60.0; 31P NMR (D2O, 121 MHz) δ 4.79 (t,
J ) 81 Hz); 19F NMR (D2O, 282 MHz) δ -114.2 (dd, J ) 287, 81
Hz), -115.4 (dd, J ) 287, 81 Hz); LR-ESIMS m/z (relative
intensity) 357 (M - 2Na + 1, 100), 224 (44); HREIMS m/z: calcd
for C10H12F2N2O8P, 357.0299; found, 357.0297.
1
30 °C; H NMR (300 MHz) 8.13 (d, J ) 4.5 Hz, 1H), 7.20-7.47
(m, 5H), 6.88 (d, J ) 4.5 Hz, 1H), 6.80 (s, 1H), 5.37 (s, 2H), 4.32
(s, 2H); 13C NMR (75 MHz) 164.0, 148.78, 147.3, 137.1, 128.4,
127.9, 127.8, 117.1, 110.9, 67.7, 30.5; LREIMS m/z (relative
intensity) 277 (M, 55), 198 (100), 91 (54); HREIMS m/z: calcd
for C13H12ONBr, 277.0102; found, 277.0107.
9428 J. Org. Chem., Vol. 71, No. 25, 2006

Uridine- and 3-Deazauridine-4-phosphate Analogues
2-Hydroxy-isonicotinic Acid (25). To 2-chloroisonicotinic acid
(5.00 g, 31.7 mmol) was added 90 mL of concd HCl. The
suspension was heated to 140-150 °C (oil bath). The reaction
initially became a solution upon heating, though after heating for
24 h again it became a suspension. Another 20 mL of concd HCl
was added, the reaction became clear, heating was continued for
another 24 h, after which an additional 20 mL of concd HCl was
added, and the reaction was heated for 12 h. The reaction was
cooled in an ice bath during which compound 25 precipitated out
of solution. The mixture was diluted with an equal volume of ice-
cold water and filtered, and the filter cake was washed with cold
water. The precipitate was dried under high vacuum to give pure
25 as a white solid (3.92 g, 89%). NMR spectra were identical to
those reported.30 1H NMR (DMSO-d6, 300 MHz) δ 12.67 (bs, 2H),
7.43 (d, J ) 6.6 Hz, 1H), 6.77 (s, 1H), 6.47 (d, J ) 6.5 Hz, 1H);
13C NMR (DMSO-d6, 75 MHz) δ 166.2, 162.9, 143.0, 136.7, 121.4,
104.0.
LREIMS m/z (relative intensity) 295 (M, 73), 217 (100), 121 (21).
HREIMS m/z: calcd for C9H14NO6PS, 295.0279; found, 295.0285.
(2-Hydroxypyridin-4-ylmethyl)phosphonic Acid Dimethyl
Ester (16). To a solution of phosphonate 28 (1.36 g, 4.61 mmol)
in THF (35 mL) at 0 °C (ice bath) was added an ice cold solution
of LiOH hydrate (386 mg, 9.22 mmol) in H2O (35 mL) over a
period of 10 min. After the addition, the solution was stirred for 5
min, and then the ice bath was removed and stirred for 4.5 h at
room temperature. The reaction was neutralized with 0.3 N HCl
and concentrated by high vacuum rotary evaporation, and the
residue was purified by flash chromatography (20% MeOH-80%
CHCl3) to give compound 16 as a white solid (0.758 g, 76%). Mp
132-134 °C; 1H NMR (CD3OD, 300 MHz) δ 7.36 (d, J ) 6.7 Hz,
1H), 6.47 (s, 1H), 6.38 (d, J ) 6.7 Hz, 1H), 3.74 (d, J ) 11.1 Hz,
6H), 3.21 (d, J ) 22.8 Hz, 2H); 13C NMR (CD3OD, 75 MHz) δ
164.0, 148.2 (d, J ) 9.1 Hz), 134.3, 119.8 (d, J ) 8.3 Hz), 109.4
(d, J ) 4.0 Hz), 52.6 (d, J ) 7.1 Hz), 31.2 (d, J ) 136.1 Hz); 31
P
NMR (CD3OD, 121 MHz) δ 29.4; LREIMS m/z (relative intensity)
217 (M, 100), 185 (15), 121 (44), 109 (37). HREIMS m/z: calcd
for C8H12NO4P, 217.0504; found, 217.0497.
