7-Deazapurine (Pyrrolo[2,3- d]pyrimidine) 2 ... Organic Chemistry, 2012, 16, 161-223 161...
Transcript of 7-Deazapurine (Pyrrolo[2,3- d]pyrimidine) 2 ... Organic Chemistry, 2012, 16, 161-223 161...
Current Organic Chemistry, 2012, 16, 161-223 161
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7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides: Syntheses and
Transformations
Frank Seela,1,2
* Simone Budow1 and Xiaohua Peng
1
1Laboratory of Bioorganic Chemistry and Chemical Biology, Center for Nanotechnology, Heisenbergstraße 11, 48149 Münster,
Germany and 2Laboratorium für Organische und Bioorganische Chemie, Institut für Chemie, Universität Osnabrück, Barbarastraße
7, 49069 Osnabrück, Germany
Abstract: This review reports on the synthesis of 7-deazapurine (pyrrolo[2,3-d]pyrimidine) 2’-deoxyribonucleosides, including -D- and
-L-enantiomers, fluoro derivatives, and 2’,3’-dideoxyribonucleosides. It covers the various aspects of convergent nucleoside synthesis.
Stereochemically defined -D and -L 2’-deoxyribonucleosides as well as sugar derivatives were prepared by nucleobase anion glycosy-
lation. This glycosylation reaction is regioselective for the pyrrole nitrogen and stereoselective for -nucleoside formation. Common gly-
cosylation protocols lead to 7-deazapurine 2’-deoxyribonucleosides with unusual glycosylation sites. 7-Deazapurine 2’,3’-
dideoxyribonucleosides were also obtained from 2’-deoxy- or 3’-deoxyribonucleosides by Barton-McCombie deoxygenation, by elimina-
tion of sugar hydroxyl groups or by anion glycosylation. Another aspect of the review is the functionalization of pyrrolo[2,3-d]pyrimidine
nucleosides. A broad range of reporter groups were introduced by the Sonogashira cross coupling or the copper(I)-catalyzed Huisgen-
Meldal-Sharpless “click” reaction. The application of 7-deazapurine nucleosides as antiviral or anticancer agents, and the use of 7-
deazapurine nucleoside triphosphates in the Sanger dideoxy DNA-sequencing are also reported.
Keywords: 7-Deazapurine, pyrrolo[2,3-d]pyrimidines, nucleosides, glycosylation, enantiomers, halogenation, cross-coupling, click reaction,
triphosphates, sequencing.
1. INTRODUCTION
7-Deazapurine (pyrrolo[2,3-d]pyrimidine) 2’-deoxyribonucle-
osides and their derivatives have found widespread applications in
chemistry, physics, and biology [1-6]. In contrast to the naturally
occurring 7-deazapurine ribonucleosides [3, 5, 7], the correspond-
ing 2'-deoxyribonucleosides are not found in nature. However, a
few deoxyribonucleosides with very particular sugar structure have
been isolated from natural sources. Kanagawamicin (1, AB-116)
was obtained from Actinoplanes kanagawaensis [8], and 5’-deoxy-
7-iodotubercidin (2) [9] as well as the mycalisines A (3a) and B
(3b) [10] were identified in marine organisms (Fig. 1). (If not oth-
erwise stated purine numbering is used throughout the review).
However, no “true” 7-deazapurine 2’-deoxyribonucleoside with the
general structure of motif I was found in nature.
*Address correspondence to this author at the Laboratory of Bioorganic Chemistry and
Chemical Biology, Center for Nanotechnology, Heisenbergstraße 11, 48149 Münster,
Germany; Tel: +49 (0)251 53406 500; Fax: +49 (0)251 53406 857; E-mail:
[email protected], Homepage: www.seela.net
Current address of X. Peng: Department of Chemistry and Biochemistry, University of
Wisconsin-Milwaukee, 3210 North Cramer Street, Milwaukee, Wisconsin 53211,
United States.
It had been recognized that the shape of 7-deazapurine 2’-
deoxyribonucleosides resembles closely to that of purine nucleo-
sides found in duplex DNA [6, 11, 12]. Stable Watson-Crick base
pairs are formed between 7-deazapurines and complementary
pyrimidines within oligonucleotide duplexes, showing similar base
stacking as those of canonical DNA bases (Fig. 2, motifs II and III).
Substituents introduced at the 7-position of 7-deazapurine nucleo-
sides do not change the conformation around the glycosylic bond
significantly while 8-substituents of purines force the nucleosides
into the syn-conformation [13-15]. 7-Substituted purines are posi-
tively charged, while 7-substituted 7-deazapurines are neutral being
well accepted by DNA polymerases [13-15]. Bulky substituents
introduced at the 7-position of the 7-deazapurine base within the
oligonucleotide chain are well accommodated in the major groove
of B-DNA. This position is also suitable for the introduction of
reporter groups in DNA with the use of building blocks such as
triphosphates or phosphoramidites. While the latter are employed in
solid-phase oligonucleotide synthesis, the former are substrates of
DNA polymerases [11, 12, 16-18]. 7-Deazapurine nucleoside
O
HO OH
N
N N
H3C
NH2 I
2
O
HO
HN
N N
HO
O
H2N
COMe
O
R
1: R = NH2
O
MeO OH
N
N N
RCN
3a: R = NH2
b: R = OH
systematic numberingpurine numbering
1
2
3
45
6
7
4a
7a
1
2
3
4
56
7
8
9
O
HO
N
N N
HO
R
R
R = various substituents
R
R
Motif I
Fig (1). Structures of naturally occurring 7-deazapurine deoxyribonucleosides and general formula of 7-deazapurine 2’-deoxyribonucleosides.
162 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
triphosphates such as 7-deaza-2'-deoxyadenosine triphosphate
(c7dATP) and 7-deaza-2'-deoxyguanosine triphosphate (c
7dGTP, 4)
are efficiently incorporated in the growing DNA chain [16]. Com-
pound 4 can replace dGTP in the Sanger dideoxy sequencing when
dG-dC rich DNA-fragments have to be resolved by electrophoresis
[19, 20]. Corresponding 7-deazapurine 2’,3’-dideoxyribonucleoside
triphosphates, as compound 5, are utilized as fluorescent chain ter-
minators in automated sequencing machines [21, 22].
Contrary to purine 2’-deoxyribonucleosides, the 7-deazapurine
nucleosides are extremely stable at their N-glycosyl bond. Glycosyl
bond cleavage does not take place in strong acid even at elevated
temperatures; instead isomerization of the sugar moiety is observed
yielding anomeric mixtures of furanosides and pyranosides [23].
Due to the high stability of the glycosylic bond, the 7-deaza-2’-
deoxyadenosine derivative 6b of the Ca2+
-mobilizing second mes-
senger cyclic adenosine 5’-diphosphate ribose (cADPR; 6a) was
synthesized as a hydrolysis-resistant agonist candidate (Fig. 3) [24];
however its biological activity needs to be evidenced. Moreover,
the glycosylic bond stability of 7-deazapurine nucleosides has been
exploited to overcome depurination reactions occurring during
MALDI-TOF mass spectrometry thereby leading to better detection
sensitivity [25]. 7-Deazapurine nucleosides such as 7-deaza-2'-
deoxyguanosine (c7Gd) or 7-deaza-2'-deoxyisoguanosine (c
7iGd) are
more easily oxidized than their purine counterparts. Quenching of
fluorescent dyes e.g. DNA-bound ethidium bromide has been ob-
served in oligonucleotide duplexes containing c7Gd, c
7iGd, or their
7-substituted derivatives [26, 27]. This phenomenon was used to
study the electron transfer within duplex DNA. Very recently, al-
kynylated 7-deazapurine nucleosides were employed in cross-
linking reactions utilizing the copper-catalyzed azide-alkyne Huis-
gen-Meldal-Sharpless cycloaddition “click” reaction.
The widespread applicability of 7-deazapurine nucleosides in
chemistry, physics, and biology has generated intensive studies on
their synthesis and incorporation into nucleic acids. These topics
have already been subjects of several reviews [1-6, 12, 28-32],
which include the synthesis of 7-deazapurine ribonucleosides [31]
and of oligonucleotides containing 7-deazapurines as monomeric
constituents [12]. The current review focuses on the convergent
syntheses of 7-deazapurine (pyrrolo[2,3-d]pyrimidine) 2’-
deoxyribonucleosides and describes various glycosylation protocols
which make use of activated 2'-deoxyribofuranose derivatives. The
review includes work on 7-deazapurine 2’-deoxy- -D-
ribonucleosides and -L-enantiomers; it also reports on fluoronu-
cleosides and 2’,3’-dideoxyribonucleosides. Transformation reac-
tions, leading to 7-halogenated derivatives which stabilize duplex
DNA are described along with the conversion into 7-alkynyl deriva-
tives by the Sonogashira cross coupling reaction. Functionalization
of alkynylated 7-deazapurine nucleosides by the copper-catalyzed
Huisgen-Meldal-Sharpless “click” reaction with various reporter
groups and cross-linking of alkynyl 7-deazapurine nucleosides are
also covered by this review.
2. GENERAL COMMENTS ON THE CONVERGENT SYN-
THESIS OF 7-DEAZAPURINE NUCLEOSIDES
In 1983, our laboratory developed the method for the stereose-
lective nucleobase anion glycosylation which was exemplified by
the 7-deazapurine system [33, 34]. Initially, the synthesis of 7-
deazapurine nucleosides was encountered with difficulties due to
the low nucleophilicity of the 7-deazapurine pyrrole nitrogen for
electrophilic glycosylation reactions. Thus, the inertness of the pyr-
N
N
N
N
N
O
N
H
H
CH3
N
N
O
N
N
N
N
N O
H
CH3
O
H
H
H
H
H
Motif II: 'c7Ad' - dT (aps) Motif III: 'c7Gd' - dC (aps)
major groove
minor groove
RR
major groove
minor groove
O
HN
N N
O
O
H2N
4HO
POPOPHO
O O O
OH OH OH
O
N
N N
O
NH2
POPOPHO
O O O
OH OH OH
CH2NH DYE
5
R = variable substituents
Fig (2). Watson-Crick base pairs formed by 7-deazapurines and pyrimidine bases in anti-parallel stranded (aps) duplexes (upper part); 7-deazapurine triphos-
phates used in the Sanger dideoxy sequencing (lower part).
N
N
X
NH2
N 6a: X = N, R = OH
b: X = CH, R = H
O
O
OH
O
P
OP
O
HO
HO
OO
O
O
R
Fig (3).
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 163
role nitrogen directs the glycosylation towards the pyrimidine moi-
ety or takes place on the more nucleophilic pyrrole carbons. This
problem was overcome by generating the pyrrolyl anion which is
highly reactive and allows glycosylation at ambient temperature.
Consequently, the anomerically defined sugar halides employed in
the glycosylation do not isomerise. The generation of the pyrrolyl
anion was originally applied to ribonucleoside synthesis [35, 36].
Later, it was used by our laboratory to synthesize 7-deazapurine
arabinonucleosides [37], and presently we employ this method in
the synthesis of 7-deazapurine 2’-deoxyribonucleosides.
The stereoselective nucleobase anion glycosylation [33, 34, 38],
which is now the most widely used protocol for the synthesis of
purine or purine related 2'-deoxyribonucleosides, utilizes sugar
halides with defined configuration at the anomeric center. As sev-
eral sugar halides (halogenoses) are accessible as pure -D- or -L-
anomers, the synthesis of 7-deazapurine 2’-deoxyribonucleosides
proceeds under stereochemical control with the exclusive formation
of -D-2’-deoxyribonucleosides or -L-2’-deoxyribonucleosides. A
large number of 7-deazapurine nucleosides have been synthesized
under these conditions [30, 38, 39]. Many of them show antiviral or
anticancer activity and several have been incorporated into oligonu-
cleotides [12].
