A constrained tricyclic nucleic acid analogue of α-l-LNA: Investigating the effects of dual conformational restriction on duplex thermal stability

Article abstract of DOI:10.1021/jo401170y

A constrained tricyclic analogue of α-l-LNA (2), which contains dual modes of conformational restriction about the ribose sugar moiety, has been synthesized and characterized by X-ray crystallography. Thermal denaturation experiments of oligonucleotide se

Full text of DOI:10.1021/jo401170y

Article
pubs.acs.org/joc
A Constrained Tricyclic Nucleic Acid Analogue of α‑L‑LNA:
Investigating the Effects of Dual Conformational Restriction on
Duplex Thermal Stability
Stephen Hanessian,*,† Jernej Wagger,† Bradley L. Merner,† Robert D. Giacometti,†
Michael E. Østergaard,‡ Eric E. Swayze,‡ and Punit P. Seth*,‡
†Department of Chemistry, Universite
́ ́ ́
de Montreal, Montreal, QC, H3C 3J7, Canada
‡Department of Medicinal Chemistry, Isis Pharmaceuticals, Inc., Carlsbad, California 92010, United States
S
* Supporting Information
ABSTRACT: A constrained tricyclic analogue of α-L-LNA (2), which contains dual modes of conformational restriction about
the ribose sugar moiety, has been synthesized and characterized by X-ray crystallography. Thermal denaturation experiments of
oligonucleotide sequences containing this tricyclic α-L-LNA analogue (α-L-TriNA 2, 5) indicate that this modification is
moderately stabilizing when paired with complementary DNA and RNA, but less stabilizing than both α-L-LNA (2) and α-L-
TriNA 1 (4).
thermal stability.7 However, it was not clear whether the duplex
destabilization observed for analogue 3 was a consequence of
the steric bulk enforced by the appended six-membered ring,
INTRODUCTION
■
The synthesis of modified nucleic acids that demonstrate
improved target recognition and affinity, through standard
the conformational equilibrium of the cis-fused bicyclic ring
Watson−Crick base pairing, is an important area of research for
RNA-targeted therapeutics.1 Antisense oligonucleotides that
incorporate these synthetic nucleic acid modifications show
improved duplex thermal stability versus DNA and RNA
complements. The 2′,4′-bridged nucleic acid (BNA) mod-
ifications, such as LNA2 and α-L-LNA3 (2, Figure 1), are
exemplary in this regard and have demonstrated marked
increases in duplex thermal stability (LNA, ΔTm = +5 °C/mod
vs RNA; α-L-LNA, ΔTm = +5 °C/mod vs RNA) when
compared to unmodified or natural nucleic acids (e.g., 1). The
key structural modification in these two important nucleic acid
monomers is the incorporation of a short oxamethylene bridge
between the 2′- and 4′-positions, which ultimately locks the
sugar pucker of the ribose moiety into an N-type, C3′-endo
conformation. Other important modifications to natural nucleic
acids involve constraining the dihedral or torsion angles of the
deoxyribose moiety.4,5
We recently showed that tricyclic nucleic acid analogue 4 (α-
L-TriNA 1),6 which restricts conformational mobility via spiro-
annulation around the torsion angle γ in α-L-LNA, provides
unprecedented improvements in oligonucleotide duplex
stability in thermal denaturation (Tm) experiments (Figure
1). In the preceding article we demonstrated that constraining
the torsion angle γ in a cis-α-L-[4.3.0]bicyclo-DNA monomer
(3), which lacks the 2′,4′-bridging group present in α-L-LNA
(2) and tricyclic analogue 4, had a destabilizing effect on duplex
system, or the absence of the 2′,4′-bridging group. To address
these questions we designed α-L-TriNA 2 (5), a tricyclic
analogue of α-L-LNA (2), which introduces a 2′,4′-bridging
group into bicyclic analogue 3. Introduction of this oxa-
methylene bridge not only locks the furanose ring in an N-type
sugar pucker, but also restricts conformational mobility of the
appended six-membered ring by locking the cyclohexane
system in a single chair conformation (dual conformational
restrictions). In this article we report the stereocontrolled
synthesis of α-L-TriNA 2, a constrained tricyclic nucleoside
modification represented by monomer 5, as well as the duplex
stabilizing properties of oligonucleotides that incorporate this
modification.
At the outset of this work, there were very few reports of
TriNA syntheses,6,8 especially of systems containing a
selectively protected and architecturally complex tricyclic core
such as that found in 6 (Scheme 1). Molecular models
indicated that formation of the 2′,4′-anhydrobridge in 6 might
present a formidable synthetic challenge if carried out at a late
stage in the synthesis of the designed nucleoside, given the
need to invert the C2′-OH group. However, we were also
cognizant that this could be done by way of a 2,2′-
Received: June 4, 2013
© XXXX American Chemical Society
A
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Scheme 1. Retrosynthetic Analysis of Tricyclic Nucleoside 6
this approach is shown in Scheme 1, with readily available
bicyclic alcohol 12 as the synthetic precursor.10
RESULTS AND DISCUSSION
■
Initial forays into delineating a viable synthetic route to tricyclic
nucleic acid monomer 5 were pursued for a uridine analogue of
6. The synthesis of 6 (R1, R2 = H; Bx = uracil) commenced
with alcohol 12 (Scheme 2), since it was available in multigram
quantities from a process developed during efforts toward the
synthesis of related nucleoside monomers for antisense
applications.10 Oxidation to the aldehyde, followed by a BF3·
Et2O-mediated Sakurai allylation9 reaction furnished (S)-
configured homoallylic alcohol 13 in 85% yield over two
steps (dr >20:1). Unfortunately, 13 proved to be resistant to
standard benzylation procedures, and in all cases only starting
material was recovered. In fact, the hindered nature of this
secondary, neopentylic-like alcohol limited the use of
protecting groups to either acetate or silyl ethers (TBS or
TES). Accordingly, 13 was converted to acetate 14 to avoid
compatibility issues that could arise with the introduction of a
second silyl ether protecting group.
Cleavage of the isopropylidene group followed by acetylation
gave a mixture of anomeric acetates that was used in a
Vorbruggen glycosylation reaction to give nucleoside 15 in 73%
̈
Figure 1. Rational design of tricyclic analogues of α-L-LNA (2).
yield for the three-step sequence.11 Chemoselective hydrolysis
of the C2′-OAc group followed by mesylation afforded 16 in
90% yield. Passage to tricyclic intermediate 17 was achieved
through DBU-induced formation of the 2,2′-anhydronucleo-
side, followed by cleavage of the TBDPS ether under AcOH-
buffered TBAF conditions and subsequent conversion of the
resulting primary alcohol to the corresponding mesylate. Upon
treatment of 17 with a 2 M solution of NaOH, formation of the
2′,4′-bridged nucleoside occurred with concomitant cleavage of
the homoallylic acetate group to give 18. Alternatively, removal
of the TBDPS group and mesylation, followed by intra-
molecular cyclization and cleavage of the acetate also led to 18
in 65% overall yield (Scheme 2). Protection of the resulting
alcohol as a TBS ether was followed by oxidative cleavage of the
2-naphthylmethyl (Nap) ether in the presence of DDQ,
furnishing 19 in 76% yield. Disappointingly, the C3′-OH group
proved to be resistant to further oxidation under a variety of
conditions, and, consequently, we were unable to test the
proline-catalyzed intramolecular aldol cyclization. To the best
anhydronucleoside intermediate (8, Scheme 1), which would
ultimately be followed by intramolecular cyclization of the
inverted C2′-OH group onto a suitably activated C5′ sulfonate
ester (7). With this strategy in mind, we focused on forming
the oxabicyclic ring system of 7, which already contained a
pyrimidine nucleobase. Intramolecular aldol carbocyclization of
10 was planned to construct the six-membered carbocyclic
component of the oxabicyclic ring system present in 9. Our
successful implementation of this strategy in the synthesis of cis-
α-L-[4.3.0]bicyclo-DNA monomer 37 provided the basis for
following a similar approach, with the added challenge of
establishing the 2′,4′-anhydrobridge present in 6 later in the
synthetic sequence. The stereochemistry of the secondary
alcohol on the cyclohexane ring in 11 was envisaged to
originate from a stereoselective Sakurai allylation reaction on
the corresponding aldehyde of 12.9 The retrosynthetic plan for
B
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a
Scheme 2. Synthesis of Nucleoside 19 and Attempted Synthesis of Ketone 20
a
Reagents and conditions: (a) PCC, CH2Cl2, 40 °C, 12 h; (b) BF3·Et2O, allyltrimethylsilane, CH2Cl2, −40 °C, 1 h, dr >20:1; (c) Ac2O, pyridine, 100
°C, 16 h; (d) AcOH/H2O (4:1), 90 °C, 6 h; (e) Ac2O, pyridine, 23 °C, 12 h; (f) uracil, BSA, MeCN, 90 °C, 1 h then 0 °C, TMSOTf, 50 °C, 24 h;
(g) K2CO3, MeOH, 23 °C, 1 h; (h) MsCl, pyridine, 23 °C, 2 h; (i) DBU, MeCN, 23 °C, 3 h; (j) TBAF, AcOH, 4 °C, 3 d; (k) MsCl, pyridine, 23 °C,
2 h; (l) NaOH, H2O/EtOH (1:1), 90 °C, 1 h; (m) TBSOTf, CH2Cl2, pyridine, 23 °C, 12 h; (n) DDQ, H2O/CH2Cl2 (1:9), 23 °C, 48 h.
of our knowledge, the oxidation of endo-disposed secondary
alcohols of related 1,4-dioxa-bicyclo[2.2.1]heptane systems is
unprecedented. Nevertheless, despite these shortcomings, the
stereochemical outcome of the Sakurai allylation reaction was
corroborated by an X-ray crystal structure of alcohol 19.12
In our second approach, we once again planned to employ a
proline-catalyzed intramolecular aldol reaction, which was
successfully used in the synthesis of nucleoside 3 to construct
the cyclohexane ring in 6.7 The synthesis began with the
formation of TBS ether 21 (Scheme 3), which was readily
converted to ketone 22 in 85% yield over two steps.
Application of the Wacker oxidation gave the desired methyl
ketone derivative of 22, which underwent a smooth intra-
molecular aldol reaction in the presence of a catalytic amount
of L-proline to furnish tricyclic ketone 23 in 78% overall yield.
Reduction of the ketone13 followed by thiocarbonyl imidazole
ester formation and a Barton−McCombie deoxygenation14
reaction led to 24. With the carbocyclic ring of 6 now intact,
the next stage in the synthetic plan called for the introduction
of the thymine nucleobase. This necessitated the removal of the
isopropylidene group present in 24, which, surprisingly, proved
to be incompatible with the TBS protecting group at this
juncture in the synthesis. Virtually all standard and modified
procedures for the acidic hydrolysis of the acetonide present in
24 led to simultaneous cleavage of the TBS ether and resulted
in the formation of 25. The formation of 25 presumably
proceeds through an oxocarbenium ion intermediate that is
subsequently attacked by the proximal secondary alcohol of the
cyclohexane ring system (see proposed mechanism, Scheme
3).15
Scheme 3. Synthesis of Tricyclic Intermediate 24 and
Undesired Acetonide Hydrolysis Side Product 25
a
a
Reagents and conditions: (a) TBSOTf, pyridine, CH2Cl2, 0 to 23 °C,
3 h; (b) DDQ, H2O/CH2Cl2 (1:9), 23 °C, 6 h; (c) DMP, NaHCO3,
CH2Cl2, 23 °C, 18 h; (d) O2 (balloon), PdCl2, Cu(OAc)2, DMA/H2O
(7:1), 40 °C, 24 h; (e) L-proline, DMSO, 55 °C, 54 h; (f)
Me4NBH(OAc)3, AcOH, MeCN, 0 to 23 °C, 7 h; (g) Im2CS, DMAP,
CH2Cl2, 40 °C, 19 h; (h) Bu3SnH, AIBN, PhMe, 110 °C, 15 min.
In the third and final approach (Scheme 4), we exploited
differences in reactivity between the TBS and TBDPS
protecting groups in the presence of TBAF. Gratifyingly,
selective cleavage of the TBDPS group was possible at 0 °C in
THF, furnishing the corresponding primary alcohol, which was
C
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a
Scheme 4. Completion of the Synthesis and Preparation of Phosphoramidite 33
a
Reagents and conditions: (a) TBAF, THF, 0 °C, 1 h; (b) NaH, NapBr, TBAI, THF/DMF (1:1), 0 °C, 2 h; (c) TBAF, THF, 23 °C, 24 h; (d) NaH,
BnBr, TBAI, THF/DMF (1:1), 23 °C, 3 h; (e) FeCl3·6H2O, CH2Cl2, 0 to 23 °C, 1 h; (f) Ac2O, DMAP, pyridine, CH2Cl2, 0 to 23 °C, 1 h; (g) BSA,
thymine, 1,2-dichloroethane, 80 °C, 1 h, then TMSOTf, 0 to 80 °C, 13 h; (h) K2CO3, MeOH, 23 °C, 18 h; (i) MsCl, DMAP, pyridine, 0 to 23 °C,
16 h; (j) DBU, MeCN, 80 °C, 3 d; (k) DDQ, H2O/CH2Cl2 (1:9), 23 °C, 3 h; (l) MsCl, DMAP, pyridine, 0 to 23 °C, 12 h; (m) 2 M NaOH, EtOH/
H2O (1:1), 100 °C, 30 min; (n) H2, Pd(OH)2/C, EtOH/MeOH (1:1), 23 °C, 15 h; (o) LevOH, Mukaiyama reagent, NEt3, DMAP, CH2Cl2, 23 °C,
16 h; (p) NC(CH2)2OPClN(i-Pr)2, N-methylimidazole, EtN(i-Pr)2, DMF, 23 °C, 2 h.
Table 1. Sequence and Duplex Stabilizing Properties of DNA Oligonucleotides Modified with α-L-LNA (2), cis-α-L-[4.3.0]bc-
DNA (3), α-L-TriNA 1 (4), and, α-L-TriNA 2 (5) versus Complementary DNA and RNA
a
b
b
ODN
mod
mass calcd
mass found
sequence (5′ to 3′)
ΔTm/mod (°C) vs DNA
ΔTm/mod (°C) vs RNA
c
A1
A2
A3
A4
A5
B1
B2
B3
B4
DNA
3633.4
3661.4
3673.4
3701.5
3701.5
3645.2
3673.5
3713.5
3713.5
3632.9
3660.6
3672.9
3700.5
3700.9
3645.5
3672.6
3712.7
3712.9
d(GCGTTTTTTGCT)
d(GCGTTTTTTGCT)
d(GCGTTTTTTGCT)
d(GCGTTTTTTGCT)
d(GCGTTTTTTGCT)
d(CCAGTGATATGC)
d(CCAGTGATATGC)
d(CCAGTGATATGC)
d(CCAGTGATATGC)
ref
ref
c
2
+1.4
−8.8
+2.6
−2.6
ref
+5.7
−3.7
+7.1
+1.2
ref
d
3
c
4
5
c
DNA
c
2
+6.5
+7.4
+2.3
+6.3
+8.3
c
4
5
+4.4
a
b
Bold and underlined letters indicates modified nucleotide, base code: T = thymine, U = uracil, C = cytosine, A = adenine, and G = guanine. Tm
values were measured in 10 mM sodium phosphate buffer (pH 7.2) containing 100 mM NaCl and 0.1 mM EDTA. Sequence of DNA complements:
5′-d(AGCAAAAAACGC)-3′ for A1−A5 and 5′-d(GCATATCACTGG)-3′ for B1−B4. Sequence of RNA complements: 5′-r(AGCAAAAAACGC)-
c
d
3′ for A1−A5 and 5′-r(GCAUAUCACUGG)-3′ for B1−B4; Reference 6. Reference 7.
selectively reprotected as the Nap ether to give 26 in 88% yield
for the two-step sequence. Removal of the TBS ether and
subsequent protection of both the secondary and tertiary
alcohols as benzyl ethers afforded 27. Having taken a somewhat
circuitous (but necessary) route to a fully O-protected
intermediate (27), we were able to selectively cleave the
isopropylidene group without incident in the presence of FeCl3·
6H2O. Acetylation of the mixture of diasteromeric diols,
moiety and subsequent formation of tricyclic nucleoside 31 in
96% yield.
Hydrogenolysis afforded 32, which could not be converted to
the corresponding 4,4′-dimethoxytrityl (DMTr) derivative,
presumably due to the congested tricyclic core and hindered
nature of the C5′-OH group. Fortunately, levulinyl (Lev) esters
have emerged as suitable replacements of DMTr ethers,6,16 and
conversion of 32 to the corresponding Lev ester gave key
nucleoside monomer 6 (R1 = Lev, R2 = H, Bx = thymine) for
Tm studies. Single crystals of 32 suitable for X-ray diffraction
analysis were obtained, which allowed us to unequivocally
determine the absolute stereochemistry of the tricyclic
nucleoside.12 Furthermore, the X-ray structure of 32 shows
that the ribose sugar pucker is locked in an N-type
followed by a Vorbruggen glycosylation reaction gave bicyclic
̈
nucleoside 28. Cleavage of the C2′-OAc group and mesylation
of the resulting alcohol furnished 29 in 87% yield. Three
straightforward synthetic operations led to 2,2′-anhydronucleo-
side intermediate 30, which upon treatment with 2 M NaOH at
100 °C underwent rapid hydrolysis of the 2,2′-anhydrouridine
D
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conformation and that the dihedral angle γ is 177° (cf., 169° for
4).6
Table 2. Mismatch Discrimination Properties of DNA, α-L-
LNA (2), α-L-TriNA 1 (4), and α-L-TriNA 2 (5)
Oligonucleotide synthesis was carried out using an
automated DNA synthesizer on a Unylinker-loaded polystyrene
resin followed by purification using ion exchange chromatog-
raphy. DNA phosphoramidites were incorporated using
standard coupling, oxidation, and capping reagents. The
modified phosphoramidite (33) was incorporated with >95%
efficiency (judged by analysis of failure sequences after cleavage
from the resin), using manual conditions that were previously
described in detail.6
a
ΔTm/ mod vs RNA (°C)
mismatch
discrimination
[Tm(mismatch) −
Tm(match)]
b
ODN
mod
X = A
X = G
X = C
X = U
A1
A2
A4
A5
DNA
ref
−4.1
−4.7
−5.5
−5.7
−13.0
−14.8
−16.7
−14.3
−13.2
−13.5
−17.0
−12.1
2
4
5
+5.7
+7.1
+1.2
a
We measured the ability of the α-L-TriNA 2 motif (5, Figure
1) to stabilize oligonucleotide duplexes versus complementary
DNA and RNA using two different sequences (A and B). In
sequence A, motif 5 was incorporated into a stretch of dT
nucleotides, whereas it was flanked on either side by dA
nucleotides in sequence B (Table 1). In sequence A, a single
incorporation of α-L-TriNA 2 (5) (A5, ΔTm +1.2 °C/mod vs
RNA and −2.6 °C/mod vs DNA) had a modest stabilizing
effect on duplex thermal stability versus complementary RNA
and a slightly destabilizing effect versus complementary DNA.
In contrast, both α-L-LNA (2) (A2, ΔTm +5.7 °C/mod vs RNA
and +1.4 °C/mod vs DNA) and α-L-TriNA 1 (4) (A4, ΔTm
+7.1 °C/mod vs RNA and +2.6 °C/mod vs DNA) had a
significantly higher stabilizing effect on duplex thermal stability.
However, tricyclic analogue 5 was clearly more stabilizing than
its bicyclic cis-α-L-[4.3.0]bicyclo-DNA counterpart (3) (A3,
ΔTm −3.7 °C/mod vs RNA and −8.8 °C/mod vs DNA), which
underscores the importance of the 2′,4′-bridging group in 5,
and its contribution to improving duplex thermal stability. In
sequence B, a single incorporation of 5 showed improved
duplex thermal stability (B4, ΔTm +4.4 °C/mod vs RNA and
+2.3 °C/mod vs DNA) versus complementary RNA and DNA.
However, the increase in Tm was still less than that observed
when employing α-L-LNA (2) (B2, ΔTm +6.5 °C/mod vs RNA
and +6.3 °C/mod vs DNA) and α-L-TriNA 1 (4) (B3, ΔTm
+8.3 °C/mod vs RNA and +7.4 °C/mod vs DNA) in the same
sequence. Unfortunately, a single incorporation of bicyclic
analogue 3 into sequence B was not available for comparison.
Lastly, we measured the ability of tricyclic analogue 5 to
discriminate between matched and mismatched RNA comple-
ments (Table 2). We found that 5 exhibited excellent mismatch
discrimination properties, which were comparable to those
observed for DNA, α-L-LNA (2), and α-L-TriNA 1 (4).
Tm values were measured in 10 mM sodium phosphate buffer (pH
7.2) containing 100 mM NaCl and 0.1 mM EDTA using 5′
d(GCGTTTTTTGCT)-3′ where the bold and underlined nucleotide
indicates the site of modification; sequence of RNA complement 5′-
b
r(AGCAAAXAACGC)-3′. Mismatch discrimination values were
calculated by subtracting the Tm measured versus the mismatched
RNA complement (X = G, C, or U) from the Tm versus the matched
RNA complement (X = A) for each modification.
imparted by the 2′,4′-bridging oxamethylene group.6 Both
tricyclic nucleoside modifications 4 and 5 restrict rotation
around the C4′−C5′ bond (torsion angle γ) in almost the same
orientation (see overlay structures, Figure 1). However, while
the added bulk of the six-membered ring in 4 is expected to lie
at the edge of the major groove of the modified duplex, the
added bulk of the appended six-membered ring in 5 is directed
into the minor groove where it could disrupt the water of
hydration network around the sugar−phosphate backbone.17
Interestingly, we have previously shown that introducing a
methyl group in the (S)-configuration at the C5′-position of α-
L-LNA had a destabilizing effect on duplex thermal stability in
comparison to α-L-LNA (2) and the (R)-5′-Me-α-L-LNA
epimer.18 The absolute stereochemistry of the 5′-carbon atom
in modification 5 is also in the (S)-configuration. However, it
remains unclear at this time as to how, or if, the configuration
of the 5′-carbon atom in 5 affects duplex stability. Inverting the
configuration of this stereogenic center in 5 would position the
5′-hydroxyl group in an axial orientation and drastically alter
the orientation around γ from that observed in the NMR
structure solution of an α-L-LNA (2) modified DNA/RNA
duplex.19 For these reasons, constrained nucleic acid analogues
such as 4 and 5 provide additional opportunities to further
probe the conformational requirements around the backbone
torsion angles for efficient hybridization with complementary
nucleic acids.
CONCLUSIONS
■
This work demonstrates that incorporating a 2′,4′-anhydro-
bridge into the cis-α-L-[4.3.0]bicyclo-DNA framework of
nucleic acid analogue 38 results in a significant increase in the
duplex thermal stability of oligonucleotides that incorporate
tricyclic analogue 5 versus those containing bicyclic analogue 3
(ΔTm = +4.9 °C/mod vs RNA and +6.2 °C/mod vs DNA).
This observation highlights the pivotal role of the 2′,4′-bridging
group in restricting the conformation of the nucleoside
furanose ring and consequently enhancing the thermal stability
of oligonucleotide duplexes. However, the increase in duplex
thermal stability produced by tricyclic nucleoside 5 was
significantly lower than that seen with α-L-LNA (2), which
suggests that the linearly annulated six-membered, carbocyclic
ring in 5 has a net destabilizing contribution. This is in contrast
to our previous observation with tricyclic analogue 4, where the
spiro-annulated six-membered, carbocyclic ring had an additive
effect on enhancing duplex thermal stability beyond that
EXPERIMENTAL SECTION
■
General Procedure. All nonaqueous reactions were run in oven-
(120 °C) or flame-dried glassware under a positive pressure of argon,
with exclusion of moisture from reagents and glassware, using standard
techniques for manipulating air-sensitive compounds, unless otherwise
stated. Anhydrous tetrahydrofuran, diethyl ether, toluene, and
dichloromethane were obtained by passing these solvents through
activated columns of alumina, while all other solvents were used as
received from chemical suppliers. Reagents were purchased and used
without further purification. Yields refer to chromatographically and
spectroscopically (1H NMR) homogeneous material, unless otherwise
stated. Reactions were monitored by thin layer chromatography
(TLC) carried out on 0.25 mm silica plates that were visualized using a
UV lamp (254 nm) and developed with iodine vapor, an aqueous
solution of cerium ammonium molybdate, or an ethanolic solution of
p-anisaldehyde. Flash chromatography20 was performed using 40−63
μm (230−400 mesh) silica gel, and all column dimensions are
reported as height × diameter in centimeters. NMR spectra were
E
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recorded at 300 or 400 MHz, calibrated using residual undeuterated
solvent as an internal reference (CHCl3, δ = 7.26 ppm; CHD2OD =
3.31 ppm), and reported in parts per million relative to trimethylsilane
(TMS δ = 0.00 ppm) as follows: chemical shift (multiplicity, coupling
constant (Hz), integration). The following abbreviations were used to
explain multiplicities: s = singlet, d = doublet, t = triplet, q = quartet, m
= multiplet, br = broad, dd = doublet of doublets, dt = doublet of
triplets. High-resolution mass spectra (HRMS) were recorded on a
TOF mass spectrometer using electrospray ionization time-of-flight
reflectron experiments. Melting points are given as ranges and are
reported in °C. Specific rotation measurements are reported in units of
deg· cm3·g−1·dm−1.
colorless oil: Rf = 0.27 (1:9 EtOAc/hexanes), [α]20 +15 (c 0.16,
CHCl3); H NMR (400 MHz, CDCl3) δ 7.88−7.81 (m, 4H), 7.67−
D
1
7.64 (m, 2H), 7.62−7.57 (m, 2H), 7.53−7.31 (m, 9H), 5.88 (d, J = 4.0
Hz, 1H), 5.83 (dd, J = 7.7, 3.0 Hz, 1H), 5.81−5.68 (m, 1H), 5.02−
4.91 (m, 3H), 4.80−4.76 (m, 1H), 4.65 (d, J = 11.9 Hz, 1H), 4.50 (d, J
= 5.4 Hz, 1H), 3.87 (d, J = 11.0 Hz, 1H), 3.80 (d, J = 11.0 Hz, 1H)
2.89−2.83 (m, 1H), 2.34−2.27 (m, 1H), 1.73 (s, 3H), 1.72 (s, 3H)
1.41 (s, 3H), 0.98 (s, 9H); 13C NMR (100 MHz, CDCl3) δ 169.8,
135.8, 135.73, 135.71, 135.3, 135.2, 133.4, 133.25, 133.23, 133.1,
129.97, 129.93, 128.3, 128.1, 127.92, 127.90, 127.88, 127.0, 126.3,
126.14, 126.06, 117.0, 114.0, 105.0, 87.9, 79.5, 77.9, 73.0, 72.0, 64.3,
35.4, 26.9, 26.6, 21.1, 19.3; HRMS (ESI) calcd for C41H48O7SiNa [M
+ Na]+ m/z = 703.3062, found 703.3068.
1,2-O-Isopropylidene-3-O-(2-naphthylmethyl)-4-C-(tert-
butyldiphenylsilyl)oxymethyl-6-deoxy-6-C-vinyl-β-L-manno-
furanose (13). PCC (5.41 g, 25.2 mmol) was added to a stirred
solution of 12 (5.02 g, 8.39 mmol) in dichloromethane (60 mL). The
reaction was heated to 40 °C for 13 h, cooled to room temperature,
and diluted with diethyl ether (60 mL). Silica gel (ca. 6 g) was added,
and the resulting slurry was filtered through a short pad of silica gel (4
cm), which was subsequently washed with diethyl ether (100 mL).
The filtrate was evaporated under reduced pressure, and the residue
was purified by flash chromatography (20 × 5 cm; 1:9 EtOAc/
hexanes) to give the corresponding aldehyde (34) as a white solid
1-(2,5-O-Diacetyl-3-O-(2-naphthylmethyl)-4-C-(tert-
butyldiphenylsilyl)oxymethyl-6-deoxy-6-C-vinyl-α-L-
mannofuranosyl)uracil (15). A mixture of 14 (3.42 g, 5.02 mmol)
and 4:1 AcOH/water (25 mL) was heated to 90 °C. After 3 h, the
reaction mixture was poured directly into ice−water (200 mL), and
the resulting solution was extracted with dichloromethane (3 × 40
mL). The combined organic extracts were washed with water (100
mL) and a saturated solution of NaHCO3 (100 mL), dried over
MgSO4, filtered, and concentrated under reduced pressure. The
residue was then dissolved in pyridine (20 mL), and acetic anhydride
(2.60 g, 25.1 mmol) was added. After 12 h, the reaction mixture was
diluted with ethyl acetate (50 mL), and the resulting solution was
washed with 1 M HCl (3 × 100 mL). The aqueous phases were
combined and extracted with EtOAc (2 × 40 mL). The combined
organic extracts were then washed with 1 M HCl (100 mL) and a
saturated solution of NaHCO3 (2 × 100 mL), dried over MgSO4,
filtered, and concentrated under reduced pressure. The residue was
purified by flash chromatography (20 × 4 cm; 10% to 15% EtOAc/
hexanes) to give the peracetylated product (35) as a light yellow oil
(2.91 g, 80% over two steps). A stirred slurry of uracil (0.901 g, 8.02
mmol) and N,O-bis(trimethylsilyl)acetamide (4.08, 20.1 mmol) in
acetonitrile (20 mL) was heated to 90 °C. After 1 h, the resulting
mixture was cooled to 0 °C, and a solution of the preceding product
(34, 2.91 g, 4.01 mmol) in acetonitrile (15 mL), followed by TMSOTf
(1.57, 6.82 mmol), was added. The cooling bath was removed, and the
reaction mixture was heated to 50 °C. After 16 h, the reaction mixture
was poured into an ice-cold solution of saturated NaHCO3 (40 mL)
and extracted with dichloromethane (3 × 30 mL). The combined
organic extracts were washed with brine (50 mL), dried over MgSO4,
filtered, and concentrated under reduced pressure. The residue was
purified by flash chromatography (20 × 4 cm; 1:2 EtOAc/hexanes) to
give nucleoside 15 as a white solid (2.82 g, 91%): Rf = 0.25 (1:2
(4.73 g, 95%): Rf = 0.32 (1:9 EtOAc/hexanes); [α]20 +5.4 (c 0.25,
D
CHCl3); 1H NMR (400 MHz, CDCl3) δ 9.93 (s, 1H), 7.84−7.75 (m,
4H), 7.60−7.53 (m, 4H), 7.51−7.45 (m, 3H), 7.43−7.35 (m, 2H),
7.35−7.28 (m, 4H), 5.85 (d, J = 3.3 Hz, 1H), 4.90 (d, J = 12.3 Hz,
1H), 4.78 (d, J = 12.3 Hz, 1H), 4.67−4.65 (m, 1H), 4.57 (d, J = 4.4
Hz, 1H), 3.89 (d, J = 11.4 Hz, 1H), 3.85 (d, J = 11.4 Hz, 1H), 1.64 (s,
3H), 1.37 (s, 3H), 0.91 (s, 9H); 13C NMR (100 MHz, CDCl3) δ
200.5, 135.8, 135.7, 134.6, 133.5, 133.4, 133.0, 132.8, 130.1, 130.0,
128.7, 128.1, 127.99, 127.95, 127.2, 126.5, 126.4, 125.9, 114.4, 105.1,
90.9, 79.2, 78.8, 73.2, 63.3, 26.9, 26.4, 19.4; HRMS (ESI) calcd for
C36H40NaO6Si [M + Na]+ m/z = 619.2486, found 619.2500. Boron
trifluoride diethyl etherate (1.42 g, 10.0 mmol) was added to a stirred
−41 °C solution of the preceding aldehyde (34, 3.01 g, 5.05 mmol) in
dichloromethane (45 mL). After 5 min, allyltrimethylsilane (1.20 mL,
7.6 mmol) was added. The reaction was kept at −41 °C for an
additional 2 h and then poured into a 0 °C saturated solution of
NaHCO3. The layers were separated, and the aqueous phase was
extracted with dichloromethane (2 × 40 mL). The combined organic
extracts were washed with a saturated solution of NaHCO3 (50 mL),
dried over MgSO4, filtered, and concentrated under reduced pressure.
The residue was purified by column chromatography (20 × 4 cm; 8%
EtOAc/hexanes) to give 13 as a white foam (2.83 g, 88%): Rf = 0.31
EtOAc/hexanes); mp 82−84 °C (CHCl3); [α]20 +29 (c 0.32,
1
(1:9 EtOAc/hexanes); [α]20 +9.9 (c 0.15, CHCl3); H NMR (400
D
D
CHCl3); 1H NMR (400 MHz, CDCl3) δ 9.17 (s, 1H), 7.88−7.75 (m,
3H), 7.74−7.55 (m, 6H), 7.54−7.29 (m, 10H), 6.29 (d, J = 5.7 Hz,
1H), 5.70−5.58 (m, 1H), 5.56−5.42 (m, 2H), 5.37 (d, J = 7.8 Hz,
1H), 5.01−4.87 (m, 2H), 4.69 (d, J = 11.1 Hz, 1H), 4.58−4.45 (m,
2H), 3.96 (d, J = 11.3 Hz, 1H), 3.86 (d, J = 11.3 Hz, 1H), 2.71−2.65
(m, 1H), 2.28−2.14 (m, 1H), 2.09 (s, 3H), 1.64 (s, 3H), 1.10 (s, 9H);
13C NMR (100 MHz, CDCl3) δ 170.4, 169.5, 163.2, 150.5, 139.8,
MHz, CDCl3) δ 7.86−7.76 (m, 4H), 7.61−7.55 (m, 2H), 7.55−7.46
(m, 5H), 7.43−7.35 (m, 2H), 7.34−7.25 (m, 4H), 5.89−5.77 (m, 2H),
5.07−4.95 (m, 2H), 4.77−4.65 (m, 3H), 4.46 (td, J1 = 11.0, 2.1 Hz,
1H), 3.95 (d, J = 11.2 Hz, 1H), 3.79 (d, J = 11.2 Hz, 1H), 3.33 (d, J =
2.2 Hz, 1H), 2.51 (dd, J1 = 14.4, 6.7 Hz, 1H), 1.95−1.82 (m, 1H), 1.63
(s, 3H), 1.38 (s, 3H), 0.90 (s, 9H); 13C NMR (100 MHz, CDCl3) δ
136.6, 135.8, 135.7, 134.5, 133.5, 133.4, 133.3, 133.2, 129.94, 129.85,
128.9, 128.2, 127.97, 127.95, 127.93, 127.88, 127.8, 127.5, 126.6,
126.5, 125.9, 116.4, 114.0, 104.9, 88.4, 79.4, 78.3, 73.3, 72.8, 62.7, 34.8,
27.3, 26.9, 26.7, 19.3; HRMS (ESI) calcd for C39H46NaO6Si [M +
Na]+ m/z = 661.2956, found 661.2970.
1,2-O-Isopropylidene-3-O-(2-naphthylmethyl)-4-C-(tert-
butyldiphenylsilyl)oxymethyl-5-O-acetyl-6-deoxy-6-C-vinyl-β-
L-mannofuranose (14). Acetic anhydride (2.86 g, 28.1 mmol) was
added to a stirred solution of 13 (3.82 g, 5.98 mmol) in pyridine (12
mL), and the resulting mixture was heated to 110 °C. After 24 h, the
reaction mixture was cooled to room temperature and diluted with
EtOAc (50 mL), and the resulting solution was washed with 1 M HCl
(4 × 100 mL). The combined aqueous phases were extracted with
EtOAc (2 × 50 mL), and the organic extracts were combined, washed
with 1 M HCl (100 mL) and a saturated solution of NaHCO3 (2 ×
100 mL), dried over MgSO4, filtered, and concentrated under reduced
pressure. The light brown residue was purified by flash chromatog-
raphy (20 × 4 cm; 1:9 EtOAc/hexanes) to give 14 (3.42 g, 84%) as a
135.9, 135.6, 134.5, 134.2, 133.34, 133.28, 132.7, 131.9, 130.52,
130.45, 128.5, 128.3, 128.1, 127.9, 127.0, 126.5, 126.4, 125.9, 117.8,
103.2, 88.5, 86.6, 78.0, 77.4, 75.3, 75.0, 72.0, 64.7, 35.0, 27.23, 20.9,
19.5; HRMS (ESI) calcd for C44H48N2O9SiNa [M + Na]+ m/z =
799.3021, found 799.3018.
1-(2-O-Methanesulfonyl-3-O-(2-naphthylmethyl)-4-C-(tert-
butyldiphenylsilyl)oxymethyl-5-O-acetyl-6-deoxy-6-C-vinyl-α-
L-mannofuranosyl)uracil (16). Potassium carbonate (0.348 g, 2.52
mmol) was added to a stirred solution of 15 (1.86 g, 2.40 mmol) in
methanol (40 mL). After 1 h, the reaction mixture was poured into
water (40 mL) and further diluted with dichloromethane (30 mL).
The layers were separated, and the aqueous phase was extracted with
dichloromethane (2 × 30 mL). The combined organic extracts were
washed with brine (40 mL), dried over MgSO4, filtered, and
concentrated under reduced pressure to give the corresponding
alcohol (36) as a colorless oil (1.70 g, 97%). This material was used in
the next step without further purification: Rf = 0.16 (1:2 EtOAc/
F
dx.doi.org/10.1021/jo401170y | J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
hexanes); [α]20D +32 (c 0.22, CHCl3); 1H NMR (400 MHz, CDCl3) δ
9.82 (s, 1H), 7.87−7.81 (m, 4H), 7.64−7.60 (m, 4H), 7.50−7.31 (m,
10 H), 6.09 (d, J = 5.6 Hz, 1H), 5.74−5.63 (m, 1H) 5.55 (dd, J = 9.8,
2.6 Hz, 1H), 5.40 (dd, J = 8.0, 1.6 Hz, 1H), 4.99−4.93 (m, 2H), 4.88
(d, J = 11.4 Hz, 1H) 4.67 (d, J = 11.4 Hz, 1H), 4.46−4.41 (m, 1H),
4.36 (d, J = 6.1 Hz, 1H), 3.94−3.91 (m, 2H), 3.84 (d, J = 11.2 Hz,
1H), 2.77−2.68 (m, 1H), 2.34−2.23 (m, 1H), 1.75 (s, 3H), 1.09 (s,
9H); 13C NMR (100 MHz, CDCl3) δ 170.0, 140.2, 135.9, 135.6,
134.2, 133.9, 133.44, 133.36, 132.7, 132.0, 130.53, 130.47, 128.9,
128.3, 128.2, 128.0, 127.6, 126.65, 126.61, 126.1, 117.9, 103.0, 90.0,
88.3, 79.2, 77.4, 75.5, 75.2, 72.5, 65.1, 35.0, 29.9, 27.2, 21.0, 19.5;
HRMS (ESI) calcd for C42H46N2O8SiNa [M + Na]+ m/z = 757.2916,
found 757.2921. Methanesulfonyl chloride (0.357 g, 3.12 mmol) was
added to a stirred solution of the preceding alcohol (36) in pyridine
(20 mL). After 2 h, the reaction was diluted with dichloromethane (40
mL) and washed with 1 M HCl (3 × 50 mL). The aqueous phases
were combined and extracted with dichloromethane (30 mL). The
combined organic extracts were washed with a saturated solution of
NaHCO3 (2 × 50 mL), dried over MgSO4, filtered, and concentrated
under reduced pressure. The residue was purified by flash
chromatography (18 × 3 cm; 2:3 EtOAc/hexanes) to give nucleoside
(100 MHz, CDCl3) δ 173.1, 170.1, 160.2, 135.6, 134.4, 134.2, 133.2,
133.1, 128.4, 128.1, 127.8, 127.0, 126.4, 126.3, 125.8, 117.6, 109.5,
92.9, 89.7, 88.0, 85.2, 73.6, 72.5, 62.6, 35.8, 21.1; HRMS (ESI) calcd
for C26H27N2O7 [M + H]+ m/z = 479.1758, found 479.1756.
1-((1S,3R,4S,7S)-1-((S)-1-Hydroxybut-3-en-1-yl)-7-(naphtha-
len-2-ylmethoxy)-2,5-dioxabicyclo[2.2.1]heptan-3-yl)-
pyrimidine-2,4(1H,3H)-dione (18). Methanesulfonyl chloride
(0.171 g, 1.06 mmol) was added to a stirred solution of nucleoside
17 (0.410 g, 0.709 mmol) in pyridine. After 2 h, dichloromethane was
added, and the resulting solution was washed with 1 M HCl (3 × 50
mL). The aqueous phases were combined and extracted with
dichloromethane (2 × 30 mL). The combined organic extracts were
washed with a saturated solution of NaHCO3 (2 × 50 mL), dried over
MgSO4, filtered, and concentrated under reduced pressure to give the
corresponding mesylate (38, 0.464 g) as a light yellow oil, which was
used in the next step without purification. A 2 M solution of NaOH
(0.90 mL, 1.8 mmol) was added to a stirred 80 °C solution of the
preceding mesylate (38, 0.464 g, 0.709 mmol) in 1:1 water/EtOH (30
mL). After 1 h, the reaction was cooled and diluted with water (30
mL), and the resulting solution was extracted with dichloromethane (3
× 30 mL). The combined organic extracts were washed with a
saturated solution of NaHCO3 (50 mL), dried over MgSO4, filtered,
and concentrated under reduced pressure. The residue was purified by
flash chromatography (15 × 3 cm; 3:2 EtOAc/hexanes) to give
16 as a colorless oil (1.76 g, 93%): Rf = 0.29 (2:3 EtOAc/hexanes);
1
[α]20 +41 (c 0.30, CHCl3); H NMR (400 MHz, CDCl3) δ 9.09 (s,
D
1H), 7.88 (d, J = 7.3 Hz, 2H), 7.81 (s, 1H), 7.72 (d, J = 8.0 Hz, 1H),
7.68−7.59 (m, 2H), 7.58−7.34 (m, 6H), 6.34 (d, J = 6.2 Hz, 1H),
5.52−5.45 (m, 2H), 5.12−4.81 (m, 2H), 3.97 (d, J = 11.4 Hz, 1H),
3.86 (d, J = 11.4 Hz, 1H), 3.04 (s, 2H), 2.72−2.65 (m, 2H), 2.28−2.19
(m, 2H), 1.75 (s, 3H), 1.29 (s, 4H). 13C NMR (100 MHz, CDCl3) δ
169.3, 163.0, 150.6, 139.6, 135.9, 135.6, 134.2, 134.1, 133.3, 132.4,
131.7, 130.60, 130.55, 128.6, 128.4, 128.2, 127.9, 127.5, 126.5, 126.4,
126.1, 117.8, 103.5, 88.9, 86.2, 77.94, 77.89, 77.3, 75.2, 71.9, 65.3, 38.8,
35.0, 30.0, 27.2, 20.9, 19.5; C43H48N2O10SSiNa [M + Na]+ m/z =
835.2691, found 835.2702.
(S)-1-((2S,3S,3aS,9aR)-2-(Hydroxymethyl)-3-(naphthalen-2-
ylmethoxy)-6-oxo-2,3,3a,9a-tetrahydro-6H-furo[2′,3′:4,5]-
oxazolo[3,2-a]pyrimidin-2-yl)but-3-en-1-yl Acetate (17). DBU
(0.192 g, 1.26 mmol) was added to a stirred solution of 16 (0.893 g,
1.10 mmol) in acetonitrile (25 mL). After 3 h, the reaction was poured
into 1 M HCl (30 mL) and further diluted with dichloromethane (30
mL). The layers were separated, and the aqueous phase was extracted
with dichloromethane (2 × 30 mL). The combined organic extracts
were washed with a saturated solution of NaHCO3 (30 mL), dried
over MgSO4, filtered, and concentrated under reduced pressure to give
the corresponding 2,2′-anhydronucleoside (37), which was used in the
next step without further purification: Rf = 0.31 (3% MeOH/CH2Cl2);
1H NMR (400 MHz, CDCl3) δ 7.85−7.79 (m, 3H), 7.58−7.24 (m,
14H), 6.13 (d, J = 6.4 Hz, 1H), 5.99 (d, J = 7.5 Hz, 1H), 5.73−5.62
(m, 1H), 5.49−5.37 (m, 2H), 5.14−4.93 (m, 3H), 4.81−4.70 (m, 2H),
2.54−2.6 (m, 2H), 1.96 (s, 3H), 0.85 (s, 9H); 13C NMR (100 MHz,
CDCl3) δ 171.5, 169.9, 159.3, 135.64, 135.60, 134.3, 133.9, 133.6,
133.43, 133.37, 132.4, 130.2, 128.9, 128.3, 128.1, 128.0, 127.9, 127.6,
126.7, 126.6, 125.7, 118.2, 110.8, 91.3, 88.7, 87.5, 84.0, 77.4, 73.8, 73.4,
63.5, 35.8, 26.7, 21.2, 19.1; HRMS (ESI) calcd for C42H44N2O7SiNa
[M + Na]+ m/z = 739.2810, found 739.2801. A 1.0 M solution of
TBAF (2.5 mL, 2.5 mmol) in THF was added to a stirred solution of
acetic acid (2 mL) and the above 2,2′-anhydronucleoside (37, 1.26
mmol) in THF (10 mL). After 3 d, the reaction was diluted with
dichloromethane (20 mL) and poured into water (20 mL). The layers
were separated, and the aqueous phase was extracted with dichloro-
methane (2 × 10 mL). The combined organic extracts were washed
with a saturated solution of NaHCO3 (20 mL) and brine (20 mL),
dried over MgSO4, filtered, and concentrated under reduced pressure.
The residue was purified by flash chromatography (18 × 2 cm; 5%
MeOH/CH2Cl2) to give nucleoside 17 as a colorless oil (0.536 g, 74%
over two steps): Rf = 0.20 (5% MeOH/CH2Cl2); 1H NMR (400 MHz,
CDCl3) δ 7.83−7.77 (m, 4H), 7.51−7.38 (m, 3H), 7.24 (d, J = 6.2 Hz,
1H), 6.10 (d, J = 6.2 Hz, 1H), 5.95 (d, J = 7.1 Hz, 1H), 5.78−5.62 (m,
1H), 5.38−5.31 (m, 1H), 5.22−5.17 (m, 1H), 5.03−4.92 (m, 2H),
4.87−4.76 (m, 3H), 3.69, (d, J = 11.9 Hz, 1H), 3.52 (d, J = 11.9 Hz,
1H), 2.61−2.50 (m, 1H), 2.42−2.30 (m, 1H), 1.93 (s, 3H); 13C NMR
1
nucleoside 18 as white foam: Rf = 0.31 (3:2 EtOAc/hexanes); H
NMR (400 MHz, CDCl3) δ 9.13 (s, 1H), 7.89−7.81 (m, 4H), 7.65 (d,
J = 8.2 Hz, 1H), 7.52−7.48 (m, 3H), 6.27 (s, 1H), 5.92−5.81 (m, 1H),
5.74 (dd, J = 8.1, 1.6 Hz, 1H), 5.22 (s, 1H), 5.19−5.16 (m, 1H), 4.92
(d, J = 12.1 Hz, 1H), 4.85, (d, J = 12.0 Hz, 1H), 4.50 (dd, J = 2.4, 0.9
Hz, 1H), 4.39, (d, J = 2.4 Hz, 1H), 4.14−4.09 (m, 2H), 3.89 (d, J = 8.8
Hz, 1H), 2.67−2.59 (m, 1H), 2.22−2.11 (m, 1H); 13C NMR (100
MHz, CDCl3) δ 163.6, 150.4, 140.3, 134.8, 134.3, 133.44, 133.39,
128.9, 128.2, 128.0, 127.4, 126.53, 126.48, 126.0, 117.9, 101.4, 91.2,
90.5, 81.7, 75.5, 73.4, 71.9, 67.1, 38.4, 26.0, 18.2, −3.7, −4.7; HRMS
(ESI) calcd for C24H24N2O6Na [M + Na]+ m/z = 459.1527, found
459.1532.
1-((1R,3R,4S,7S)-1-((S)-1-((tert-Butyldimethylsilyl)oxy)but-3-
en-1-yl)-7-hydroxy-2,5-dioxabicyclo[2.2.1]heptan-3-yl)-
pyrimidine-2,4(1H,3H)-dione (19). TBSOTf (0.740 mmol) was
added to a stirred 0 °C solution of pyridine (0.058 g, 0.74 mmol) and
18 (0.215 g, 0.493 mmol) in dichloromethane (10 mL). The cooling
bath was removed, and the reaction was allowed to warm to room
temperature. After 12 h, 1 M HCl (20 mL) was added, and the layers
were separated. The aqueous phase was extracted with dichloro-
methane (2 × 10 mL). The combined organic extracts were washed
with a saturated solution of NaHCO3 (20 mL), dried over MgSO4,
filtered, and concentrated under reduced pressure. The residue was
purified by flash chromatography (18 × 2 cm; 2:3 EtOAc/hexanes) to
give the corresponding TBS ether (39) as a colorless oil: Rf = 0.29 (2:3
EtOAc/hexanes); 1H NMR (400 MHz, CDCl3) δ 9.08 (s, 1H), 7.89−
7.81 (m, 4H), 7.65 (d, J = 8.2 Hz, 1H), 7.52−7.48 (m, 3H), 6.18 (s,
1H), 5.95−5.84 (m, 1H), 5.74 (dd, J = 8.1, 1.6 Hz, 1H), 5.20−5.12
(m, 2H), 4.86 (d, J = 11.3 Hz, 1H), 4.80 (d, J = 11.3 Hz, 1H), 4.52
(dd, J = 2.2, 0.9 Hz, 1H), 4.35−4.31, (m, 2H), 4.15−4.11 (m, 2H),
3.99 (d, J = 8.8 Hz, 1H), 2.13−2.07 (m, 1H), 1.98−1.91 (m, 1H), 0.90
(s, 9H), 0.11 (s, 6H); 13C NMR (100 MHz, CDCl3) δ 163.6, 150.4,
134.8, 134.3, 133.44, 133.39, 128.9, 128.2, 128.0, 127.4, 126.53,
126.48, 126.0, 117.9, 101.4, 91.2, 90.5, 81.7, 75.5, 73.4, 71.9, 67.2, 38.4,
26.0, 18.2, −3.7, −4.7; HRMS (ESI) calcd for C30H38N2O6SiNa [M +
Na]+ m/z = 573.2391, found 573.2399. DDQ (0.177 g, 0.781 mmol)
was added to a vigorously stirred solution of the preceding TBS ether
(39, 0.195 g, 0.355 mmol) in 9:1 dichloromethane/water (8 mL).
After 48 h, the reaction mixture was poured into a 10% solution of
NaHSO3 (20 mL), and the layers were separated. The aqueous phase
was extracted with dichloromethane (2 × 10 mL). The combined
organic extracts were washed with a saturated solution of sodium
bicarbonate (20 mL), dried over MgSO4, filtered, and concentrated
under reduced pressure. The residue was purified by flash
chromatography (20 × 2 cm; 3:2 EtOAc/hexanes) to give nucleoside
19 as a white solid (0.120 g, 83%): Rf = 0.32 (3:2 EtOAc/hexanes);
G
dx.doi.org/10.1021/jo401170y | J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
mp 81−83 °C (CHCl3); 1H NMR (400 MHz, CDCl3) δ 9.84 (s, 1H),
7.66 (d, J = 8.1 Hz, 1H), 6.24 (s, 1H), 6.02−5.77 (m, 1H), 5.74 (d, J =
7.8 Hz, 1H), 5.22−5.06 (m, 2H), 4.53−4.48 (m, 2H), 4.25 (dd, J =
14.5, 9.4 Hz, 1H), 4.18 (br s, 1H), 4.08 (d, J = 8.7 Hz, 1H), 3.90 (d, J
= 8.7 Hz, 1H), 2.55−2.42 (m, 1H), 2.39−2.24 (m, 1H), 0.90 (s, 9H),
0.14 (s, 3H), 0.11 (s, 3H); 13C NMR (100 MHz, CDCl3) δ 164.0,
150.8, 140.4, 134.3, 118.2, 101.5, 91.0, 90.8, 77.6, 75.4, 72.4, 68.3, 38.6,
26.0, 18.2, −4.2, −4.7; HRMS (ESI) calcd for C19H30N2O6SiNa [M +
Na]+ m/z = 433.1765, found 433.1758.
1H), 4.42 (d, J = 4.2 Hz, 1H), 3.92 (dd, J = 8.0, 4.2 Hz, 1H), 3.85 (d, J
= 10.4 Hz, 1H), 3.82 (d, J = 10.4 Hz, 1H), 2.39−2.22 (m, 2H), 1.55 (s,
3H), 1.38 (s, 3H), 0.99 (s, 9H), 0.69 (s, 9H), −0.09 (s, 3H), −0.25 (s,
3H); 13C NMR (100 MHz, CDCl3) δ 209.6, 135.8, 135.7, 132.5,
132.3, 130.14, 130.09, 128.1, 117.6, 115.3, 103.1, 93.3, 78.1, 73.9, 68.1,
38.5, 27.1, 27.0, 26.9, 26.0, 19.3, 18.2, −3.3, −4.6; HRMS (ESI) calcd
for C34H50NaO6Si2 [M + Na]+ m/z = 633.3038, found 633.3059.
(3aR,4aS,5S,8aR,8bR)-5-((tert-Butyldimethylsilyl)oxy)-4a-
(((tert-butyldiphenylsilyl)oxy)methyl)-8a-hydroxy-2,2-
dimethylhexahydro[1,3]dioxolo[4,5-b]benzofuran-7(4aH)-one
(23). PdCl2 (0.065 g, 0.37 mmol) and Cu(OAc)2 (0.134 g, 0.738
mmol) were added to a stirred 40 °C solution of 22 (2.23 g, 3.66
mmol) in 7:1 N,N-dimethylacetamide/water (45 mL). The reaction
mixture was purged with and maintained under an atmosphere of
oxygen gas (via an oxygen-filled balloon) for 24 h, at which point it
was filtered through a pad of Celite and washed with diethyl ether
(100 mL). The filtrate was then washed with a saturated solution of
NaHCO3 (200 mL), the layers were separated, and the aqueous phase
was extracted with diethyl ether (3 × 50 mL). The combined organic
extracts were dried over MgSO4, filtered, and concentrated under
reduced pressure. The residue was purified by flash chromatography
(20 × 3 cm; 5% EtOAc/hexanes) to give the corresponding ketone
(41) as a light yellow oil (2.18 g, 95%): Rf = 0.52 (10% EtOAc/
hexanes); [α]20D +27 (c 0.090, CHCl3); 1H NMR (400 MHz, CDCl3)
δ 7.67−7.63 (m, 2H), 7.60−7.56 (m, 2H), 7.46−7.33 (m, 6H), 6.21
(d, J = 4.2 Hz, 1H), 4.56 (dd, J = 7.2, 2.8 Hz, 1H), 4.41 (d, J = 4.2 Hz,
1H), 3.98 (d, J = 10.5 Hz, 1H), 3.74 (d, J = 10.5 Hz, 1H), 2.73 (dd, J =
18.0, 2.8 Hz, 1H), 2.56 (dd, J = 18.0, 7.5 Hz, 1H), 2.08 (s, 3H), 1.59
(s, 3H), 1.38 (s, 3H), 0.99 (s, 9H), 0.68 (s, 9H), −0.09 (s, 3H), −0.17
(s, 3H); 13C NMR (100 MHz, CDCl3) δ 208.3, 205.7, 135.8, 135.6,
132.6, 132.2, 130.2, 130.10, 128.1, 115.3, 102.8, 91.4, 78.2, 69.5, 66.7,
48.0, 27.2, 26.9, 26.8, 26.1, 19.2, 18.2, −3.3, −4.6; HRMS (ESI) calcd
for C34H50NaO7Si2 [M + Na]+ m/z = 649.2987, found 649.3001. L-
Proline (0.116 g, 1.0 mmol) was added to a stirred 55 °C solution of
the preceding ketone (41, 2.11 g, 3.35 mmol) in DMSO (90 mL).
After 54 h, the reaction mixture was cooled to room temperature and
poured into a saturated solution of NaHCO3 (200 mL), and the
resulting mixture was extracted with EtOAc (4 × 100 mL). The
combined organic extracts were washed with water (2 × 200 mL) and
brine (200 mL), dried over MgSO4, filtered, and concentrated under
reduced pressure. The residue was purified by flash chromatography
(22 × 3 cm; 10% EtOAc/hexanes) to give the preceding ketone
starting material (41, 0.155 g, 7%) and 23 (1.72 g, 82%) as a colorless
1,2-O-Isopropylidene-3-O-(2-naphthylmethyl)-4-C-(tert-
butyldiphenylsilyl)oxymethyl-5-O-(tert-butyldimethylsilyl)-6-
deoxy-6-C-vinyl-β-L-mannofuranose (21). Pyridine (0.982 g, 12.4
mmol) and TBSOTf (1.61 g, 6.10 mmol) were added to a stirred 0 °C
solution of 13 (2.60 g, 4.08 mmol) in dichloromethane (50 mL). The
cooling bath was removed, and the reaction was allowed to warm to
room temperature. After 3 h, the reaction mixture was poured into a
saturated solution of NaHCO3 (70 mL), and the layers were separated.
The aqueous phase was extracted with dichloromethane (2 × 50 mL),
and the organic extracts were combined and washed with brine (50
mL), dried over MgSO4, filtered, and concentrated under reduced
pressure. The residue was purified by flash chromatography (20 × 4
cm; 5% EtOAc/hexanes) to give 21 as a colorless oil (3.01 g, 98%): Rf
= 0.56 (10% EtOAc/hexanes); [α]20D +7.7 (c 0.11, CHCl3); 1H NMR
(400 MHz, CDCl3) δ 7.87−7.79 (m, 4H), 7.66−7.58 (m, 4H), 7.57−
7.53 (m, 1H), 7.52−7.46 (m, 2H), 7.45−7.39 (m, 2H), 7.38−7.32 (m,
4H), 6.02 (d, J = 4.1 Hz, 1H), 5.91−5.78 (m, 1H), 5.06 (d, J = 12.0
Hz, 1H), 5.04 (br d, J = 16.8 Hz, 1H), 4.90 (br d, J = 10.1 Hz, 1H),
4.85 (dd, J = 5.2 Hz, 4.4 Hz, 1H), 4.67 (d, J = 12.0 Hz, 1H), 4.56 (dd,
J = 5.6 Hz, 4.1 Hz, 1H), 4.38 (d, J = 5.6 Hz, 1H), 3.92 (d, J = 11.1 Hz,
1H), 3.79 (d, J = 11.1 Hz, 1H), 2.60−2.68 (m, 1H), 2.38−2.49 (m,
1H), 1.62 (s, 3H), 1.41 (s, 3H), 0.96 (s, 9H), 0.70 (s, 9H), −0.07 (s,
3H), −0.16 (s, 3H); 13C NMR (100 MHz, CDCl3) δ 136.8, 135.8,
135.7, 133.4, 133.3, 133.19, 133.15, 129.9, 129.8, 128.2, 128.1, 127.94,
127.90, 126.9, 126.3, 126.0, 116.7, 113.9, 105.6, 91.2, 81.0, 78.1, 77.4,
72.5, 72.0, 66.6, 38.6, 27.4, 27.3, 27.0, 26.1, 25.9, 19.3, 18.3, −3.5,
−4.6; HRMS (ESI) calcd for C45H60NaO6Si2 [M + Na]+ m/z =
775.3821, found 775.3821.
1,2-O-Isopropylidene-3-keto-4-C-(tert-butyldiphenylsilyl)-
oxymethyl-5-O-(tert-butyldimethylsilyl)-6-deoxy-6-C-vinyl-β-L-
mannofuranose (22). DDQ (1.81 g, 7.97 mmol) was added and to a
vigorously stirred solution of 21 (3.02 g, 4.01 mmol) in 9:1
dichloromethane/water (70 mL). After 6 h, the reaction mixture
was poured into a saturated solution of NaHCO3 (100 mL), and the
layers were separated. The aqueous phase was extracted with
dichloromethane (3 × 50 mL), and the combined organic extracts
were dried over MgSO4, filtered, and concentrated under reduced
pressure. The residue was purified by flash chromatography (20 × 3
cm; 2% EtOAc/hexanes) to give the corresponding alcohol (40) as a
solid: Rf = 0.25 (1:9 EtOAc/hexanes); mp 40−43 °C; [α]20 +52 (c
D
1
0.12, CHCl3); H NMR (400 MHz, CDCl3) δ 7.70−7.66 (m, 2H),
7.64−7.60 (m, 2H), 7.48−7.36 (m, 6H), 6.19 (d, J = 3.9 Hz, 1H), 4.62
(s, 1H), 4.61 (d, J = 3.9 Hz, 1H), 4.07 (d, J = 10.8 Hz, 1H), 3.89 (dd, J
= 9.9, 3.9 Hz, 1H), 3.76 (d, J = 10.8 Hz, 1H), 2.70 (dd, J = 18.2, 9.9
Hz, 1H), 2.38 (d, J = 16.3 Hz, 1H), 2.34 (dd, J = 18.2, 3.8 Hz, 1H),
1.49 (s, 3H), 1.29 (s, 3H), 1.09 (s, 9H), 0.69 (s, 9H), −0.09 (s, 3H),
−0.18 (s, 3H); 13C NMR (100 MHz, CDCl3) δ 206.3, 135.7, 135.6,
131.6, 131.3, 130.6, 130.5, 128.4, 128.3, 114.6, 104.0, 91.2, 90.8, 80.7,
69.1, 68.0, 46.5, 43.0, 27.2, 26.7, 25.8, 19.2, 18.2, −4.4, −4.8; HRMS
(ESI) calcd for C34H50NaO7Si2 [M + Na]+ m/z = 649.2987, found
649.3002.
colorless oil (2.14 g, 88%): Rf = 0.62 (5% EtOAc/hexanes); [α]20
D
+39 (c 0.070, CHCl3); 1H NMR (400 MHz, CDCl3) δ 7.68−7.62 (m,
4H) 7.46−7.34 (m, 6H), 6.14 (d, J = 4.5 Hz, 1H), 5.85−5.70 (m, 1H),
5.00−4.87 (m, 2H), 4.91 (br d, J = 10.1 Hz, 1H), 4.30 (dd, J = 7.6 Hz,
3.4 Hz, 1H), 4.23 (dd, J = 6.4 Hz, 4.1 Hz, 1H), 3.91 (d, J = 10.9 Hz,
1H), 3.82 (d, J = 10.9 Hz, 1H), 2.92 (d, J = 4.0 Hz, 1H), 2.53−2.45
(m, 1H), 2.09−1.99 (m, 1H), 1.60 (s, 3H), 1.44 (s, 3H), 1.05 (s, 9H),
0.74 (s, 9H), 0.06 (s, 3H), 0.01 (s, 3H); 13C NMR (100 MHz, CDCl3)
δ 136.8 135.7, 132.8, 130.1, 130.0, 128.1, 116.7, 114.8, 106.4, 94.1,
82.4, 72.0, 70.6, 67.3, 37.8, 27.9, 27.4, 27.2, 26.2, 19.3, 18.4, −3.9,
−4.4; HRMS (ESI) calcd for C34H52NaO6Si2 [M + Na]+ m/z =
635.3195, found 635.3204. Dess−Martin periodinane (2.34 g, 5.52
mmol) was added to a stirred slurry of NaHCO3 (0.51 g, 5.9 mmol)
and the preceding alcohol (40, 2.23 g, 3.52 mmol) in dichloromethane
(55 mL). After 18 h, the reaction mixture was filtered through a pad of
silica gel (6 × 2.5 cm), which was washed with dichloromethane (50
mL). The filtrate was concentrated under reduced pressure, and the
residue was purified by flash chromatography (15 × 3 cm; 3% EtOAc/
hexanes) to give 22 as a colorless oil (2.18 g, 97%): Rf = 0.61 (5%
EtOAc/hexanes); [α]20D +25 (c 0.050, CHCl3); 1H NMR (400 MHz,
CDCl3) δ 7.66−7.61 (m, 2H), 7.58−7.53 (m, 2H), 7.45−7.32 (m,
6H), 6.29 (d, J = 4.2 Hz, 1H), 5.03−4.97 (m, 1H), 4.97−4.92 (m,
(3aR,4aS,5S,8aR,8bR)-5-((tert-Butyldimethylsilyl)oxy)-4a-
(((tert-butyldiphenylsilyl)oxy)methyl)-2,2-dimethylhexahydro-
[1,3]dioxolo[4,5-b]benzofuran-8a(4aH)-ol (24). Acetic acid (1.11
mL, 19.4 mmol) was added to a stirred 0 °C slurry of
tetramethylammonium triacetoxyborohydride (2.55 g, 9.69 mmol) in
MeCN (45 mL). After 15 min, a solution of 23 (1.21 g, 1.93 mmol) in
MeCN (80 mL) was added. The cooling bath was removed, and the
reaction was allowed to warm to room temperature. After 7 h, the
reaction mixture poured into water (200 mL), and the resulting
solution was extracted with dichloromethane (3 × 100 mL). The
combined organic extracts were washed with a saturated solution of
NaHCO3 (2 × 100 mL), dried over MgSO4, filtered, and concentrated
under reduced pressure. The residue was purified by flash
chromatography (3:7 EtOAc/hexanes) to give the corresponding
alcohol (42) as a white solid (1.13 g, 93%): Rf = 0.20 (3:7 EtOAc/
H
dx.doi.org/10.1021/jo401170y | J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
hexanes); mp 39−42 °C; [α]20D +33 (c 0.14, CHCl3); 1H NMR (400
MHz, CDCl3) δ 7.71−7.61 (m, 4H), 7.45−7.32 (m, 6H), 6.14 (d, J =
4.8 Hz, 1H), 4.85 (d, J = 4.8 Hz, 1H), 3.97−3.88 (m, 2H), 3.68 (d, J =
10.8 Hz, 1H), 3.57 (dd, J = 9.7 Hz, 3.6 Hz, 1H), 3.44 (s, 1H), 2.34−
2.28 (m, 1H), 2.11 (dd, J1 = 14.8 Hz, 6.0 Hz, 1H), 2.00−1.80 (m, 3H),
1.60 (s, 3H), 1.31 (s, 3H), 1.08 (s, 9H), 0.72 (s, 9H), −0.06 (s, 3H),
−0.17 (s, 3H); 13C NMR (100 MHz, CDCl3) δ 135.8, 135.7, 132.1,
131.9, 130.4, 130.2, 128.2, 114.2, 104.1, 90.8, 90.1, 80.9, 69.4, 68.2,
65.1, 39.66, 37.74, 27.1, 26.9, 26.6, 19.2, 18.2, −4.3, −4.6; HRMS
(ESIMS) calcd for C34H52NaO7Si2 [M + Na]+ m/z = 651.3144, found
651.3172. DMAP (0.024 g, 0.20 mmol) and 1,1′-thiocarbonyldiimi-
dazole (0.892 g, 4.51 mmol) were added to a stirred solution of the
preceding alcohol (42, 2.41 g, 3.83 mmol) in dichloromethane (125
mL). The reaction mixture was then heated to 40 °C for 19 h, cooled
to room temperature, and poured into a 0.5 M solution of HCl (200
mL). The layers were separated, and the aqueous phase was extracted
with dichloromethane (3 × 50 mL). The combined organic extracts
were washed with with a saturated solution of NaHCO3 (100 mL),
dried over MgSO4, filtered, and concentrated under reduced pressure.
The residue was purified by flash chromatography (3:7 EtOAc/
hexanes) to give the corresponding thioimidazole carbamate (43) as a
white solid (2.35 g, 83%): Rf = 0.23 (1:2 EtOAc/hexanes); mp 88−90
°C; [α]20D +23 (c 0.070, CHCl3); 1H NMR (400 MHz, CDCl3) δ 8.34
(s, 1H), 7.72−7.64 (m, 4H), 7.59 (s, 1H), 7.47−7.35 (m, 6H), 7.01 (s,
1H), 6.19 (d, J = 5.1 Hz, 1H), 5.74−5.64 (m, 1H), 4.99 (d, J = 5.1 Hz,
1H), 3.92 (d, J = 11.0 Hz, 1H), 3.73 (d, J = 11.0 Hz, 1H), 3.65 (dd, J =
8.6 Hz, 3.6 Hz, 1H), 3.56 (s, 1H), 2.33 (s, 1H), 2.30 (s, 1H), 2.20−
2.12 (m, 1H), 2.11−2.01 (m, 1H), 1.57 (s, 3H), 1.32 (s, 3H), 1.09 (s,
9H), 0.73 (s, 9H), −0.07 (s, 3H), −0.17 (s, 3H); 13C NMR (100
MHz, CDCl3) δ 183.5, 137.1, 135.8, 135.7, 132.1, 131.9, 130.8, 130.4,
130.3, 128.3, 118.0, 114.2, 104.3, 90.8, 89.8, 80.4, 77.4, 68.6, 68.1, 35.0,
34.5, 27.1, 26.6, 19.2, 18.2, −4.4, −4.7; HRMS (ESIMS): calcd for
C38H54N2O7SSi2 [M + H]+ = 739.3263, found 739.3274. A suspension
of Bu3SnH (6.49 mL, 24.5 mmol) and AIBN (0.050 mg, 0.31 mmol)
in toluene (10 mL) was added dropwise, via a syringe, to a stirred 110
°C solution of the preceding thioimidazole carbamate (43, 2.26 g, 3.10
mmol) in toluene (175 mL). After 15 min, from the start of the
addition, the reaction mixture was concentrated under reduced
pressure, and the residue was purified by flash chromatography (20
× 3 cm; 5−10% EtOAc/hexanes) to give 24 as a colorless oil (1.65 g,
hydride (60% dispersion in mineral oil, 0.101 g, 2.50 mmol), 2-
(bromomethyl)naphthalene (0.568 g, 2.57 mmol), and tetrabutylam-
monium iodide (0.432 g, 1.16 mmol) were added sequentially to a
stirred 0 °C solution of the preceding alcohol (44, 0.875 g, 2.34 mmol)
in 1:1 DMF/THF (32 mL). After 2 h, methanol (7 mL) was added,
and the resulting mixture was further diluted with dichloromethane
(30 mL) and water (30 mL). The layers were separated, and the
aqueous phase was extracted with dichloromethane (3 × 30 mL). The
combined organic extracts were washed with a saturated solution of
NaHCO3 (2 × 40 mL) and brine (40 mL), dried over MgSO4, filtered,
and concentrated under reduced pressure. The residue was purified by
flash chromatography (20 × 3 cm; 1:9 EtOAc/hexanes) to give 26 as a
colorless oil (1.15 g, 96%): Rf = 0.17 (1:9 EtOAc/hexanes); [α]20
D
+16 (c 0.050, CHCl3); 1H NMR (400 MHz, CDCl3) δ 7.87−7.79 (m,
3H), 7.74 (s, 1H), 7.52−7.45 (m, 2H), 7.42 (br d, J = 8.3 Hz, 1H),
5.96 (d, J = 4.9 Hz, 1H), 4.70 (s, 3H), 3.87−3.81 (m, 3H), 3.47 (s,
1H), 1.95−1.85 (m, 1H), 1.77−1.65 (m, 3H), 1.55−1.52 (m, 4H),
1.30 (s, 3H), 0.85 (s, 9H), 0.03 (s, 3H), 0.02 (s, 3H); 13C NMR (100
MHz, CDCl3) δ 134.5, 133.4, 133.3, 128.8, 128.1, 127.9, 127.2, 126.5,
125.7, 114.1, 103.8, 90.1, 89.9, 80.8, 74.4, 74.0, 70.7, 29.9, 29.7, 28.3,
26.8, 26.4, 26.1, 18.3, 18.2, −3.8, −4.6; HRMS (ESI) calcd for
C29H42NaO6Si [M + Na]+ m/z = 538.2672, found 538.2671.
(3aR,4aS,5S,8aR,8bR)-5,8a-Bis(benzyloxy)-2,2-dimethyl-4a-
((naphthalen-2-ylmethoxy)methyl)octahydro[1,3]dioxolo[4,5-
b]benzofuran (27). A 1.0 M solution of TBAF (5.5 mL, 5.5 mmol)
in THF was added to a stirred solution of 26 (0.948 g, 1.84 mmol) in
THF (40 mL). After 24 h, the reaction was diluted with
dichloromethane and (40 mL) and water (100 mL). The layers
were separated, and the aqueous phase was extracted with dichloro-
methane (2 × 30 mL). The combined organic extracts were washed
with brine (80 mL), dried over MgSO4, filtered, and concentrated
under reduced pressure. The residue was purified by flash
chromatography (20 × 3 cm; 2:3 EtOAc/hexanes) to give the
corresponding alcohol (45) as a light yellow solid (0.700 g, 95%): Rf =
0.27 (2:3 EtOAc/hexanes); mp 101−102 °C; [α]20 +33 (c 0.16,
D
CHCl3); 1H NMR (400 MHz, CDCl3) δ 7.85−7.80 (m, 3H), 7.73 (s,
1H), 7.51−7.46 (m, 2H), 7.41 (dd, J = 8.4 Hz, 1.5 Hz, 1H), 5.96 (d, J
= 4.5 Hz, 1H), 4.70 (s, 2H), 4.62 (d, J = 4.5 Hz, 1H), 3.77−3.70 (m,
3H), 3.52 (s, 1H), 2.63 (d, J = 5.1 Hz, 1H), 2.16−2.06 (m, 1H), 1.84−
1.65 (m, 3H), 1.62−1.44 (m, 5H), 1.29 (s, 3H); 13C NMR (100 MHz,
CDCl3) δ 134.6, 133.34, 133.27, 128.7, 128.1, 127.9, 126.5, 125.6,
114.0, 104.2, 90.5, 89.9, 80.5, 74.3, 73.9, 70.0, 31.3, 28.0, 26.3, 26.1,
16.9; HRMS (ESI) calcd for C23H28NaO6 [M + Na]+ m/z = 423.1778,
found 423.1770. Sodium hydride (60% dispersion in mineral oil, 0.251
g, 6.25 mmol), benzyl bromide (0.775 g, 4.53 mmol), and
tetrabutylammonium iodide (0.602 g, 1.63 mmol) were added
sequentially to a stirred 0 °C solution of the preceding alcohol (45,
0.652 g, 1.63 mmol) in 1:1 DMF/THF (40 mL). The cooling bath was
removed, and the reaction was allowed to warm to room temperature.
After 3 h, methanol (10 mL) was added, and the resulting solution was
diluted further with dichloromethane (40 mL) and water (40 mL).
The layers were separated, and the aqueous phase was extracted with
dichloromethane (2 × 30 mL). The combined organic extracts were
washed with a saturated solution of NaHCO3 (2 × 40 mL) and brine
(50 mL), dried over MgSO4, filtered, and concentrated under reduced
pressure. The residue was purified by flash chromatography (20 × 3
87%): Rf = 0.66 (15% EtOAc/hexanes); [α]20 +37 (c 0.10, CHCl3);
D
1H NMR (400 MHz, CDCl3) δ 7.70−7.62 (m, 4H), 7.45−7.35 (m,
6H), 6.13 (d, J = 5.0 Hz, 1H), 4.90 (d, J = 5.0 Hz, 1H), 4.06 (d, J =
10.9 Hz, 1H), 3.72 (d, J = 10.9 Hz, 1H), 3.60 (dd, J = 10.0 Hz, 5.2 Hz,
1H), 3.00 (s, 1H), 2.03−1.94 (m, 1H), 1.77−1.44 (m, 8H), 1.29 (s,
3H), 1.09 (s, 9H), 0.66 (s, 9H), −0.11 (s, 3H), −0.26 (s, 3H); 13C
NMR (100 MHz, CDCl3) δ 135.8, 135.7, 132.3, 132.2, 130.2, 130.1,
128.1, 113.7, 104.0, 92.0, 90.1, 80.6, 70.7, 68.6, 29.9, 28.4, 27.2, 26.8,
26.2, 19.2, 18.0, −3.9, −4.9; HRMS (ESI) calcd for C34H52NaO6Si2
[M + Na]+ m/z = 635.3195, found 635.3214.
(3aR,4aS,5S,8aR,8bR)-5-((tert-Butyldimethylsilyl)oxy)-2,2-di-
methyl-4a-((naphthalen-2-ylmethoxy)methyl)hexahydro[1,3]-
dioxolo[4,5-b]benzofuran-8a(4aH)-ol (26). A 1.0 M solution of
TBAF (3.23 mL, 3.23 mmol) in THF was added to a stirred 0 °C
solution of 24 (1.72 g, 2.81 mmol) in THF (50 mL). After 1 h,
dichloromethane (30 mL) and water (40 mL) were added. The layers
were separated, and the aqueous phase was extracted with dichloro-
methane (2 × 30 mL). The combined organic extracts were washed
with brine (50 mL), dried over MgSO4, filtered, and concentrated
under reduced pressure. The residue was purified by flash
chromatography (20 × 3 cm; 35% EtOAc/hexanes) to give the
cm; 1:9 EtOAc/hexanes) to give 27 as a colorless oil (0.861 g, 91%):
1
Rf = 0.70 (1:1 EtOAc/hexanes); [α]20 +120 (c 0.350, CHCl3); H
D
NMR (400 MHz, CDCl3) δ 7.83−7.79 (m, 1H), 7.72−7.70 (m, 2H),
7.70 (s, 1H), 7.48−7.43 (m, 2H), 7.40 (dd, J = 8.5, 1.2 Hz, 1H), 7.32−
7.17 (m, 10H), 6.14 (d, J = 5.4 Hz, 1H), 5.17 (d, J = 5.4 Hz, 1H), 4.70
(d, J = 12.0 Hz, 1H), 4.63 (d, J = 7.8 Hz, 1H), 4.61 (d, J = 7.8 Hz,
1H), 4.53 (d, J = 11.7 Hz, 1H), 4.48 (d, J = 11.7 Hz, 1H), 4.42 (d, J =
12.0 Hz, 1H), 4.02 (d, J = 9.9 Hz, 1H), 3.75 (d, J = 9.9 Hz, 1H), 3.35
(dd, J = 11.3, 5.0 Hz, 1H), 2.10−2.02 (m, 1H), 1.97−1.89 (m, 1H),
1.78−1.63 (m, 3H), 1.56 (s, 3H), 1.42−1.33 (m, 1H), 1.29 (s, 3H);
13C NMR (100 MHz, CDCl3) δ 139.2, 138.6, 136.2, 133.5, 133.1,
128.4, 128.2, 128.1, 127.9, 127.7, 127.5, 127.2, 126.3, 126.2, 126.0,
125.7, 114.0, 105.1, 91.7, 85.5, 85.4, 74.1, 73.2, 71.3, 66.8, 27.0, 26.1,
corresponding alcohol (44) as a colorless oil (0.97 g, 92%): Rf = 0.33
1
(2:3 EtOAc/hexanes); [α]20 +23 (c 0.11, CHCl3); H NMR (400
D
MHz, CDCl3) δ 5.89 (d, J = 4.7 Hz, 1H), 4.77 (d, J = 4.7 Hz, 1H),
3.95−3.85 (m, 1H), 3.83−3.74 (m, 2H), 3.35−3.25 (m, 1H), 1.90−
1.82 (m, 1H), 1.72−1.60 (m, 3H), 1.50 (s, 3H), 1.52−1.42 (m, 2H),
1.27 (s, 3H), 0.83 (s, 9H), 0.02 (s, 3H), 0.01 (s, 3H); 13C NMR (100
MHz, CDCl3) δ 114.3, 103.4, 89.6, 89.3, 81.1, 70.3, 65.0, 30.7, 29.8,
28.8, 26.8, 26.1, 26.0, 18.5, 18.2, −4.1, −4.7; HRMS (ESI) calcd for
C18H34NaO6Si [M + Na]+ m/z = 397.2017, found 397.2010. Sodium
I
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25.8, 18.9; HRMS (ESI) calcd for C37H40NaO6 [M + Na]+ m/z =
603.2717, found 603.2714.
nate (29). Potassium carbonate (0.141 g, 1.02 mmol) was added to
a stirred solution of nucleoside 28 (0.561 g, 0.812 mmol) in methanol
(25 mL). After 18 h, a solution of saturated NH4Cl (25 mL), water
(35 mL), and dichloromethane (80 mL) were added. The layers were
separated, and the aqueous phase was extracted with dichloromethane
(2 × 40 mL). The combined organic extracts were washed with brine
(40 mL), dried over MgSO4, filtered, and concentrated under reduced
pressure to afford the corresponding alcohol (48) as a colorless solid
(0.542 g, 93%), which was used in the next step without further
(2R,3R,3aR,7S,7aS)-3a,7-Bis(benzyloxy)-2-(5-methyl-2,4-
dioxo-3,4-dihydropyrimidin-1(2H)-yl)-7a-((naphthalen-2-
ylmethoxy)methyl)octahydrobenzofuran-3-yl Acetate (28).
FeCl3·6H2O (0.419 g, 1.55 mmol) was added to a stirred 0 °C
solution of 27 (0.901 g, 1.55 mmol) in dichloromethane (40 mL). The
cooling bath was removed, and the reaction was allowed to warm to
room temperature. After 1 h, a saturated solution of NaHCO3 (50
mL) was added, and the layers were separated. The aqueous phase was
extracted with dichloromethane (3 × 30 mL), and the combined
organic extracts were dried over MgSO4, filtered, and concentrated
under reduced pressure. The residue was purified by flash
chromatography (15 × 3 cm; 35% EtOAc/hexanes) to afford the
corresponding diol (46) as a colorless oil. Acetic anhydride (0.63 g, 6.2
mmol) was added to a stirred 0 °C solution of the preceding diol (46),
pyridine (0.024 g, 0.31 mmol), and DMAP (19 mg, 0.15 mmol) in
dichloromethane (30 mL). The cooling bath was removed, and the
reaction mixture was allowed to warm to room temperature. After 1 h,
1.0 M HCl (8 mL) and water (60 mL) were added. The layers were
separated, and the aqueous phase was extracted with dichloromethane
(3 × 30 mL). The combined organic extracts were washed with a
saturated solution of NaHCO3 (50 mL) and brine (50 mL), dried over
MgSO4, filtered, and concentrated under reduced pressure. The
residue was purified by flash chromatography (18 × 2.5 cm; 1:4
EtOAc/hexanes) to give the corresponding diacetate (47) as a
colorless oil (0.698 g, 72% over 2 steps): Rf = 0.27 (1:4 EtOAc/
hexanes); [α]20D +25 (c 0.075, CHCl3); 1H NMR (400 MHz, CDCl3)
δ 7.92−7.77 (m, 4H), 7.52−7.45 (m, 3H), 7.32−7.20 (m, 10H), 6.79
(d, J = 6.0 Hz, 1H), 6.29 (d, J = 6.0 Hz, 1H), 4.85 (d, J = 12.6 Hz,
1H), 4.72 (d, J = 12.6 Hz, 1H), 4.67 (d, J = 11.9 Hz, 1H), 4.59 (d, J =
11.1 Hz, 1H), 4.49 (d, J = 11.1 Hz, 1H), 4.42 (d, J = 11.9 Hz, 1H),
3.99 (d, J = 9.8 Hz, 1H), 3.79 (d, J = 9.8 Hz, 1H), 3.37 (dd, J = 10.8,
4.4 Hz, 1H), 2.34−2.26 (m, 1H), 2.11 (s, 3H), 2.10 (s, 3H), 2.02−1.92
(m, 2H), 1.78−1.68 (m, 2H), 1.53−1.41 (m, 1H); 13C NMR (100
MHz, CDCl3) δ 169.4, 169.0, 138.5, 138.2, 135.8, 133.5, 133.0, 128.4,
128.3, 128.2, 128.0, 127.7, 127.63, 127.57, 127.4, 126.04 126.01, 125.7,
125.5, 93.3, 86.3, 85.5, 76.9, 76.6, 73.8, 71.7, 66.1, 25.5, 24.7, 21.1,
20.7, 18.7; HRMS (ESI) calcd for C38H40NaO8 [M + Na]+ m/z =
647.2615, found 647.2610. A stirred mixture of thymine (0.311 g, 2.52
mmol) and N,O-bis(trimethylsilyl)acetamide (5.02 g, 24.7 mmol) in
1,2-dichloroethane (30 mL) was heated at 90 °C for 1 h until a clear
colorless solution persisted. The resulting solution was cooled to 0 °C,
and a solution of the preceding diacetate (47, 0.687 g, 0.995 mmol) in
1,2-dichloroethane (15 mL) and TMSOTf (0.550 g, 2.49 mmol
mmol) were added sequentially. The cooling bath was removed, and
the reaction mixture was heated at 55 °C. After 13 h, the reaction
mixture was cooled to room temperature and poured into a saturated
solution of NaHCO3 (40 mL). The layers were separated, and the
aqueous phase was extracted with dichloromethane (2 × 30 mL). The
combined organic extracts were washed with brine (50 mL), dried
over MgSO4, filtered, and concentrated under reduced pressure. The
residue was purified by flash chromatography (15 × 3 cm; 1:1 EtOAc/
hexanes) to give nucleoside 28 as a white solid (0.615 g, 81%): Rf =
purification: Rf = 0.26 (2:3 EtOAc/hexanes); mp 79−83 °C; [α]20
D
1
+26 (c 0.060, CHCl3); H NMR (400 MHz, CDCl3) δ 8.67 (s, 1H),
7.83−7.79 (m, 1H), 7.78−7.73 (m, 2H), 7.66 (s, 1H), 7.49−7.44 (m,
4H), 7.40−7.29 (m, 7H), 7.25−7.18 (m, 3H), 7.14−7.10 (m, 2H),
6.07 (d, J = 2.0 Hz, 1H), 4.77−4.67 (m, 4H), 4.60 (d, J = 12.1 Hz, H),
4.53 (d, J = 12.0 Hz, 1H), 4.42−4.36 (m, 1H), 4.31 (d, J = 12.0 Hz,
1H), 4.17 (br d, J = 3.8 Hz, 1H), 3.94 (d, J = 11.4 Hz, 1H), 3.86 (d, J =
11.4 Hz, 1H), 2.32−2.22 (m, 1H), 1.84−1.75 (m, 4H), 1.60−1.51 (m,
2H), 1.33 (s, 3H); 13C NMR (100 MHz, CDCl3) δ 163.9, 151.0,
137.8, 137.0, 136.8, 135.2, 133.3, 133.2, 128.9, 128.6, 128.5, 128.4,
128.0, 127.9, 127.6, 126.8, 126.5, 126.3, 125.8, 93.8, 90.7, 84.9, 81.1,
73.9, 73.2, 71.0, 64.5, 29.9, 27.4, 25.7, 18.1, 12.2; HRMS (ESI) calcd
for C39H40N2NaO7 [M + Na]+ m/z = 671.2728, found 671.2738.
Methanesulfonyl chloride (0.287 g, 2.51 mmol) was added to a stirred
0 °C solution of the preceding alcohol (48, 0.542 g, 0.835 mmol) and
DMAP (0.010 g, 0.082 mmol) in anhydrous pyridine (25 mL). The
cooling bath was removed, and the reaction mixture was allowed to
warm to room temperature. After 16 h, a saturated solution of
NaHCO3 (40 mL) and dichloromethane (40 mL) were added. The
layers were separated, and the aqueous phase was extracted with
dichloromethane (3 × 30 mL). The combined organic extracts were
washed with 1.0 M HCl (2 × 40 mL) and brine (40 mL), dried over
MgSO4, filtered, and concentrated under reduced pressure. The
residue was purified by flash chromatography (20 × 2.5 cm; 1:1
EtOAc/hexanes) to give nucleoside 29 as a colorless solid (0.569 g,
94%): Rf = 0.33 (1:1 EtOAc/hexanes); mp 83−86 °C; [α]20 +48 (c
D
1
0.070, CHCl3); H NMR (400 MHz, CDCl3) δ 8.78 (s, 1H), 7.82−
7.75 (m, 4H), 7.55 (s, 1H), 7.49−7.45 (m, 2H), 7.42−7.40 (m, 1H),
7.39−7.35 (m, 2H), 7.34−7.29 (m, 2H), 7.28−7.25 (m, 1H), 7.23−
7.15 (m, 5H), 6.45 (d, J = 4.5 Hz, 1H), 5.53 (d, J = 4.4 Hz, 1H), 4.81
(d, J = 12.0 Hz, 1H), 4.72−4.61 (m, 3H), 4.58−4.53 (m, 2H), 4.00−
3.96 (m, 2H), 3.85 (d, J = 11.0 Hz, 1H), 2.84 (s, 3H), 2.23−2.13 (m,
1H), 2.06−1.91 (m, 2H), 1.89−1.80 (m, 1H), 1.68−1.59 (m, 1H),
1.55−1.46 (m, 1H), 1.23 (d, J = 1.1 Hz, 3H); 13C NMR (100 MHz,
CDCl3) δ 163.6, 151.0, 138.2, 137.4, 136.2, 135.0, 133.5, 133.2, 128.7,
128.62, 128.58, 128.56, 128.12, 128.08, 128.0, 127.9, 127.8, 126.7,
126.5, 126.3, 125.7, 111.7, 89.4, 87.6, 85.2, 83.6, 77.2, 73.9, 72.3, 71.6,
65.1, 39.1, 27.4, 25.3, 17.5, 12.0; HRMS (ESI) calcd for
C40H42N2NaO9S [M + Na]+ m/z = 749.2503, found 749.2521.
((5aR,6aS,7S,10aR,10bS)-7,10a-Bis(benzyloxy)-3-methyl-2-
oxo-7,8,9,10,10a,10b-hexahydro-2H-benzofuro[2′,3′:4,5]-
oxazolo[3,2-a]pyrimidin-6a(5aH)-yl)methyl Methanesulfonate
(30). DBU (0.435 mL, 3.13 mmol) was added to a stirred solution of
29 (0.569 g, 0.783 mmol) in acetonitrile (45 mL). The reaction
mixture was heated at 80 °C for 75 h, cooled to 0 °C, and diluted with
dichloromethane (45 mL). The resulting solution was poured into a
solution of 0.6 M HCl (40 mL), and the layers were separated. The
aqueous phase was extracted with dichloromethane (2 × 25 mL), and
the combined organic extracts were washed with brine (30 mL), dried
over MgSO4, filtered, and concentrated under reduced pressure. The
residue was purified by flash chromatography (18 × 2.5 cm; 3%
EtOH/EtOAc) to give the corresponding 2,2′-anhydronucleoside (49)
as a colorless solid (0.347 g, 71%): Rf = 0.35 (7% EtOH/EtOAc); mp
0.15 (2:3 EtOAc/hexanes); mp 71−74 °C; [α]20 +28 (c 0.14,
D
CHCl3); 1H NMR (400 MHz, CDCl3) δ 8.96 (s, 1H), 7.87−7.75 (m,
5H), 7.51−7.42 (m, 3H), 7.37−7.17 (m, 10H), 6.39 (d, J = 5.9 Hz,
1H), 6.13 (d, J = 5.9 Hz, 1H), 4.78 (d, J = 12.2 Hz, 1H), 4.70−4.61
(m, 2H), 4.58−4.52 (m, 2H), 4.44 (d, J = 10.8 Hz, 1H), 4.04 (d, J =
10.6 Hz, 1H), 3.77 (d, J = 10.6 Hz, 1H), 3.70−3.64 (m, 1H), 2.21−
2.12 (m, 1H), 2.06 (s, 3H), 2.05−1.95 (m, 1H), 1.91−1.72 (m, 3H),
1.51−1.40 (m, 1H), 1.29 (s, 3H); 13C NMR (100 MHz, CDCl3) δ
170.3, 163.9, 151.2, 138.1, 137.8, 136.8, 135.0, 133.5, 130.1, 128.6,
128.5, 128.4, 128.3, 128.0, 127.93, 127.90, 127.8, 127.6, 126.4, 126.3,
126.1, 125.4, 111.0, 88.4, 86.7, 83.2, 78.8, 77.3, 73.8, 72.5, 72.2, 65.5,
26.8, 25.5, 20.9, 17.9, 11.9; HRMS (ESI) calcd for C41H42N2NaO8 [M
+ Na]+ m/z = 713.2833, found 713.2847.
79−83 °C; [α]20 −21 (c 0.061, CHCl3); 1H NMR (400 MHz,
D
CDCl3) δ 7.85−7.76 (m, 3H), 7.67 (s, 1H), 7.52−7.43 (m, 2H),
7.37−7.25 (m, 6H), 7.20−7.13 (m, 4H), 7.12−7.05 (m, 2H), 5.93 (d, J
= 6.2 Hz, 1H), 5.33 (d, J = 6.2 Hz, 1H), 4.75 (d, J = 12.1 Hz, 1H), 4.68
(d, J = 11.7 Hz, 1H), 4.59−4.53 (m, 3H), 4.49 (d, J = 11.2 Hz, 1H),
4.15 (d, J = 3.0 Hz, 1H), 3.82 (d, J = 11.0 Hz, 1H), 3.64 (d, J = 11.0
Hz, 1H), 2.35−2.26 (m, 1H), 1.93−1.84 (m, 4H), 1.77−1.65 (m, 2H),
1.64−1.55 (m, 1H), 1.40−1.30 (m, 1H); 13C NMR (100 MHz,
(2R,3R,3aR,7S,7aS)-3a,7-Bis(benzyloxy)-2-(5-methyl-2,4-
dioxo-3,4-dihydropyrimidin-1(2H)-yl)-7a-((naphthalen-2-
ylmethoxy)methyl)octahydrobenzofuran-3-yl methanesulfo-
J
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CDCl3) δ 172.5, 159.8, 138.6, 138.0, 135.2, 130.5, 128.8, 128.5, 128.2,
128.1, 127.9, 127.8, 127.63, 127.59, 127.0, 126.4, 126.0, 118.8, 91.0,
89.5, 85.0, 83.6, 74.1, 72.6, 70.4, 67.1, 29.9, 28.1, 25.5, 16.2, 14.3;
HRMS (ESI) calcd for C39H38N2NaO6 [M + Na]+ m/z = 653.2622,
found 653.2627. DDQ (0.315 g, 1.39 mmol) was added to a stirred
solution of the preceding 2,2′-anhydronucleoside (49, 0.286 g, 0.461
mmol) in 1:9 water/dichloromethane (28 mL). After 3 h, the reaction
mixture was poured into a solution of saturated NaHCO3 (50 mL).
The layers were separated, and the aqueous phase was extracted with
dichloromethane (2 × 50 mL). The combined organic extracts were
dried over MgSO4, filtered, and concentrated under reduced pressure.
The residue was purified by flash chromatography (15 × 2 cm; 7%
EtOH/EtOAc) to give the corresponding alcohol as a colorless solid
added to a stirred solution of tricyclic nucleoside 31 (0.184 g, 0.375
mmol) in a 1:1 mixture of EtOH/MeOH (30 mL), which was placed
under an atmosphere of hydrogen gas via a hydrogen-filled balloon.
After 15 h, the reaction mixture was filtered through sintered glass
funnel, and the filtrate was concentrated under reduced pressure. The
residue was purified by flash chromatography (15 × 2 cm; 5% EtOH/
EtOAc) to give tricyclic nucleoside 32 as a colorless solid (0.114 g,
98%): Rf = 0.22 (5% EtOH/EtOAc); mp 209−210 °C; [α]20D +158 (c
1
0.060, CHCl3); H NMR (400 MHz, CD3OD) δ 7.98 (d, J = 1.0 Hz,
1H), 5.91 (d, J = 1.0 Hz, 1H), 4.28 (d, J = 1.1 Hz, 1H), 4.09 (s, 2H),
3.89 (dd, J = 10.7, 4.8 Hz, 1H), 1.92 (d, J = 1.1 Hz, 3H), 1.87−1.73
(m, 6H); 13C NMR (100 MHz, CD3OD) δ 166.7, 152.2, 138.2, 109.9,
90.7, 90.0, 82.0, 81.8, 74.8, 68.3, 30.6, 28.6, 21.5, 12.4; HRMS (ESI)
calcd for C14H18N2NaO6 [M + Na]+ m/z = 333.1057, found 333.1061.
(2R,3S,3aR,7S,7aS)-3a-Hydroxy-2-(5-methyl-2,4-dioxo-3,4-
d i h y d r o p y r i m i d i n - 1 ( 2 H ) - y l ) h e x ah y d r o - 2 H - 3 , 7 a -
(50, 0.220 g, 96%): Rf = 0.43 (7% EtOH/EtOAc); mp 105−107 °C;
1
[α]20 −14 (c 0.060, CHCl3); H NMR (400 MHz, CDCl3) δ 7.24−
D
7.10 (m, 11H), 5.65 (d, J = 5.7 Hz, 1H), 5.08 (d, J = 5.7 Hz, 1H),
4.39−4.53 (m, 4H), 3.94−3.88 (m, 1H), 3.75−3.64 (m, 1H), 3.62−
3.55 (m, 1H), 2.68−2.64 (m, 1H), 2.04−1.95 (m, 1H), 1.78−1.68 (m,
4H), 1.65−1.37 (m, 4H); 13C NMR (100 MHz, CDCl3) δ 172.5,
160.0, 137.8, 137.1, 128.6, 128.5, 128.0, 127.8, 118.5, 89.3, 88.2, 85.9,
84.3, 77.0, 72.7, 67.4, 63.1, 29.7, 28.6, 25.4, 17.7, 14.0; HRMS (ESI)
calcd for C28H30N2NaO6 [M + Na]+ m/z = 513.1996, found:
513.1997. Methanesulfonyl chloride (0.130 g, 1.14 mmol) was added
to a stirred 0 °C solution of the preceding alcohol (50, 0.221 g, 0.451
mmol) and DMAP (5.5 mg, 0.045 mmol) in anhydrous pyridine (20
mL). The cooling bath was removed, and the reaction was allowed
warm to room temperature. After 12 h, the reaction mixture was
diluted with dichloromethane (50 mL) and poured into a saturated
solution of NaHCO3 (100 mL). The layers were separated, and the
aqueous phase was extracted with dichloromethane (2 × 50 mL). The
combined organic extracts were washed with 1.0 M HCl (50 mL) and
brine (50 mL), dried over MgSO4, filtered, and concentrated under
reduced pressure. The residue was purified by flash chromatography
(17 × 2 cm; 7% EtOH/EtOAc) to give 2,2′-anhydronucleoside 30 as a
colorless solid (0.226 g, 93%): Rf = 0.28 (10% EtOH/EtOAc); mp =
(epoxymethano)benzofuran-7-yl 4-Oxopentanoate (6: R1
=
Lev, R2 = H, Bx = thymine). Mukaiyama’s reagent (0.075 g, 0.29
mmol) was added to a stirred solution of tricyclic nucleoside 32 (0.070
g, 0.23 mmol) and levulinic acid (28 μL, 0.27 mmol) in acetonitrile
(11 mL). After 15 min, triethylamine (94 μL, 0.68 mmol) and DMAP
(0.008 g, 0.07 mmol) were added. After an additional 16 h, the
reaction mixture was poured into an ice-cold solution of 1 M HCl (20
mL) and further diluted with dichloromethane (15 mL). The layers
were separated, and the aqueous phase was extracted with dichloro-
methane (3 × 10 mL). The combined organic extracts were washed
with a saturated aqueous solution of NaHCO3 (15 mL), dried through
a phase separator cartridge, and concentrated under reduced pressure.
The residue was purified by flash column chromatography (15 × 1.5
cm; 1:20 MeOH/CH2Cl2) to afford tricyclic nucleoside 6 (R1 = Lev,
R2 = H, Bx = thymine) as a colorless solid (0.070 g, 76% yield): Rf =
20
0.24 (1:15 MeOH/CH2Cl2); mp 238−239 °C (CH2Cl2); [α]D +34
(c = 0.25, CHCl3); 1H NMR (400 MHz, CD2Cl2) δ 8.32 (s, 1H), 7.80
(s, 1H), 5.99 (s, 1H), 5.33 (s, 1H), 5.12−4.99 (m, 1H), 4.29 (s, 1H),
4.03 (d, J = 9.8 Hz, 1H), 3.99 (d, J = 9.7 Hz), 2.86 (ddd, J = 18.7, 9.1,
4.4 Hz, 1H), 2.75 (ddd, J = 18.6, 6.4, 4.1 Hz, 1H), 2.57 (ddd, J = 16.9,
9.1, 4.1 Hz, 1H), 2.45 (ddd, J = 17.0, 6.6, 4.4 Hz, 1H), 2.16 (s, 3H),
1.98 (s, 3H), 1.96−1.78 (m, 6H); 13C NMR (75 MHz, CD2Cl2) δ
207.7, 172.9, 164.1, 150.7, 136.6, 110.1, 89.3, 88.3, 82.4, 80.2, 74.3,
70.0, 38.8, 29.9, 28.7, 27.9, 26.6, 20.9, 12.5; HRMS (ESI) calcd for
C19H25N2O8 [M + H]+ m/z = 409.1605, found 409.1596.
99−101 °C; [α]20 +30 (c 0.064, CHCl3; 1H NMR (400 MHz,
D
CDCl3) δ 7.37−7.16 (m, 11H), 6.05 (d, J = 5.9 Hz, 1H), 5.30 (d, J =
6.0 Hz, 1H), 4.68−4.54 (m, 3H), 4.51−4.42 (m, 2H), 4.26 (d, J = 11.1
Hz, 1H), 3.94 (d, J = 4.0 Hz, 1H), 2.76 (s, 3H), 2.23−2.06 (m, 1H),
1.92−1.89 (m, 5H), 1.75−1.48 (m, 3H); 13C NMR (100 MHz,
CDCl3) δ 172.2, 159.6, 137.5, 137.4, 130.5, 128.6, 128.1, 128.0, 127.9,
127.8, 118.7, 89.6, 88.3, 84.4, 84.2, 75.9, 72.6, 69.1, 67.3, 37.0, 30.0,
27.9, 25.1, 16.3, 14.1; HRMS (ESI) calcd for C29H32N2NaO8S [M +
Na]+ m/z = 591.1772, found 591.1768.
(2R,3S,3aR,7S,7aS)-3a-(((2-Cyanoethoxy)(diisopropylamino)-
phosphanyl)oxy)-2-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-
1(2H)-yl)hexahydro-2H-3,7a-(epoxymethano)benzofuran-7-yl
4-Oxopentanoate (33). 2-Cyanoethyl N,N-diisopropylchlorophos-
phoramidite (90 μL, 0.39 mmol) was added to a stirred solution of
nucleoside 6 (R1 = Lev, R2 = H, Bx = thymine) (0.064 g, 0.16 mmol),
N,N-diisopropylethylamine (70 μL, 0.39 mmol), and N-methylimida-
zole (1 drop) in dichloromethane (1.5 mL). After 2 h, additional
portions of N,N-diisopropylethylamine (35 μL) and 2-cyanoethyl N,N-
diisopropylchlorophosphoramidite (30 μL) were added, and the
stirring was continued for another 10 min. The reaction was then
cooled in an ice bath and quenched by the addition of
diisopropylethylamine (0.1 mL) and methanol (0.8 mL). After
another 10 min of stirring in the ice bath, the solvents were
evaporated under reduced pressure, and the residue was purified by
column chromatography (20% to 30% acetone/dichloromethane) to
give phosphoramidite 33 (57 mg, 60%): Rf = 0.34 (1:4 acetone/
1-((2R,3S,3aR,7S,7aS)-3a,7-Bis(benzyloxy)hexahydro-2H-
3,7a-(epoxymethano)benzofuran-2-yl)-5-methylpyrimidine-
2,4(1H,3H)-dione (31). A 2.0 M solution of NaOH (0.42 mL, 0.84
mmol) was added to stirred suspension of 2,2′-anydronucleoside 30
(0.220 g, 0.387 mmol) in 1:1 water/ethanol (50 mL). The reaction
was heated at 100 °C for 30 min, cooled to room temperature, and
diluted with dichloromethane (50 mL). The layers were separated, and
the aqueous phase was extracted with dichloromethane (2 × 30 mL).
The combined organic extracts were washed with a saturated solution
of NaHCO3 (100 mL), dried over MgSO4, filtered, and concentrated
under reduced pressure. The residue was purified by flash
chromatography (20 × 2 cm 3:2 EtOAc/hexanes) to give tricyclic
nucleoside 31 as a colorless solid (0.181 g, 96%): Rf = 0.92 (5%
1
EtOH/EtOAc); mp 96−99 °C; [α]20 +121 (c 0.100, CHCl3); H
1
dichloromethane); H NMR (300 MHz, CDCl3) δ = 8.02 (br s, 1H),
D
NMR (400 MHz, CDCl3) δ 9.61 (s, 1H), 7.58 (s, 1H), 7.48−7.24 (m,
10H), 6.05 (s, 1H), 4.78−4.66 (m, 2H), 4.57−4.46 (m, 3H), 4.21 (d, J
= 8.0 Hz, 1H), 4.16 (d, J = 8.1 Hz, 1H), 3.81−3.74 (m, 1H), 2.18 (br
d, J = 11.6 Hz, 1H), 2.08−2.02 (m, 1H), 1.98 (s, 3H), 1.90−1.84 (m,
1H), 1.82−1.64 (m, 3H); 13C NMR (100 MHz, CDCl3) δ 164.3,
150.8, 138.2, 137.8, 135.8, 128.6, 128.5, 128.0, 127.9, 127.72, 127.68,
109.7, 89.7, 89.1, 86.3, 77.3, 74.8, 74.4, 71.5, 67.5, 26.1, 23.3, 20.4,
12.8; HRMS (ESI) calcd for C28H30N2NaO6 [M + Na]+ m/z =
513.1996, found 513.1993.
7.85 (d, J = 5.0 Hz, 1H), 6.09 (br s, 1H), 5.07 (br s, 1H), 4.71 (d, J =
3.8 Hz, 1H), 4.29−3.99 (m, 4H), 3.84 (d, J = 11.5 Hz, 2H), 3.70−3.40
(m, 5H), 2.97−2.55 (m, 9H), 2.49 (br s, 1H), 2.33−2.13 (m, 4H),
2.09−1.94 (m, 5H), 1.86 (br s, 5H), 1.41−1.01 (m, 33H); 31P NMR
(121 MHz, CDCl3) δ 145.8, 144.8; LRMS (ESI) calcd for
C28H41N4O9P [M + H]+ m/z = 608.3, found 608.3.
ASSOCIATED CONTENT
■
S
* Supporting Information
1-((2R,3S,3aR,7S,7aS)-3a,7-Dihydroxyhexahydro-2H-3,7a-
(epoxymethano)benzofuran-2-yl)-5-methylpyrimidine-2,4-
(1H,3H)-dione (32). Pearlman’s catalyst (0.026 g, 0.038 mmol) was
1H and 13C NMR spectra of new compounds, as well as X-ray
crystallography reports and CIF files for compounds 19 and 32.
K
dx.doi.org/10.1021/jo401170y | J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
(16) Romieu, A.; Gasparutto, D.; Cadet, J.; Romieu, J. J. Chem. Soc.,
Perkin Trans. 1 1999, 1257−1263.
This material is available free of charge via the Internet at
http://pubs.acs.org.
(17) Egli, M.; Usman, N.; Rich, A. Biochemistry 1993, 32, 3221−
3237.
AUTHOR INFORMATION
■
(18) (a) Seth, P. P.; Allerson, C. R.; Østergaard, M. E.; Swayze, E. E.
Bioorg. Med. Chem. Lett. 2012, 22, 296−299. (b) Seth, P. P.; Jazayeri,
A.; Yu, J.; Allerson, C. R.; Bhat, B.; Swayze, E. E. Mol. Ther. Nucleic
Acids 2012, 1, e47.
(19) Nielsen, J. T.; Stein, P. C.; Petersen, M. Nucleic Acids Res. 2003,
31, 5858−5867.
(20) Chromatography procedures and conditions were followed as
described in Still, W. C.; Kahn, M.; Mitra, A. J. Org. Chem. 1978, 43,
2923−2925.
Corresponding Authors
*E-mail: stephen.hanessian@umontreal.ca.
*E-mail: pseth@isisph.com.
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
We are grateful for financial support from NSERC and
FQRNT. R.D.G. thanks NSERC for a graduate scholarship.
̂
We thank Benoit Deschen
analyses.
̂
es-Simard for X-ray crystallographic
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dx.doi.org/10.1021/jo401170y | J. Org. Chem. XXXX, XXX, XXX−XXX