4-Hydroxymethyl-pyridin-2-ol (26). To a solution of compound
25 (4.80 g, 34.5 mmol) in dry THF (170 mL) at 0 °C (ice bath)
was added a 1 M solution of BH3-THF (150 mL, 150 mmol). The
ice bath was removed, and the reaction was stirred for 90 min.
The reaction was cooled with an ice bath, and MeOH (75 mL)
was added slowly. The reaction was concentrated to dryness. MeOH
(100 mL) was added to the solid residue, stirred for 15 min, and
then concentrated to dryness. The residue was suspended in 20%
MeOH-80% CHCl3, stirred for 5 min, and then filtered. The
precipitate was subjected to this again; the two filtrates were
combined and concentrated to give crude 26 as a pale yellow solid.
Recrystallization in absolute ethanol gave pure 26 as a white solid
(1.52 g). The filtrate from the recrystallization was concentrated,
and the residue was purified by flash chromatography (20% EtOH-
80% EtOAc and then 20% MeOH-80% CHCl3) to get an additional
{1-[3,4-Bis-(benzoyloxy)-5-benzoyloxymethyltetrahydro-furan-
2-yl]-2-oxo-1,2-dihydro-pyridin-4-ylmethyl}phosphonic Acid Dim-
ethyl Ester (29). Phosphonate 16 (0.300 g, 1.37 mmol, 1 equiv)
and acetyl 2,3,5-tri-O-benzoyl-â-D-ribofuranose (0.691 g, 1.37
mmol) were suspended in dry CH3CN (6 mL) under Ar at 0 °C
(ice bath). To this was added BTMSTFA (0.40 mL, 1.5 mmol, 1.1
equiv); the ice bath was removed, and the solution was stirred for
1.5 h. TMSOTf (0.247 mL, 1.37 mmol, 1 equiv) was added, and
the solution was stirred for 3 h. The reaction was diluted with
EtOAc (200 mL) and ether (100 mL) and washed with saturated
NaHCO3- and saturated brine. The organic layer was dried (MgSO4)
and concentrated, and the resulting residue was subjected to flash
chromatography (5% EtOH-95% EtOAc) to give pure compound
29 as a white foam (0.59 g, 65%). 1H NMR (300 MHz) δ 8.09 (d,
J ) 7.6, 2H), 7.46 (d, J ) 8.0 Hz, 2H), 7.90 (d, J ) 7.3 Hz, 2H),
7.30-7.63 (m, 9H), 6.56 (d, J ) 3.9 Hz), 6.40 (s, 1H, 3H), 6.10
(d, J ) 7.2 Hz, 1H), 5.89 (overlapping dd, J ) 5.8 Hz), 5.78
(overlapping dd, J ) 5.1 Hz, 1H), 4.60-4.90 (m, 3H), 3.72 (d, J
) 11.2 Hz, 6H), 2.93 (d, J ) 22.5 Hz, 2H); 13C NMR (75 MHz)
δ 166.0, 165.2, 165.0, 161.5 (d, J ) 2.2 Hz), 145.2 (d, J ) 8.6
Hz), 133.5, 133.4, 132.1, 129.8, 129.7, 129.6, 129.3, 128.6, 128.5,
128.3, 121.1 (d, J ) 9.9 Hz), 108.4 (d, J ) 4.2 Hz), 88.3, 80.0,
74.8, 70.8, 63.6, 53.0 (d, J ) 7.1 Hz), 52.9 (d, J ) 6.3 Hz), 32.5
(d, J ) 137.5 Hz); 31P NMR (121 MHz) δ 27.6; LREIMS m/z
(relative intensity) 661 (M, 1), 539 (24), 445 (28), 282 (79), 105
(100). HREIMS m/z: calcd for C34H32NO11P, 661.1717; found,
661.1707.