3. SYNTHESIS OF 7-DEAZAPURINE – PYRROLO[2,3-
d]PYRIMIDINE – 2’-DEOXYRIBONUCLEOSIDES
A few 7-deazapurine 2’-deoxyribonucleosides, such as 7-deaza-
2’-deoxyadenosine, were prepared from the naturally occurring
ribonucleosides by chemical deoxygenation (see section 3.3) or by
reduction of nucleoside triphosphates with ribonucleotide reductase
from Lactobacillus leichmannii [40]. These routes are only applica-
ble to 7-deazapurine 2’-deoxyribonucleosides when corresponding
ribonucleosides are available. Thus, considerable efforts have been
devoted towards their convergent synthesis. After the nucleobase
anion glycosylation was developed by our laboratory, 7-
deazapurine 2’-deoxyribonucleosides became easily accessible. The
generation of the nucleobase anion, which is the key step in the
stereocontrolled glycosylation, can be performed in different ways:
(a) the anion is formed at the interphase of a bi-layered mixture of
aqueous NaOH and an organic solvent in the presence of a quater-
nary ammonium salt (liquid-liquid conditions); (b) the anion is
generated in the organic solvent (MeCN) in the presence of pow-
dered KOH containing 15% of water using a phase-transfer catalyst
such as tris-[2-(2-methoxyethoxy)ethyl]amine (TDA-1) (solid-
liquid conditions); (c) the nucleobase is suspended or dissolved in
MeCN, and the anion is generated with NaH (sodium salt glycosy-
lation). As halogenoses, 2-deoxy-3,5-di-O-(p-toluoyl)- -D-erythro-
pentofuranosyl chloride (7) and the corresponding -L-enantiomer
8 were employed for the glycosylation. Both are stereochemically
assigned -anomers and can be isolated as crystalline compounds
(Fig. 4). The halogenose 7 was first described by Hoffer [41] and its
L-enantiomer 8 by orm [42]. The reaction time of the glycosyla-
tion should be short (15-60 min), and the reaction temperature
should be kept below 25°C to avoid anomerization of the halo-
genose.
87
O
TolO
TolO
Cl
O
OTol
OTol
Cl
Fig (4).
3.1. Nucleobase Anion Glycosylation Performed under Liquid-
liquid Conditions
This glycosylation reaction is performed in a biphasic mixture
of saturated aq. NaOH or KOH (as aqueous phase) and an organic
phase (usually dichloromethane or benzene) containing the halo-
genose and a quaternary ammonium salt as phase-transfer catalyst.
The reaction mixture is stirred with a vibromixer. It was observed
that the anomeric ratio can be changed by the concentration of so-
dium hydroxide or by the amount of phase-transfer catalyst used
during the glycosylation. Both have an influence on the reaction
rate and therefore on the anomeric ratio [43, 44]. The first conver-
gent synthesis of a 7-deazapurine 2’-deoxyribonucleoside under
stereocontrolled conditions utilizing this protocol employed 50%
aq. NaOH, benzene/dimethoxyethane and Aliquat 336 (methyltrioc-
tylammonium chloride) as a phase-transfer catalyst [33].
Glycosylation of 2-amino-7-deaza-6-methoxypurine (9) [45]
with the sugar chloride 7 furnished the anomerically pure 2-amino-
7-deaza-6-methoxypurine -D-nucleoside 10 (Scheme 1) [33]. This
N
N
OMe
H2N
N
N NH
OMe
H2N
O
HO
N
HO
+
9 7
11a
HN
N
O
H2N
O
HO
N
HO
12a
Aliquat 336, 50% aq. NaOH,
benzene/dimethoxyethane
sodium p-thiocresolate,
hexamethylphosphoric
triamide, toluene
48% 91%
N
N
OMe
H2N
O
TolO
N
TolO
10
aq. NH3
O
TolO
TolO
Cl
Scheme 1.
164 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
compound was deblocked with aq. ammonia to give the methoxy
nucleoside 11a (48% yield); 37% of nucleobase 9 was recovered.
The methoxy group of 11a was displaced with sodium p-
thiocresolate [46] in hexamethylphosphoric triamide/toluene under
nitrogen atmosphere furnishing 7-deaza-2’-deoxyguanosine (12a)
in 91% yield. The glycosylation reaction was regioselective for
nitrogen-9 and proceeds to inversion of configuration. As the crys-
talline halogenose 7 was the pure -D-anomer, the -D-nucleoside
was formed exclusively.
The conditions described above were applied later to the syn-
thesis of a series of 7-deazapurine 2’-deoxyribonucleosides. 7-
Deaza-2’-deoxy-7-methylguanosine (13) [47], 2'-deoxytubercidin
(14a) [48], 7-deaza-2’-deoxyinosine (15a) [49], 7-deaza-2’-
deoxynebularine (16a) and its 2-substituted derivatives 16b,c [50-
52], 7-deaza-2’-deoxy-6-thioguanosine (17) [53], 2-amino-2’-
deoxytubercidin (18a) [53] (Fig. 5) and related nucleosides were
prepared by our laboratory and others.
2’-Deoxytubercidin (14a) was synthesized according to Scheme
2. The glycosylation of nucleobase 19 [54] with halogenose 7 pro-
duces the -D-anomer 20 exclusively [48]. Alkaline deblocking of
the sugar protecting groups of 20 yielded 2’-deoxy-2-
methylthiotubercidin (21). Desulfurization with Raney-Nickel cata-
lyst afforded 2’-deoxytubercidin (14a).
7-Deaza-2’-deoxyinosine (15a) was prepared by the same route
(Scheme 3). Glycosylation of compound 22 [37] with 7 performed
in CH2Cl2 in the presence of 50% aq. NaOH and PhCH2(Et)3NCl
was followed by deprotection, leading to the formation of -D-
nucleoside 23 [49]. The cleavage of the 6-methoxy group was per-
formed with hydrochloric acid yielding the methylthio nucleoside
24, which after desulfurization (Raney-Nickel catalyst), gave 7-
deaza-2’-deoxyinosine (15a). As the N-glycosyl bond of 7-
dezapurine 2’-deoxyribonucleosides is rather stable, acidic condi-
tions can be used for the cleavage of the methyl group. However, in
some cases, strong acid can lead to the isomerization of the sugar
moiety [23]. In such cases, conversion under alkaline conditions is
performed. Different to the imidazole ring of the purine skeleton,
the pyrrole ring of pyrrolo[2,3-d]pyrimidine is not opened in the
presence of strong base.
Similarly, 7-deaza-2’-deoxynebularine (16a), its 2-methylthio
(16b), and the 2-amino derivative 16c have been prepared via liq-
uid-liquid phase-transfer glycosylation of 25a, 25b, and 25c, re-
spectively, employing halogenose 7 (Scheme 4) [50-52]. Due to its
high hydrophilicity, the glycosylation of 25a was carried out in
THF, and formation of an anomeric mixture (26a and its -anomer)
was observed, while the corresponding reactions of 25b and 25c
were performed in CH2Cl2 and gave almost exclusively the -D-
HN
N
O
H2N
O
HO
N
HO
13
N
N
NH2
O
HO
N
HO
14a
HN
N
O
O
HO
N
HO
15a
Me
N
N
O
HO
N
HO
R
16a: R = H
b: R = SCH3
c: R = NH2
N
N
NH2
O
HO
N
HO
18a
H2N
HN
N
S
H2N
O
HO
N
HO
17
Fig (5).
N
N
NH2
H3CS
N
N NH
NH2
H3CSO
TolO
N
TolO
19 7
20
PhCH2(Et)3NCl, 50% aq. NaOH,
benzene/dimethoxyethane
40%
79%
1M NaOMe/MeOH
N
N
NH2
H3CS
O
HO
N
HO
21
N
N
NH2
O
HO
N
HO
14a
Ra-Ni, N,N-dimethylacetamide (DMA)
67%
+O
TolO
TolO
Cl
Scheme 2.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 165
anomers 26b,c accompanied by an increased glycosylation yield.
The increased yields of nucleosides 26b and 26c can be attributed
to the better solubility of the nucleobases 25b,c in CH2Cl2. Depro-
tection of compounds 26a-c afforded nucleosides 16a-c. Compound
16c is highly fluorescent with a quantum yield of = 0.47. The
fluorescence spectrum of 16c is shown in Fig (6). The emission
maximum of 16c is located at 395 nm upon excitation at 311 nm
[55].
Glycosylation of 2-amino-6-chloro-7-deazapurine (27a) with
halogenose 7 was performed under liquid-liquid conditions
(CH2Cl2/30% KOH/tBu4NHSO4) to yield -D-nucleoside 28a
(Scheme 5). The latter was deprotected to give 29a, which was
further aminated (NH3/MeOH) furnishing 2-amino-7-deaza-2’-
deoxyadenosine (18a). Nucleophilic displacement of the 6-chloro
substituent of 28a by a mercapto group resulted in the formation of
6-thionucleoside 30. Deprotection furnished 7-deaza-2'-deoxy-6-
thioguanosine (17) [53]. Very recently, thionucleoside 17, as a con-
stituent of single-stranded oligonucleotides, was used as a molecu-
lar anchor for gold nanoparticles (AuNPs). The covalent attachment
of 17 via the thiol group makes use of the strong affinity of the
sulphur atom for the noble metal gold. This method was employed
for the preparation of oligonucleotide gold nanoparticle conjugates
[56].
N
N
OCH3
H3CS
N
N NH
OCH3
H3CS
O
HO
N
HO
22
723
(i) 50% aq. NaOH,
PhCH2(Et)3NCl,
CH2Cl2
(ii) NaOMe/MeOH
40% 60%
HN
N
O
H3CS
O
HO
N
HO
24
HN
N
O
O
HO
N
HO
15a
Ra-Ni,
DMA
48%+
7N HCl
O
TolO
TolO
Cl
Scheme 3.
7
O
TolO
TolO
Cl
N
NRN
N NH
RO
TolO
N
TolO
25a-c
26a-c
50% aq. NaOH,
PhCH2(Et)3NCl,
a: THF; b, c: CH2Cl2
a: 36%
b: 70%
c: 77%
+
N
NR
O
HO
N
HO
16a-c
a: R = H; b: R = SCH3; c: R = NH2
NaOMe,
MeOH
Scheme 4.
Fig (6). Fluorescence spectra of the nebularine derivative 16c in water. The concentration of 16c was 10-5
M.
N
NH2N
O
HO
N
HO
16c
166 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
3.2. Nucleobase Anion Glycosylation Performed Under Solid-
Liquid Conditions
The glycosylation yields obtained under liquid-liquid conditions
were limited to 40-50% and depended very much on the solubility
of the nucleobases. When lipophilic nucleobases were used, the
yields were acceptable and the -D anomers were the only glycosy-
lation products. The situation changed when more hydrophilic nu-
cleobases, such as 7-deazapurine (25a) or 2-amino-6-chloro-7-
deazapurine (27a) were used [34]. Unfavorable partition between
the organic and the aqueous phase led to an extension of the reac-
tion time resulting in partial sugar deprotection. Side reactions were
observed on the nucleobase as well as on the halogenose 7. Under
strongly alkaline conditions, the nucleophilic substitution took
place in dichloromethane and two nucleobases were linked by a
methylene group via the pyrrole nitrogen resulting in the formation
of bis-(pyrrolo[2,3-d]pyrimidines) such as 31 (Scheme 6) [57, 58].
The second side reaction is due to the partial deprotection of the
sugar moiety. The anion of the protecting group (4-methylbenzoate)
acts as nucleophile and competes with the nucleobase anion in the
glycosylation reaction thereby forming the tris-toluoylated sugars
32a and 32b (Scheme 7) [53]. These reactions consume the halo-
genose 7 as well as the nucleobases, and reduce the glycosylation
yield.
Consequently, the protocol was changed. Powdered KOH was
employed as reagent for the nucleobase anion generation, MeCN as
solvent, and the cryptand, tris-[2-(2-methoxyethoxy)ethylamine
(TDA-1) [59] was selected as catalyst. TDA-1 is an aminopolyether
and combines properties of an amine with those of a crown ether. It
forms cavities in anhydrous aprotic solvents such as MeCN, thereby
chelating monovalent and divalent cations as well as transition
metal salts. TDA-1 shows great complex affinities for ionic com-
pounds containing large polarizable anions, e.g. nucleobase anions.
Powdered KOH contains 15% of water which is able to form an
interphase between the solid and the liquid phase thereby promoting
the glycosylation reaction. In most cases, the reaction was carried
out at room temperature. Instead of powdered KOH, sodium- or
cesium hydroxides or carbonates are sometimes advantageous as
well as organic bases (DBU). The glycosylation performed under
solid-liquid conditions results in significantly higher yields than
those obtained in the liquid-liquid system. For instance, the glyco-
sylation of nucleobase 27a with halogenose 7 formed compound
N
N
Cl
H2N
N
N NH
Cl
H2N
O
TolO
N
TolO
27a
28a
N
N
Cl
H2N
O
HO
N
HO
29a
N
N
NH2
H2N
O
HO
N
HO
18a
NaOMe,
MeOH
68%
thiourea,
1-propanol
79% 63%
NH3,
MeOH
HN
N
S
H2N
O
TolO
N
TolO
30
NaOMe,
MeOH
17
58%
+
7
tBu4NHSO4,
30% KOH,
CH2Cl2
45%O
TolO
TolO
Cl
Scheme 5.
