2'-Amino-2'-deoxyuridine via an intramolecular cyclization of a trichloroacetimidate

Full text of DOI:10.1021/jo9510548

J . Org. Chem. 1996, 61, 781-785
781
Sch em e 1
2′-Am in o-2′-d eoxyu r id in e via a n
In tr a m olecu la r Cycliza tion of a
Tr ich lor oa cetim id a te
Danny P. C. McGee,* Alecia Vaughn-Settle,
Chandra Vargeese, and Yansheng Zhai
NeXstar, inc., 2860 Wilderness Place,
Boulder, Colorado 80301
Received J une 9, 1995
In tr od u ction
It has been demonstrated that incorporation of 2′-
amino-2′-deoxypyrimidines1 into oligoribonucleotides
(RNA) results in increased stability against chemical and
nuclease degradation. We are interested in evaluating
random pools of stabilized RNA utilizing an enrichment
strategy called SELEX2 against a number of therapeutic
and diagnostic targets and were thus interested in
preparing 2′-amino-2′-deoxyuridine (9a ). 2′-Aminouri-
dine 9a was first prepared3 in 1971 by lithium azide
opening of 2,2′-O-anhydrouridine 1 in approximately 50%
yield followed by catalytic reduction to the amine. To
this day all subsequent preparations of 9a have followed
this first report with minor variations,4 e.g., substitu-
tion of NH4Cl/NaN3 for LiN3. Although the LiN3 pro-
cedure is satisfactory, the cost and intermitant avail-
ability of LiN3 as well as the toxicity, instability, and
disposal of azide prompted us to seek an alternative
process.
Our initial approach (Scheme 1) was to make use of
the 3′-hydroxyl of anhydrouridine 1 to deliver intramo-
lecularly an appropriate amine nucleophile and thus
overcome the tendency of amines to attack at the 2-posi-
tion of the pyrimidine.5 The opening of a 2,2′-O-anhydro
nucleoside by a 3′-O-benzoate has been described whereby
3′,5′-di-O-benzoyl-2,2′-O-anhydro-L-uridine upon treat-
ment with boron trifluoride etherate afforded a mixture
of 3′,5′- and 2′,5′-dibenzoates of L-uridine in 80% yield.6
Circumstantial reports of this type of intramolecular
transformation have been implicated where a 3′-phos-
phate7 and a 3′-N-phenylcarbamate8 have opened the
2,2′-anhydrouridine linkage. The use of trichloroaceto-
nitrile for this purpose is not readily evident from the
chemical literature. Trichloroacetonitrile reacts readily
with hydroxyls under base catalysis to afford the corre-
sponding imidate which has been used for the activation
of the anomeric position of sugars.9 Additionally some
isolated examples of trichloroacetonitrile use include
opening of a 2,3-epoxy alcohol via acid-catalyzed activa-
tion of the preformed trichloroacetimidate resulting in
1,3 opening of the epoxide10 and the Claisen rearrange-
ment of allylic trichloroacetimidates.11
Methods for the synthesis of the 2′-aminopurines12 (A,
G) and the protected forms13 suitable for the chemical
synthesis of oligonucleotides by phosphoramidite chem-
istry have been described. Comparable methodology for
the synthesis of protected 2′-aminopyrimidines is either
implied or incomplete;1,14 all of which use 2′-azido-2′-
deoxyuridine3 as a starting point. With our novel ap-
proach to the synthesis of 2′-amino-2′-deoxyuridine, we
thought it would be necessary to outline the elaboration
of intermediate 4a to protected forms of 2′-aminouridine
8a and 2′-aminocytidine 13a suitable for use in the
chemical synthesis of oligonucleotides by the phosphora-
midite method.15
Resu lts a n d Discu ssion
Uridine, a relatively inexpensive starting material
(∼$500/kg) is easily converted to anhydrouridine16 1 by
reaction with diphenyl carbonate and sodium bicarbonate
in DMF. We report on an improved isolation of 1 from
the reaction mixture. This is achieved simply by con-
ducting the reaction in a minimal amount of DMF leading
to crystallization of the product and an easy filtration
and wash to give reproducibly high isolated yields (75-
85%). Compound 1 was easily dimethoxytritylated in the
usual way (DMT-Cl, DMAP, Pyr, DMF) to give the 5′-
(dimethoxytrityl)anhydrouridine 2. Observation of the
crude reaction mixture by HPLC showed a >90% conver-
sion to 2. Isolation of this compound by silica gel
(1) (a) Hobbs, J .; Sternbach, H.; Spinzl, M.; Eckstein, F. Biochemistry
1973, 12(25), 5138-5145. (b) Pieken, W. A.; Olsen, D. B.; Benseler,
F.; Aurup, H.; Eckstein, F. Science 1991, 253, 314-317. (c) Aurup, H.;
Tuschl, T.; Bensler, F.; Ludwig, J .; Eckstein, F. Nucleic Acids Res. 1994,
22(1), 20-24.
(2) Tuerk, C.; Gold, L. Science 1990, 249, 505-510.
(3) Verheyden, J . P. H.; Wagner, D.; Moffatt, J . G. J . Org. Chem.
1971, 36, 250-254.
(9) Schmidt, R. R. Angew. Chem., Int. Ed. Engl. 1986, 25, 212-235
and references therein.
(10) Schmidt, U.; Respondek. M.; Lieberknecht, A.; Werner, J .;
Fisher, P. Synthesis 1989, 256-261.
(11) Overman, L. E. Acc. Chem. Res. 1980, 13, 218-224. J . Am.
Chem. Soc. 1976, 98, 2901-2910.
(4) (a) Sasaki, T.; Minamoto, K.; Sugiura, T.; Masanao, N. J . Org.