1
0.7 g of pure 26 (total yield: 2.22 g, 54%). Mp 144-145 °C; H
NMR (CD3OD, 300 MHz) δ 7.40 (d, J ) 6.8 Hz, 1H), 6.60 (s,
1H), 6.40 (d, J ) 6.8 Hz, 1H), 4.54 (s, 2H); 13C NMR (CD3OD,
75 MHz) δ 164.6, 158.2, 133.8, 114.3, 105.5, 61.8; LREIMS m/z
(relative intensity) 125 (M, 100), 96 (24), 80 (22). HREIMS m/z:
calcd for C6H7NO2, 125.0477; found, 125.0478.
Methanesulfonic Acid 4-Bromomethyl-pyridin-2-yl Ester (27).
To a solution of compound 26 (0.125 g, 1.00 mmol) and 2,6-lutidine
(0.670 g, 2.50 mmol, 2.50 equiv) in dry CH2Cl2 (4 mL) was added
methanesulfonylbromide (0.397 g, 2.5 mmol, 2.5 equiv). The
reaction was stirred for 48 h, diluted with CH2Cl2 (40 mL), and
washed with 0.3 N HCl and H2O. The organic layer was dried
(MgSO4) and concentrated, and the resulting residue was purified
by flash chromatography (70% hexane-30% EtOAc) to give pure
1
27 as a colorless oil (187 mg, 71%). H NMR (300 MHz) δ 8.26
{1-[3,4-Bis-(hydroxy)-5-hydroxymethyltetrahydrofuran-2-yl]-
2-oxo-1,2-dihydropyridin-4-ylmethyl}phosphonic Acid (3). Com-
pound 3 was prepared from 115 mg (0.242 mmol) of 29 using the
general procedure. After RP-HPLC (90% TFA (0.1%) in water-
1% CH3CN, isocratic, tR ) 9.0 min), compound 3 was obtained as
(d, J ) 5.0 Hz, 1H), 7.25 (d, J ) 5.0 Hz, 1H), 7.07 (s, 1H), 4.36
(s, 1H), 345 (s, 1H); 13C NMR (75 MHz) δ 157.6, 151.1, 148.3,
122.7, 115.1, 40.7, 29.2; LREIMS m/z (relative intensity) 267 (M
(81Br), 40), 265 (M (79Br), 40), 187 (100), 108 (35), 80 (38);
HREIMS m/z: calcd for C7H8BrNO3S, 264.9408; found, 264.9408.
1
a white powder (69.8 mg, 90% yield). H NMR (D2O, 300 MHz)
Methanesulfonic Acid 4-(Dimethoxyphosphorylmethyl)py-
ridin-2-yl Ester (28). A solution of compound 27 (1.38 g, 5.20
mmol) in P(OMe)3 (20 mL) was heated at 100 °C for 2.5 h. The
reaction was concentrated by high vacuum rotary evaporation, and
the resulting residue was purified by flash chromatography (5%
MeOH-95% CHCl3) to give pure 28 as a pale yellow oil which
solidified after we subjected the oil to high vacuum overnight (1.43
δ 7.70 (d, J ) 6.7 Hz, 1H), 6.36 (d, J ) 6.7 Hz, 1H), 6.35 (s, 1H),
5.88 (s, 1H), 4.06 (s, 1H), 3.97 (s, 2H), 3.76 (d, J ) 12.7 Hz, 1H),
3.63 (d, J ) 12.7 Hz), 2.90 (d, J ) 21.9 Hz, 2H); 13C NMR (D2O,
125 MHz) δ 163.1, 150.1 (d, J ) 7.2 Hz), 132.7, 118.4 (d, J ) 6.5
Hz), 111.5, 90.2, 83.7, 74.8, 68.9, 60.4, 34.8 (d, J ) 28.5 Hz); 31
P
NMR (D2O, 121 MHz) δ 23.0; LR-ESIMS m/z (relative intensity)
320 (M - 1, 100). HR-ESIMS m/z: calcd for C11H15NO8P,
320.0535; found, 320.0540.
