Synthesis of 2′-deoxy-2′-C-α-methylpurine nucleosides

Article abstract of DOI:10.1055/s-2005-872204

2′-Deoxy-2′-C-α-methylribonucleosides provide valuable biochemical probes with which to study RNA structure and function. Using methyl 2-acetoxymethyl-3,5-di-O-(tert-butyldimethylsilyl)-D-ribofuranoside (1) as a glycosylating agent, we achieved in four st

Full text of DOI:10.1055/s-2005-872204

PAPER
2865
Synthesis of 2¢-Deoxy-2¢-C-a-methylpurine Nucleosides
2
¢
-
Deoxy-2
¢
-
C
-
a
-m
a
ethylpurin
n
e
N
ucleoside
-
s
Sheng Li,* Joseph A. Piccirilli*
Howard Hughes Medical Institute, Department of Biochemistry & Molecular Biology and Department of Chemistry, University of Chicago,
5841 S. Maryland Ave., MC 1028, Chicago, IL 60637, USA
Fax +1(773)7020271; E-mail: nli@uchicago.edu; E-mail: jpicciri@uchicago.edu
Received 15 April 2005; revised 26 May 2005
a)
Abstract: 2¢-Deoxy-2¢-C-a-methylribonucleosides provide valu-
creates "cavity"
fills "cavity"
able biochemical probes with which to study RNA structure and
function. Using methyl 2-acetoxymethyl-3,5-di-O-(tert-butyldi-
methylsilyl)-D-ribofuranoside (1) as a glycosylating agent, we
achieved in four steps an improved synthesis of 2¢-deoxy-2¢-C-a-
methyladenosine (8) and the first synthesis of 2¢-deoxy-C-a-meth-
ylguanosine (9) in 25% and 17% overall yield, respectively.
B
B
O
B
O
O
O
O
O
O
CH3
O
OH
O
Key words: nucleosides, deoxygenation, glycosylation, deoxy-
methyladenosine, deoxymethylguanosine
b)
X = CH3, SH, NH2, OH, Cl,
increasing electronegativity
F
Ribonucleic acids (RNA) adopt complex three-dimen-
sional architectures that mediate biological functions.
These architectures and their accompanying functions de-
pend inextricably on the presence of the 2¢-hydroxyl
group. This integral component of the RNA backbone in-
fluences ribose conformation and helix geometry, medi-
ates tertiary interactions via metal ion coordination and
hydrogen bonding, and provides a scaffold for the hydra-
tion network that accompanies the folded molecule.1
Understanding RNA biology therefore hinges on identifi-
cation of the residues that bear important hydroxyl groups
and elucidation of the underlying mechanisms by which
those hydroxyl groups impart function.
HO
B
O
X = CH3, Cl, SH, F, NH2, OH
decreasing hydrophobicity
OH
X
c)
controls for
extends OH
the alkyl group
away from ring
B
O
B
B
O
O
O
O
O
O
CH3
O
O
H
O
O
H
CH2
O
2¢-C-a-Methylnucleosides, aside from their potential bio-
technological and therapeutic value,2 serve as valuable an-
alogues by which to investigate how hydroxyl groups
engender functional behavior within RNA molecules. (1)
They complement deoxynucleosides as analogues with
which to evaluate the effect of eliminating the hydrogen
bond capacity of the hydroxyl group (Figure 1a).3 (2)
They impart unique chemical diversity to an array of 2¢-
modified nucleoside analogues, empowering the use of
quantitative structure activity relationships (QSAR) in the
analysis of RNA biology (Figure 1b).4 (3) They serve as
‘deoxynucleoside controls’ for 2¢-a-hydroxymethylnucle-
osides, thereby revealing the specific functional contribu-
tion of a hydroxyl group upon extending it away from the
ribose ring by one methylene unit (Figure 1c). The effect
of hydroxymethyl nucleoside substitution on an RNA-
mediated process relative to that of methyl nucleoside
substitution gives a measure of whether a 2¢-hydroxyl
group imparts its functional contribution via inductive
effects or through-space interactions with solvent
H
H
d)
B
HO
O
OH
X
X = OH, Cl, CH3, SH
2'-OH
12 Å3
2'-Cl
27 Å3
2'-CH3
32 Å3
2'-SH
37 Å3
increasing molecular volume
Figure 1 Applications for 2¢-C-a-methylnucleosides in the study of
RNA structure and function. a) Probing the importance of the 2¢-hy-
droxyl group’s space-filling capacity. b) Quantitative structure-activi-
ty relationship analysis. c) Probing the importance of solvent
interactions with the 2¢-hydroxyl group. d) Probing the spatial envi-
ronment (the packing density) surrounding a 2¢-hydroxyl group
within a structured RNA. See text for further description and explana-
tion.
SYNTHESIS 2005, No. 17, pp 2865–2870
x
x.
x
x
.
2
0
0
5
Advanced online publication: 23.08.2005
DOI: 10.1055/s-2005-872204; Art ID: M02705SS
© Georg Thieme Verlag Stuttgart · New York

2866
N.-S. Li, J. A. Piccirilli
PAPER
(Figure 1c).4 (4) Together with 2¢-chloro and 2¢-mercapto- report the practical utility of this approach for the synthe-
nucleosides, 2¢-methylnucleosides form a series of ana- sis of the purine analogues.
logues that span a narrow range (~10 Å3) of molecular
Sugar reagent 1 glycosylates persilylated uracil or cy-
volumes so as to provide a sensitive measure of the spatial
tosine in acetonitrile in the presence of SnCl4 to give the
environment (the packing density) surrounding a 2¢-hy-
corresponding nucleoside derivatives in about 60% yield
droxyl group within a structured RNA molecule. The re-
with b/a selectivity of 94:6 and 90:10, respectively.6
sulting packing density metric offers a strategy to evaluate
However, the analogous glycosylation reaction of persily-
the functional validity of crystallographic and biochemi-
lated N6-benzoyladenine or N2-acetylguanine gives com-
cal RNA structure models (Figure 1d).5
plex results, presumably due to reaction at both the N-7
The preceding experimental approaches for the study of and N-9 positions. Consequently, we employed two pre-
RNA biology require access to analogues of the four nat- viously established experimental strategies to enhance the
ural nucleosides, adenosine, guanosine, cytidine, and uri- yield of the N-9 isomer.11 (1) We used trimethylsilyl trif-
dine. We previously reported efficient syntheses of the late as a catalyst in combination with a nonpolar solvent,
pyrimidine nucleosides: 2¢-deoxy-2¢-C-a-methyluridine either 1,2-dichloroethane or toluene; and (2) we conduct-
and 2¢-deoxy-2¢-C-a-methylcytidine.6 Here we report the ed the reactions at reflux temperature to favor formation
synthesis of the purine nucleoside analogues: 2¢-deoxy-2¢- of the thermodynamically stable product (N-9 isomer).
C-a-methyladenosine and 2¢-deoxy-2¢-C-a-methylgua-
nosine.
In the presence of trimethylsilyl triflate, N6-benzoyl
bis(trimethylsilyl)adenine reacted with 1 to give only the
The literature contains no report of the synthesis of 2¢- N-9 isomer (Scheme 1). However, the yield and anomeric
deoxy-2¢-C-a-methylguanosine and only one report of the ratio depended on the conditions, including reaction tem-
synthesis of 2¢-deoxy-2¢-C-a-methyladenosine.7 Novak et perature and time (Table 1). Reaction in refluxing toluene
al. obtained 2¢-deoxy-2¢-C-a-methyladenosine from the (~110 °C) or p-xylene (~138 °C) resulted in decomposi-
reaction of 2-a-methyl-3,5-di-O-(4-methylbenzoyl)ribo- tion of the products (entries 1 and 2). In contrast, reaction
furanosyl chloride with chloromercuri-N-benzoyladenine, in refluxing dichloroethane for 1 hour gave the N-9 iso-
followed by removal of the protecting groups. The glyco- mer in 61% yield with b/a anomeric ratio of ~3.7:1 (entry
sylation reaction gave a mixture of anomers, and no infor- 3). Allowing the reaction to proceed at room temperature
mation about yield or selectivity was provided.7 for 2 or 42 hours before initiating reflux decreased the ste-
Moreover, the synthesis of the glycosylating agent, 2-a- reoselectivity (entries 4 and 5). Refluxing for 19 hours re-
methyl-3,5-di-O-(4-methylbenzoyl)ribofuranosyl chlo- sulted in product decomposition, reducing the yield of 2b
ride, required six steps from 2-C-b-methyl-D-ribonolac- to only 8% (entry 6).
tone and gave no stereoselectivity, rendering this
published synthesis unattractive for accessing 2¢-deoxy-
2¢-C-a-methylpurine nucleosides.
XO
XO
i)
O
O
OMe
CH2OAc
To synthesize 2¢-deoxy-2¢-C-a-methylpurine nucleosides,
we considered adapting approaches used previously for
the synthesis of 2¢-deoxy-2¢-C-a-methylpyrimidine nucle-
osides. These approaches include both a linear strategy in-
volving nucleoside transformation and a convergent
strategy involving glycosylation of the pyrimidine nucle-
obases with the appropriate modified sugar derivative.
The linear strategy from nucleosides usually lacks effi-
ciency, as the desired 2¢-a-methyl isomers emerge as mi-
nor products.8,9 The convergent strategy using modified
sugar reagents generally has proven more efficient than
nucleoside transformation, as glycosylation of persilylat-
ed nucleobases gives the desired 1¢-b-isomers predomi-
nantly.10
ABz
13% (2a)
48% (2b)
XO
XO
CH2OAc
2a
1
+
X = TBDMS
XO
ABz
NHBz
N
O
N
ABz
=
CH2OAc
XO
N
N
2b
Scheme 1 Reagents and conditions: i) N6-benzoyl bis(trimethylsi-
lyl)adenine (2.0 equiv), 1,2-dichloroethane, TMSOTf (1.0 equiv), re-
flux, 1 h
Recently we exploited the glycosylation strategy to devel-
op an efficient synthesis of 2¢-C-a-methyl-2¢-deoxypyri-
midine nucleosides via radical deoxygenation of 2¢-C-a- Compared to persilylated N6-benzoyladenine, the persily-
hydroxymethylnucleosides.6 We accessed the hydroxy- lated N2-acetylguanine reacted with 1 to produce the de-
methylnucleosides directly and efficiently from the readily sired isomer with even weaker regio- and stereo-
prepared ribose derivatives, methyl 2-a-acetoxymethyl- selectivity. In dichloroethane at room temperature with
3,5-di-O-(4-chlorobenzyl)-2-deoxy-D-ribofuranoside and SnCl4 as a catalyst, the reaction yielded N7-isomer 3c pre-
methyl 2-a-acetoxymethyl-3,5-di-O-(tert-butyldimethyl- dominately. However, reaction in refluxing dichloroeth-
silyl)-2-deoxy-D-ribofuranoside (1). These reagents gly- ane for one hour using trimethylsilyl triflate (1.0 equiv) as
cosylate pyrimidines with high b-selectivity.6 Here we a catalyst gave the best results, producing the desired 3b
Synthesis 2005, No. 17, 2865–2870 © Thieme Stuttgart · New York

PAPER
2¢-Deoxy-2¢-C-a-methylpurine Nucleosides
2867
XO
XO
deoxy-2¢-hydroxymethylpurines 4 and 5 in 86 and 89%
yields, respectively (Scheme 3). Reaction of 4 and 5 with
phenyl chlorothionoformate followed by reaction with
tributyltin hydride in refluxing benzene produced the
methylnucleoside derivatives 6 and 7 in 64 and 53%
yields, respectively. Desilylation gave the final product
2¢-deoxy-2¢-C-a-methyladenine (8) and 2¢-deoxy-2¢-C-a-
methylguanosine (9) in 94 and 95% yield, respectively.
i)
O
O
OMe
CH2OAc
12% (3a)
38% (3b)
8% (3c)
GAc
CH2OAc
XO
XO
1
3a
+
XO
GAc
O
XO
XO
B
B
i)
O
O
X = TBDMS
CH2OAc
XO
86% (4)
89% (5)
O
CH2OH
CH2OAc
XO
XO
3b
N
NH
GAc
=
2b: X = TBDMS, B = ABz
3b: X = TBDMS, B = GAc
4: X = TBDMS, B = A
5: X = TBDMS, B = G
+
N
N
N
NHAc
NHAc
XO
7GAc
O
N
XO
HO
7GAc
=
B
B
ii)
iii)
O
O
N
N
CH2OAc
XO
94% (8)
95% (9)
64% (6)
O
53% (7)
3c
CH3
CH3
XO
HO
Scheme 2 Reagents and conditions: i) N2-acetyl tris(trimethylsi-
lyl)guanine (1.5 equiv), 1,2-dichloroethane, TMSOTf (1.0 equiv), re-
flux, 1 h
6: X = TBDMS, B = A
7: X = TBDMS, B = G
8: B = A
9: B = G
Scheme 3 Reagents and conditions: i) NH3, MeOH, 0–4 °C, 72 h
for 4; 0 °C to r.t., 24 h for 5; ii) 1) PhOCSCl, DMAP, CH2Cl2, r.t., 3–
5 h; 2) Bu3SnH/AIBN, benzene, reflux, 5–7 h; iii) TBAF, THF, r.t.,
3.5–12 h
Table 1 Glycosylation of N6-Benzoyl Bis(trimethylsilyl)adenine
with Reagent 1
Runa Solvent
1d,e
p-xylene
2d,e,f toluene
Reaction conditions Yield (%)b
2b/2ac
–
We confirmed the structures of 2a and 2b by NOESY ex-
periments. For 2a, 1¢-H (d = 6.70) exhibited a stronger
NOE with 2¢-H (d = 3.20) than with 2¢-CH2 (d = 3.95–
3.89); additionally, 8-H (d = 8.50) exhibited stronger
NOEs with 2¢-CH2 (d = 3.95-3.89) and 4¢-H (d = 4.45)
than with 1¢-H (d = 6.70) or 2¢-H (d = 3.20). We also ob-
served a strong NOE between 2¢-H (d = 3.20) and 3¢-H
(d = 4.56). These results suggest that 2a has the a-config-
uration at the 1¢-carbon. In contrast, the NOESY spectra
for 2b exhibited a pattern consistent with the b-configura-
tion; we observed that 1¢-H (d = 6.27) exhibited a stronger
NOE with 2¢-CH2 (d = 4.18) than with 2¢-H (d = 3.24).
Conversely, the 8-H (d = 8.28) exhibited a strong NOE
with 2¢-H (d = 3.24) and no NOE with 2¢-CH2 (d = 4.38,
4.18). We also observed a strong NOE between 2¢-H (d =
3.24) and 3¢-H (d = 4.55). These results suggest that 2b
has the b-configuration at the 1¢-carbon. Comparison of
the 13C NMR data for the C-4, C-5 and C-8 carbons of 2a
and 2b with 7- and 9-methyladenine supported our regio-
chemical assignment of 2a and 2b as 9-glycosylad-
enines.12
reflux, 2 h
reflux, 3 h
reflux, 1 h
decomposed
decomposed
–
3
ClCH2CH2Cl
61
–
3.7:1
2.9:1
4e
ClCH2CH2Cl
ClCH2CH2Cl
ClCH2CH2Cl
0 °C to r.t., 2 h;
then reflux, 1 h
5e
6
0 °C to r.t., 42 h;
then reflux, 1 h
–
8
2.1:1
reflux, 19 h
2b
only
a All experiments were carried out with N6-benzoyl bis(trimethylsi-
lyl)adenine (2.0 equiv) in the presence of trimethylsilyl triflate (1.0
equiv) under argon.
b Isolated yield.
c Ratio was estimated by 1H NMR of the crude product.
d Ratio not calculated, and product was decomposed.
e Product not isolated.
f Reaction was incomplete, and product was partially decomposed.
Analogously, we confirmed the structures of 3a and 3c by
NOESY experiments. For 3a, we observed that 1¢-H (d =
6.28) exhibited a stronger NOE cross peak with 2¢-H (d =
3.05) than with 2¢-CH2 (d = 4.02, 3.83), and that 3¢-H (d =
4.48) exhibited strong NOE cross peaks with 2¢-H (d =
3.05) and 5¢-H (d = 3.70, 3.58). These results suggest that
3a has the a-configuration at 1¢-carbon. For 3c, we ob-
served that 1¢-H (d = 6.46) exhibited stronger NOE cross
in 38% yield along with smaller amounts of 3a (12%) and
3c (8%) (Scheme 2). As described for N6-benzoyladenine
above, refluxing in higher boiling solvents such as p-xy-
lene decomposed the product.
To transform 2b and 3b to the corresponding 2¢-C-a-
methylnucleosides, we first treated them with ammonia in
methanol to obtain 3¢,5¢-di-O-(tert-butyldimethylsilyl)-2¢-
Synthesis 2005, No. 17, 2865–2870 © Thieme Stuttgart · New York

2868
N.-S. Li, J. A. Piccirilli
PAPER
eluting with 40% EtOAc in hexane, to give compound 2a (87 mg,
13%, the slow isomer) and 2b (317 mg, 48%, the fast isomer).
peaks with 2¢-CH2 (d = 4.42, 4.28) than with 2¢-H (d =
2.95); we also observed strong NOE cross peaks between
8-H (d = 8.25) and 2¢-H (d = 2.95). These results suggest
that 3c has the b-configuration at the 1¢-carbon. Compari-
son of the 13C NMR data for the C-4, C-5, and C-8 carbons
of 3a, 3b and 3c with N2-acetyl-2¢,3¢,5¢-tri-O-acetyl-7-
and 9-guanosine supported our regiochemical assign-
ments of 3a and 3b as 9-glycosylguanines and 3c as a 7-
glycosylguanine.13 We confirmed the structure of 3b indi-
rectly by the NOESY spectra of the final product: 2¢-
deoxy-2¢-C-a-methylguanosine (9). We observed that 1¢-
H (d = 5.64) exhibited a stronger NOE cross peak with 2¢-
CH3 (d = 0.90) than with 2¢-H (d = 2.76). We also ob-
served a strong NOE between 8-H (d = 7.84) and 2¢-H
(d = 2.76), but no NOE between 8-H (d = 7.84) and 2¢-
CH3 (d = 0.90), 3¢-H (d = 4.24), 4¢-H (d = 4.03), or 5¢-H
(d = 3.65). These results support our assignment of com-
pound 9 as the b-nucleoside.
2a
1H NMR (CDCl3/TMS): d = 9.22 (br s, 1 H, 6-NH), 8.78 (s, 1 H, 2-
H), 8.50 (s, 1 H, 8-H), 8.01 (d, 2 H, J = 7.2 Hz), 7.56 (m, 1 H), 7.49
(t, 2 H, J = 7.7 Hz), 6.70 (d, 1 H, J = 7.4 Hz, 1¢-H), 4.56 (m, 1 H, 3¢-
H), 4.45 (m, 1 H, 4¢-H), 3.95–3.89 (m, 2 H, 2¢-CH2), 3.74 (dd, 1 H,
J = 3.3, 10.9 Hz, 5¢-H), 3.63 (m, 1 H, 5¢-H), 3.20 (m, 1 H, 2¢-H),
1.90 (s, 3 H), 0.92 (s, 9 H), 0.91 (s, 9 H), 0.166 (s, 3 H), 0.115 (s, 3
H), 0.100 (s, 3 H), 0.095 (s, 3 H).
13C NMR (CDCl3): d = 170.2, 164.7, 152.6, 151.7, 149.3, 142.8,
133.8, 132.6, 128.7, 127.8, 122.2, 90.2, 85.2, 73.7, 63.5, 58.9, 46.7,
25.8, 25.7, 20.6, 18.3, 18.0, –4.7, –5.3, –5.4, –5.6.
HRMS: m/z calcd for C32H50N5O6Si2 [MH+]: 656.3300; found:
656.3321.
2b
1H NMR (CDCl3/TMS): d = 9.44 (br s, 1 H, 6-NH), 8.74 (s, 1 H, 2-
H), 8.28 (s, 1 H, 8-H), 7.99 (d, 2 H, J = 7.4 Hz), 7.54 (t, 1 H, J = 7.3
Hz), 7.45 (t, 2 H, J = 7.6 Hz), 6.27 (d, 1 H, J = 8.5 Hz, 1¢-H), 4.55
(m, 1 H, 3¢-H), 4.38 (dd, 1 H, J = 7.4, 11.2 Hz, 2¢-CH2), 4.18 (dd, 1
H, J = 7.2, 11.2 Hz, 2¢-CH2), 4.09 (m, 1 H, 4¢-H), 3.88 (dd, 1 H,
J = 4.7, 11.2 Hz, 5¢-H), 3.75 (dd, 1 H, J = 2.7, 11.2 Hz, 5¢-H), 3.24
(m, 1 H, 2¢-H), 1.72 (s, 3 H), 0.91 (s, 9 H), 0.90 (s, 9 H), 0.101 (s, 3
H), 0.094 (s, 3 H), 0.088 (s, 3 H), 0.080 (s, 3 H).
In summary, we have established synthetic access to 2¢-
deoxy-2¢-C-a-methylpurine nucleosides starting from the
glycosylating agent, methyl 3,5-di-O-(tert-butyldimethyl-
silyl)-2-a-acetoxyribofuranoside. As expected, this re-
agent and its precursor exhibit greater stereo- and
regioselectivity with pyrimidines than with purines. Nev-
ertheless, we have optimized conditions that give the 2¢-
deoxy-2¢-C-a-methylpurine nucleosides in yields that are
practically useful. Moreover, our overall strategy offers
both the methyl and hydroxymethyl analogues of the four
natural nucleosides in a highly convergent manner from a
common sugar reagent. These analogues may be convert-
ed either to the corresponding phosphoramidite for site-
specific incorporation by solid phase synthesis or to the
corresponding nucleoside triphosphates (or a-thiotriphos-
phates) for incorporation into RNA by transcription.
13C NMR (CDCl3): d = 170.4, 164.6, 152.7, 151.8, 149.5, 141.6,
133.7, 132.8, 128.8, 127.9, 123.2, 88.1, 86.6, 73.5, 63.3, 60.2, 48.2,
25.9, 25.7, 20.5, 18.4, 18.0, –4.6, –5.1, –5.3, –5.5.
HRMS: m/z calcd for C32H50N5O6Si2 [MH+]: 656.3300; found:
656.3292.
2¢-C-a-Acetoxymethyl-3¢,5¢-di-O-(tert-butyldimethylsilyl)-N2-
acetyl-a-guanosine (3a), 2¢-C-a-Acetoxymethyl-3¢,5¢-di-O-(tert-
butyldimethylsilyl)-N2-acetyl-b-guanosine (3b) and 2¢-C-a-Ace-
toxymethyl-3¢,5¢-di-O-(tert-butyldimethylsilyl)-N2-acetyl-b-(N-
7)-guanosine (3c)
Persilylated N2-acetylguanine was prepared by the reaction of N2-
acetylguanine (580 mg, 3.0 mmol) with hexamethyldisilazane (10
mL) in anhyd pyridine (4.0 mL) as described for the persilylation of
N6-benzoyladenine in the synthesis of 2a and 2b. Under argon, the
persilylated guanine was dissolved into anhyd 1,2-dichloroethane
(20 mL), and 1 (896 mg, 2.0 mmol) and TMSOTf (0.36 mL, 2.0
mmol) were added. The mixture was then refluxed for 1 h, allowed
to cool, and then quenched with sat. aq NaHCO3. The organic phase
was separated, and the aqueous phase was extracted with CH2Cl2.
The organic layers were combined, washed with brine and dried
(MgSO4). After removal of the solvent, the residue was purified by
silica gel chromatography, eluting with 2.5% MeOH in CHCl3, to
give compound 3a (152 mg, 12% yield, the slow isomer), 3b (462
mg, 38% yield, the middle isomer), and 3c (100 mg, 8% yield, the
fast isomer).
1H and 13C NMR spectra were recorded on GE 500, Bruker 500 or
Bruker 400 MHz NMR spectrometer. 1H chemical shifts are report-
ed in d (ppm) relative to tetramethylsilane and 13C chemical shifts d
(ppm) relative to the solvent used. High Resolution Mass spectra
were obtained from the Department of Chemistry, University of
California at Riverside on the VG-ZAB instrument. Methyl 2-a-ac-
etoxymethyl-3,5-di-O-(tert-butyldimethylsilyl)-2-deoxy-D-ribo-
furanoside (1) was prepared according to our reported procedure.6
2¢-C-a-Acetoxymethyl-3¢,5¢-di-O-(tert-butyldimethylsilyl)-N6-
benzoyl-a-adenosine (2a) and 2¢-C-a-Acetoxymethyl-3¢,5¢-di-O-
(tert-butyldimethylsilyl)-N6-benzoyl-b-adenosine (2b)
A mixture of N6-benzoyladenine (478 mg, 2.0 mmol), 1,1,1,3,3,3-
hexamethyldisilazane (5.0 mL) and pyridine (2.5 mL) was stirred
under argon at reflux for 3 h to obtain a clear solution. The solution
was evaporated to dryness, and the residue was dried under vacuum
overnight. The persilylated adenine was dissolved into anhyd 1,2-
dichloroethane (10 mL) under argon, and compound 1 (448 mg, 1.0
mmol) was added, followed by slow addition of TMSOTf (0.18 mL,
1.0 mmol). The mixture was then refluxed for 1 h. TLC showed that
1 was completely consumed. The mixture was cooled and quenched
with sat. aq NaHCO3. The organic phase was separated, and the
aqueous phase was extracted with CH2Cl2. The organic layers were
combined, washed with brine and dried (MgSO4). After solvent was
removed, the residue was purified by silica gel chromatography,
3a
1H NMR (CDCl3/TMS): d = 9.73 (br s, 1 H), 8.12 (s, 1 H, 8-H), 6.28
(d, 1 H, J = 7.2 Hz, 1¢-H), 4.48 (m, 1 H, 3¢-H), 4.41 (m, 1 H, 4¢-H),
4.02 (m, 1 H, 2¢-CH2), 3.83 (dd, 1 H, J = 8.8, 10.8 Hz, 2¢-CH2), 3.70
(m, 1 H, 5¢-H), 3.58 (m, 1 H, 5¢-H), 3.05 (m, 1 H, 2¢-H), 2.28 (s, 3
H), 1.94 (s, 3 H), 0.90 (s, 9 H), 0.88 (s, 9 H), 0.139 (s, 3 H), 0.084
(s, 3 H), 0.079 (s, 3 H), 0.076 (s, 3 H).
13C NMR (CDCl3): d = 172.2, 170.3, 155.9, 148.2, 147.4, 138.9,
120.4, 90.2, 85.1, 73.3, 63.6, 58.5, 46.8, 25.8, 25.7, 24.4, 20.7, 18.3,
18.0, –4.7, –5.4.
HRMS: m/z calcd for C27H48N5O7Si2 [MH+]: 610.3092; found:
610.3119.
Synthesis 2005, No. 17, 2865–2870 © Thieme Stuttgart · New York

PAPER
2¢-Deoxy-2¢-C-a-methylpurine Nucleosides
2869
3b
3¢,5¢-Di-O-(tert-butyldimethylsilyl)-2¢-C-a-methyl-b-adenosine
(6)
1H NMR (CDCl3/TMS): d = 10.15 (br s, 1 H), 7.92 (s, 1 H, 8-H),
5.91 (d, 1 H, J = 8.4 Hz, 1¢-H), 4.48 (m, 1 H, 3¢-H), 4.25 (m, 1 H, 2¢-
CH2), 4.15 (m, 1 H, 2¢-CH2), 4.00 (m, 1 H), 3.73 (m, 2 H, 5¢-H), 2.95
(m, 1 H, 2¢-H), 2.25 (s, 3 H), 1.77 (s, 3 H), 0.86 (s, 9 H), 0.83 (s, 9
H), 0.06–0.02 (m, 12 H).
To a solution of 4 (94 mg, 0.19 mmol) and DMAP (147 mg, 1.2
mmol) in anhyd CH2Cl2 (10 mL) at 0 °C was added phenyl chlo-
rothionoformate (28 mL, 0.20 mmol). After stirring the mixture at
r.t. for 3 h, the mixture was diluted with EtOAc, and the EtOAc lay-
er was washed consequently with sat. aq NaHCO3 and brine. The
organic layer was dried (MgSO4). The solvent was removed, and the
residue was dried under vacuum and used directly in the next step
without purification. To the solution of dried crude thionocarbonyl
ester in benzene (15 mL) under argon, were added Bu3SnH (0.30
mL, 1.1 mmol) and AIBN (10 mg, 0.060 mmol). The mixture was
refluxed under stirring for 5 h. The solvent was removed and the
residue was purified by silica gel chromatography, eluting with
50% EtOAc in hexane, to give compound 6 as a white foam (58 mg,
64%).
1H NMR (CDCl3/TMS): d = 8.34 (s, 1 H), 8.08 (s, 1 H), 6.06 (br s,
2 H), 6.01 (d, 1 H, J = 8.4 Hz), 4.36 (m, 1 H), 4.06 (m, 1 H), 3.85
(dd, 1 H, J = 4.8, 10.8 Hz), 3.75 (dd, 1 H, J = 3.2, 11.2 Hz), 2.82 (m,
1 H), 1.05 (d, 3 H, J = 6.8 Hz), 0.920 (s, 9 H), 0.917 (s, 9 H), 0.09
(s, 12 H).
13C NMR (CDCl3): d = 155.5, 153.0, 150.1, 139.0, 119.8, 89.3,
87.7, 74.9, 63.5, 43.9, 26.0, 25.8, 18.4, 18.1, 9.3, –4.6, –4.9, –5.4,
–5.5.
13C NMR (CDCl3): d = 172.1, 170.4, 156.0, 148.4, 147.4, 137.0,
120.8, 87.8, 85.9, 73.1, 63.2, 59.9, 48.7, 25.7, 25.5, 24.2, 20.4, 18.2,
17.8, –4.8, –5.3, –5.6, –5.7.
HRMS: m/z calcd for C27H48N5O7Si2 [MH+]: 610.3092; found:
610.3108.
3c
1H NMR (CDCl3/TMS): d = 11.51 (br s, 1 H), 8.25 (s, 1 H, 8-H),
6.46 (d, 1 H, J = 7.7 Hz, 1¢-H), 4.51 (m, 1 H, 3¢-H), 4.42 (m, 1 H, 2¢-
CH2), 4.28 (m, 1 H, 2¢-CH2), 4.07 (m, 1 H, 4¢-H), 3.90 (m, 1 H, 5¢-
H), 3.79 (m, 1 H, 5¢-H), 2.95 (m, 1 H, 2¢-H), 2.40 (s, 3 H), 1.86 (s,
3 H), 0.923 (s, 9 H), 0.917 (s, 9 H), 0.12 (s, 3 H), 0.11 (s, 3 H), 0.10
(s, 3 H), 0.09 (s, 3 H).
13C NMR (CDCl3): d = 173.4, 170.4, 156.8, 152.8, 148.2, 141.6,
111.4, 88.5, 87.7, 72.7, 63.0, 60.1, 50.2, 25.9, 25.6, 24.6, 20.6, 18.3,
17.9, –4.7, –5.1, –5.5, –5.6.
HRMS: m/z calcd for C27H48N5O7Si2 [MH+]: 610.3092; found:
610.3114.
HRMS: m/z calcd for C23H44N5O3Si2 [MH+]: 494.2983; found:
494.2987.
3¢,5¢-Di-O-(tert-butyldimethylsilyl)-2¢-C-a-hydroxymethyl-b-
adenosine (4)
3¢,5¢-Di-O-(tert-butyldimethylsilyl)-2¢-C-a-methyl-b-guanosine
(7)
Ammonia was bubbled into a solution of 2b (114 mg, 0.17 mmol)
in MeOH (10 mL) at 0 °C for 30 min. The flask was sealed and re-
frigerated (0–4 °C) for 3 d. The solvent was removed, and the resi-
due was purified by silica gel chromatography, eluting with 7.5%
MeOH in CHCl3, to give compound 4 as a white solid powder (76
mg, 86%).
1H NMR (CDCl3/TMS): d = 8.23 (s, 1 H), 8.06 (s, 1 H), 6.39 (br s,
2 H), 6.30 (d, 1 H, J = 7.4 Hz), 4.60 (dd, 1 H, J = 2.2, 5.9 Hz), 4.06
(m, 1 H), 3.93 (m, 2 H), 3.86 (dd, 1 H, J = 5.1, 11.1 Hz), 3.73 (dd,
1 H, J = 3.6, 11.1 Hz), 3.02 (m, 1 H), 0.91 (s, 9 H), 0.88 (s, 9 H),
0.112 (s, 3 H), 0.109 (s, 3 H), 0.06 (s, 6 H).
13C NMR (CDCl3): d = 155.6, 152.7, 149.3, 139.0, 119.8, 87.7,
87.6, 74.2, 62.9, 59.0, 50.9, 25.9, 25.7, 18.3, 17.9, –4.7, –5.1, –5.5,
–5.6
HRMS: m/z calcd for C23H44N5O4Si2 [MH+]: 510.2932; found:
510.2925.
To a solution of 5 (245 mg, 0.47 mmol) and DMAP (374 mg, 3.1
mmol) in anhyd CH2Cl2 (10 mL) at 0 °C was added phenyl chlo-
rothionoformate (71 mL, 0.51 mmol). After stirring the mixture at
r.t. for 5 h, the reaction was quenched with MeOH (0.50 mL). The
mixture was diluted with EtOAc, and the EtOAc layer was conse-
quently washed with sat. aq NaHCO3 and brine, The organic layer
was dried (MgSO4). The solvent was removed, and the residue was
dried under vacuum overnight. The dried crude thionocarbonyl es-
ter was dissolved into benzene (20 mL) under argon, and Bu3SnH
(0.67 mL, 2.5 mmol) and AIBN (20.0 mg, 0.12 mmol) were added.
The mixture was refluxed under stirring for 7 h. The solvent was re-
moved and the residue was purified by silica gel chromatography,
eluting with 6% MeOH in CH2Cl2 to give compound 7 as a white
foam (126 mg, 53%).
1H NMR (CDCl3/TMS): d = 7.75 (s, 1 H), 6.52 (br s, 1 H), 5.77 (d,
1 H, J = 4.0 Hz), 4.34 (m, 1 H), 4.03 (m, 1 H), 3.76 (m, 2 H), 2.72
(m, 1 H), 1.10–0.85 (m, 21 H), 0.20–0.01 (m, 12 H).
13C NMR (CDCl3): d = 159.2, 153.9, 152.0, 135.5, 117.3, 88.2,
87.5, 74.9, 63.5, 43.0, 26.0, 25.8, 18.4, 18.1, 9.3, –4.6, –4.8, –5.3,
–5.5.
3¢,5¢-Di-O-(tert-butyldimethylsilyl)-2¢-C-a-hydroxymethyl-b-
guanosine (5)
Ammonia was bubbled into a solution of 3b (158 mg, 0.26 mmol)
in MeOH (15 mL) at 0 °C for 30 min. The flask was sealed and kept
at r.t. for 24 h. TLC showed that 3b was completely consumed. The
solvent was removed, and the residue was purified by silica gel
chromatography, eluting with 10% MeOH in CHCl3, to give com-
pound 5 as a white solid powder (121 mg, 89%).
1H NMR (CDCl3/TMS): d = 7.76 (s, 1 H), 6.48 (br s, 2 H), 6.08 (d,
1 H, J = 6.4 Hz), 4.57 (m, 1 H), 4.00–3.80 (m, 3 H), 3.80–3.65 (m,
2 H), 2.94 (m, 1 H), 0.91 (s, 9 H), 0.87 (s, 9 H), 0.12 (s, 3 H), 0.11
(s, 3 H), 0.08 (s, 3 H), 0.07 (s, 3 H).
HRMS: m/z calcd for C23H43N5O4Si2Na [MNa+]: 532.2751; found:
532.2753.
2¢-C-a-Methyl-b-adenosine (8)
To a solution of 6 (58.0 mg, 0.12 mmol) in THF (15 mL) was added
TBAF (1.0 M in THF, 0.50 mL, 0.50 mmol). The mixture was
stirred at r.t. for 12 h. The solvent was removed and the residue was
purified by silica gel chromatography, eluting with 12.5% MeOH in
CHCl3 to give the product as a white solid (30 mg, 94%).
13C NMR (CDCl3): d = 158.9, 153.7, 151.1, 135.8, 116.9, 87.2,
86.8, 73.7, 62.8, 59.1, 50.4, 25.9, 25.7, 18.4, 18.0, –4.8, –5.0, –5.4,
–5.5.
1H NMR (D2O): d = 8.07 (s, 1 H), 7.90 (s, 1 H), 5.71 (d, 1 H, J = 9.0
Hz), 4.24 (m, 1 H), 4.05 (m, 1 H), 3.66 (m, 2 H), 2.68 (m, 1 H), 0.86
(d, 3 H, J = 6.9 Hz).
HRMS: m/z calcd for C23H43N5O5Si2Na [MNa+]: 548.2700; found:
13C NMR (D2O): d = 155.2, 152.1, 148.2, 140.1, 118.7, 89.8, 87.3,
73.6, 62.0, 42.9, 7.8.
548.2700.
Synthesis 2005, No. 17, 2865–2870 © Thieme Stuttgart · New York

2870
N.-S. Li, J. A. Piccirilli
PAPER
HRMS: m/z calcd for C11H16N5O3 [MH+]: 266.1253; found:
266.1266.
(4) Gordon, P. M.; Fong, R.; Deb, S.; Li, N.-S.; Schwans, J. P.;
Ye, J.-D.; Piccirilli, J. A. Chem. Biol. 2004, 11, 237.
(5) Schwans, J. P.; Li, N.-S.; Piccirilli, J. A. Angew. Chem. Int.
Ed. 2004, 43, 3033.
2¢-C-a-Methyl-b-guanosine (9)
To a solution of 7 (42.0 mg, 0.082 mmol) in THF (10 mL) was add-
ed TBAF (1.0 M in THF, 0.40 mL, 0.40 mmol). The mixture was
stirred at r.t. for 3.5 h. The solvent was removed, and the residue
was purified by silica gel chromatography, eluting with 15% MeOH
in CHCl3 to give the product as a white solid (22 mg, 95%).
1H NMR (D2O/CD3OD): d = 7.81 (s, 1 H, 8-H), 5.61 (d, 1 H, J = 9.2
Hz, 1¢-H), 4.21 (m, 1 H, 3¢-H), 4.00 (m, 1 H, 4¢-H), 3.65 (m, 2 H, 5¢-
H), 2.71 (m, 1 H, 2¢-H), 0.88 (d, 3 H J = 7.2 Hz, 2-CH3).
13C NMR (D2O/CD3OD): d = 159.1, 153.9, 151.3, 137.6, 116.5,
89.0, 86.9, 73.5, 61.9, 42.2, 7.7.
HRMS: m/z calcd for C11H15N5O4Na [MNa+]: 304.1022; found:
304.1026.
(6) Li, N.-S.; Piccirilli, J. A. J. Org. Chem. 2004, 69, 4751.
(7) Novak, J. J. K.; Smejkal, J.; Sôrm, F. Tetrahedron Lett.
1969, 1627.
(8) Cicero, D. O.; Neuner, P. J. S.; Franzese, O.; D’Onofrio, C.;
Iribarren, A. M. Bioorg. Med. Chem. Lett. 1994, 4, 861.
(9) (a) Matsuda, A.; Takenuki, K.; Sasaki, T.; Ueda, T. J. Med.
Chem. 1991, 34, 234. (b) Matsuda, A.; Itoh, H.; Takenuki,
K.; Sasaki, T.; Ueda, T. Chem. Pharm. Bull. 1988, 36, 945.
(10) Schmit, C. Synlett 1994, 238.
(11) (a) Vorbruggen, H.; Krolikiewicz, K.; Bennua, B. Chem.
Ber. 1981, 114, 1234. (b) Garner, P.; Ramakanth, S. J. Org.
Chem. 1988, 53, 1294. (c) Vorbruggen, H.;Ruh-Pohlenz, C.
Handbook of Nucleoside Synthesis; Wiley: New York, 2001,
94–98. (d) Robins, M. J.; Zou, R.; Guo, Z.; Wnuk, S. F. J.
Org. Chem. 1996, 61, 9207.
Acknowledgment
(12) The 13C NMR data of the adenine ring are assigned as
follows: For 2a (CDCl3): d = 152.6 (C-6), 151.7 (C-2), 149.3
(C-4), 142.8 (C-8), 122.2 (C-5) and 2b (CDCl3): d = 152.7
(C-6), 151.8 (C-2), 149.5 (C-4), 141.6 (C-8), 123.2 (C-5); cf.
experimental section. For 7-methyladenine (DMSO-d6): d =
159.82 (C-4), 152.31 (C-2), 151.91 (C-6), 145.94 (C-8),
111.77 (C-5) and 9-methyladenine (DMSO-d6): d = 155.98
(C-6), 152.50 (C-2), 149.94 (C-4), 141.47 (C-8), 118.72 (C-
5); see: Chenon, M.-T.; Pugmire, R. J.; Grant, D. M.;
Panzica, R. P.; Townsend, L. B. J. Am. Chem. Soc. 1975, 97,
4627.
N.-S.L. is a Research Specialist and J.A.P. is an Associate Investi-
gator of the Howard Hughes Medical Institute. We thank J. Ye, J.
Hougland, S. Koo and R. Fong for helpful discussions and critical
comments on the manuscript.
References
(1) (a) Auffinger, P.; Westhof, E. J. Mol. Biol. 1997, 274, 54;
and references cited therein. (b) Simons, R. W.; Grunberg-
Manago, M. RNA Structure and Function; Cold Spring
Harbor Laboratory Press: Cold Spring Harbor, New York,
1998.
(2) (a) Breaker, R. R. Nature 2004, 432, 838. (b) Uhlmann, E.;
Peyman, A. Chem. Rev. 1990, 90, 543. (c) Cooke, S. T.
Ann. Rev. Pharmacol. Toxicol. 1992, 32, 329. (d) Schmit,
C.; Bevierre, M.-O.; Mesmaeker, A. D.; Altmann, K.-H.
Bioorg. Med. Chem. Lett. 1994, 4, 1969.
(3) (a) Gesteland, R. F.; Cech, T. R.; Atkins, J. F. The RNA
World, 2nd ed.; Cold Spring Harbor Laboratory Press: Cold
Spring Harbor New York, 1999. (b) Eckstein, F.; Lilley, D.
M. J. Catalytic RNA; Springer-Verlag: Berlin Germany,
1997.
(13) The 13C NMR data of the guanine ring are assigned as
follows: For 3a (CDCl3): d = 155.9 (C-6), 148.2 (C-4), 147.4
(C-2), 138.9 (C-8), 120.4 (C-5), 3b (CDCl3): d = 156.0 (C-
6), 148.4 (C-4), 147.4 (C-2), 137.0 (C-8), 120.8 (C-5) and 3c
(CDCl3): d = 156.8 (C-6), 152.8 (C-4), 148.2 (C-2), 141.6
(C-8), 111.4 (C-5); cf. experimental section. For N2-acetyl-
2¢,3¢,5¢-tri-O-acetyl-9-guanosine (DMSO-d6): d = 154.68
(C-6), 148.58 (C-4), 148.22 (C-2), 137.79 (C-8), 120.38 (C-
5) and N2-acetyl-2¢,3¢,5¢-tri-O-acetyl-7-guanosine (DMSO-
d6): d = 158.37 (C-6), 152.17 (C-4), 147.51 (C-2), 144.12 (C-
8), 110.56 (C-5); see: Boryski, J. J. Chem. Soc., Perkin
Trans. 2 1997, 649.
Synthesis 2005, No. 17, 2865–2870 © Thieme Stuttgart · New York