Chem. 1976, 41, 3138-3143. (b) Kirschenheuter, G. P.; Zhai, Y.;
Pieken, W. A. Tetrahedron Lett. 1994, 35(46), 8517-8520.
(5) Moffatt, J . G. In Nucleoside Analoques; Walker, R. T., De Clercq,
E., Eckstein, F., Eds.; Plenum Press: New York, 1979; pp 71-164 and
references therein.
(6) (a) Holy, A. Collect. Czech. Chem. Commun. 1973, 38, 423-427.
(b) Holy, A. Collect. Czech. Chem. Commun. 1975, 40, 739-752.
(7) Ogilvie, K. K.; Iwacha, D. Can. J . Chem. 1970, 48, 862-864.
(8) Mikhailopulo, I. A.; Zaitseva, G. V.; Vaaks, E. V.; Balzarini, J .;
De Clercq, E.; Rosemeyer, H.; Seela, F. Liebigs Ann. Chem. 1993, 513-
519.
(12) (a) Hobbs, J .; Eckstein, F. J . Org. Chem. 1977, 42(4), 714-719.
(b) Imazawa, M.; Eckstein, F. J . Org. Chem. 1979, 44(12), 2039-2041.
(13) Benseler, F.; Williams, D. M.; Eckstein, F. Nucleosides Nucle-
otides 1992, 11(7), 1333-1351.
(14) (a) Miller, P. S.; Bhant, P.; Kan, L.-S. Nucleosides Nucleotides
1993, 12(8), 785-792. (b) Smith, L. M.; Fund, S.; Kaiser, R. J . U.S.
Patent 5,015,733, filed May 1991. (c) Cook, A. F.; Rao, K. V. B.
International Patent WO 93/23570; filed Nov 1993.
(15) Beaucage, S. L.; Iyer, R. P. Tetrahedron 1993, 49(28), 6123-
6194.
(16) Hampton, A.; Nichol, A. W. Biochemistry 1966, 5, 2076-2081.
Ogilvie, K. K.; Iwacha, D. Can. J . Chem. 1969, 47, 495-497.
0022-3263/96/1961-0781$12.00/0 © 1996 American Chemical Society

782 J . Org. Chem., Vol. 61, No. 2, 1996
Notes
Sch em e 2
readily be protected as the trifluoroacetamide 8a (ethyl
trifluoroacetate, acetonitrile, TEA, quantitative by TLC)
which for convenience is isolated in 56% yield by simple
filtration of the crystalline product from the reaction
mixture. Alternatively, the completely deprotected 2′-
aminouridine nucleoside 9a can be obtained in 80%
isolated yield by treatment of oxazoline 4a with 80%
acetic acid.
We next turned our attention to the uridine to cytidine
conversion starting with oxazoline 4a (Scheme 2) using
the method of Divakar and Reese.17 The method was
modified by filtration of the precipitated triethylamine
hydrochloride from the reaction, which when present
caused some detritylation to occur. The two-step syn-
thesis (POCl3, TEA, 1,2,4-triazole; then NH4OH, dioxane)
gave a clean conversion of 4a to oxazoline cytidine 4b
isolated in 74% yield. When we attempted to deprotect
the 2′,3′-positions of 4b with ethanolic sodium hydroxide,
we obtained approximately 40% of the desired aminocy-
tidine 7b and up to 25% of the aminouridine 7a , the
result of attack of hydroxide at the 4-position of the
pyrimidine. This problem was eliminated when cesium
carbonate was used as the base to afford 7b in 94% yield
with no appreciable formation of 7a (uridine) detected.
The resultant aminocytidine 7b when treated with ethyl
trifluoroacetate gave a 94% yield of 2′-N-trifluoroaceta-
mide 8b without reaction at the N4 of the pyrimidine.
N4-Benzoylation of trifluoroacetamido cytidine 8b with
benzoic anhydride in DMF according to the procedure of
Bhat et al.18 afforded the fully protected 2′-aminocytidine
nucleoside 13a in 66% yield. The major side product in
this reaction was the 3′-O,N4-bis-benzoate. With the
intention of designing the most efficient approach, we
also looked at a branch point to cytidine further along in
the cascade of intermediates depicted in Scheme 3,
namely 8a . The use of uridine 8a first required a
protection step for the 3′-hydroxyl. A one-pot conversion
of uridine 8a to cytidine 8b was evaluated by first
silylating the 3′-hydroxyl (2 equiv of TMS-Cl, TEA)
followed by addition of the phosphorus oxy tris-triazolide
reagent17 which had been filtered free of precipitated
triethylamine hydrochloride, followed by an extractive
workup and treatment of the residue with NH4OH in
dioxane to afford 8b in 80% yield for the three steps. Both
routes to 8b were impacted by the lower yield (66%) for
the benzoylation reaction described above. In an attempt
to improve this last step we investigated the use of the
tert-butyldimethylsilyl group for protection of the 3′-
hydroxyl of 8a for both the uridine to cytidine reaction
and the benzoylation. Compound 8a was first silylated
(TBDMS-Cl, TEA) and converted by the procedure de-
scribed above to the silylated cytidine 12b in 71% yield
(Scheme 4). Compound 12b was then benzoylated to give
13b in 93% isolated yield, followed by removal of the silyl
group using triethylamine hydrofluoride to give the fully
protected cytidine 13a in 95% yield.