1
g, 93%). Mp 80-81 °C; H NMR (300 MHz) δ 8.15 (d J ) 4.9
Hz, 1H), 7.14 (bs, 1H), 6.94 (s, 1H), 3.61 (d, J ) 11.4 Hz, 6H),
3.38 (s, 3H), 3.10 (d, J ) 22.5 Hz, 2H); 13C NMR (75 MHz) δ
157.4 (d, J ) 2.7 Hz), 147.7 (d, J ) 2.5 Hz), 146.0 (d, J ) 8.6
Hz), 123.9 (d, J ) 5.6 Hz), 116.2 (d, J ) 6.2 Hz), 52.9 (d, J ) 6.9
Hz), 40.4, 32.0 (d, J ) 137 Hz); 31P NMR (121 MHz) δ 27.2;
(2-Benzyloxypyridin-4-ylmethyl)phosphonic Acid Diethyl Es-
ter (31). To a solution of bromide 23 (10.7 mmol) in dry benzene
(130 mL) was added P(OEt)3 (18.4 mL, 107 mmol), and the mixture
was heated to 95 °C (oil bath) for 16 h. P(OEt)3 (9.20 mL, 53.0
mmol) was added, the mixture was heated to 105 °C, and the
benzene was slowly distilled off. When approximately 85 mL of
benzene was removed, the temperature was lowered to 95 °C, and
heating continued for another 2 h. The benzene was removed by
(30) Rollet, F.; Richard, C.; Pilichowski, J.-F.; Aboab, B. Org. Biomol.
Chem. 2004, 2, 2253.
J. Org. Chem, Vol. 71, No. 25, 2006 9429

Taylor et al.
rotary evaporator, and excess phosphate and diethyl ethylphospho-
nate were removed using a high vacuum rotary evaporator.
Purification of the residue by flash chromatography (20% hexane-
80% EtOAc to 10% hexane-90% EtOAc) gave pure 31 as a
Diethyl Ester (34). To a solution of phosphonate 33 (0.10 g, 0.36
mmol) and acetyl 2,3,5-tri-O-acetyl-â-D-ribofuranose (0.113 g, 0.36
mmol) in dry CH3CN (2 mL) was added BTMSTFA (0.113 mL,
0.42 mmol), and it was stirred at rt for 2 h. To this was added
TMSOTf (0.077 mL, 0.42 mmol), and that was stirred for 3 h. The
reaction was diluted with EtOAc (40 mL), washed with saturated
NaHCO3 and brine, dried (MgSO4), and concentrated. Purification
of the residue by flash chromatography (20% hexane-80% EtOAc)
gave pure 34 as a white foam (0.15 g, 78%). 1H NMR (300 MHz)
δ 7.57 (d, J ) 7.4 Hz, 1H), 6.65 (s, 1H), 6.36 (d, J ) 7.4 Hz, 1H),
6.17 (d, J ) 4.1 Hz, 1H), 5.20-5.35 (m, 2H), 4.10-4.38 (m, 6H),
2.05 (s, 3H), 2.30 (s, 3H), 2.00 (s, 3H), 1.3 (d, J ) 7.3 Hz, 6H);
13C NMR (75 MHz) δ 170.0, 169.3, 169.1, 160.9, 144.5 (dt, J )
22.6, 14.1 Hz), 132.8, 118.9 (dt, J ) 5.6, 3.2 Hz), 115.9 (dt, J )
263.9, 214.4 Hz), 103.0 (t, J ) 5.3 Hz), 88.1, 79.5, 73.8, 69.4,
65.1 (d, J ) 6.8 Hz), 62.5, 20, 20.3, 16.1 (d, J ) 5.4 Hz); 31P
NMR (121 MHz) δ 6.1 (t, J ) 108 Hz); 19F NMR (282 MHz) δ
-112.7 (d, J ) 108 Hz); LREIMS m/z (relative intensity) 539 (M,
10), 321 (45), 259 (100), 139 (73). HREIMS m/z: calcd for
C21H28F2NO11P, 539.1368; found, 539.1371.
[Difluoro-{1-[3,4-bis-(hydroxy)-5-hydroxymethyltetrahydro-
furan-2-yl]-2-oxo-1,2-dihydro-pyridin-4-yl}-methyl]phosphon-
ic Acid (4). Compound 4 was prepared from 88 mg (0.164 mmol)
of 34 using the general procedure. However, the TMSBr was added
in portions starting with 4 equiv of TMSBr at the beginning of the
reaction. After 24 h, another 2 equiv of TMSBr was added followed
by an additional 2 equiv after 48 h. After RP-HPLC (99% TFA
(0.1%) in water-1% CH3CN, isocratic, tR ) 9.4 min) and
lyophilization, compound 4 was obtained as a white powder (26.3
mg, 45% yield). 1H NMR (D2O, 300 MHz) δ 7.88 (d, J ) 7.4 Hz,
1H), 6.62 (s, 1H), 6.54 (d, J ) 7.4 Hz, 1H), 5.96 (s, 1H), 4.12 (s,
1H), 4.03 (s, 2H), 3.83 (d, J ) 12.8 Hz, 1H), 3.69 (d, J ) 12.8 Hz,
1H); 13C NMR (D2O, 125 MHz) δ 163.2, 148.3 (dt, J ) 22.8, 12.0
Hz), 133.6, 117.9 (dt, J ) 163.0, 71.4 Hz), 116.7 (t, J ) 7.4 Hz),
105.6 (t, J ) 5.3 Hz), 90.5, 83.7, 74.8, 68.8, 60.4; 31P NMR (D2O,
121 MHz) δ 4.0 (t, J ) 96.0 Hz); 19F NMR (D2O, 282 MHz) δ
-113.0 (d, J ) 97.0 Hz); LR-ESIMS m/z (relative intensity) 356
(M - 1, 100), 224 (20). HR-EIMS m/z: calcd for C11H13F2NO8P,
356.0347; found, 356.0359.
1
colorless oil (3.34 g, 93%). H NMR (300 MHz) δ 8.01 (d, J )
4.9 Hz), 7.20-7.40 (m, 5H), 6.79 (bs, 1H), 6.67 (s, 1H), 5.29 (s,
2H), 3.90-4.05 (m), 3.00 (d, J ) 21.8 Hz), 1.18 (t, J ) 7.9 Hz,
6H); 13C NMR (75 MHz) δ 163.8 (d, J ) 2.2 Hz), 146.7 (d, J )
3.7 Hz), 143.7 (d, J ) 8.7 Hz), 137.1, 128.2, 127.7 (CHarom), 127.6
(CHarom), 118.4 (d, J ) 5.4 Hz), 112.0 (d, J ) 7.5 Hz), 67.4
(PhCH2), 62.2 (d, J ) 6.5 Hz), 33.2 (d, J ) 137.4 Hz), 16.2 (d, J
) 6.6 Hz); 31P NMR (121 MHz) δ 25.9; LREIMS m/z (relative
intensity) 335 (M, 100), 258 (28), 229 (25), 91 (51). HREIMS
m/z: calcd for C17H22NO4P, 335.1286; found, 335.1292.
[(2-Benzyloxypyridin-4-yl)difluoromethyl]phosphonic Acid
Diethyl Ester (32). To a solution of phosphonate 31 (3.10 g, 9.25
mmol) in dry THF (75 mL) was added NFSi (7.29 g, 23.1 mmol).
The solution was cooled to -78 °C, and KHMDS (40.7 mL of a
0.5 M solution in toluene, 20.3 mmol) was added dropwise over 2
min. The mixture was stirred for 30 min at -78 °C; the cold bath
was removed, and the mixture was stirred for 1 h. The reaction
was quenched with water (300 mL) and then extracted with CH2-
Cl2 (3 × 300 mL). The organics were dried (MgSO4) and
concentrated. Purification of the residue by flash chromatography
(30% hexane-70% EtOAc) gave pure 32 as a colorless oil (2.97
1
g, 87%). H NMR (300 MHz) δ 8.24 (d, J ) 5.2 Hz, 1H), 7.20-
7.48 (m, 5H), 7.08 (d, J ) 4.8 Hz), 7.01 (s, 1H), 5.38 (s, 2H),
4.10-4.30 (m, 4H), 1.30 (t, J ) 7.3 Hz, 6H); 13C NMR (75 MHz)
δ 163.7, 147.4, 143.8 (dt, J ) 22.5, 13.9 Hz), 136.8, 128.3, 127.8,
126.1, 116.6 (dt, J ) 264.3, 214.3 Hz), 113.6 (t, J ) 6.1 Hz), 108.7
(dt, J ) 2.2, 7.8 Hz), 67.9, 66.0 (d, J ) 6.3 Hz), 16.2 (d, J ) 5.4
Hz); 31P NMR (121 MHz) δ 6.8 (t, J ) 10.2 Hz); 19F NMR (282
MHz) δ -110.7 (d, J ) 90.3 Hz); LREIMS m/z (relative intensity)
371 (M, 100), 265 (20), 233 (19), 91 (59). HREIMS m/z: calcd
for C17H20F2NO4P, 371.1098; found, 371.1100.
[Difluoro-(2-hydroxy-pyridin-4-yl)-methyl]-phosphonic Acid
Diethyl Ester (33). To a solution of 32 (2.1 g, 5.7 mmol) in MeOH
(30 mL) was added 10% Pd/C (200 mg), and the reaction was stirred
for 3 h. The reaction was filtered through Celite, and the filtrate
was concentrated. Purification of the residue by flash chromatog-
raphy (5% MeOH-95% CHCl3) gave pure 33 as a white solid (1.58
1
Acknowledgment. This work was supported by a Discovery
Grant to S.D.T. and a NSERC Collaborative Health Research
Project Grant to S.L.B. and S.D.T.
g, 99%). Mp 74-76 °C; H NMR (300 MHz) δ 7.43 (d, J ) 6.9
Hz, 1H), 6.79 (s, 1H), 6.50 (d, J ) 6.9 Hz, 1H), 4.10-4.34 (m,
4H), 1.34 (t, J ) 7.4 Hz, 6H); 13C NMR (75 MHz) δ 164.5, 146.4
(dt, J ) 22.3, 13.8 Hz), 135.5, 118.2, 116.1 (dt, J ) 214.0, 264.2
Hz), 103.9 (t, J ) 5.0 Hz), 65.3 (d, J ) 6.8 Hz), 16.3 (d, J ) 5.0
Hz); 31P NMR (121 MHz) δ 6.3 (t, J ) 108.2 Hz); 19F NMR (282
MHz) δ 112.4 (d, J ) 108.2 Hz); LREIMS m/z (relative intensity)
281 (M, 90), 225 (90), 145 (100), 109 (64). HREIMS m/z: calcd
for C10H14F2NO4P, 281.0629; found, 281.0628.
Supporting Information Available: NMR spectra (1H, 13C, 31
P
(when applicable), and 19F spectra (when applicable)) for com-
pounds 1-5, 7, 9, 10, 13, 16, 21-23, 25-29, and 31-34. This
material is available free of charge via the Internet at http://
pubs.acs.org.
[Difluoro-{1-[3,4-bis-(acetoxy)-5-acetoxymethyltetrahydrofu-
ran-2-yl]-2-oxo-1,2-dihydro-pyridin-4-yl}-methyl]phosphonic Acid
JO0617666
9430 J. Org. Chem., Vol. 71, No. 25, 2006