N
N N
OMe
H3CS
31
N
NN
OMe
SCH3
N
N NH
OMe
H3CS
22
CH2Cl2,
aq. NaOH
+ CH2Cl2
Scheme 6.
+
C
CH3
O
O
32a7
O
TolO
TolO
Cl
O
TolO
TolO
+
OTol
32b
O
TolO
TolO
OTol
Scheme 7.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 167
28a in 45% yield under liquid-liquid conditions, but proceeds with
70% yield under solid-liquid conditions [34].
Some nucleobases are particularly useful starting materials, as
the glycosylation products can be employed as common intermedi-
ates for the synthesis of a number of 7-deazapurine 2'-
deoxyribonucleosides bearing various substituents at the nucleobase
moiety. Examples are compounds 27a and 33a (Scheme 8). Both
chlorinated 7-deazapurine bases are excellent substrates for the
solid-liquid glycosylation (TDA-1/KOH/MeCN). When employing
halogenose 7, compound 28a is obtained in 70% [34] and com-
pound 34a in 81% yield [60, 61]. Compound 28a can be used to
access 7-deazapurine 2'-deoxyribonucleosides related to dG, its 6-
thio derivative, 2-amino-dA, 2’-deoxyxanthosine, and isoGd (see
section 3.2.1). The 6-chloro nucleoside 34a functions in an analo-
gous manner to access nucleosides related to dA and dI (see section
3.2.2).
By employing the nucleobase anion glycosylation under solid-
liquid conditions, a number of novel nucleosides as well as their 7-
substituted derivatives have been prepared, such as 7-deaza-2’-
deoxyguanosines 12a-h, 7-deaza-2’-deoxyadenosines 14a-e,g, 7-
deaza-2’-deoxyinosines 15a-e, 7-deaza-2’-deoxyxanthosines 35c,d,
2-amino-7-deaza-2’-deoxyadenosines 18a-f, 7-deaza-2’-
deoxyisoguanosines 36a-c, 36e, 7-deaza-2’-deoxyisoinosine (37),
2-halogenated 7-deaza-2’-deoxyadenosines 38a-e, 38i, and others
(Fig. 7).
3.2.1. Synthesis of 7-Deazapurine 2'-Deoxyribonucleosides Re-
lated to 2'-Deoxyguanosine and 2'-Deoxyxanthosine and Deriva-
tives Thereof (12b-h and 35c,d)
Using compound 28a as staring material, several nucleosides
functionalized at the 6-position were prepared by nucleophilic dis-
placement of the 6-chloro substituent. It has already been described
in Scheme 5 of section 3.1 that nucleoside 28a can be converted to
2-amino-2’-deoxytubercidin (18a) and 7-deaza-2’-deoxy-6-
thioguanosine (17). The 6-methoxy compound 11a was also pre-
pared from 28a (NaOMe in MeOH) (Scheme 9). The displacement
of the methoxy group of 11a by a hydroxyl function furnished 7-
deaza-2’-deoxyguanosine (12a) [34]. Here, an alkaline solution (0.2
M NaOH) was used for the conversion of OMe to the OH group,
which resulted in a fast and clean reaction.
Compound 11a is also a central intermediate for the synthesis
of 7-halogenated 7-deaza-2’-deoxyguanosines and halogenated 2’-
deoxyxanthosines. For this synthesis, 11a was converted to the fully
protected nucleoside 39a with N,N-dimethylformamide diethy-
lacetal followed by isobutyrylation (Scheme 10) [62]. Regioselec-
tive halogenation at position-7 was performed with N-
halosuccinimides (NXS, X = Cl, Br, and I) yielding the 7-
halogenated nucleosides 39b-d [63]. The 7-deazaguanosine analogs
N
N
Cl
H2NN
N NH
Cl
H2N
O
TolO
N
TolO
27a28a
TDA-1, KOH,
MeCN
70%
+ 7
N
N
Cl
N
N NH
Cl
O
TolO
N
TolO
33a
34a
TDA-1, KOH,
MeCN
81%
+ 7
Scheme 8.
HN
N
O
H2N
O
HO
N
HO
12a-h
N
N
NH2
O
HO
N
HO
14a-e,g
HN
N
O
O
HO
N
HO
15a-e
HN
NH
O
O
HO
N
HO
O
35c,d 36a-c,e
N
N
NH2
O
HO
N
HO
18a-f
H2N
HN
N
NH2
O
O
HO
N
HO
R R R R
R
R
a: R = H; b: R = Cl; c: R = Br; d: R = I; e: R = F; f: R = CN; g: R = NO2; h: R = CO2H
37
HN
NO
O
HO
N
HO
38i
N
NF
O
HO
N
HO
NH2
N
NCl
O
HO
N
HO
NH2 R
38a-e
Fig (7).
168 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
12b-d were accessible from 39b-d by methoxy/hydroxy substituent
exchange in refluxing 2 M NaOH [63]. Compounds 39b-d were
deprotected in 0.5 M NaOMe/MeOH to give 7-halogenated 4-
methoxy derivatives 11c,d. Deamination of compounds 11c,d with
NaNO2/AcOH yielded the derivatives 40c,d. Finally, the methoxy
group was displaced by a hydroxy group in 2 M NaOH under reflux
conditions (72 h), affording the 7-deaza-2’-deoxyxanthosines 35c,d
(Scheme 10) [64].
Treatment of the 7-iodo compound 12d with copper cyanide in
pyridine (method I) gave the 7-cyano derivative 12f in 86% yield
(Scheme 10) [65]. As these conditions require a long reaction time
(7 h) for the completion of the iodo-nitrile exchange, the conversion
was also performed under microwave conditions (method II) [66].
By applying method II, compound 12f was isolated in 59% yield
after 1 h. The 7-cyanonucleoside 39f was prepared from 39d in a
similar way, with (method I) and without (method II) microwave
assistance [65, 66]. Heating of 39f under mild alkaline conditions
(1.0 M NaOH at 60°C) provided 7-carbamoyl-7-deaza-2’-
deoxyguanosine (12j), while strong alkaline conditions (5 M KOH,
100°C) furnished 2’-deoxycadeguomycin (12h) (Scheme 11).
Also, 7-fluoro compounds were prepared. The synthesis of 7-
deaza-2’-deoxy-7-fluoroguanosine (12e) started from the 7-
fluorinated 7-deazapurine base 41e [67-69] (Scheme 12). The latter
was obtained from the 2-pivaloyl derivative 41a [70], which was
N
N
Cl
H2N
O
TolO
N
TolO
28a 12a
N
N
OMe
H2N
O
HO
N
HO
11a
NaOMe/MeOH,
reflux
88% 85%
0.2M NaOH,
reflux
HN
N
O
H2N
O
HO
N
HO
Scheme 9.
O
(i-Bu)O
N
N N
(i-Bu)O
OMe
HCHN
O
R
39b-d
O
(i-Bu)O
N
N N
(i-Bu)O
OMe
HCHN
O
39a
O
HO
N
N N
HO
OMe
H2N
O
HO
N
NH
N
HO
OMe
O
R R
c: 78%
d: 80%
c: 75%
d: 79%
R
O
HO
N
N N
HO
OMe
H2N
11a
NXS, DMF, rt
b: 71%
c: 78%
d: 92%
2M NaOH
reflux
b: 96%
c: 91%
d: 92%
NaOMe,
MeOH
AcOH/H2O,
NaNO2, rt
(i) Me2NHC(OEt)2, DMF
(ii) i-Bu2O, MeCN, rt
i: 90%
ii: 81%
39b-d 11c,d 35c,d40c,d
c: 87%
d: 65%O
HO
HN
NH
N
HO
O
O
R
O
HO
HN
N N
HO
O
H2N
R
12b-d
O
HO
HN
N N
HO
O
H2N
CN(I) CuCN, pyridine,
100°C, 7h
(II) CuCN, DMF,
130°C, 1h, MW
(I) 86%
(II) 59%
12f
a: R = H; b: R = Cl; c: R = Br; d: R = I
R
2M NaOH
Scheme 10.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 169
fluorinated regioselectively at the 7-positon using Selectfluor in
MeCN in the presence of acetic acid. The glycosylation of 41e with
the sugar halide 7 (MeCN, KOH, TDA-1) furnished the protected
nucleoside 42e. Deprotection and displacement of the 6-chloro
group yielded the 6-methoxy compound 11e, which was trans-
formed afterwards with 2 N NaOH to the 2’-deoxyguanosine analog
12e [67-69]. The synthesis of 7-deaza-2’-deoxy-7-methylguanosine
(13) shown in Scheme 13 made use of the unprotected base 43
which was glycosylated ( 44) and transformed into 13 by a simi-
lar route as described for the 7-fluoronucleoside 12e [71].
3.2.2. Synthesis of 7-Deazapurine 2'-Deoxyribonucleosides Re-
lated to 2'-Deoxyadenosine and 2'-Deoxyinosine and Derivatives
Thereof (14a-g and 15a-e)
While the 7-deazaguanosine or 7-deazaxanthosine derivatives
were prepared from the nucleobase intermediates 27a and 41a, the
7-deazaadenosine analogs or 7-deazainosine derivatives were syn-
thesized employing 6-chloro-7-deazapurine (33a) and its 7-
substituted derivatives 33b-e (Scheme 14 and 15). Nucleobase an-
ion glycosylation of 33a-d [72] or 33e [73] with halogenose 7 per-
formed in MeCN in the presence of KOH and TDA-1 afforded the
O
(i-Bu)O
N
N N
(i-Bu)O
OMe
HCHN
O
I
39d
(I) CuCN, pyridine,
100°C, 7h
(II) CuCN, DMF,
130°C, 1h, MW
O
(i-Bu)O
N
N N
(i-Bu)O
OMe
HCHN
O
CN
39f
O
HO
HN
N N
HO
O
H2N
R
12h: R = COOH (57%)
j: R = CONH2 (47%)
12h: 5M KOH, 100°C, 1h
12j: 1M NaOH, 60°C, 15h
(I) 78%
(II) 58%
Scheme 11.
N
N NH
Cl
PivHN
41a
Selectfluor, MeCN,
AcOH, 50°C
30%
41e
N
N NH
Cl
PivHN
F
42e
N
N
Cl
PivHN
F
O
TolO
N
TolO
11e
N
N
OMe
H2N
F
O
HO
N
HO
NaOMe,
MeOH
87%
12e
HN
N
O
H2N
F
O
HO
N
HO
2N NaOH
70%
7, MeCN, KOH, TDA-1
64%
Scheme 12.
43
N
N NH
Cl
H2N
Me
44
N
N
Cl
H2N
Me
O
TolO
N
TolO
45
N
N
OMe
H2N
Me
O
HO
N
HO
NaOMe,
MeOH
84%
13
HN
N
O
H2N
Me
O
HO
N
HO
2N NaOH
80%
MeCN,
KOH, TDA-1
73%+
7
O
TolO
TolO
Cl
Scheme 13.
170 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
toluoyl-protected -D-nucleosides 34a-e in 65-81% yield (Scheme
14) [61, 67, 74, 75]. Compounds 34a-e were converted to the 2’-
deoxytubercidin derivatives 14a-e using conc. aq. ammonia at ele-
vated temperature (120°C) in a steel vessel (Scheme 15). The syn-
thesis of the corresponding 7-deaza-2’-deoxyinosine derivatives
15a-e used the same starting materials (34a-e). Sugar deprotection
in 0.5 M NaOMe in MeOH yielded the methoxy nucleosides 46a-e;
displacement of the methoxy group in 2 N NaOH furnished nucleo-
sides 15a-e [67, 76].
7-Deaza-2’-deoxyadenosine (14a) has been converted to its 7-
nitro derivative as shown in Scheme 16 [77-79]. For this purpose,
the amino group as well as the sugar hydroxy groups were protected
by acetylation (Ac2O/pyridine) yielding the intermediate 47. Nitra-
tion of 47 (HNO3/H2SO4 48) followed by deprotection
(NaOMe/MeOH) gave 7-deaza-2’-deoxy-7-nitroadenosine (14g). 7-
Deaza-7-nitropurine nucleosides, such as 14g and 7-deaza-2’-
deoxy-7-nitroguanosine (12g) have been utilized for DNA genotyp-
ing (Incorporation and Complete Chemical Cleavage, ICCC) [80]
and a footprinting technique (Template Directed Interference, TDI)
[81]. The synthesis of 12g was performed as described in section
3.2.4 (Scheme 29).
From 2’-deoxytubercidin (14a), the novel fluorescent and
chemically stable 1,N6-etheno-7-deaza-2'-deoxyadenosine 49 was
prepared by the reaction of 14a with 2-chloroacetaldehyde (Scheme
17) [82]. Compound 49 exhibits similar fluorescence properties as
its parent 1,N6-etheno-2'-deoxyadenosine. Its chemical stability
under acidic and alkaline conditions is significantly higher than that
of 1,N6-etheno-2'-deoxyadenosine. The use of 1,N
6-etheno-7-deaza-
2'-deoxyadenosine (49) is highly advantageous and superior to the
use of its purine congener, particularly for oligonucleotide synthe-
sis.
The solid-liquid nucleobase anion glycosylation was also ap-
plied to the synthesis of 7-deaza-2’-deoxy-2-fluoroadenosine (38i)
(Scheme 18). A reaction sequence starting with the fluorination of
the diamino nucleoside 18a under strongly acid conditions
(HF/pyridine) failed as the 2,6-diamino nucleoside 18a is unstable
under these conditions. Therefore, the corresponding nucleobase 50
was used, which was prepared from 2-amino-6-chloro-7-
deazapurine (27a) (see Scheme 8) by treatment with aq.
NH3/dioxane at 100°C in an autoclave. The diazotiza-
tion/fluorination reaction was performed in HF/pyridine by drop-
wise addition of tBuNO2 resulting in the 2-fluoro base 51 in 30%
yield. Glycosylation of 51 with halogenose 7 afforded the toluoyl-
protected -D-nucleoside 52 (74% yield). Compound 52 was depro-
tected in methanolic ammonia at room temperature to give 7-deaza-
2’-deoxy-2-fluoroadenosine (38i) [83].
In analogy to the 2-fluoronucleoside 38i, the 2-chloro com-
pound 38a and its 7-halogenated derivatives 38b-e were prepared
(Scheme 19). Glycosylation of 7-deaza-2,6-dichloropurine (53a)
[58] and its 7-halogenated derivatives 53b-e with sugar halide 7
(TDA-1/KOH/MeCN) furnished the intermediates 54a-e in 60-74%
O
TolO
N
N N
TolO
ClR
N
N NH
ClR
O
TolO
TolO
Cl
+
a: 81%
b: 70%
c: 75%
d: 65%
e: 67%
33a-e
734a-e
TDA-1, KOH,
MeCN, rt
a: R = H; b: R = Cl; c: R = Br; d: R = I; e: R = F
Scheme 14.
a: 70%
b: 75%
c: 86%
d: 70%
e: 89%
14a-e
O
HO
N
N N
HO
OMeR
46a-e
a: 90%
b: 90%
c: 90%
d: 95%
e: 85%
a: R = H; b: R = Cl; c: R = Br; d: R = I; e: R = F
0.5M NaOMe,
MeOH, reflux
2N NaOH
reflux
a-d: aq. NH3,
dioxane, 120°C
e: NH3/MeOH
a: 75%
b: 75%
c: 79%
d: 80%
e: 83%
O
HO
N
N N
HO
NH2 R
O
HO
HN
N N
HO
OR
15a-e
34a-e
Scheme 15.
14a
Ac2O,
pyridine
97%O
AcO
N
N N
AcO
NHAc
47
O
AcO
N
N N
AcO
NHAc
48
HNO3/H2SO4 (1:1)
91%
NO2
O
HO
N
N N
HO
NH2 NO2
NaOMe,
MeOH
79%
14g
Scheme 16.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 171
yield [84]. Selective displacement of the 6-chloro substituent by an
amino group (NH3/MeOH, 100°C) gave 2-chloro-7-deaza-2’-
deoxyadenosine (38a) or its 7-halogented derivatives 38b-e. The
photoreaction of 38a or 38e in diluted aq. NH3 yielded 7-deaza-2’-
deoxyisoguanosine (36a) or its 7-fluorinated derivative 36e [84,
85]. Compounds 54a-e were converted to the 6-methoxy derivatives
55a-e in NH3/MeOH at room temperature. In the case of 54a, com-
pound 55a (48%) was formed together with the 7-deaza-2,6-
dichloropurine 2’-deoxyribonucleoside 56 (30%). In a 0.5 M or
higher concentrated NaOMe/MeOH solutions and at elevated tem-
perature, both chloro substituents were displaced by methoxy
groups yielding the 2,6-dimethoxy derivatives 57a-e [84].
14a +
O
HCl
H2O
O
HO
N
N N
HO
N
H
Cl
O
HO
N
N N
HO
N
H
H
H
+
-Cl-
O
HO
N
N N
HO
49
N
-H+
Scheme 17.
N
N NH
NH2
H2N
N
N NH
NH2
F
N
N N
NH2
F
O
TolO
TolO
aq. NH3,
dioxane,
100°C
27a
50 51
52
91% 30% 74% 85%
38i
HF/pyridine,
tBuNO2, -50°C 7, TDA-1,
KOH, MeCN
NH3/MeOH,
rt
Scheme 18.
N
N NH
Cl
Cl
N
N N
Cl
Cl
O
TolO
TolO
53a-e
7, TDA-1,
KOH, MeCN
R
R
N
N N
NH2
Cl
O
HO
HO
38a-e
R
N
N N
OMe
Cl
O
HO
HO
55a-e
R
NH3,
MeOH,
100°C
NH3, MeOH, rt
a: R = H; b: R = Cl; c: R = Br; d: R = I; e: R = F
N
N N
OMe
MeO
O
HO
HO
57a-e
R
a: 74%
b: 73%
c: 63%
d: 71%
e: 60%
a: 48%
b: 86%
c: 89%
d: 80%
a: 88%
b: 84%
c: 83%
d: 83%
a: 79%
b: 89%
c: 83%
d: 92%
0.5M NaOMe/MeOH, 60°C
HN
N N
NH2
O
O
HO
HO
36a,e
R
dilute aq. NH3,
hv
a: 54%
e: 35%
54a-e
N
N N
Cl
Cl
O
TolO
TolO
R
54a-e
Scheme 19.
172 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
3.2.3. Synthesis of 7-Deazapurine 2'-Deoxyribonucleosides Re-
lated to 2-Amino-2’-deoxyadenosine, 2'-Deoxyisoguanosine, 2'-
Deoxyisoinosine, and Derivatives Thereof (18a-e, 36a-c and 37)
Compound 28a was transformed to the 2,6-diamino 7-
deazapurine nucleoside 18a (Scheme 20). The selective deamina-
tion of 18a at position-2 with NaNO2/AcOH/H2O furnished 7-
deaza-2’-deoxyisoguanosine (36a) [86].
The syntheses of the 7-halogenated 2-amino-7-deazaadenine
nucleosides 18b-d and 7-deaza-2’-deoxyisoguanine derivatives
36b,c were carried out according to Scheme 21 [70, 87, 88]. As
direct halogenation of 2-amino-6-chloro-7-deazapurine (27a) takes
place at the 8-position, the 2-amino group of 27a was protected
with a pivaloyl residue giving the 2-pivaloyl derivative 41a. The
halogenation reactions of 41a with N-halosuccinimides (NXS, X =
Cl, Br, and I) were performed in CH2Cl2 yielding 7-halogenated
compounds 41b-d exclusively [70, 87, 88]. Subsequent solid-liquid
nucleobase anion glycosylations of 41b-d with halogenose 7 (TDA-
1/KOH/MeCN) afforded the toluoyl-protected nucleosides 42b-d in
62-68% yield (Scheme 21) [70, 88]. Removal of the toluoyl protect-
ing groups and displacement of the 6-chloro substituents of 42b-d
were performed in 25% aq. ammonia in a steel bomb. The 2,6-
diamino nucleosides 18b-d were formed, and the 7-halogen sub-
stituents were not displaced under these conditions. Selective
deamination of 18b or 18c with sodium nitrite in AcOH/H2O (V/V,
1:5) furnished 7-chloro-7-deaza-2'-deoxyisoguanosine (36b) and
the corresponding 7-bromo derivative 36c.
When the deprotection/displacement conditions used for the
conversion of 42b-d were applied to the corresponding 7-
fluorinated derivative 42e, decomposition of the molecule was ob-
served [84]. Under more moderate reaction conditions (NH3,
MeOH, 70°C, 5 d), the diamino compound 18e (33%) was formed
together with 29e (26%) and 11e (21%) as by-products (Scheme
22). Efforts to convert 18e into 7-deaza-2’-deoxy-7-fluoroiso-
guanosine (36e) by deamination conditions which were employed
for the other halogenated derivatives 18b-d led to the decomposi-
tion of 18e. Instead, nucleoside 36e was obtained from 38e by pho-
tochemical conversion as shown in Scheme 19 (see section 3.2.2).
Isoguanosine (iG) as well as its 2'-deoxyribonucleoside (iGd)
form a significant proportion of enol tautomers in aqueous solution
(10%). They are responsible for non-selective base recognition
during hybridization, which finally results in mutagenicity [89-91].
The replacement of iGd by 7-deaza-2'-deoxyisoguanosine (36a)
increases the content of the keto tautomer significantly with KTAUT
= [keto]/[enol] = 1000 [92]. The introduction of 7-halogen substitu-
ents shifts the tautomeric keto-enol equilibrium of 36b and 36c
further towards the keto form with KTAUT [keto]/[enol] 104 com-
ing close to that of 2’-deoxyguanosine (104
105) [93] (Fig. 8). As a
result, nucleosides 36b and 36c show a much better mismatch dis-
crimination against dT than iGd in antiparallel as well as in parallel
DNA [93]. Therefore, they can be expected to increase selectivity
of base incorporation opposite to isoCd in the polymerase-catalyzed
elongation of triphosphates reaction.
N
N
Cl
H2N
O
TolO
N
TolO
N
N
NH2
H2N
O
HO
N
HO
18a
HN
N
NH2
O
O
HO
N
HO
36a
aq. NH3,
dioxane
AcOH/H2O,
NaNO2, rt
88% 67%
28a
Scheme 20.
N
N NH
Cl
PivHN
27a 41a 41b-d
N
N NH
Cl
H2N
t-BuCOCl,
pyridine NXS, CH2Cl2
84% b: 81%
c: 80%
d: 85%
7, MeCN,
KOH, TDA-1
42b-d
HN
N
NH2
O
R
aq. NH3/dioxane,
120°C, 24h
.
b: 85%
c: 88%
d: 89%
AcOH/H2O,
NaNO2, rt, 0.5h
N
N NH
Cl
PivHN
R
N
N
Cl
PivHN
R
O
TolO
N
TolO
O
HO
N
HO
N
N
NH2
H2N
R
O
HO
N
HO
18b-d 36b,c
b: R = Cl; c: R = Br; d: R = I
b: 62%
c: 68%
d: 58%
b: 61%
c: 65%
Scheme 21.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 173
a: R = H; b: R =Cl; c: R = Br
KTAUT
HN
N
NH2
OHO
HO
NO
N
N
NH2
OHO
HO
NHO
36a-c (keto) 36a-c (enol)
36a: K TAUT = 103
36b: KTAUT = 104
36c: KTAUT = 104
R R
Fig (8). Tautomeric equilibrium of 7-deaza-2’-deoxyisoguanosines 36a-c.
Single-crystal X-ray analysis of 7-deaza-2’-deoxyisoguanosine
(36a) and its 7-bromo derivative 36c confirms that both nucleosides
form the N1-H, 2-keto-6-amino tautomer in the crystalline state [93,
94] (Fig. 9). The sugar ring adopts a C2’-endo sugar conformation
with S-type pucker in both crystals. The predominant conformation
in aqueous solution is also S [70].
7-Deaza-2’-deoxyisoinosine (37), which is the 6-deamino ana-
log of 2'-deoxyisoguanosine, is a highly fluorescent nucleoside with
a quantum yield of 0.22 [95]. It was synthesized utilizing 6-chloro-
7-deaza-2-methoxypurine (58) as a starting material [96] (Scheme
23). Glycosylation of 58 with halogenose 7 under solid-liquid con-
ditions (KOH, TDA-1, MeCN) yielded 59 [97]. Catalytic hydro-
genation of the latter furnished the protected 2-methoxy compound
60, which was deblocked (NH3/MeOH) to give nucleoside 61. The
latter was treated with 2 N NaOH in the presence of DMSO under
reflux conditions furnishing nucleoside 37. The fluorescence spec-
trum of 37 is shown in Fig (10). Compound 37 shows an emission
maximum at 440 nm upon excitation at 328 nm [98].
3.2.4. Nucleobase Anion Glycosylation - Sodium Salt Procedure
The sodium salt procedure generates 7-deazapurine anions with
sodium hydride. It goes back to a protocol developed by Goto for
the ribonucleoside Q synthesis in 1977 [35, 36]. As 2,3,5-tri-O-
benzyl-D-ribofuranosyl bromide, which was used for the glycosyla-
tion, exists as an anomeric mixture and the benzyl groups do not
assist in neighboring group participation from the -side, the out-
come of this glycosylation reaction is an anomeric mixture of -
and -nucleosides. Later, a number of 7-deazapurine -D-
arabinonucleosides were prepared in our laboratory by this tech-
nique [37, 54, 99]. In 1984, Robins and Kazimierczuk used NaH to
generate the nucleobase anion with MeCN as solvent and employed
the sugar halide 7 in the glycosylation reaction [38]. This method
was applied to the synthesis of a number of 7-dezapurine 2'-
deoxyribonucleosides, such as 7-deaza-2’-deoxyguanosine (12a),
its derivatives 12d, 12g, 12h, 12j [100-103], and related 2’-
deoxytubercidin derivatives 14a, 14f, 14j, 18f, 21, 38a, and 62-68
(Fig. 11) [38, 39, 104-106].
42e
N
N
Cl
PivHN
F
O
TolO
N
TolO
N
N
NH2
H2N
F
O
HO
N
HO
18e (33%)
N
N
Cl
H2N
F
O
HO
N
HO
29e (26%)
N
N
OMe
H2N
F
O
HO
N
HO
11e (21%)
NH3, MeOH
70°C, 5d+ +
Scheme 22.
(A)
(B)
Fig (9). Perspective views showing the displacement ellipsoids obtained from the single-crystal X-ray analyses of compounds 36a (A) and 36c (B).
174 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
The synthesis of the 2-substituted 2’-deoxytubercidin deriva-
tives 38a and 21 by the sodium salt protocol is shown in Scheme 24
[38]. The nucleobase anion of 53a or 69 [107] which was generated
with NaH, was glycosylated with the halogenose 7 yielding -D-
nucleosides 54a or 70. They were deprotected and aminated at posi-
tion-6 to give compounds 38a or 21 (aq. NH3, 110°C).
Our laboratory used the sodium salt protocol for the synthesis
of 7-deaza-2’-deoxyxanthosine (35a). The glycosylation of 7-
deaza-2,6-dichloropurine (53a) with 7 was performed in DMF in
the presence of NaH yielding the -D-nucleoside 54a (53%)
(Scheme 25). The latter was transformed to 2,6-dimethoxy nucleo-
side 57a. Demethylation of 57a with 33% HBr/AcOH furnished the
xanthosine derivative 35a [108].
In the crystalline state, compound 35a forms water-filled nano-
tubes with C-H· · · O hydrogen bonds (Fig. 12A and Fig. 12B)
[109]. In addition to the intermolecular hydrogen bonds (N3-H· · ·
OH-5’), an array of further hydrogen bonds stabilizes a su-
pramolecular aggregate of four molecules of 35a. These interac-
tions cause the formation of an almost flat tetramer (Fig. 12A) with
an oval cavity at the center that has the approximate dimensions of
9.5 6.5 3.0 Å3 (± 0.5 Å
3). A pile of completely stacked tetramers
forms a nanotube-like structure (Fig. 12B). Compound 35a repre-
58
N
N NH
Cl
MeO
MeCN, KOH, TDA-1
59
NH3,
MeOH
N
N
Cl
MeO
O
TolO
N
TolO
60
N
NMeO
O
TolO
N
TolO
Pd/C, H2
61
N
NMeO
O
HO
N
HO
HN
NO
O
HO
N
HO
2N NaOH,
DMSO
+ 7
37
86% 92%
81% 85%
Scheme 23.
Fig (10). Fluorescence spectra of nucleoside 37 in water. The concentration of 37 was 10-5
M.
HN
NO
O
HO
N
HO
37
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 175
sents a new nanostructure which binds water molecules inside a
nanotube.
The sodium salt glycosylation was also applied to the synthesis
of 2’-deoxytoyocamycin (14f) and 2’-deoxysangivamycin (14j)
using 2-amino-5-bromopyrrol-3,4-dicarbonitrile (71) as starting
material (Scheme 26). After protection of the amino group with
diethoxymethyl acetate in MeCN, compound 71 was glycosylated
with halogenose 7 in the presence of NaH yielding the intermediate
72. This was cyclized in NH3/MeOH to give 8-bromo-7-cyano-7-
deaza-2’-deoxyadenosine (73). 2’-Deoxytoyocamycin (14f) was
prepared from 73 by a three step procedure (acetylation, debromi-
nation, and deacetylation) [104, 106]. Then, compound 14f was
converted to 2’-deoxysangivamycin (14j) using H2O2/NH3·H2O
[104].
Similarly, 7-cyano-7-deaza-2,6-diaminopurine 2’-deoxyribo-
nucleoside (18f) was prepared [87] using 7-cyano-7-deaza-2,6-
diaminopurine (77) as starting material (Scheme 27). Nucleobase
77 was synthesized from 71 by debromination (H2/Pd/BaCO3,
76), followed by ring-closure with chloroformamidine hydrochlo-
ride at 170°C. In order to increase solubility, compound 77 was
transformed to the more soluble 78. Glycosylation of 78 with halo-
genose 7 (NaH/MeCN) gave -D-nucleoside 79, which was depro-
tected using aq. NaOH/pyridine/MeOH ( 80) followed by 40%
aq. MeNH2 resulting in nucleoside 18f.
N
N
NH2
O
HO
N
HO
R
N
N
NH2
O
HO
N
HO
RN
N
NH2
O
HO
N
HO
R
38a: R = Cl
21: R = SCH3
62: R = CH3
63: R = CH3
64: R = Cl
N
N
O
HO
N
HO
CH3
65
N
N
O
O
HO
N
HO
R
H2N
12a: R = H
d: R = I
g: R = NO2
h: R = CO2H
j: R = CONH2
14a: R = H
f: R = CN
j: R = CONH2
N
N
O
HO
N
HO
CH3
66
N
N
O
HO
N
HO
68
H3CS
S
N
N
NH2
O
HO
N
HO
H2N
18f
CN
N
N
NH2
O
HO
N
HO
Cl
H3C
67
H
Fig (11).
54a
68%
N
N
OMe
MeO
O
HO
N
HO
57a
HN
NH
O
O
HO
N
HO
35a
60%
NaOMe, MeOH O HBr, AcOH
53a + 7NaH, DMF
53%
Scheme 25.
O
TolO
N
N N
TolO
Cl
N
N NH
Cl
O
TolO
TolO
Cl
+ NaH, MeCN
53a: R = Cl
69: R = SCH3
7
NH3/H2O,
110°C
O
HO
N
N N
HO
NH2
R
R R
54a: 60%
70: 66%
38a: 61%
21: 72%
54a: R = Cl
70: R = SCH3
38a: R = Cl
21: R = SCH3
Scheme 24.
176 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
(A) (B)
Fig (12). Crystalline nanotubes of stacked tetramers of 7-deaza-2’-deoxyxanthosine (35a) units.
O
TolO
TolOO
TolO
TolO
Cl
+
71
772
O
HO
N
N N
HO
NH2
73
NH
NC
H2N
CN
Br
N
NC
N
CN
Br
EtO
CN
Br
NH3/MeOH, rt
(i) diethoxymethylacetate,
MeCN, reflux
(ii) NaH, MeCN
75% 77%
O
AcO
N
N N
AcO
NH2
74
CN
Br
O
AcO
N
N N
AcO
NH2 CN
O
HO
N
N N
HO
NH2 R
Ac2O,
pyridine
BSA, MeCN, KF,
18-crown-6 ether NH3/MeOH
97% 56% 10%
7514f: R = CN
j: R = CONH2
H2O2/aq. NH3
BSA = N,O-bis(trimethylsilyl)acetamide
Scheme 26.
7, NaH,
MeCN
H2/Pd/BaCO3,
DMF/MeOH
NH
NC
H2N
CN
NH2
ClH2N
Cl
Dowtherm A,
170°C, 80%
N
N NH
NH2 CN
H2N
N
N NH
NHPivCN
PivHN
O
TolO
N
N N
TolO
NHPivCN
PivHN
O
HO
N
N N
HO
NH2 CN
H2N
40% aq. MeNH2,
55°C, 18h
pivaloyl chloride,
pyridine
60% 72%
70%
71
76 77 78
79 18f
O
HO
N
N N
HO
NHPivCN
PivHN
80
aq. NaOH,
pyridine, MeOH
96%
Scheme 27.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 177
7, NaH, MeCN
74%
28f
N
N
Cl
H2N
CN
O
TolO
N
TolO
12h
HN
N
O
H2N
CO2H
O
HO
N
HO
64%
HN
N
O
H2N
C
NH
HN
N
O
H2N NH2
OMe
O
Cl
CHO
HN
N
O
H2N N
OMe
O
OCMe3O
NH2
O
N
N
Cl
H2N
CN
NH
+ HN
N
O
H2N NH
OMe
O
50%
DBDC,
DMF-TEA
50%
NH3/MeOH,
120°C
73%
POCl3,
DEA
40%
12j
HN
N
O
H2N
C
O
HO
N
HO
NH3/MeOH
88%
29f
N
N
Cl
H2N
CN
O
HO
N
HO
NH2
O
NaOAc,
100°C
H2O2,
NH3/H2O
56%
81 82 83 84
85 27f
5M KOH
Scheme 28.
The synthesis of 2’-deoxycadeguomycin (12h) by the sodium
salt procedure is shown in Scheme 28 [101, 110]. The condensation
of 81 with 82 was performed in aq. NaOAc solution to give the 7-
deazaguanine derivative 83. After protection of the pyrrole nitrogen
with a t-BOC group ( 84), the methyl ester was converted to an
amide with concomitant deprotection of the t-BOC group yielding
nucleobase 85. Chlorination with POCl3 in the presence of N,N-
diethylaniline (DEA) gave 6-chloro-7-cyano-7-deazapurine (27f).
Glycosylation of 27f with halogenose 7 (NaH/MeCN) furnished the
protected nucleoside 28f. Deprotection of 28f yielded compound
29f, which was converted to 7-carbamoyl-7-deaza-2’-
deoxyguanosine (12j) using H2O2/NH3/H2O. The amide hydrolysis
of 12j with 5 M KOH afforded 2’-deoxycadeguomycin (12h).
The sodium salt procedure was also applied to the synthesis of
7-deaza-2’-deoxy-7-nitroguanosine (12g) (Scheme 29). The nitro
group was introduced in 41a (see Scheme 21) with HNO3/H2SO4
(1:1) yielding the 7-nitro derivative 41g, which was converted to 7-
nitro-7-deazapurine 9g. Compound 9g was glycosylated with 2-
deoxy-3,5-di-O-(p-chlorobenzoyl)- -D-erythro-pentofuranosyl
chloride (86) in DMF in the presence of NaH resulting in -D-
nucleoside 87. Demethylation (Me3SiCl/NaI/MeCN, 88) was
followed by deprotection with K2CO3/MeOH, yielding 7-deaza-2’-
deoxy-7-nitroguanosine (12g) [78, 79].
3.3. 7-Deazapurine 2’-Deoxyribonucleosides From Ribonucleo-
sides by Deoxygenation
As mentioned in the introduction of section 3, chemical deoxy-
genation of ribonucleosides to their corresponding 2’-
deoxyribonucleosides offers an expedient route when other syn-
thetic or enzymatic protocols fail. Different deoxygenation proce-
dures were applied to purine or pyrimidine ribonucleosides with
variable success [111-113].
7-Deazapurine ribonucleoside 2’-deoxyribonucleoside con-
version is reported in an early publication (1976) of Robins and
Muhs, in which the transformation of the antibiotic tubercidin (89a)
to its 2’-deoxyribo derivative (14a) is described [114]. The reaction
sequence includes hydroxyl and amino group protection, introduc-
tion of a mesyloxy group followed by catalytic hydrogenation to
yield eventually the deoxygenated target compound 2’-
deoxytubercidin (14a) in 27% overall yield over eight steps
(Scheme 30).
In the mid 1970s, Barton and McCombie developed a versatile
method for the deoxygenation of alcohols by radical reduction
[115]. This protocol comprises the conversion of the respective
alcohol to a thiocarbonyloxy derivative as a first step. Second, the
thiocarbonyl oxy derivative is treated with tri-n-butyltin hydride (n-
Bu3SnH) together with a radical initiator to finally yield the deoxy-
genated compound by a radical-mediated mechanism.
Later, Robins and Wilson made use of the Barton-McCombie
deoxygenation to establish a four-step procedure for the specific
conversion of ribonucleosides to their corresponding 2’-
deoxyribonucleosides [116]. This protocol was also applied for the
conversion of tubercidin (89a) and toyocamycin (89f) into the cor-
responding 2’-deoxyribonucleosides 14a,f,j as shown in Scheme 31
[117]. First, the respective ribonucleoside 89a or 89f was treated
with Markiewicz’s reagent (1,3-dichloro-1,1,3,3-tetraisopropyl-
178 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
NaH, DMF, 0°C
61%
87
N
N
OMe
H2N
NO2
O
pCl BzO
N
pCl BzO
88
HN
N
O
H2N
NO2
O
pCl BzO
N
pCl BzO
12g
HN
N
O
H2N
NO2
O
HO
N
HO
79%
Me3SiCl,
NaI, MeCN
73%
86
O
pCl BzO
pCl BzOCl
41a
41g
N
N NH
Cl
PivHN
NO2
9g
N
N NH
OMe
H2N
NO2
HNO3/H2SO4
78%
NaOMe,
MeOH
65%
K2CO3, MeOH
Scheme 29.
N
N
NH2
O
HO
N
HO
14a
N
N
NH2
O
N
HO
90
N
N
NBz2
O
N
BzO
N
N
NHBz
O
N
BzO SBn
O O
OH
N
N
NH2
O
HO
N
HO
89a
OH
N
N
NHBz
O
N
HO SBn
OH
N
N
NHBz
O
N
BzO SBn
OMs
+
91
92a (68%) 92b (22%) 93
N
N
NH2
O
N
HO SBn
OH
94
N
N
NH2
O
N
HO BnS
95
benzoylation
sodium benzylthiolate,
hot THF
quantitatively
92a: mesylation
1) Na benzoate, hot DMF
2) NaOMe, MeOH
HO
+
95: Ra-Ni,
DMF, 100°C
77%
Scheme 30.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 179
O
HO
HO
89a,f
OH
O
O
O
96a,f
OH
Si
O
Si
i-Pr i-Pr
i-Pr
i-Pr
(i-Pr)2ClSiOSi(i-Pr)2Cl,
pyridine, rt
O
O
O
OCOPh
Si
O
Si
i-Pr i-Pr
i-Pr
i-Pr
S
PhOC(S)Cl, DMAP,
MeCN, rt
O
O
O
98a,f
Si
O
Si
i-Pr i-Pr
i-Pr
i-Pr
(n-Bu)3SnH, AIBN,
toluene, 75°C 1M TBAF/THF
N
NN
NH2
N
N N
NH2
N
NN
NH2
N
N N
NH2
R
97a,f
97a,f
R R
R
O
HO
HO
14a: 68% overall yield
f: 69% overall yield
N
N N
NH2 R
14f: Dowex 1-X2 (OH-)
14j (65% overall yield)
a: R = H; f: R = CN
O
HO
HO
N
N N
NH2 CONH2
Scheme 31.
disiloxane) [118] to simultaneously protect the 3’- and 5’-hydroxyl
groups of the sugar moiety ( 96a,f). Reaction of the silylated
nucleoside 96a,f with phenoxythiocarbonyl chloride in the presence
of dimethylaminopyridine (DMAP) using acetonitrile as solvent
gave the 2’-O-phenoxythiocarbonyl derivative 97a or 97f. Reduc-
tive deoxygenation of 97a or 97f was performed with tri-n-
butyltin(IV) hydride in toluene at 75°C using , ’-
azoisobutyronitrile (AIBN) as radical initiator and furnished the
silylated nucleosides 98a,f. The last step comprised desilylation
with tetra-n-butylammonium fluoride (TBAF) in THF to give the
free 2’-deoxyribonucleoside 14a or 14f. 2’-Deoxytubercidin (14a)
and 2’-deoxytoyocamycin (14f) were obtained in 68% and 69%
yield, respectively which is superior to any other chemical trans-
formation protocol employing the corresponding parent ribonucleo-
sides as precursors [117]. 2’-Deoxytoyocamycin (14f) was further
converted to 2’-deoxysangivamycin (14j; 65% overall yield over 5
steps) by passing an aqueous solution of 14f through a column of
Dowex 1-X2 (OH-) resin (Scheme 31).
Depending on the structure of the target nucleosides, variations
of the Barton-McCombie deoxygenation were used to synthesize
2’- and 3’-deoxyribonucleosides as well as 2’-3’-dideoxyribo-
nucleosides (see also section 4.3) [119-123].
4. FUNCTIONALIZATION OF 7-DEAZAPURINE 2'-
DEOXYRIBONUCLEOSIDES
4.1. Alkynylated 7-Deazapurine Nucleosides Prepared by the
Palladium-catalyzed Sonogashira Cross Coupling Reaction
The introduction of alkynyl- or aminoalkynyl side chains into
7-deazapurine nucleosides has a major impact on the duplex stabil-
ity when these compounds are constituents of duplex DNA or RNA
[87, 100, 124-126]. Furthermore, oligonucleotides become resistant
against enzymatic degradation [127]. Nucleosides, nucleotides or
oligonucleotides with aminoalkynyl chains are used for the func-
tionalization with reporter groups applied to nucleic acid sequenc-
ing or as tools in DNA or RNA diagnostics [128-134]. More re-
cently, 7-deazapurine 2’-deoxyribonucleosides carrying a terminal
triple bond within their side chain have been used as precursors in
the copper(I)-catalyzed Huisgen-Meldal-Sharpless alkyne-azide
cycloaddition (“click” reaction) (for more details see section 4.1.1)
[135-137]. The 7-iodo derivatives of 7-deazapurine 2’-
deoxyribonucleosides described in section 3.2 were employed as
starting materials for the introduction of alkynyl or aminoalkynyl
chains by the Pd-catalyzed Sonogashira cross-coupling reaction
[75]. Exemplified cross-coupling reactions performed on the iodo
nucleosides 12d, 14d, 18d, and 15d are depicted in Scheme 32.
The cross coupling reactions of 7-deaza-7-iodopurine 2’-
deoxyribonucleosides 12d, 14d, 15d, and 18d with the correspond-
ing alkynes were performed in anhydrous DMF in the presence of
tetrakis(triphenylphosphine)palladium(0) [Pd(0)(PPh3)4], copper(I)
iodide, and triethylamine under argon atmosphere, yielding the 7-
alkynyl- or aminoalkynyl derivatives 99a,b,d,f,h-k [63, 100, 126,
138-141], 100a-i,k [75, 100, 124, 142-144], 101b,i [87, 126], or
102f,g,i [126, 145, 146] (Scheme 32). This protocol was also em-
ployed for the construction of steroid-nucleoside conjugates (103,
104), employing 17 -ethynylestradiol or 17 -ethynyltestosterone
(Fig. 13) [75]. Some of the above mentioned nucleosides were
transformed to the corresponding phosphoramidites 105a,b,f [141,
144, 147], 106b,d,h-k [127, 138, 140, 144, 147], 107a,d,g,h [124,
142], 108b,f,i,k [139, 144, 148, 149], and 109f,g,i [126, 145, 146],
which were used in solid-phase oligonucleotide synthesis (Fig. 14).
Sonogashira cross coupling reactions were also performed in
aqueous medium (H2O/MeCN) using Pd(OAc)2 as catalyst and
3,3 ,3 -phosphinidyne tris(benzenesulfonic acid) trisodium salt
(P(PhSO3Na)3; TPPTS) as water soluble ligand in the presence of
180 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
12d
O
HO
HN
N N
HO
O
H2N
O
HO
HN
N N
HO
O
H2N
I R
100a-i,k
O
HO
N
N N
HO
NH2 R
(CH2)4
a: R =
b: R =
c: R =
d: R =
e: R =
f: R =
CH3
(CH2)3CH3
99a,b,d,f,h-k
NPhthCH2
(CH2)3NHCOCF3
(CH2)2CH3
CH2NHCOCF3g: R =
h: R =
i: R =
j: R =
k: R =
101b,i
O
HO
N
N N
HO
NH2 R
H2N
Pd(0)(PPh3)4, CuI,
Et3N, DMF, R-H
14d
O
HO
N
N N
HO
NH2 I
18d
O
HO
N
N N
HO
NH2 I
H2N
Pd(0)(PPh3)4, CuI,
Et3N, DMF, R-H
Pd(0)(PPh3)4, CuI,
Et3N, DMF, R-H
102f,g,i
O
HO
HN
N N
HO
OR
15d
O
HO
HN
N N
HO
OI
Pd(0)(PPh3)4, CuI,
Et3N, DMF, R-H
(CH2)4CH3
NPhth(CH2)2
N
H
Scheme 32.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 181
O
HO
N
N N
HO
NH2
103
C
HO
C
CH3
O
CH3
O
HO
N
N N
HO
NH2 C
HO
C
OH
CH3
104
Fig (13).
PN(i-Pr)2NCCH2CH2O
O
O
N
N N
DMTO
R1
PN(i-Pr)2NCCH2CH2O
O
O
HN
N N
DMTO
O
N
R R
105a,b,f 107a,d,g,h
PN(i-Pr)2NCCH2CH2O
O
O
N
N N
DMTO
NMe2NHC R
(CH2)4a: R =
b: R =
d: R =
CH3
(CH2)3CH3
NPhthCH2
(CH2)3NHCOCF3
CH2NHCOCF3
f: R =
g: R =
h: R =
NPhth(CH2)2
N
Me2NHC
PN(i-Pr)2NCCH2CH2O
O
O
HN
N N
DMTO
O
(i-Bu)HN
R
106b,d,h-k 108b: R1 = N=C(Me)NMe2
f: R1 = NH(i-Bu)
i: R1 = NHAc
k: R1 = NH(i-Bu)
PN(i-Pr)2NCCH2CH2O
O
O
HN
N N
DMTO
OR
109f,g,i
i: R =
j: R =
k: R =
Fig (14).
CuI and Et3N. Due to the aqueous conditions, nucleoside triphos-
phates can also be employed in the cross coupling reaction. 7-
Deaza-2’-deoxy-7-iodoadenosine (14d) or its triphosphate analog
110 were utilized as precursors together with the respective alkynes
to synthesize various nucleoside or nucleoside triphosphate conju-
gates with bipyridine ligands (111a,b), ruthenium bipyridine com-
plexes (111c,d, 112d), ferrocene (112e), amino acids (111f, 112f)
or bile acids (111g, 112g) attached to position-7 of the nucleobase
(see Scheme 33 for selected examples) [150-154]. Alternatively, the
aqueous-phase Suzuki-Miyaura cross coupling reaction was applied
to introduce similar reporter groups into position-7 of the 7-
deazapurine moiety as shown in Scheme 34. Reactions of the corre-
sponding boronate precursors 113a-c or 114d-i with iodo nucleo-
side 14d or iodo nucleotide 110 in the presence of Pd(OAc)2-
TPPTS and Cs2CO3 in MeCN/H2O afforded the 7-substituted com-
pounds 115a-g [153, 154] as well as 116g-i [153, 155, 156]. Nu-
cleoside triphosphate conjugates 112d-g and 116g-i obtained by
either of the above mentioned coupling reactions have been suc-
cessfully subjected to enzymatic incorporation into oligonucleotides
[150-153, 155, 156].
Single-crystal X-ray analyses of 7-deaza-2’-deoxy-7-propynyl-
guanosine (99b) and 7-deaza-2’-deoxy-7-propynyladenosine (100b)
were reported by our laboratory [157, 158] (Fig. 15). In both com-
pounds, the orientation of the nucleobase relative to the sugar moi-
ety is anti with the torsion angle (O4’-C1’-N9-C4) of -117.1 (5)°
for 99b and of -130.7 (2)° for 100b. The sugar moieties of these
nucleosides adopt the S-conformation (99b: P = 152.5° with an
amplitude m = 41.9°; 100b: P = 185.9°, m = 39.1°). The linear
propynyl group is inclined by 4.6° to the nucleobase for 99b and by
1.6° for 100b. The triple bond length is 1.184 (7) Å for 99b and
1.185 (3) Å for 100b, which is in the range of non-conjugated tri-
ple-bonds.
In the multi-layered network of nucleoside 99b, the nucleobases
are stacked head-to-head with the closest distance of 3.728 (1) Å.
This is slightly higher than the average base pair stacking distance
in B-DNA (3.5 Å) (Fig. 16A). In contrast, a head-to-tail stacking of
182 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
O
HO
N
N N
R2O
NH2
14d: R = H
O
HO
N
N N
RO
NH2 I
111a: R1 = a, R2 = H
111b: R1 = b, R2 = H
111c: R1 = c, R2 = H
111d: R1 = d, R2 = H
112d: R1 = d, R2 = P3O104 -
112e: R1 = e, R2 = P3O104 -
111f: R1 = f, R2 = H
112f: R1 = f, R2 = P3O104 -
111g: R1 = g, R2 = H
112g: R1 = g, R2 = P3O104 -
R1
N N N N
N N
RuII(bpy)2
2 PF6-
N N
RuII(bpy)2
2 PF6-
Fe
Pd(OAc)2, TPPTS, Et3N,
CuI, DMF or H2O/MeCN
a b c
d
e
NH2
COOH
f
R1 =
HO
OH
O
O
OHH
g
R1
P O
O
O
P O
O
O
P
O
O
O110: R =
Scheme 33.
O
HO
N
N N
R2O
NH2
14d: R = H
O
HO
N
N N
RO
NH2 I
115a: R1 = a, R2 = H
115b: R1 = b, R2 = H
115c: R1 = c, R2 = H
115d: R1 = d, R2 = H
115e: R1 = e, R2 = H
115f: R1 = f, R2 = H
115g: R1 = g, R2 = H
116g: R1 = g, R2 = P3O104 -
116h: R1 = h, R2 = P3O104 -
116i: R1 = i, R2 = P3O104 -
N N
N N
RuII(bpy)2
2 PF6-
Pd(OAc)2, TPPTS, Cs2CO3
or Na2CO3, H2O/MeCN
a
d
NH2
COOH
g
R1 =
NNN
N
N
RuII(bpy)2
2 PF6-
RuII(tpy)
2 PF6-
N N N
N
N
NH2 NO2
R1
R1 B
O
O
113a-c
R1 B
OH
OH
114d-i
b c
e f
h i
113a-c or 114d-i,
P
O
O
O P
O
O
O P
O
O
O
110: R =
Scheme 34.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 183
(A) (B)
Fig (15). Perspective views of (A) 7-deaza-2’-deoxy-7-propynylguanosine (99b) and (B) 7-deaza-2’-deoxy-7-propynyladenosine (100b). Displacement ellip-
soids are drawn at the 50% probability level.
(A) (B)
Fig (16). Views showing (A) head-to-head base stacking of nucleoside 99b and (B) head-to-tail base stacking of nucleoside 100b.
nucleoside nucleoside"click" reaction
reporter groupreporter groupN3
N N
N
Fig (17). Schematic illustration of the “click” reaction.
nucleobases is observed for nucleoside 100b (Fig. 16B). The closest
distance of the stacked bases is 3.197 (1) Å for 100b, which is
smaller than the average base pair stacking distance in B-DNA.
4.1.1. Functionalization of 7-Deazapurine 2’-Deoxyribonucleo-
sides Empolying the “Click” Reaction
The copper(I)-catalyzed Huisgen-Meldal-Sharpless 1,3-dipolar
cycloaddition of organic azides and alkynes (“click” reaction,
CuAAC reaction) has emerged as one of the ideal bio-orthogonal
protocols for the preparation of rich chemical diversity [159-161].
This cycloaddition is driven by the high energy content of the com-
ponents (azides and alkynes), yielding less reactive 1,2,3-triazoles,
which are highly stable to oxygen, light, and aqueous environment
[159, 160]. In nucleic acid chemistry, the CuAAC reaction has been
performed on nucleoside, nucleotide as well as on oligonucleotide
level in solution, on solid support, or on surfaces [for e.g. see 137,
162-177].
The synthesis of azide-modified DNA is encountered with dif-
ficulties. As azides are prone to reduction during solid-phase oli-
gonucleotide synthesis, the alkyne component is usually attached to
the nucleoside, while the reporter group carries the azido function-
ality. To avoid perturbation of the DNA structure, the “click” reac-
tion is performed most efficiently when the ligand is introduced in
the major groove of DNA. In this regard, position-7 of 7-
deazapurine 2’-deoxyribonucleosides - as purine surrogates - has
become an ideal site for introducing side chains with terminal triple
184 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
bonds [87, 100, 124-126]. The alkynylated nucleosides 99a,f,
100a,f and 102f employed in “click” reactions are shown in section
4.1, Scheme 32. As ligands, simple residues such as benzylazide
(117) [178, 179] as well as fluorescent or fluorogenic reporter
groups have been used (Fig. 18), affording highly fluorescent nu-
cleosides. Fig. (17) schematically outlines the “click” reaction em-
ploying benzylazide 117, 3-azido-7-hydroxycoumarin 118, 3-azido-
7-methoxycoumarin 119, 1-azidomethyl pyrene 120 or 9-
azidomethyl anthracene 121 as ligands. Exemplarily, the CuAAC
reaction is shown for the “click” conjugate 122 using 7-deaza-2’-
deoxy-7-octa-1,7-diynyladenosine (100f) as precursor (Scheme 35).
Functionalization with 3-azido-7-hydroxycoumarin (118) was per-
formed in the presence of CuSO4·5 H2O using sodium ascorbate as
a reducing agent in THF/H2O/t-BuOH (3 : 1 : 1) at room tempera-
ture [180]. Likewise the “click” reaction was performed on the 7-
deazapurine 2’-deoxyribonucleosides 99a, 100a,f and 102f to afford
various fluorescent “click” conjugates (123-129) (Fig. 19) [140,
145, 146, 149, 178, 180-182].
7-Deazapurine “click” conjugates were also subjected to fluo-
rescence studies [139, 140, 145, 146, 181, 182]. It was found that 7-
deazapurine nucleoside dye conjugates show significantly lower
fluorescence than those of corresponding 8-aza-7-deazapurine or
pyrimidine nucleosides; a phenomenon which was studied with 7-
hydroxycoumarin or anthracene as reporter group [181, 182]. Two
examples illustrating these findings are shown in Fig. (20).
The fluorescence properties of compound 128 were investigated
at pH 8.5 as the 7-hydroxycoumarin dye exists predominantly in the
anionic form only under alkaline conditions [182]. The “click” con-
jugate 128 has an excitation maximum at 393 nm with an emission
at 477 nm (Fig. 20a). For comparison, the fluorescence spectrum of
the corresponding 8-aza-7-deaza-2’-deoxyguanosine coumarin
“click” conjugate 130 was measured. Fig. (20b) shows a direct
comparison of the emission spectra of 128 and 130, indicating that
the fluorescence maximum of 128 is about 10-times lower than that
of 130 [182]. Accordingly, the fluorescence properties of the 7-
deaza-2’-deoxyadenosine anthracene “click” conjugate 126 were
compared with those of the corresponding 8-aza-7-deaza-2’-
deoxyadenosine anthracene conjugate 131 [181]. In this case, the
fluorescence intensity of 126 is reduced by around 95% to the 8-
aza-7-deaza-2’-deoxyadenosine conjugate 131 (Fig. 20c). Quench-
ing within the 7-deazapurine conjugates 126 and 128 was attributed
to a charge transfer between the respective nucleobase and the dye.
Due to their low oxidation potential, 7-deazapurine nucleosides
quench the fluorescence of a dye significantly, while this is not the
case for the corresponding 8-aza-7-deazapurine conjugates (130,
131); most likely due to their higher oxidation potential [139, 181,
182].
In a very recent report, 7-deaza-2’-deoxy-7-ethynyladenosine
100a was conjugated to the spin label 4-azido-2,2,6,6-tetramethyl-
piperidine-1-oxyl (132; 4-azido TEMPO) [183] by applying the
CuAAC reaction [184]. Compound 100a was functionalized with
132 by the “click” reaction in the presence of CuI in a 3:1:1 mixture
of THF/t-BuOH/H2O (Scheme 36). In this case, CuI has been used
as copper(I) source instead of the Cu(II)SO4/ascorbic acid system
(see Scheme 35 for comparison) to avoid reduction of the nitroxide
radical by ascorbic acid to the non-paramagnetic hydroxylamine
derivative during “click” reaction [184]. Addition of N,N-
diisopropylethylamine (DIPEA) was essential for the completion of
the reaction within 4 h. The spin labeled 1,2,3-triazolyl nucleoside
conjugate 133 was obtained in 64%.
O
N3
OH
N3
O
121118
N3
120
O
N3
OMeO
119
N3
117
Fig (18). Reporter groups employed in the “click” reaction with 7-deazapurine 2’-deoxyribonucleosides.
O
HO
N
N N
HO
NH2
N
N
N
OO
OH
122
CuSO4 5 H2O
Na-ascorbate
THF:H2O:t-BuOH,
3:1:1+ 118
O
HO
N
N N
HO
NH2
100f
33%
Scheme 35.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 185
123 (25%)
O
HO
N
N N
HO
NH2
N
N
N
O
HO
HN
N N
HO
O
N
N
N
OO
OH
126 (78%) 127 (85%)
O
HO
HN
N N
HO
O
N
N
N
O
HO
HN
N N
HO
O
N
N
N
OO
OH
129 (83%)128 (53%)
H2N H2N
O
HO
N
N N
HO
NH2
N
N
N
OO
OMe
124 (48%)
O
HO
HN
N N
HO
O
N
N
N
125 (66%)
H2N
O
HO
N
N N
HO
NH2
N
N
N
OO
OH
Fig (19). Fluorescent “click” conjugates.
(a) (b)
O
HO
N
N N
X
HO
NH2
N
N
N
126: X = CH
131: X = N
O
HO
HN
N N
X
HO
O
N
N
N
OO
OH
128: X = CH
130: X = N
H2N
(c) (d)
Fig (20). (a) Excitation and emission spectra of the 7-deaza-2’-deoxyguanosine coumarin “click” conjugate 128 and (b) direct comparison of the emission
spectra of the coumarin “click” conjugates 128 and 130. Measured in a mixture of DMSO (0.5 ml) and 99.5 ml of 0.1 M Tris-HCl buffer at pH 8.5 with a con-
centration of 9.4 10-3
mol/l [182]. (c) Direct comparison of the emission spectra of anthracene “click” conjugates 126 and 131. Measurements were performed
at identical molar concentration (0.98 M) in methanol [181].
186 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
O
HO
N
N N
HO
NH2
133
+
O
HO
N
N N
HO
NH2
100a
N
N
N
NO
CuI, DIPEA, rt,
THF:H2O:t-BuOH, 3:1:1
N
O
N3
132
64%
Scheme 36.
Fig (21). (a) and (c) molecular dynamics (MD) simulation snapshots of a DNA duplex containing two spin labeled residues of 133 within one of the strands. (c)
Spin spin distance for the respective duplex determined for the oxygen atoms of the nitroxides [184].
Functionalization of 100a was performed on nucleoside level
and on 100a being a constituent of DNA oligonucleotides, allowing
accessibility to study continuous wave (cw) and pulse EPR spec-
troscopy. Interspin distances were studied on single-stranded oli-
gonucleotides and duplexes incorporating two spin labeled “click”
conjugates (133) at distant positions (Fig. 21) or when 133 was a
component of a modified ‘dA-dT’ base pair within an oligonucleo-
tide duplex [184]. By applying cw and pulse EPR spectroscopy,
interspin distances in the 1-2 nm range were obtained with high
accuracy. It was suggested that the spin labeled DNA system ob-
tained via “click” reaction has the potential to provide detailed in-
sights into structural changes caused by unusual DNA structures, by
mispairing, DNA damages and/or lesions [184].
Due to the ongoing interest of high density labelling of nucleo-
sides and oligonucleotides, tripropargylamine – a branched side
chain with two terminal triple bonds – was introduced at position-7
of 7-deaza-2’-deoxyadenosine and 7-deaza-2’-deoxyguanosine (
99k, 100k; see Scheme 32, section 4.1). As both bonds can be func-
tionalized simultaneously (“double click” reaction), the density of
labelling is increased. The “double click” reaction was performed in
THF/t-BuOH/H2O (3 : 1 : 1) in the presence of CuSO4·5 H2O and
sodium ascorbate as described for the “mono”-alkynylated com-
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 187
pounds, employing benzylazide 117, 3-azido-7-hydroxycoumarin
118, 1-azidomethyl pyrene 120 or 9-azidomethyl anthracene 121 as
ligands [140, 179]. Scheme 37 shows the “double click” reaction
using the 7-tripropargylamine derivative of 7-deaza-2’-
deoxyguanosine 99k as precursor to afford the nucleoside pyrene
conjugate 134 (79% yield) [140].
To evaluate the photophysical properties, the excitation and
emission spectra of the “double click” product 134 (tripropargy-
lamine pyrene conjugate) were measured and compared to those of
129 (octadiynyl pyrene conjugate; Fig. 19) and the abasic octyne
derivative 135. The abasic derivative 135 contains all necessary
elements of the dye conjugate except 7-deazaguanine (Fig. 22)
[140]. Fig. (22) indicates that only the “double click” product 134
with two proximal pyrenes shows strong excimer fluorescence (464
nm) and rather low monomer fluorescence at 377 nm and 394 nm,
while the conjugate 129 containing one pyrene shows only mono-
meric pyrene emission. Monomeric pyrene emission was also ob-
served for the abasic octyne derivative 135. From this, it was con-
cluded that in “double click” conjugate 134 the two pyrenes are in
proximal position thereby developing strong excimer fluorescence.
From Fig. (22) it is also apparent that both nucleoside pyrene con-
jugates (129 and 134) show rather low monomer fluorescence com-
pared to the abasic conjugate 135. These findings point to a quench-
ing of the pyrene fluorescence by the 7-deazaguanine moiety within
O
HO
HN
N N
HO
O
H2N
N
CuSO4, sodium ascorbate
THF:H2O:t-BuOH, 3:1:1, rt
79%
99k
+
O
HO
HN
N N
HO
O
H2N
N
N
N
N
NN
N
134
120
Scheme 37.
Fig (22). The excitation and emission spectra of “click” conjugates 129, 134 and 135 in methanol. All conjugates were excited at 340 nm, and the concentra-
tion of the “click” conjugates was identical (6.8 x 10-6
M.) [140].
188 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
both nucleoside conjugates which was attributed to an intramolecu-
lar charge transfer [140].
4.1.2. Cross-linking of 7-Deazapurine 2’-Deoxyribonucleosides
The “click” reaction has also been applied to crosslink DNA
strands; a “hot” topic of current research [185-189]. For that, azido
groups as well as terminal triple bonds were introduced at the ter-
mini of oligonucleotides and the “click” reaction was induced with
copper(I) salts by post-modification. Very recently, a novel proto-
col for the cross-linking of DNA strands was developed by our
laboratory [190]. Oligonucleotides incorporating constituents with
alkynylated side chains at a terminal or internal position were se-
lected for cross-linking by a “bis-click” reaction. Through this tem-
plate-free procedure, two identical strands can be linked together by
using bis-azides such as 136 or 138. A stepwise procedure was also
developed, where in the first step only one azido group was reacted
to give a triazole mono-functionalized oligonucleotide conjugate
bearing still another reactive azido group. Consequently, this inter-
mediate has the potential to be cross-linked in a second step with
another strand of any type of DNA bearing an alkynyl group. This
protocol was used to synthesize both identical as well as non-
identical cross-linked oligonucleotides [190], and can also be ap-
plied to nucleosides [149, 178, 190].
The “bis-click” reaction was performed on the 7-octadiynylated
nucleosides 99f and 100f with the respective bis-azides 136 and 138
to give the cross-linked products 137 and 139, respectively
(Scheme 38) [149, 178]. The cross-linking reaction of 99f was car-
ried out in the presence of CuSO4·5 H2O/sodium ascorbate in
THF/t-BuOH/H2O (3:1:1) and afforded 137 in 40% yield, while
CuI, DIPEA in THF/t-BuOH/H2O were used for the cross-linking
of 100f to give 139 in 54% yield.
4.1.3. Functionalization of 7-Deazapurine 2’-Deoxyribonucleo-
sides by Other Methods
Several 7-deazapurine ribonucleosides, such as queuosine
(140), epoxy-queuosine (141) or galactosyl-queuosine (142) (Fig.
HN
N N
O
H2N
+
136
HN
N N
O
H2N
N
N
N
HN
NN
O
H2N
N
N
N
CuSO4 5 H2O
sodium ascorbate,
.
THF/t-BuOH/H2O,
O
HO
HO
O
HO
HO
O
HO
HO
N3 N3
99f 137
N
N N
NH2 CuI, DIPEA, rt, 12h
N
N N
NH2
N
N
N
+
THF/t-BuOH/H2O,
3:1:1
54%
139
N
NN
NH2
N
N
N
O
HO
HO
O
HO
HO
O
HO
HO
100f
S
O
O
S
O
O
N3N3
138
3:1:1, rt, 16h, 40%
Scheme 38.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 189
23), are found in transfer RNAs (tRNAs) [191], and carry 7-
substituents which represent Mannich bases. Corresponding 2’-
deoxyribonucleosides are not naturally occurring. Therefore, it was
of interest to transfer this structural motif into DNA constituents.
Moreover, the Mannich base can be used to introduce reporter
groups into position-7 of 7-deazapurine 2’-deoxyribonucleosides.
In principle, 7-deazapurine nucleosides with Mannich bases as
side chains are accessible by two routes, (i) glycosylation of a suit-
able nucleobase precursor already carrying the Mannich side chain
or (ii) by the Mannich reaction performed on a 7-deazapurine nu-
cleoside. As the solubility of Mannich bases is low in MeCN (nu-
cleobase anion glycosylation), route (ii) was followed [192].
The Mannich reaction was performed on 7-deaza-2’-
deoxyadenosine (14a) in an aq. solution of formaldehyde, mor-
pholine and acetic acid at 60°C (conditions I) to afford compound
143 in 43% yield (Scheme 39). In a similar way, the toluoyl pro-
tected 6-methylthio nucleoside 144 was converted into the Mannich
nucleoside 145 (70% yield) [192].
When the same reaction conditions (aq. formaldehyde, mor-
pholine, AcOH as solvent, 60°C; conditions I) were applied to 7-
deaza-2’-deoxyguanosine (12a), decomposition of 12a was ob-
served. When the amount of acetic acid was reduced and the tem-
perature was strictly controlled (60°C; conditions II), the Mannich
nucleoside 146 carrying the side chain at position-8 was isolated
(Scheme 40) [192]. Next, the isobutyrylated compound 147 [193]
O
HO OH
HN
N N
HO
O
H2N
HN
HO
OH
queuosine
140
O
HO OH
HN
N N
HO
O
H2N
HN
HO
OH
141
O
O
HO OH
HN
N N
HO
O
H2N
HN
142
O
OH
O
OH
HO
OH
epoxy-queuosine galactosyl-queuosine
Fig (23).
N
N
R1
O
R2O
N
R2O
14a: R1 = NH2, R2 = H
144: R1 = SMe, R2 = Tol
N
N
R1
O
R2O
N
R2O
N O
aq. HCHO, morpholine,
AcOH, 60°C
143: R1 = NH2, R2 = H
145: R1 = SMe, R2 = Tol
143: 43%
145: 70%
Scheme 39.
HN
N
O
O
R2O
N
R2O
12a: R1 = R2 = H
147: R1 = R2 = iBu
HN
N
O
O
R2O
N
R2O
R3
aq. HCHO, morpholine,
AcOH, 60°C
146: R1 = R2 = R3 = H, R4 =
148: R1 = R2 = iBu, R3 = H, R4 =
149: R1 = R2 = iBu, R4 = H, R3 =
146: 84%
148: 72%
149: 9%
R1HN R1HN
CH2 N O
R4
CH2 N O
CH2 N O
Scheme 40.
190 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
was employed as precursor. The reaction conditions mentioned
above using AcOH as solvent (conditions I) furnished a mixture of
the 8-substituted product 148 (72%) together with a minor amount
of the 7-modified derivative 149 (9%) (Scheme 40).
The Mannich reaction was also performed on the 2-amino-6-
methoxy nucleoside 11a under conditions II (weakly acidic) as
acetic acid when utilized as solvent (conditions I) led to decomposi-
tion of 11a as it was already observed for 12a. The reaction af-
forded only the bis-product 150 while the mono-substituted com-
pound could not be detected (Scheme 41) [192]. In order to reduce
the reactivity of compound 11a, the amino group was protected
with a tosyl residue ( 151), and the reaction was carried out as
described for 11a (conditions II). In this case, the 8-funtionalized
Mannich base 152 was isolated. From these observations it was
concluded that 7-deaza-2’-deoxyguanosine derivatives cannot be
used for the introduction of a Mannich side chain at position-7
[192].
Alternatively, the protected 6-methoxy-2-methylthio-7-
deazapurine 2’-deoxyribonucleoside 153 was used as starting mate-
rial for the Mannich reaction (Scheme 42). Compound 153 was
prepared via nucleobase anion glycosylation (TDA-1, KOH,
MeCN) employing halogenose 7 [192]. Next, the Mannich reaction
(conditions I) was carried out, and nucleoside 154 with the Mannich
side chain attached to position-7 was obtained in high yield (95% of
crude product). Conversion of the OMe group into an oxo group
was performed with trimethylsilyl chloride (TMS-Cl)/NaI in MeCN
to give 155. The free nucleoside 156 was obtained in three steps
which includes treatment of 156 with 3-chloroperbenzoic acid (m-
CPBA) in CH2Cl2 followed by NH3 in dioxane. Finally, deprotec-
tion of the sugar moiety yielded the Mannich nucleoside 156 in
69% [192].
N
N
OMe
O
HO
N
HO
11a: R = H
151: R = Tos
N
N
OMe
O
HO
N
HO
R2
aq. HCHO, morpholine,
AcOH, 60°C
150: R1 = H, R2 = R3 =
152: R1 = Tos, R2 = H, R3 =
150: 39%
152: 43%
RHN R1HN
CH2 N O
R3
CH2 N O
Scheme 41.
N
N
OMe
O
TolO
N
TolO
aq. HCHO, morpholine,
AcOH, 60°C
95%
H3CS
N
N
OMe
O
TolO
NH
TolO
H3CS
N
N
OMe
O
TolO
N
TolO
H3CS
Cl
22
7
+
TDA-1, KOH,
MeCN
153
69%
N O
154
HN
N
O
O
TolO
N
TolO
H3CS
N O
155
HN
N
O
O
HO
N
HO
H2N
N O
156
TMS-Cl, NaI,
MeCN
154
1) m-CPBA, CH2Cl2
2) sat. dioxane/NH3
3) NH3, MeOH
90% 69%
Scheme 42.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 191
4.2. 7-Deazapurine 2'-Deoxy-2'-fluoroarabinonucleosides and
2',3'-Dideoxy-3'-fluororibonucleosides
It has been shown that the sugar modification of nucleosides by
a fluorine atom can enhance biological activity as well as the stabil-
ity of the glycosylic bond [194-197]. Fluorinated 7-deazapurine
nucleosides, such as 2’-deoxy-2’-fluoroarabinosangivamycin (157c)
and 2-amino-2’-deoxy-2’-fluoroarabinotubercidin (158a), can act as
antiviral agents [198-200]. Selected 7-deazapurine fluoroarabino-
nucleosides (157-161) are shown in Fig. (24). These nucleosides
have been synthesized by a convergent route using the nucleobase
anion glycosylation. This reaction utilizes a 7-deazapurine base
which is glycosylated with 3,5-di-O-benzoyl-2-deoxy-2-fluoro- -
D-arabinofuranosyl bromide (162) [201]. The nucleobase anion is
generated with sodium hydride (sodium salt glycosylation) or
KOH/TDA-1/MeCN (solid-liquid glycosylation). As the configura-
tion of sugar bromide 162 was established to be -D, and the glyco-
sylation reaction proceeds under stereoselective control, the -D-
nucleosides are the only reaction products.
A reaction sequence is outlined in Scheme 43. The nucleobase
27a was glycosylated with the sugar bromide 162 in dry MeCN in
the presence of NaH furnishing -D-nucleoside 163 exclusively
[199]. Debenzoylation of 163 with NH3/MeOH afforded compound
164a, which on treatment with 0.5 M NaOMe/MeOH gave the 6-
methoxy nucleoside 164b. Demethylation of 164b with io-
dotrimethylsilane in MeCN furnished the guanosine analog 159a.
The thioguanosine derivative 159b was obtained via thiation of 163
with thiourea followed by debenzoylation. In a similar way, the
selenoguanine nucleoside 159c was prepared. Treatment of 163
with selenourea in absolute ethanol ( 165c), followed by deben-
zoylation with NH3/MeOH yielded the selenonucleoside 159c.
Solid-liquid conditions were also applied for the glycosylation
of 7-deazapurines with the fluoro sugar 162 (Scheme 44). The con-
densation of 2-amino-6-chloro-7-deazapurine (27a) with 162 was
performed in MeCN in the presence of TDA-1 and KOH to give the
-D-anomer 163 exclusively. The latter was deblocked yielding the
intermediate 164a. The chloro substituent of compound 164a was
HN
N
X
H2N
OHO
HO
F
N
157a: R = H
b: R = CN
c: R = CONH2
d: R = CSNH2
e: R = F
160a: R = H
b: R = F
158a: R = H
b: R = Cl
c: R = Br
d: R = I
161159a: X = O
b: X = S
c: X = