Towards the synthesis of sulfonamide-based RNA mimetics

Article abstract of DOI:10.1016/j.tetasy.2010.02.005

The scalable, divergent synthesis of all four monomers required for the preparation of sulfonamide-based RNA mimetics is described. Such mimetics may combine excellent mimicry of the parent RNA with enhanced (bio)chemical robustness and convenient oligome

Full text of DOI:10.1016/j.tetasy.2010.02.005

Tetrahedron: Asymmetry 21 (2010) 469–475
Contents lists available at ScienceDirect
Tetrahedron: Asymmetry
journal homepage: www.elsevier.com/locate/tetasy
Towards the synthesis of sulfonamide-based RNA mimetics
*
Jeroen van Ameijde, Thierry K. Slot, Rob M. J. Liskamp
Medicinal Chemistry and Chemical Biology, Faculty of Science, Utrecht University, Sorbonnelaan 16, 3584 CA Utrecht, The Netherlands
a r t i c l e i n f o
a b s t r a c t
Article history:
The scalable, divergent synthesis of all four monomers required for the preparation of sulfonamide-based
RNA mimetics is described. Such mimetics may combine excellent mimicry of the parent RNA with
enhanced (bio)chemical robustness and convenient oligomerization. As a proof of principle, a dimer
resulting from the monomers is described.
Received 25 January 2010
Accepted 5 February 2010
Available online 26 February 2010
Ó 2010 Elsevier Ltd. All rights reserved.
1. Introduction
introduction of a 20-OH group necessary for true RNA mimicry
and would not be suitable for the stepwise incorporation into
In recent years short regulatory RNAs have come to the fore-
front of biological research, for example, as mediators in the
important RNA interference technology.1 Since such RNA mole-
cules are able to selectively and effectively inhibit individual genes,
they have great potential in combating diseases in which the
expression level of a certain gene is deregulated.
any RNA nucleotide sequence. Studies on sulfamoyl-based DNA
dimers, which are structurally similar to the mimics described
here, have shown that such compounds have structural properties
similar to natural DNA.7
Apart from oligomerization into small RNA mimetics, a number
of different applications of the monomers described here can be
envisaged. For instance, a similar adenine monomer may function
as an ATP analogue and be conveniently conjugated to a peptide
leading to potential bisubstrate-based inhibitors.8 Furthermore,
the primer essential for the replication of many viruses consists
of a peptide functionalized with one or more uridine groups and
However, the direct application of RNAs as therapeutics is
hampered by a number of serious drawbacks: (i) they are nega-
tively charged which makes cellular uptake difficult and often
necessitates complex transfection techniques; (ii) they are still
complex and delicate synthetic targets and (iii) they are degraded
by extra- and intracellular nucleases. In order to remedy such
drawbacks, nucleic acid mimetics may be used, arguably the most
successful of which have been the peptide nucleic acids (PNAs),2
locked nucleic acids (LNAs)3 and morpholinos.4 However, such
mimetics often lack the 20-OH group which distinguishes RNA
from DNA and may be of importance in certain biological interac-
tions. Furthermore, in particular for PNAs and morpholinos,
although they have found many successful biological applications,
the chemical backbone structure is very different from natural
nucleic acids which may also be of importance in certain biolog-
ical settings.
Herein we report an RNA mimetic in which the phosphate is
replaced by the semi-isosteric sulfonamide moiety (sRNAs,
Fig. 1). The sulfonamide moiety has a similar charge distribution
as the phosphates found in RNA, albeit not negatively charged.
In addition, they are (bio)chemically stable and can be introduced
in a stepwise solid phase oligomerization procedure analogous to
the preparation of oligosulfonamide peptidomimetics.5 Their clos-
est relatives are the DNA sulfonamide mimetics,6 however the
syntheses described for these structures would not allow the
O
O
P
S
O
O
O-
O
O
O
B
B
HN
O
O
OH
OH
P
S
O
O
O-
O
O
O
B
B
HN
OH
RNA
OH
sRNA
O
O
O
B
S
Cl
N3
OAc
* Corresponding author. Tel.: +31 (0) 30 2537396; fax: +31 (0) 30 2536655.
E-mail address: r.m.j.liskamp@uu.nl (R.M.J. Liskamp).
Figure 1. Sulfonamide-based RNA mimetics (sRNAs).
0957-4166/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetasy.2010.02.005

470
J. van Ameijde et al. / Tetrahedron: Asymmetry 21 (2010) 469–475
constructs containing the sulfonamide moieties may function as
potential anti-virals.9
At this point the sulfonyl moiety was introduced by first con-
verting the remaining free hydroxyl group into the correspond-
ing triflate, which was subsequently subjected to nucleophilic
substitution with the anion derived from isobutyl mesylate.6
Various conditions were evaluated for this reaction, such as
using nBuLi, tBuLi or LDA to generate the anion, adding HMPA,
TMEDA or a crown ether to solubilize the anion, carrying out
the reaction at ꢀ78 °C throughout or allowing the reaction mix-
ture to slowly heat to temperatures up to ꢀ20 °C or even room
temperature. However, the best yield obtained was 51%. This rel-
atively low yield can be explained by the formation of significant
amounts of dimer 4 resulting from the deprotonation of 3 upon
formation followed by reaction with a further triflate starting
compound. In spite of its relatively low yield, since this proce-
dure could be carried out on a sizeable scale (2 g), sufficient
material for further synthesis was obtained. However, we have
found an alternative route which leads to a better overall yield
for the transformation of 1 into 3 (Scheme 1). In this synthesis,
after selective deprotection of the terminal acetonide and subse-
quent oxidative cleavage as before, the resulting aldehyde was
treated with Horner–Wadsworth–Emmons (HWE) reagent 5,
which is itself accessible using a literature procedure.12 Using
The synthesis of such oligomers requires a complete and suit-
able set of monomers (Fig. 1). The azido moiety will be used as a
protected amine which can then be released under mild conditions
such as catalytic hydrogenation and the Staudinger reaction. In
addition, a sulfonyl chloride moiety is introduced which can react
rapidly with the liberated amine at the ‘N-terminus’ of the growing
oligomer.