Sch em e 3
chromatography lead to variable yields in the 50-70%
range. Reaction of compound 2 with trichloroacetonitrile
and triethylamine at rt resulted in a near quantitative
conversion to a new product as judged by TLC. This was
isolated in low yield (34%) and characterized as the 3′-
imidate 3. Treating imidate 3 in dioxane with 1 equiv
of sodium hydride at rt leads to a rapid elimination of
the trichloroimidate to regenerate the starting material
2. In an attempt to overcome the reversability of 3′-
imidate formation under basic conditions, we chose to
react compound 2 in neat trichloroacetonitrile with
catalytic triethylamine. The imidate 3 formed and was
heated to effect the thermal cyclization, with concomitant
opening of the 2,2′-anhydro linkage, to afford oxazoline
4a in 70-80% yield. This conversion represents the first
example we are aware of in which the 3′-hydroxyl of an
anhydronucleoside has intentionally been used to direct
the attack of a nuclephile regiospecifically to the 2′-
position resulting in opening the anhydro linkage to give
a 2′-substituted ribonucleoside. We decided to investi-
gate the reaction of the crude dimethoxytritylation
mixture (>90% 2 by HPLC), after a simple extractive
workup, with trichloroacetonitrile. Thus, when the crude
product 2 was refluxed with an excess of neat trichloro-
acetonitrile and catalytic triethylamine, a 70-80% yield
of oxazoline 4a was isolated. The one-pot conversion of
anhydrouridine 1 to oxazoline 4a (∼70% yield) represents
an efficient and novel entry into the production of
2′-aminopyrimidine nucleosides. All that remains is to
deprotect the 2′,3′-positions of 4a and to protect the 2′-
amine (Scheme 3). Thus oxazoline 4a when treated with
strong ethanolic sodium hydroxide initially forms 2′-N,3′-
O-oxazolidin-2-one intermediate 6a which on prolonged
heating gives a 70-80% yield of 2′-amino-5′-(dimeth-
oxytrityl)-2′-deoxyuridine 7a . The 2′-amine of 7a can
In conclusion, we have presented a novel approach to
the synthesis of 2′-aminopyrimidines that involves the
intramolecular delivery of a nucleophile regiospecifically
to the 2′-position of the sugar. This is accomplished by
tethering the approaching nucleophile to the 3′-hydroxyl
of the anhydronucleoside. The efficient approach we
(17) Divakar, K. J .; Reese, C. B. J . Chem. Soc., Perkin Trans. 1 1982,
179-183.
(18) Bhat, B.; Ugarkar, B. G.; Sayeed, V. A.; Grimm, K.; Kosora,
N.; Domenico, P. A.; Stocker, E. Nucleosides Nucleotides 1989, 8, 179-
183.

Notes
J . Org. Chem., Vol. 61, No. 2, 1996 783
1H, J ) 5.7 Hz), 5.89 (d, 1H, J ) 7.4 Hz), 5.96 (d, 1H, J ) 4.4
Hz), 6.33 (d, 1H, J ) 5.6 Hz), 6.84 and 7.16-7.28 (m, 13H), 7.96
(d, 1H, J ) 7.4 Hz). Anal. Calcd for C30H28N2O7‚0.5H2O: C,
67.03; H, 5.43; N, 5.21. Found: C, 67.02; H, 5.55; N, 4.99.
5′-O-(4,4′-Dim eth oxytr ityl)-2,2′-O-an h ydr o-1-(â-D-ar abin o-
fu r a n osyl)u r a cil 3′-O-Tr ich lor oa cetim id a te (3). To a solu-
tion of 5′-(dimethoxytrityl)anhydrouridine 2 (1.0 g, 1.9 mmol)
in dioxane (5 mL) and trichloroacetonitrile (1 mL) was added
sodium hydride (40 mg, 60% in mineral oil), and the reaction
was stirred for 16 h at room temperature and then evaporated.
The residue was purified on silica gel, eluting with 10%
methanol/dichloromethane to afford imidate 3 as an orange foam
(500 mg, 39% yield): UV λmax 230 (sh), 242; 1H NMR δ 2.91 and
3.12 (ABX, 2H, J ax ) 4.4 Hz , J bx ) 6.3 Hz, J ab ) 10.4 Hz), 3.73
(s, 6H), 4.49 (s, 1H), 5.49 (s, 1H), 5.55 (d, 1H, J ) 5.7 Hz), 5.95
(d, 1H, J ) 7.5 Hz), 6.85 and 7.13-7.29 (m, 13H), 7.95 (d, 1H,
J ) 7.5 Hz), 10.0 (s, 1H). Anal. Calcd for C32H28N3O7Cl3: C,
57.11; H, 4.19; N, 6.24. Found: C, 56.91; H, 4.27; N, 5.90.
5′-O-(4,4′-Dim et h oxyt r it yl)-2′-N,3′-O-(2-(t r ich lor om et h -
yl)oxa zolin o)-2′-d eoxy-1-(â-D-r ibofu r a n osyl)u r a cil (4a ). (a )
A mixture of 5′-(dimethoxytrityl)-2,2′-anhydrouridine 2 (1.1 g,
2.1 mmol) in neat trichloroacetnitrile (5 mL) and sodium hydride
(40 mg, 0.5 equiv, 60% in mineral oil) was heated at 90 °C for
16 h. The dark residue obtained after concentration was purified
on silica gel, eluting with 10% methanol/dichloromethane con-
taining 1% triethylamine, to afford the desired material 4a as a
yellow foam (600 mg, 43% yield). An analytical sample was
crystallized from ethanol to afford a white solid: mp 192-193
°C , UV λmax 218 (sh), 238, 261 (sh); 1H NMR δ 3.16 and 3.48
(ABX, 2H), 3.72 (s, 6H), 4.14 (m, 1H), 5.29 (dd, 1H, J 2′,3′ ) 8.3
Hz, J 2′,1′ ) 1.9 Hz), 5.43 (dd, 1H, J 3′,4′ ) 4.5 Hz), 5.65 (d, 1H, J 5,6
) 8 Hz), 5.92 (d, 1H, J 1′,2′ ) 1.8 Hz), 6.86 and 7.2-7.38 (m, 13H),
7.84 (d, 1H, J 5,6 ) 8.1 Hz), 11.44 (s, 1H); 13C NMR (75 MHz,
DMSO) δ 164.24, 162.44, 159.05, 151.09, 145.67, 144.56, 136.07,
130.47, 130.32, 128.56, 128.33, 127.45, 113.83, 102.37, 93.88,
87.16, 86.79, 86.15, 76.87, 64.31, 55.28, 55.24, 52.05. Anal.