2. Results and discussion
From the outset we decided to prepare the required azido sulfo-
nyl chloride monomers in a divergent manner by introducing the
nucleosidic base late in the synthesis, after the introduction of
the azido and sulfonate moieties (Scheme 1). Thus, the commer-
cially available diacetone-D-glucose was converted into azido com-
pound 1 in two steps using a literature procedure.10 The more
labile 5,6-O-isopropylidene was removed by mild acidolysis after
which oxidative diol cleavage and reduction with lithium borohy-
dride afforded compound
2
which has the desired
D-ribo
stereochemistry.11
O
O
O
O
O
O
O
O
O
O
O
S
(i)
(ii)
O
O
O
S
O
O
O
HO
+
O
O
O
O
O
N3
N3
N3
N3
1
2
3
O
N3
(iii)
4
O
O
O
O
(v)
(iv)
O
OAc
S
N3
OAc
PivHN
N
N
E/Z-6
NH
N
N
NHBz
O
O
O
O
OAc
S
O
O
O
O
(vii)
O
(vi)
O
O
N
N
O
S
N
S
N3
OAc
N3
OAc
N3
OAc
7
8
9
(ix)
(viii)
O
O
NHBz
O
O
O
O
O
O
O
O
O
O
N
N
N
S
(x)
S
S
NH
NH
N
Cl
O
O
O
O
N3
OAc
N3
OAc
N3
OAc
12
11
10
(xi)
NHBz
O
O
N
O
N
S
N
O
N3
OAc
13
Scheme 1. Monomer synthesis. Reagents and conditions: Method A: (i) (1) 50% HOAc (aq), 50 °C; (2) NaIO4, MeOH; (3) LiBH4, THF, ꢀ20 °C (77%); (ii) (1) Tf2O, pyr., ꢀ20 °C; (2)
isobutyl sulfonate, nBuLi, HMPA, ꢀ78 °C, THF (51%); Method B: (iii) (1) 50% HOAc (aq), 50 °C; (2) NaIO4, MeOH; (3) NaH, 5, ꢀ20 °C THF; (iv) NaBH4, ꢀ20 °C to rt, THF (67%); (v)
(1) 90% TFA (aq); (2) Ac2O, pyr (80%); (vi) N2-pivaloylguanine, bis-TMS-acetamide, SnCl4, CH3CN (58%); (vii) N6-benzoyladenine, bis-TMS-acetamide, SnCl4, CH3CN (51%); (viii)
uracil, bis-TMS-acetamide, SnCl4, CH3CN (71%); (ix) N2-benzoylcytosine, bis-TMS-acetamide, SnCl4, CH3CN (63%); (x) (1) KI, acetone,
D; (2) 20% COCl2 in toluene, DMF, DCM
(61%); (xi) (1) KI, acetone,
D
; (2) 20% COCl2 in toluene, DMF, DCM; (3) dimethylamineꢁHCl, NMM, DCM (72%).

J. van Ameijde et al. / Tetrahedron: Asymmetry 21 (2010) 469–475
471
sodium hydride as the base, the smooth formation of a 1:1 mix-
O
O
O
O
ture of two isomeric products was observed, one of which could
clearly be identified as the desired E-unsaturated product E-6
from coupling constants in the 1H NMR spectrum (3JH-5,H-6
15.2 Hz). The identity of the other product Z-6 could not be
unambiguously established at this point since it has an interme-
diate coupling constant (3JH-5,H-6 11.4 Hz), and might therefore
also be the result of epimerization of the aldehyde at C-4 under
the basic reaction conditions employed. At this point we decided
to carry out the subsequent reduction of the conjugated double
bond using sodium borohydride on the mixture even though
chromatographic separation was possible. This led to the forma-
tion of a single product 3 demonstrating that 6Z was indeed the
Z-unsaturated HWE-product, which was identical in all respects
to the material resulting from the substitution strategy described
above. The yield for this synthesis was 67%, which is a clear
improvement over the substitution route described above (yield
for the same overall transformation 39%).
In order to convert 3 towards the introduction of the nucleoside
base, the remaining isopropylidene-protecting group was removed
by acidolysis and the resulting free hydroxyl moieties were acety-
lated using a mixture of acetic anhydride and pyridine, yielding
bis-acetate 7 as a 1:4 mixture of anomers in 80% yield. The stage
was now set for the introduction of suitably protected silylated
bases using SnCl4 as a Lewis acid catalyst.13 Four bases were used:
N6-benzoyladenosine,14 N2-pivaloylguanidine,15 N4-benzoylcyto-
sine16 and uracil afforded fully protected monomer 8, 9, 10 and
11 as single anomers in 51%, 58%, 63% and 71% yields, respectively.
These building blocks are stable and can be kept for months at
ambient temperature.
N
S
NH
Cl
O
N3
OAc
12
(i)
O
O
O
O
N
S
NH
N
H
O
N3
OAc
14
(ii)
HN
S
O
O
O
N
O
NH
HN
O
OAc
N3
O
O
S
AcO
O
O
N
HN
15
O
Scheme 2. Synthesis of UU-dimer 15. Reagents and conditions: (i) isobutyl amine,
NMM, DCM (91%); (ii) (1) H2, 10% Pd/C, HOAc, THF; (2) 12, NMM, DMF (67%).
In order for oligomerization to occur, the sulfonate ester present
in the monomers has to be deprotected and converted into the cor-
responding sulfonyl chloride. This was demonstrated using the uri-
dine building block 11. Deprotection using potassium iodide at an
elevated temperature followed by treatment with phosgene in the
presence of DMF afforded sulfonyl chloride 12 in 61% yield over
two steps. Compound 12 is relatively stable and can be purified
by column chromatography after which it can be stored for several
months at ꢀ20 °C. As an alternative, we attempted to transform
stable sulfonate ester 10 into a sulfonamide coupling product with-
out intermediate purification of the sulfonyl chloride. Thus, cyti-
dine monomer 10 was deprotected using potassium iodide and
was converted into a sulfonyl chloride using phosgene and DMF
as described above. Then, the crude sulfonyl chloride was treated
with the hydrochloric acid salt of dimethyl amine in the presence
of NMM as a base and smooth conversion into sulfonamide 13
was observed (72% yield after purification by column
chromatography).
3. Conclusions
A divergent synthesis of azido sulfonate building blocks suitable
for RNA mimicry was developed, and was carried out successfully
for all four RNA bases. The uracil monomer was subsequently
transformed into the corresponding ‘activated’ sulfonyl chloride
and subjected to a dimerization procedure resulting in a dimeric
compound. The cytidine monomer was converted into a sulfon-
amide coupling product without isolation of the intermediate sul-
fonyl chloride demonstrating the usefulness of the sulfonate
monomers as stable building blocks for the preparation of sulfon-
amide RNA mimetics.
4. Experimental
Next, we decided to demonstrate the principle of oligomeriza-
tion by the preparation of a uridine–uridine dimer as shown in
Scheme 2. First, uridine mimic 12 was coupled to isobutylamine
using NMM as a base, yielding isobutyl sulfonamide 14 in 91%
yield after purification by column chromatography. Interestingly,
this reaction demonstrates the potential for the sulfonyl chloride
monomers to be conjugated to any amine-containing molecule.
The subsequent formation of the dimer 15 was carried out by
the reduction of the azido moiety in 14 to the corresponding
amine using catalytic hydrogenolysis. During the hydrogenation
reaction, 1 equiv of acetic acid was added in order to protect
the resulting amino moiety by protonation. This was deemed
necessary because upon prolonged standing, migration of the
20-acetyl-protecting group to the liberated amine occurs to a
sizeable degree obviously leading to lower yields. The crude
amine was immediately treated with a further equivalent of sul-
fonyl chloride 9 again using NMM as a base. After purification,
the dimer 15 was obtained in 67% yield.
4.1. General procedures
Reactions were carried out at ambient temperature unless sta-
ted otherwise. All reagents were used as supplied from commercial
sources unless stated otherwise. DCM and CH3CN were stored on
molecular sieves (4 Å) prior to use. Rf values were determined by
thin layer chromatography (TLC) on Merck precoated Silica Gel
60F254 plates. Spots were visualized by UV-quenching, ninhydrin
or Hanessian’s stain (cerium molybdate). Column chromatography
was carried out using Silicycle UltraPure silica gel (40–63 l
m). 1H
NMR spectra were recorded on a Varian G-300 (300 MHz) spec-
trometer and chemical shifts are given in ppm relative to TMS
(0.00 ppm). 13C NMR spectra were recorded using the attached
proton test (APT) sequence on a Varian G-300 (75.5 MHz) spec-
trometer and chemical shifts are given in ppm relative to CDCl3
(77.0 ppm). Electrospray ionization mass spectrometry was per-
formed on a Shimadzu LCMS-QP8000 single quadrupole bench-
top mass spectrometer in positive ionization mode.

472
J. van Ameijde et al. / Tetrahedron: Asymmetry 21 (2010) 469–475
4.2. 3-Azido-3-deoxy-1,2-O-isopropylidene-b-
D
-ribose 211
-allofuranose
anes, 1/2 v/v) = 0.26 and 0.50; dH (300 MHz, CDCl3) 0.96 (2 ꢂ 6H,
2 ꢂ d, E-iBu-CH3, Z-iBu-CH3, 6.6 Hz), 1.37, 1.38, 1.58, 1.60
J
3-Azido-3-deoxy-1,2:5,6-di-O-isopropylidene-b-
D
(4 ꢂ 3H, 4 ꢂ s, E-C(CH3)2, Z-C(CH3)3), 2.01 (2 ꢂ 1H, m, E-iBu-CH, Z-
iBu-CH), 3.14 (1H, dd, Z-H-3, J 4.4 Hz, J 9.6 Hz), 3.33 (1H, dd, E-H-3,
J 4.5 Hz, 9.6 Hz), 3.89 (2H, m, E-iBu-CH2), 3.98 (2H, d, Z-iBu-CH2, J
6.6 Hz), 4.68 (1H, ddd, E-H-4, J 1.7 Hz, J 4.4 Hz, J 9.9 Hz), 4.82
(2 ꢂ 1H, m, E-H-2, Z-H-2), 5.68 (1H, t, Z-H-4, J 9.5 Hz), 5.86, 5.88
(2 ꢂ 1H, 2 ꢂ d, E-H-1, Z-H-1, J 3.6 Hz), 6.25 (1H, dd, Z-H-5, J 9.2 Hz,
J 11.4 Hz), 6.46 (1H, d, Z-H-6, J 11.4 Hz), 6.60 (1H, dd, E-H-6, J
1.7 Hz, J 15.1 Hz), 6.94 (1H, dd, E-H-5, J 4.2 Hz, J 11.5 Hz).
110 (4.31 g, 15.1 mmol) was dissolved in a mixture of acetic acid
(20 mL), water (10 mL) and MeOH (10 mL), and then stirred over-
night at 50 °C. The reaction mixture was then evaporated and co-
evaporated twice with toluene. The residue was redissolved in
MeOH (40 mL) and a solution of NaIO4 (3.89 g, 18.2 mmol) in water
(8 mL) was added. The resulting mixture was stirred for 15 min at
room temperature, after which the volatiles were evaporated. The
crude material was triturated with CHCl3 (50 mL) and after evapo-
ration the resulting oil was redissolved in dry THF (20 mL). After
cooling to ꢀ20 °C, a 2 M solution of LiBH4 in THF (7.55 mL) was
added, and this solution was stirred for 1 h at ꢀ20 °C. Then, brine
(20 mL) was added and the product was extracted with DCM
(2 ꢂ 50 mL). After drying on Na2SO4 and column chromatography
(EtOAc/hexanes, 2/1 v/v), 2 (2.89 g, 77%) was obtained as a white
The mixture was dissolved in THF (10 mL), cooled to ꢀ20 °C
after which sodium borohydride (131 mg, 3.47 mmol) was added.
The resulting suspension was allowed to slowly heat to room tem-
perature over 6 h. CHCl3 (25 mL) and water (10 mL) were added
and the layers separated. The organic layer was dried on Na2SO4
after which column chromatography (EtOAc/hexanes, 1/4 v/v)
afforded 3 (679 mg, 67%) as a clear colourless oil. Rf (EtOAc/hex-
solid. Rf (EtOAc/hexanes, 1/1 v/v) = 0.36; ½a D23
ꢃ
¼ þ124:4 (c 0.85,
anes, 1/3 v/v) = 0.29; ½a D23
¼ þ97:1 (c 0.69, CHCl3); dH (CDCl3,
ꢃ
CHCl3) {lit.11e
½
a 2D0
ꢃ
¼ þ130:4 (c 0.97)}; dH (300 MHz, CDCl3) 1.37,
300 MHz) 0.87 (6H, d, iBu-CH3, J 6.3 Hz), 1.24, 1.44 (6H, 2 ꢂ s,
C(CH3)2), 1.90 (2H, m, H-5a, H-5b), 2.19 (1H, m, iBu-CH), 3.04
(1H, dd, H-3, J 4.6 Hz, J 9.7 Hz), 3.16 (2H, m, H-6a, H-6b), 3.87
(2H, d, iBu-CH2, J 6.3 Hz), 3.99 (1H, m, H-4), 4.65 (1H, t, H-2, J
4.1 Hz), 5.68 (1H, d, H-1, J 3.6 Hz); dC (75.5 MHz, CDCl3) 18.2,
27.9 (2 ꢂ iBu-CH3), 25.8, 25.9 (C(CH3)2), 26.1 (C-5), 36.6 (iBu-CH),
46.0 (C-6), 63.8, 74.8, 79.7 (C-2, C-3, C-4), 75.5 (iBu-CH2), 103.7
(C-1), 112.7 (C(CH3)2); for C13H23N3O6S calcd C, 44.69; H, 6.63; N,
12.03; found C, 44.56; H, 6.88; N, 12.15.
1.57 (6H, 2 ꢂ s, C(CH3)2), 3.24 (1H, q, OH, J 4.1 Hz), 3.58 (1H, dd,
H-3, J 4.7 Hz, J 9.6 Hz), 3.69 (1H, m, H-5a), 3.95 (1H, m, H-5b),
4.11 (1H, m, H-4), 4.76 (1H, t, H-2, J 4.1 Hz), 5.81 (1H, d, H-1, J
3.6 Hz); dC (75.5 MHz, CDCl3) 25.9 (C(CH3)3), 58.9 (C-3), 59.6 (C-
5), 77.9, 79.8 (C-2, C-4), 103.8 (C-1), 112.7 (C(CH3)2); for
C8H13N3O4 calcd C, 44.65; H, 6.09; N, 19.53; found C, 44.42; H,
5.91; N, 19.27.
4.3. 3-Azido-3,5,6-trideoxy-6-(isobutyloxysulfon)-1,2-O-
isopropylidene-b-
D-ribo-hexofuranose 3
4.4. 1,2-Di-O-acetyl-3-azido-3,5,6-trideoxy-6-
(isobutyloxysulfon)-a/b-D-ribo-hexofuranose 7
Method A: 3-Azido-3-deoxy-1,2-O-isopropylidene-b-
D
-ribose 2
(2.32 g, 11.6 mmol) was dissolved in dry DCM (30 mL) and pyridine
(1.42 mL, 17.4 mmol) was added. After cooling to ꢀ20 °C, trifluoro-
methanesulfonic anhydride (2.15 mL, 12.8 mmol) was added in por-
tions. The mixture was stirred for 30 min at ꢀ20 °C, after which it
was washed with 1 M HCl (30 mL) and water (30 mL). After drying
on Na2SO4, the crude triflate was obtained as an off-white crystalline
solid which was used without further purification. Isobutyl methyl-
sulfonate (1.95 g, 12.8 mmol) and HMPA (2.0 mL) were dissolved in
dry THF (20 mL) under a nitrogen atmosphere and cooled to ꢀ78 °C.
Then, a 2.5 M solution of nBuLi in hexanes (5.12 mL) was added
dropwise. The resulting yellow solution was stirred at ꢀ78 °C for
1 h, after which a solution of the triflate in dry THF (5 mL) was added
and the resulting mixture was stirred for 2 h at ꢀ78 °C. The reaction
was quenched with acetic acid (4.0 mL) and after heating to room
temperature, saturated aqueous NaHCO3 (20 mL) was added. The
crude product was extracted with DCM (2 ꢂ 30 mL) and dried on
Na2SO4. Column chromatography (EtOAc/hexanes, 1/4 v/v) yielded
3 (2.07 g, 51%) as a clear, colourless oil.
3-Azido-3,5,6-trideoxy-6-(isobutyloxysulfon)-1,2-O-isopropyli-
dene-b- -ribo-hexofuranose 3 (1.25 g, 3.58 mmol) was dissolved in
D
a mixture of TFA (2.7 mL) and water (0.3 mL) and stirred for 30 min
at room temperature, after which the volatiles were evaporated
and the residue co-evaporated twice with toluene. Pyridine
(5.0 mL) and acetic anhydride (5.0 mL) were added and the result-
ing mixture was stirred overnight at room temperature. The sol-
vents were evaporated and the residue was co-evaporated twice
with toluene. Column chromatography (EtOAc/hexanes, 1/2 v/v)
afforded a mixture of anomers 7 (1.12 g, 80%) as a clear, colourless
oil. Rf (EtOAc/hexanes, 1/2 v/v) = 0.40;
CHCl3); dH (300 MHz, CDCl3) 0.99 (6H, d,
½
a 2D3
a
ꢃ
¼ þ23:9 (c 0.81,
-iBu-CH3, b-iBu-CH3, J
-H-5b, b-H-5b), 2.08
-Ac-CH3), 2.12 (0.6H, s, b-Ac-CH3), 3.24 (2H, m, -H-6a,
-H-6b, b-H-6b), 3.85 (0.2H, dd, -H-3, J 4.9 Hz, J 7.7 Hz),
-iBu-CH2,
J 6.6 Hz), 4.01 (1.6H, d, b-iBu-CH2, J 6.6 Hz), 4.12 (0.8H, m, b-H-
4), 4.19 (0.2H, m, -H-4), 5.26 (0.2H, dd, -H-2, J 4.6 Hz, J
7.5 Hz), 5.35 (0.8H, d, b-H-2, J 4.7 Hz), 6.11 (0.8H, s, b-H-1), 6.38
(0.2H, d, -H-1, J 4.6 Hz); dC (75.5 MHz, CDCl3) 18.2, 19.8, 2.1,
20.6 ( -iBu-CH3, b-iBu-CH3,
27.8, 27.9 ( b-Ac-CH3), 28.4 (b-C-5), 45.8 (b-C-6), 46.0 (
61.0, 71.4, 80.1 ( -C-2, -C-3,
3, b-C-4), 75.5 ( b-iBu-CH2), 93.3 (
b-C@O); for C14H23N3O8S calcd C,
6.6 Hz), 2.01–2.39 (2H, m,
a-H-5a, b-H-5a, a
(2.4H, s,
a
a
b-H-6a,
a
a
3.94 (0.8H, dd, b-H-3, J 4.8 Hz, J 7.5 Hz), 4.00 (0.4H, d,
a
a
a
Method B: 3-Azido-3-deoxy-1,2:5,6-di-O-isopropylidene-b-D-
allofuranose 1 (622 mg, 2.89 mmol) was dissolved in a mixture of
acetic acid (5.0 mL), H2O (2.5 mL) and MeOH (2.5 mL) and stirred
at 50 °C overnight. After evaporation of the solvents and co-evapora-
tion with toluene, the crude diol was redissolved in a mixture of
MeOH (10 mL) and water (2.0 mL), and sodium periodate (742 mg,
3.47 mmol) was added. The resulting mixture was stirred at room
temperature for 20 min after which all volatiles were evaporated.
Trituration with CHCl3 afforded the crude aldehyde. THF (10 mL)
was cooled to ꢀ20 °C, 60% NaH in mineral oil (127 mg, 3.17 mmol)
and isobutyl (diphenoxyphosphoryl) methanesulfonate 5 (1.10 g,
2.89 mmol) were added, and the resulting mixture was stirred at
ꢀ20 °C for 15 min, after which the crude aldehyde was added in
THF (5.0 mL). After stirring for three hours at ꢀ20 °C, CHCl3
(25 mL) and water (10 mL) were added and the layers separated.
The organic layer was dried on Na2SO4 affording isomers E-6 and
Z-6 as a 1:1 mixture as judged by 1H NMR integration. Rf (EtOAc/hex-
a
a
a-iBu-CH, b-iBu-CH), 27.6 (
a
a
-C-5),
-C-6),
a
a
a
a-C-4), 62.9, 75.6, 79.4 (b-C-2, b-C-
a
a-C-1), 97.8 (b-C-1), 168.5,
169.1, 169.2, 169.3 (2 ꢂ
a
42.74; H, 5.89; N, 10.68; found C, 42.45; H, 5.81; N, 10.91.
4.5. 3-Azido-3,5-dideoxy-5-(isobutyloxysulfonmethyl)-N6-
benzoyl-adenosine 8
To a suspension of N6-benzoyladenine (589 mg, 2.46 mmol) in
CH3CN (5.0 mL) N,O-bis(trimethylsilyl)acetamide (1.80 mL,
7.38 mmol) was added and this mixture was stirred at room temper-
ature for 30 min. The resulting clear solution was cooled to ꢀ20 °C
after which 1,2-di-O-acetyl-3-azido-3,5,6-trideoxy-6-(isobutyloxy-

J. van Ameijde et al. / Tetrahedron: Asymmetry 21 (2010) 469–475
473
sulfon)-
a
/b-
D
-ribo-hexofuranose 7 (484 mg, 1.23 mmol) and SnCl4
m, ArH, H-5), 7.71 (1H, d, H-6, J 7.5 Hz), 7.91 (2H, m, ArH), 9.24
(1H, br s, NH); dC (75.5 MHz, CDCl3) 18.5 (iBu-CH3), 20.6 (iBu-
CH), 26.7 (C-50), 28.2 (C(O)CH3), 46.1 (C-60), 62.5, 75.5, 79.4 (C-20,
C-30, C-40), 75.8 (iBu-CH2), 94.0 (C-10), 97.1 (C-5), 127.7, 128.8,
132.8, 133.2 (ArC), 146.3 (C-6), 154.3, 163.1, 166.7, 170.0 (C@O,
C@N); for C23H28N6O8S calcd C, 50.36; H, 5.14; N, 15.32; found C,
50.41; H, 5.38; N, 15.07.
(722
l
L, 6.15 mmol) were added. After stirring at ꢀ20 °C to room
temperature for 72 h, water (10 mL) was added and the crude prod-
uct was quickly extracted with CHCl3 (20 mL). After drying on
Na2SO4 and column chromatography (MeOH/DCM, 3/97 v/v), 8
(292 mg, 51%) was obtained as a clear colourless oil. Rf (MeOH/
DCM, 1/9 v/v) = 0.66; ½a D23
¼ þ6:6 (c 2.20, CHCl3); dH (300 MHz,
ꢃ
CDCl3) 0.94 (6H, d, iBu-CH3, J 6.6 Hz), 2.00 (1H, m, iBu-CH), 2.18
(3H, s, C(O)CH3), 2.43 (2H, m, H-50a, H-50b), 3.26 (2H, m, H-60a, H-
60b), 3.96 (2H, d, iBu-CH2, J 6.6 Hz), 4.21 (1H, m, H-40), 4.72 (1H, dd,
H-30, J 5.8 Hz, 7.4 Hz), 6.00 (1H, dd, H-20, J 3.3 Hz, 5.8 Hz), 6.03 (1H,
d, H-10, J 3.3 Hz), 7.54 (3H, m, ArH), 8.02 (2H, d, ArH, J 7.2 Hz), 8.11
(1H, s, H-2/8), 8.74 (1H, s, H-2/8), 9.39 (1H, br s, NH); dC (75.5 MHz,
CDCl3) 18.3 (iBu-CH3), 20.2 (iBu-CH), 26.9 (C-50), 28.0 (C(O)CH3),
45.9 (C-60), 62.6, 75.1, 79.5 (C-20, C-30, C-40), 75.6 (iBu-CH2), 87.9
(C-10), 123.9, 127.8, 128.5, 132.6, 133.1, 149.9, 151.0, 164.9, 169.6
(2 ꢂ C@O, C-2, C-4, C-5, C-6, C-8); for C24H28N8O7 calcd C, 50.34; H,
4.93; N, 19.57; found C, 50.05; H, 4.86; N, 19.37.
4.8. 3-Azido-3,5-dideoxy-5-(isobutyloxysulfonmethyl)-uridine 11
To a suspension of uracil (479 mg, 4.27 mmol) in CH3CN
(5.0 mL), N,O-bis(trimethylsilyl)acetamide (3.17 mL, 12.8 mmol)
was added and this mixture was stirred at room temperature for
30 min. The resulting clear solution was cooled to ꢀ20 °C after
which 1,2-di-O-acetyl-3-azido-3,5,6-trideoxy-6-(isobutyloxysul-
fon)-a/b-D-ribo-hexofuranose 7 (1.11 g, 2.28 mmol) and SnCl4
(1.32 mL, 11.3 mmol) were added. After stirring at ꢀ20 °C to room
temperature for 24 h, water (10 mL) was added and the crude
product was quickly extracted with CHCl3 (20 mL). After drying
on Na2SO4 and column chromatography (EtOAc/hexanes, 2/1 v/v),
11 (888 mg, 71%) was obtained as a clear, colourless oil. Rf
4.6. 3-Azido-3,5-dideoxy-5-(isobutyloxysulfonmethyl)-N2-
pivaloyl-guanosine 9
(EtOAc/hexanes, 3/1 v/v) = 0.30; ½a D23
¼ þ31:5 (c 1.38, CHCl3); dH
ꢃ
To a suspension of N2-pivaloylguanine (385 mg, 1.64 mmol) in
CH3CN (5.0 mL), N,O-bis(trimethylsilyl)acetamide (1.21 mL,
4.91 mmol) was added and this mixture was stirred at room temper-
ature for 30 min. The resulting clear solution was cooled to ꢀ20 °C
after which 1,2-di-O-acetyl-3-azido-3,5,6-trideoxy-6-(isobutyloxy-
(300 MHz, CDCl3) 0.98 (6H, d, iBu-CH3, J 6.9 Hz), 2.04 (1H, m,
iBu-CH), 2.19 (3H, s, Ac-CH3), 2.31 (2H, m, H-50a, H-50b), 3.31
(2H, m, H-60a, H-60b), 4.01 (3H, m, iBu-CH2, H-40), 4.27 (1H, dd,
H-30, J 6.5 Hz, J 8.2 Hz), 5.54 (1H, d, H-10, J 2.8 Hz), 5.62 (1H, dd,
H-20, J 2.8 Hz, J 6.5 Hz), 5.79 (1H, d, H-5, J 8.1 Hz), 7.31 (1H, d, H-
6, J 8.1 Hz), 10.3 (1H, s, H-3); dC (75.5 MHz, CDCl3) 18.3 (iBu-
CH3), 20.2 (iBu-CH), 26.5 (C-50), 28.0 (Ac-CH3), 45.7 (C-60), 62.2,
75.0, 78.8 (C-20, C-30, C-40), 75.7 (iBu-CH2), 92.4 (C-10), 102.6 (C-
5), 142.3 (C-6), 149.9, 163.6, 169.7 (3 ꢂ C@O); for C16H23N5O8S
calcd C, 43.14; H, 5.20; N, 15.72; found C, 42.88; H, 5.15; N, 15.59.
sulfon)-a/b-D-ribo-hexofuranose 7 (428 mg, 1.09 mmol) and SnCl4
(512
l
L, 4.36 mmol) were added. After stirring at ꢀ20 °C to room
temperature for 24 h, water (10 mL) was added and the crude prod-
uct was quickly extracted with CHCl3 (20 mL). After drying on
Na2SO4 and column chromatography (EtOAc/hexanes, 3/1 v/v), 9
(358 mg, 58%) was obtained as a white solid. Rf (EtOAc/hexanes, 3/
1 v/v) = 0.22; ½a 2D3
ꢃ
¼ þ47:7 (c 0.21, DMF); dH (300 MHz, CDCl3)
4.9. 3-Azido-3,5-dideoxy-5-(chlorosulfonmethyl)-uridine 12
0.99 (6H, d, iBu-CH3, J 6.8 Hz), 1.39 (6H, s, C(CH3)2), 2.05 (1H, m,
iBu-CH), 2.27 (3H, s, Ac-CH3), 2.47 (2H, m, H-50a, H-50b), 3.36 (1H,
m, H-60a), 3.54 (1H, m, H-60b), 4.04 (2H, d, iBu-CH2, J 6.7 Hz), 4.15
(1H, m, H-40), 4.85 (1H, dd, H-30, J 6.0 Hz, J 8.2 Hz), 6.07 (1H, d, H-
10, J 2.8 Hz), 6.34 (1H, dd, H-20, J 2.7 Hz, J 5.8 Hz), 7.81 (1H, s, H-8),
8.98 (1H, s, H-1), 12.3 (1H, s, 6-NH); dC (75.5 MHz, CDCl3) 18.5
(iBu-CH3), 20.6 (iBu-CH), 26.6 (C(CH3)3), 26.9 (C-50), 28.1 (Ac-CH3),
40.2 (C(CH3)3), 46.1 (C-60), 62.0, 77.4, 79.8 (C-20, C-30, C-40), 75.8
(iBu-CH2), 90.0 (C-10), 110.4, 147.5, 152.1, 157.8, 170.4, 181.1
(3 ꢂ C@O, C-2, C-4, C-5, C-6, C-8); for C22H32N8O8S calcd C, 46.47;
H, 5.67; N 19.71; found C, 46.20; H, 5.62; N, 19.84.
To a solution of 3-azido-3,5-dideoxy-5-(isobutyloxysulfon)-uri-
dine 11 (861 mg, 1.93 mmol) in acetone (10 mL), sodium iodide
(579 mg, 3.86 mmol) was added, and the resulting mixture was
stirred at reflux for 48 h. The resulting white precipitate was re-
moved by filtration and suspended in DCM (10 mL), after which
DMF (300 lL) and a 20% solution of phosgene in toluene (3.0 mL)
were added. This mixture was stirred for 5 h at room temperature
after which the solids were removed by filtration. The filtrate was
evaporated and subjected to column chromatography (EtOAc/hex-
anes, 2/1 v/v), after which 12 (480 mg, 61%) was obtained as a
white foam. Rf (EtOAc/hexanes, 1/1 v/v) = 0.19; ½a D23
¼ þ42:1 (c
ꢃ
4.7. 3-Azido-3,5-dideoxy-5-(isobutyloxysulfonmethyl)-N4-
benzoyl-cytidine 10
0.21, CHCl3); dH (300 MHz, CDCl3) 2.21 (3H, s, Ac-CH3), 2.44 (1H,
m, H-50a), 2.55 (1H, m, H-50b), 3.85 (2H, m, H-60a, H-60b), 4.04
(2H, m, H-30, H-40), 4.37 (1H, dd, H-20, J 6.3 Hz, J 8.5 Hz), 5.65
(1H, d, H-10, J 6.3 Hz), 5.78 (1H, d, H-5, J 8.0 Hz), 7.13 (1H, d, H-6,
J 8.0 Hz), 9.61 (1H, s, H-3); dC (75.5 MHz, CDCl3) 20.4 (Ac-CH3),
27.1 (C-50), 60.8 (C-60), 62.2, 75.2, 78.6 (C-20, C-30, C-40), 94.3 (C-
10), 102.9 (C-5), 143.2 (C-6), 150.0, 163.8, 170.1 (3 ꢂ C@O); for
C12H14ClN5O7S calcd C, 35.34; H, 3.46; N, 17.17; found C, 35.49;
H, 3.25; N, 16.92.
To a suspension of N4-benzoylcytosine (123 mg, 0.57 mmol) in
CH3CN
(2.0 mL),
N,O-bis(trimethylsilyl)acetamide
(418 lL,
1.71 mmol) was added and stirred at room temperature for
30 min. The resulting clear solution was cooled to ꢀ20 °C after
which 1,2-di-O-acetyl-3-azido-3,5,6-trideoxy-6-(isobutyloxysul-
fon)-a/b-
D-ribo-hexofuranose 7 (223 mg, 0.57 mmol) and SnCl4
(201
l
L, 1.71 mmol) were added. The mixture was allowed to
slowly heat to room temperature and stirred overnight, after which
water (10 mL) was added and the product was extracted quickly
with CHCl3 (10 mL). After drying on Na2SO4 and column chroma-
tography (EtOAc/hexanes, 1/1 v/v), 10 (198 mg, 63%) was obtained
4.10. 3-Azido-3,5-dideoxy-5-(dimethylaminosulfonmethyl)-N4-
benzoyl-cytidine 13
3-Azido-3,5-dideoxy-5-(isobutyloxysulfonmethyl)-N4-benzoyl-
cytidine 10 (66 mg, 0.12 mmol) and potassium iodide (20 mg,
0.12 mmol) were added to acetone (3.0 mL) and stirred at reflux
temperature for 48 h, after which the volatiles were evaporated.
The residue was suspended in DCM (2.5 mL) and a 20% solution
of phosgene in toluene (1.0 mL) was added. DMF (0.5 mL) was
added dropwise followed by stirring for 3 h at room temperature,
as a white foam. Rf (EtOAc/hexanes, 1/1 v/v) = 0.14; ½a D23
¼ þ42:3 (c
ꢃ
0.26, CHCl3); dH (300 MHz, CDCl3) 0.98 (6H, d, CH(CH3)2, J 6.9 Hz),
2.04 (1H, m, CH(CH3)2), 2.18 (3H, s, C(O)CH3), 2.36 (2H, m, H-50a,
H-50b), 3.34 (2H, m, H-60a, H-60b), 4.01 (2H, d, OCH2, J 6.6 Hz),
4.08 (1H, m, H-40), 4.33 (1H, dd, H-30, J 6.2 Hz, J 8.9 Hz), 5.61 (1H,
d, H-10, J 2.2 Hz), 5.76 (1H, dd, H-20, J 2.2 Hz, J 6.2 Hz), 7.49 (4H,

474
J. van Ameijde et al. / Tetrahedron: Asymmetry 21 (2010) 469–475
after which a clear, light brown solution was obtained. The sol-
References
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0.12 mmol) and NMM (33 lL, 0.30 mmol) were added. After stir-
ring for 1 h at room temperature, TLC analysis indicated complete
transformation into a new product and the reaction mixture was
washed with 1 M HCl (aq) and brine. After drying on Na2SO4 and
column chromatography (EtOAc/hexanes, 3/1 v/v), 13 (45 mg,
72%) was obtained as an off-white foam. Rf (EtOAc/hexanes, 3/1
v/v) = 0.18; ½a 2D3
¼ þ38:7 (c 0.61, CHCl3); dH (300 MHz, CDCl3)
ꢃ
2.20 (3H, s, C(O)CH3), 2.33 (2H, m, H-50a, H-50b), 2.90 (6H, s,
N(CH3)2), 3.09 (2H, m, H-60a, H-60b), 4.10 (1H, m, H-40), 4.33 (1H,
dd, H-30, J 6.2 Hz, J 8.8 Hz), 5.55 (1H, d, H-10, J 2.2 Hz), 5.72 (1H,
dd, H-20, J 2.2 Hz, J 6.2 Hz), 7.59 (5H, m, ArH, H-5, H-6), 7.93 (2H,
m, ArH), 9.63 (1H, br s, NH); dC (75.5 MHz, CDCl3) 20.6 (C(O)CH3),
26.3 (C-50), 37.5, 37.6 (N(CH3)2), 54.8 (C-60), 62.7, 75.7, 79.9 (C-20,
C-30, C-40), 94.8 (C-10), 97.4 (C-5), 127.8, 129.0, 132.7, 133.4 (ArC),
156.8 (C-6), 154.1, 163.2, 169.9 (3 ꢂ C@O); for C21H27N7O7S calcd
C, 48.36; H, 5.22; N, 18.80; found C, 48.61; H, 5.02; N, 18.69; m/z
(ESI +ve) 522.2 [M+H]+, C21H28N7O7S requires 522.2.
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4.11. 3-Azido-3,5-dideoxy-5-(isobutylaminosulfonmethyl)-
uridine 14
3-Azido-3,5-dideoxy-5-(chlorosulfon)-uridine
mol) was dissolved in DCM (1.0 mL) after which NMM
(10.1 L, 91 mol) and isobutylamine (6.7 L, 67 mol) were added.
12
(25 mg,
61
l
l
l
l
l
The resulting mixture was stirred for 90 min at room temperature,
after which it was washed with 1 M HCl (aq) and brine, dried on
Na2SO4 and purified by column chromatography (EtOAc/hexanes,
3/1 v/v) yielding 14 (24.6 mg, 91%) as a white foam. (EtOAc/hexanes,
3/1 v/v) = 0.55; ½a D23
¼ þ48:8 (c 0.32 g/100 mL, CHCl3); dH (300 MHz,
ꢃ
CDCl3) 0.97 (6H, d, CH(CH3)2, J 6.6 Hz), 1.79 (1H, m, CH(CH3)3), 2.20
(3H, s, C(O)CH3), 2.32 (2H, m, H-50a, H-50b), 2.93 (2H, m, NHCH2),
3.18 (2H, m, H-60a, H-60b), 3.99 (1H, m, H-40), 4.24 (1H, dd, H-30, J
6.3 Hz, J 8.2 Hz), 4.84 (1H, br s, CH2NH), 5.44 (1H, d, H-10, J 2.8 Hz),
5.60 (1H, dd, H-20, J 2.8 Hz, J 6.3 Hz), 5.78 (1H, d, H-5, J 8.2 Hz), 7.21
(1H, d, H-6, J 8.2 Hz), 9.62 (1H, br s, H-3); dC (75.5 MHz, CDCl3) 19.8
(CH(CH3)2), 20.5 (C(O)CH3), 27.1 (CH(CH3)2), 28.9 (C-50), 48.4
(NHCH2), 50.6 (C-60), 62.6, 75.4, 79.6 (C-20, C-30, C-40), 93.2 (C-10),
103.0 (C-5), 142.5 (C-6), 149.9, 163.5, 170.1 (C@O); for C16H24N6O7S
calcdC, 43.24;H, 5.44;N, 18.91;foundC, 43.28; H, 5.19;N, 18.73;m/z
(ESI +ve) 444.9 [M+H]+, C16H25N6O7S requires 445.1.
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4.12. UU-dimer 15
3-Azido-3,5-dideoxy-5-(isobutylaminosulfon)-uridine
(20.0 mg, 45 mol) was dissolved in THF (1.0 mL) after which acetic
acid (3.9 L, 68 mol) and 10% Pd/C (5.0 mg) were added. The result-
14
l
l
l
ing mixture was stirred under a H2 atmosphere for 2 h when TLC
indicated complete consumption of the starting material. The sol-
vents were evaporated and co-evaporated twice with toluene. The
residue was dissolved in DMF (1.0 mL) after which 12 (18.4 mg,
45 lmol) and NMM (14.8 lL, 135 lmol) were added. The reaction
mixture was stirred overnight at room temperature, after which
the volatiles were evaporated and the residue was purified by col-
umn chromatography (MeOH/DCM, 5/95 to 20/80 v/v), yielding 15
(23.8 mg, 67%) as a white foam. Rf (MeOH/DCM, 9/1 v/v) = 0.26; dH
(300 MHz, CDCl3) 0.98 (6H, d, CH(CH3)2, J 6.6 Hz), 1.81 (1H, m,
CH(CH3)2), 2.15 (6H, s, 2 ꢂ C(O)CH3), 2.31 (4H, m, 2 ꢂ H-50a, 2 ꢂ H-
50b), 2.90 (2H, NHCH2), 3.10 (4H, m, 2 ꢂ H-60a, 2 ꢂ H-60b), 3.99
(2H, m, 2 ꢂ H-40), 4.30 (1H, dd, H-30, J 6.4 Hz, J 8.0 Hz), 4.54 (1H, m,
H-30), 4.80 (1H, br s, NHCH2), 5.45 (1H, d, H-10, J 2.7 Hz), 5.58 (3H,
m, H-10, 2 ꢂ H-20), 5.78 (2H, m, 2 ꢂ H-5), 7.19 (2H, m, 2 ꢂ H-6),
9.50 (2H, br s, 2 ꢂ H-3); for C28H39N9O14S2 calcd C, 42.58; H, 4.98;
N, 15.96; found C, 42.29; H, 5.20; N, 15.77; m/z (ESI +ve) 789.9
[M+H]+, C28H40N9O14S2 requires 790.2.
12. Carretero, J. C.; Demillequand, M.; Ghosez, L. Tetrahedron 1987, 43, 5125.

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