Calcd for C32H28N3O7Cl3: C, 57.11; H, 4.19; N, 6.24; Cl, 15.80.
Found: C, 57.43; H, 4.78; N, 6.08; Cl, 15.44.
(b) On e-P ot P r oced u r e fr om Com p ou n d 1. To a 1 L
Erlenmeyer flask containing anhydrouridine 1 (120.6 g, 0.534
mol), 4,4′-dimethoxytrityl chloride (190.2 g, 1.05 equiv) and
DMAP (0.5 g, catalytic) was added a mixture of pyridine (400
mL) and DMF (300 mL), and the reaction was stirred for 16 h
at rt and then partially evaporated. The concentrated reaction
mixture was partitioned between dichloromethane and water
(1×) and then dilute sodium bicarbonate/sodium chloride, and
the organic phase was dried over magnesium sulfate, filtered,
and evaporated. The residue was then coevaporated once with
toluene to afford a sticky oil. To this reddish oil were added
trichloroacetonitrile (540 g) and triethylamine (10 mL), and the
solution was refluxed for 16 h. The black reaction mixture was
then concentrated on a Buchii (the distilled trichloroacetonitrile/
triethylamine distillate can be reused), and the resulting black
oil was purified by chromatography on silica gel, loading the
compound with toluene/ethyl acetate and eluting with 20-80%
ethyl acetate in hexanes to afford the desired compound 4a as
a foam (289.4 g, 80% yield), identical to the material described
above.
Sch em e 4
describe offers an alternative to the use of azide ion (and
associated explosive hazard) for this type of tranforma-
tion, especially when scale-up to commercial quantities
is considered. The elaboration of anhydrouridine to fully
protected 2′-aminouridine suitable for conversion to the
phosphoramidite is accomplished in three steps with an
approximate overall yield of 50%. The preparation of the
corresponding 2′-aminocytidine analog 13a was investi-
gated by three different approaches. Both the TBDMS
protection (i.e. 10b) and oxazoline C (i.e. 7b) routes afford
the desired protected 2′-aminocytidine in 44% overall
yield from 4a , while the TMS protection route (i.e. 11a )
afforded a 38% overall yield. We are continuing to look
at the utility of this approach for the efficient introduction
of other 2′-substituents.19
Exp er im en ta l Section
Gen er a l. All chromatography was done using Silica gel (J .T.
Baker, 40 mM, 7024-R); NMR were conducted in DMSO on a
300 MHz instrument unless indicated otherwise. UV spectra
were measured in absolute ethanol. Melting points are reported
uncorrected.
2,2′-O-An h yd r o-1-(â-D-a r a bin ofu r a n osyl)u r a cil (1). To a
stirred suspension of diphenyl carbonate (964 g, 1.1 equiv) and
N,N-dimethylformamide (1.1 L) in a 4 L beaker was added
uridine (1 kg, 4.09 mol), and the slurry was heated to 80 °C at
which point sodium bicarbonate was added (4 g) and the reaction
was heated at 110-120 °C (gas evolution, then a brown solution
results which soon deposits the solid product). Once the
evolution of gas subsides (4-5 h), the reaction suspension is
allowed to cool to rt, the precipitate obtained is isolated by
filtration on a large Buchner funnel, and the solids are washed
with ∼2 bed volumes of methanol. The solid is then slurried in
∼1.5 L of methanol, heated for 3 h, and cooled to rt, and the
product is isolated by filtration and dried at 100 °C for 16 h
under high vacuum to afford 1 as a tan solid (785.5 g, 84% yield).
1H NMR, mp, and UV are identical to published data.16
5′-O-(4,4′-Dim eth oxytr ityl)-2,2′-O-an h ydr o-1-(â-D-ar abin o-
fu r a n osyl)u r a cil (2). A suspension of 2,2′-O-anhydrouridine
1 (10.1 g, 0.045 mol), dimethoxytrityl chloride (17.5 g, 1.1 equiv)
and catalytic DMAP (∼50 mg) in pyridine (100 mL) was stirred
16 h at rt and then evaporated. The residue was washed with
dichloromethane/water, and the organic phases were washed
with dilute sodium bicarbonate, dried with magnesium sulfate,
and evaporated. The resulting foam was purified on silica gel,
eluting with 0-20% methanol/ethyl acetate, to afford the desired
material 2 as a foam (13.3 g, 56% yield): UV λmax 218 (sh), 238;
1H NMR δ 2.81 and 2.85 (ABX, 2H, J ab ) 10.2 Hz, J ax ) 4.2 Hz,
J bx ) ∼1 Hz), 3.73 (s, 6H), 4.22 (m, 1H), 4.31 (m, 1H), 5.21 (d,
5′-O-(4,4′-Dim et h oxyt r it yl)-2′-N,3′-O-(2-(t r ich lor om et h -
yl)oxa zolin o)-2′-d eoxycytid in e (4b). Compound 4a (66.5 g,
98.9 mmol) was coevaporated twice with pyridine, then dissolved
in anhydrous acetonitrile (100 mL), and cooled in an ice bath.
In a separate flask, phosphorus oxychloride (27.6 mL, 3 equiv)
was added to an ice-bath-cooled solution of 1,2,4-triazole (11.5
g, 10.7 equiv) in dry acetonitrile (100 mL). After stirring for
∼15 min triethylamine (160 mL, 11.6 equiv) was added and
stirring continued for 20 min, at which time the suspension was
filtered directly into the previously cooled acetonitrile solution
of 4a . The reaction mixture was allowed to warm to rt over 2 h
and then evaporated. The residue was purified by chromatog-
raphy on silica gel using 20-80% ethyl acetate in hexanes
containing 1% triethylamine as the eluent to afford the crude
N4-triazolide 5 as a yellow foam: UV λmax 222, 236, 318; 1H NMR
δ 3.33 and 3.52 (ABX, 2H, J ) 7.9, 3.3, 10.4 Hz), 3.65 (s, 3H),
3.68 (s, 3H), 4.31 (dt, 1H, J ) 4, 7.7 Hz), 5.39 (dd, 1H, J ) 1.5,
8.2 Hz), 5.51 (dd, 1H, J ) 4, 8.2 Hz), 6.06 (d, 1H, J ) 1.4 Hz),
6.79-6.86 and 7.17-7.38 (m, 13H), 6.95 (d, 1H, J ) 7.1 Hz),
8.44 (s, 1H), 8.56 (d, 1H, J ) 7.3 Hz), 9.45 (s, 1H). Intermediate
(19) McGee, D. P. C.; Sebesta, D. P.; O’Rourke, S.; Martinez, M. L.;
J ung, M. E.; Pieken, W. A. Tetrahedron. Lett. 1995, submitted. Patent
Application No. 08/264,029, filed J une 1994.

784 J . Org. Chem., Vol. 61, No. 2, 1996
Notes
5 was dissolved in a mixture of dioxane (350 mL) and ammonium
2′-Am in o-5′-O-(4,4′-dim eth oxytr ityl)-2′-deoxycytidin e (7b).
A mixture of compound 4b (0.672 g, 1 mmol) and cesium
carbonate (0.65 g, 2 mmol) in EtOH:H2O (9 mL, 2:1) was heated
under reflux for 18 h at which point TLC showed complete
conversion of 4b to the product 7b. The solvent was evaporated
under reduced pressure, and the residue was taken up in
dichloromethane, washed once each with saturated NH4Cl,
water, and brine, and then dried (MgSO4). Removal of the
solvent under reduced pressure gave a gel, which was coevapo-
rated twice with methanol to afford the product 7b (0.51 g, 94%)
as a white powder: UV λmax 224 (sh), 236, 276; 1H NMR δ 3.27
(m, 2H), 3.35 (m, 1H), 3.74 (s, 6H), 3.98 (m, 2H), 5.35 (br s,1H),
5.54 (d, 1H, J ) 7.38 Hz), 5.73 (d, 1H, J ) 6.03 Hz), 6.9 and
7.10-7.4 (m, 13H), 7.65 (d, 1H, J ) 7.4 Hz). Anal. Calcd for
hydroxide (250 mL), stirred for 1.5 h at rt, and then evaporated
1
to
/ volume. The reaction mixture was partitioned between
3
dichloromethane and water (1×) and the organic phase dried
over magnesium sulfate and evaporated. The residue was
purified by chromatography on silica gel, eluting first with 40-
80% ethyl acetate in hexanes and then 5-8% methanol in ethyl
acetate (all eluting solvents contain 1% triethylamine). The
desired fractions were pooled and evaporated to afford 4b as a
yellow foam (49.4 g, 74% yield): UV λmax 226, 246, 272 (sh); 1H
NMR (400 MHz) δ 3.22 and 3.52 (ABX, 2H, J ) 4.3, 7.6, 10.2
Hz), 3.73 (s, 3H), 3.74 (s, 6H), 4.07 (m, 1H), 5.19 (dd, 1H, J )
2.14, 8.52 Hz), 5.46 (dd, 1H, J ) 4.2, 8.1 Hz), 5.72 (d, 1H, J )
7.2 Hz), 5.81 (d, 1H, J ) 2.1 Hz), 6.83-7.38 (m, 15H), 7.76 (d,
1H, J ) 7.2 Hz). Anal. Calcd for C32H30N4O6‚0.5 H2O: C, 56.44;
H, 4.44; N, 8.23; Cl, 15.62. Found: C, 56.44; H, 4.46; N, 8.14;
Cl, 16.6.
2′-Am in o-2′-d eoxyu r id in e (9a ). Dimethoxytrityl oxazoline
4a (3.6 g, 5.35 mmol) was treated with 80% aqueous acetic acid
(50 mL) for 16 h at room temperature and then evaporated. The
residue was coevaporated with methanol and then partitioned
between dichloromethane/water, the water phase was evapo-
rated, and the residue was chromatographed on silica gel, eluting
with 20% methanol in dichloromethane to afford 9a (1.1 g, 84%
yield) as a foam. A sample was crystallized from ethanol: mp
197.5-199.5 °C (lit.3 mp 197-198 °C); UV λmax 210, 262; 1H NMR
δ 3.27 (q, 1H, J ) 7.9, 5.2 Hz), 3.5 (s, 2H), 3.85 (m, 1H), 3.89 (d,
1H, J ) 4.9 Hz), 5.06 (t, 1H), 5.38 (br s,1H), 5.65 (d, 2H, J ) 7.4
Hz), 7.82 (d, 1H, J ) 8.1 Hz).
2′-Am in o-5′-O-(4,4′-dim eth oxytr ityl)-2′-deoxyu r idin e (7a).
(a ) A solution of dimethoxytrityl oxazoline 4a (1.5 g, 2.23 mmol)
in dioxane (30 mL) containing 2.72 N sodium hydroxide (1 mL)
is refluxed for 10 h and then evaporated. The residue is
partitioned between water/dichloromethane, dried with magne-
sium sulfate, and evaporated. The residue was purified on silica
gel, eluting with 5-10% methanol/dichloromethane to afford first
5′-O-(4,4′-dimethoxytrityl)-2′-N,3′-O-(2-oxooxazolidin)-2′-de-
oxyuridine (6a ) as a yellow foam (900 mg, 58% yield) [1H NMR
δ 3.15 and 3.36 (ABX, 2H), 3.7 (s, 6H), 4.20 (m, 1H), 4.51 (d,
1H), 4.98 (q, 1H), 5.59 (d, 1H, J ) 8 Hz), 5.76 (br s, 2H, H-1′),
6.67 and 7.23-7.4 (m, 13H), 7.68 (d, 1H, J ) 8 Hz), 8.27 (s, 1H),
11.47 (s, 1H). Anal. Calcd for C31H29N3O8‚0.5 H2O: C, 64.13;
H, 5.20; N, 7.23. Found: C, 64.14; H, 5.23; N, 6.87.] followed
by the free amino compound 7a (95 mg, 8% yield) as a foam . 1H
NMR identical with material prepared via the traditional 2′-
azido route.
C
30H32N4O6‚H2O: C, 64.04; H, 6.09; N, 9.96. Found: C, 63.98;
H, 5.87; N, 9.62.
5′-O-(4,4′-Dim eth oxytr ityl)-2′-tr iflu or oa ceta m id o-2′-d e-
oxycytid in e (8b). To a solution of amine 7b (32.7 g, 60.0 mmol)
in dry acetonitrile (300 mL) and triethylamine (1.8 mL, 2.4
mmol) was added ethyl trifluoroacetate (14.3 mL, 120 mmol),
and the reaction was stirred for 16 h at rt. The resulting
suspension was filtered, and the filtrate washed with acetonitrile
and dried to afford 8b as a white solid (20.6 g, 53.5% yield). The
mother liquors and washes were pooled and evaporated. The
residue was purified by chromatography on silica gel, eluting
with 5% methanol in dichloromethane containing 1% triethyl-
amine, to give an additional quantity of 8b as a foam (16.7 g,
total yield 96%). An analytical sample was crystallized from
DMF/dichloromethane: mp 164-166 °C; UV λmax 222, 242, 274;
1H NMR δ 3.23 (m, 2H), 3.74 (s, 6H), 4.04 (m, 1H), 4.26 (m, 1H),
4.52 (m, 1H), 5.59 (d,1H, J ) 7.4 Hz), 5.70 (d, 1H, J ) 5.2 Hz),
6.09 (d, 1H, J ) 5.9 Hz), 6.89 and 7.21-7.41 (m, 15H), 7.65 (d,
1H, J ) 7.4 Hz), 9.6 (d, 1H, J ) 8.5 Hz). Anal. Calcd for
C
32H31N4O7F3: C, 60.0; H, 4.88; N, 8.47. Found: C, 59.90; H,
4.67; N, 8.63.
U to C Con ver sion : TMS a s a Tr a n sien t P r otectin g
Gr ou p . 5′-O-(4,4′-Dim eth oxytr ityl)-2′-tr iflu or oa ceta m id o-
2′-d eoxycytid in e (8b). To an ice-bath-cooled solution of the
nucleoside 8a (1.635 g, 2.55 mmol) in dry CH3CN (12 mL) and
triethylamine (2.95 mL, 20 mmol) was added trimethylsilyl
chloride (0.65 mL, 5.10 mmol), and the reaction was stirred for
1 h. The reaction mixture was then concentrated under reduced
pressure, and the residue was dissolved in dichloromethane and
washed with water and brine and dried (MgSO4). Evaporation
of the solvent afforded 10a as a foam (1.82 g). In a separate
flask the phosphorus oxy tris-triazolide was prepared as de-
scribed for compound 4b using POCl3 (0.71 mL, 7.65 mmol),
1,2,4-triazole (1.88 g, 27.29 mmol), and triethylamine (4.62 mL,
33.15 mmol). The white triethylamine hydrochloride precipitate
was quickly filtered off and washed once with dry CH3CN (10
mL). To the stirred ice-bath-cooled filtrate was added dropwise
a solution of the silylated nucleoside 10a (1.82 g, 2.55 mmol) in
dry CH3CN (5 mL) containing triethylamine (0.5 mL), and the
reaction mixture was slowly warmed up to rt. After 4 h of
stirring at ambient temperature, TLC showed complete conver-
sion of the starting material to the product. The solvent was
removed under reduced pressure, and the reaction was quenched
with ice water. The aqueous layer was extracted with dichlo-
romethane (2 × 15 mL), and the organic phase was washed with
water and brine and dried (MgSO4). Evaporation of the solvent
under reduced pressure gave 11a as a slightly yellowish foam
(2 g) which was used as such in the next step. An analytical
sample can be prepared by chromatography on silica gel, eluting
with 30-80% ethyl acetate in hexanes, pooling and evaporating
the desired fractions to a foam which was crystallized from ethyl
(b) Meth od Usin g Eth a n ol/Wa ter . To a solution of oxazo-
line 4a (267.6 g, 0.398 mol) in ethanol (1 L) was added a 6 N
sodium hydroxide solution (500 mL), and the reaction was
refluxed for 16 h, then cooled, and evaporated partially. The
residue was partitioned between dichloromethane and satu-
rated ammonium chloride (2×), the aqueous phase was back
washed 1× with dichloromethane, and the combined organic
phase was dried over magnesium sulfate, filtered, and evapo-
rated to a foam. The foam was purified by chromatography on
silica gel, eluting with 5-10% methanol in dichloromethane
containing 1% triethylamine, to afford compound 7a (172 g, 79%
yield) as a foam. An analytical sample was crystallized from
ethyl acetate/hexanes to afford a white solid: mp 116-118 °C;
UV λmax 212, 236, 266; 1H NMR δ 3.18 and 3.22 (ABX, 2H, J )
2.8, 4.6, 14.4 Hz), 3.38 (t, 2H, J ) 6.1 Hz), 3.7 (s, 6H), 3.97 (m,
2H), 5.41 (d, 1H, J ) 8 Hz), 5.68 (d, 1H, J ) 7.2 Hz), 6.88 and
7.23-7.39 (m, 13 H), 7.64 (d, 1H, J ) 8.1 Hz). Anal. Calcd for
C
30H31N3O7‚0.5 H2O: C, 64.97; H, 5.82; N, 7.57. Found: C,
65.23; H, 5.97; N, 7.40.
1
5′-O-(4,4′-Dim eth oxytr ityl)-2′-tr iflu or oa ceta m id o-2′-d e-
oxyu r id in e (8a ). A solution of amine 7a (118.7 g, 0.217 mol),
triethylamine (10 mL), and ethyl trifluoroacetate (52 mL, 2
equiv) in dry acetonitrile (800 mL) was stirred for 16 h at rt.
The resulting suspension was filtered, washed with fresh
acetonitrile (∼40 mL), and dried to afford 8a (76.9 g, 55% yield).
An analytical sample was recrystallized from ethanol to afford
acetate to afford 11a as white solid: mp 137-139 °C; H NMR
δ 3.34 and 3.39 (ABX, 2H), 3.75 (s, 6H), 4.21 (m, 1H), 4.43 (m,
1H), 4.68 (m, 1H), 5.82 (d, J ) 4.7 Hz, 1H), 6.09 (d, J ) 3.3 Hz,
1H), 6.74 (d, J ) 7.1 Hz, 1H), 6.92 and 7.26-7.45 (m, 13H), 8.43
(s, 1H), 8.52 (d, J ) 7.1 Hz, 1H), 9.48 (s, 1H), 9.2 (d, J ) 7.8
Hz,1H). Anal. Calcd for C34H31N6O7F3‚H2O: C, 57.46; H, 4.68;
N, 11.82; F, 8.02. Found: C, 52.81; H, 4.55; N, 11.81; F, 7.16. A
solution of the crude compound 11a (2 g) obtained above was
stirred at rt in THF:NH4OH (10 mL, 9:1.5) for 4 h at which time
TLC showed complete conversion to a more polar compound. The
solvent was evaporated under reduced pressure, and the residue
was taken up in dichloromethane, washed with water and brine,
and dried (MgSO4). The residue obtained after concentration
1
a white solid: mp 227-229 °C; UV λmax 218, 236, 264 (sh); H
NMR δ 3.20 and 3.30 (ABX, 2H), 3.74 (s, 6H), 4.04 (m, 1H), 4.27
(m, 1H), 4.61 (m, 1H), 5.46 (d, J ) 8.1 Hz, 1H), 5.79 (br d, 1H),
6.9 and 7.22-7.41 (m, 13H), 7.69 (d, J ) 8.1 Hz, 1H), 9.55 (br d,
1H), 11.41 (br s, 1H). Anal. Calcd for C32H30N3O8F3: C, 59.90;
H, 4.71; N, 6.55. Found: C, 59.89; H, 4.93; N, 6.44.

Notes
J . Org. Chem., Vol. 61, No. 2, 1996 785
was purified by silica gel column chromatography, eluting with
5% methanol in ethyl acetate to afford the product 8b as a foam
(1.34 g, 82% yield). The overall yield in three steps was >80%.
Data as described previously for 8b.
Less polar spots are eluted with ethyl acetate and 1% triethyl-
amine and the compound eluted with 3% methanol/ethyl acetate
and 1% triethylamine to afford compound 12b as a foam (2.1 g,
71% yield): UV λmax 206, 236, 274; 1H NMR (400 MHz) δ -0.19
(s, 3H), -0.10 (s, 3H), 0.69 (s, 9H), 3.15 and 3.29 (ABX, 2H, J )
2, 5.4, 10.6 Hz), 3.74 (s, 6H), 4.08 (m, 1H), 4.37 (dt, 1H, J ) 0,
6.6 Hz), 4.51 (m, 1H), 5.65 (d, 1H, J ) 7.3 Hz), 6.03 (d, 1H, J )
4.4 Hz), 6.88 and 7.22-7.40 (m, 15H), 7.74 (d, 1H, J ) 7.3 Hz),
9.73 (d, 1H, J ) 9.2 Hz). Anal. Calcd for C38H45N4O7F3-
Si‚0.5H2O: C, 59.75; H, 6.07; N, 7.33. Found: C, 60.07; H, 6.16;
N, 7.33. HRMS (MH+) 755.3074.
N4-Ben zoyl-5′-O-(4,4′-d im et h oxyt r it yl)-2′-t r iflu or oa cet -
a m id o-2′-d eoxycytid in e (13a ). To a solution of compound 8b
(54.25 g, 84.68 mmol) in dry DMF (300 mL) was added benzoic
anhydride (19.16 g, 84.69 mmol), and the reaction was stirred
for 16 h at rt. Methanol (100 mL) was added, and after stirring
for 0.5 h the reaction was evaporated under reduced pressure.
The residue was dissolved in ethyl acetate and washed with 5%
sodium bicarbonate and the organic phase dried (MgSO4) and
evaporated to dryness. The crude product was purified by
chromatography on Silica gel, eluting with 10-90% ethyl acetate
in dichloromethane, to afford after evaporation 13a as a white
foam (41.5 g, 66% yield): UV λmax 230 (sh), 248 (sh), 260, 308;
1H NMR δ 3.30 and 3.35 (ABX, 2H), 4.14 (m, 1H), 4.36 (m, 1H),
4.60 (m, 1H), 5.77 (d, J ) 5.2 Hz, 1H,), 6.08 (d, J ) 4.3 Hz, 1H),
6.92, 7.23-7.64, 8.01, 8.22 (m, 20H), 9.66 (d, J ) 7.2 Hz, 1H),
11.33 (s, 1H). Anal. Calcd for C39H35N4O8F3‚0.5 H2O: C, 62.15;
H, 4.81; N, 7.43. Found: C, 62.42; H, 4.81; N, 7.46.
N 4-B e n zo y l-3′-O-(t er t -b u t y ld im e t h y lsily l)-5′-O-(4,4′-
d im et h oxyt r it yl)-2′-t r iflu or oa cet a m id o-2′-d eoxycyt id in e
(13b). Compound 12b (2.1 g, 2.78 mmol) was dissolved in dry
N,N-dimethylformamide (6 mL). Benzoic anhydride (0.69 g, 3.06
mmol) was added and the reaction was stirred at rt overnight.
The reaction was concentrated, partitioned between dichlo-
romethane and 5% sodium bicarbonate, dried (MgSO4), and
evaporated to a yellow gum. The gum was dissolved in ethyl
acetate with 2% triethylamine and loaded onto a silica gel
column equilibrated with hexane and 2% triethylamine. The
column was eluted with 20% ethyl acetate/hexane and 1%
triethylamine followed by 40-60% ethyl acetate/hexane and 1%
triethylamine to afford compound 13b as a white foam (2.2 g
,93% yield): UV λmax 206, 236, 262, 306; 1H NMR δ -0.20 (s,
3H), -0.09 (s, 3H), 0.67 (s, 9H), 3.22 and 3.4 (ABX, 2H, J ) 2,
5.1, 11.1 Hz), 4.2 (m, 1H), 4.47 (dt, 1H, J ) 0, 7.4 Hz), 3.75 (s,
6H), 4.64 (m, 1H), 6.03 (d, 1H, J ) 3.1 Hz), 6.9 and 7.23-7.66
and 7.8 (m, 19H), 8.35 (d, 1H, J ) 7.5 Hz), 9.80 (d, 1H, J ) 9.1
Hz), 11.35 (s, 1H). Anal. Calcd for C45H49N4O8F3Si‚0.5 H2O:
C, 62.27; H, 5.81; N, 6.45. Found: C, 62.21; H, 5.89; N, 6.35.
MS (MH+) 859.
U to C Con ver sion : TBDMS a s P r otectin g Gr ou p . 3′-
O-(ter t-Bu t yld im et h ylsilyl)-5′-O-(4,4′-d im et h oxyt r it yl)-2′-
t r iflu or oa cet a m id o-2′-d eoxycyt id in e (12b ). tert-Butyldi-
methylsilyl chloride (1.21 g, 7.8 mmol) was added to a stirred
solution of 8a (2.5 g, 3.9 mmol) in dry acetonitrile (12 mL) and
triethylamine (4.3 mL, 31.2 mmol). The reaction is allowed to
stir at rt for 3 h at which time the TEA-HCl precipitate is
filtered from the reaction mixture to afford 10b. Prepared in a
separate flask, phosphorus oxychloride (1.1 mL, 11.7 mmol) is
slowly added to an ice-bath-cooled solution of 1,2,4-triazole (2.88
g, 41.7 mmol) in dry acetonitrile (12.5 mL). After 15 min
triethylamine (7.1 mL, 50.65 mmol) is added and stirring
continued for 20 min at which time the solution is filtered
directly into the flask containing 10b described above. After
15 min at rt TLC shows the required product plus a polar spot.
The reaction is allowed to stir an additional 3 h during which
time the TLC profile remains the same. The solution is
concentrated, diluted with ethyl acetate, and poured over
crushed ice with stirring. Layers are partitioned, and the
organic phase is washed once with brine and then dried (MgSO4)
and concentrated to afford intermediate 11b as an orange/brown
foam.
N4-Ben zoyl-5′-O-(4,4′-d im et h oxyt r it yl)-2′-t r iflu or oa cet -
a m id o-2′-d eoxycytid in e (13a ). Triethylamine hydrofluoride
(0.37 g, 3.07 mmol in 3 mL of dry acetonitrile) was added to a
stirred solution of 13b (2.2 g, 2.56 mmol) in dry acetonitrile (80
mL). The reaction was stirred at rt for 24 h at which time TLC
showed 50% desired product and 50% starting material. At 29
h an additional 0.09 g of triethylamine hydrofluoride was added
and the reaction stirred 42 h more. The reaction was evaporated
to a pale yellow foam (2.23 g), partitioned between dichlo-
romethane (100 mL) and water (2 × 50 mL), dried (MgSO4, trace
pink color is noted), and concentrated to afford 13a as an off
white foam (1.8 g 95% yield). Data as previously described.
Intermediate 11b (4 g crude) is stirred in THF (18 mL), and
NH4OH (2 mL) is added. After 4 h at rt an additional 1 mL of
NH4OH is added to drive the reaction to completion. At 6 h the
reaction is diluted with ethyl acetate (200 mL) and washed with
brine (2 × 100 mL) and then dried (MgSO4) and concentrated
to a yellow foam (3 g). The compound is loaded onto a silica gel
column equilibrated with ethyl acetate and 2% triethylamine.
Ack n ow led gm en t. We thank J im Reed for the
NMR experiments and Drs. Michael J ung and Wolfgang
Pieken for encouragement and suggestions.
J O9510548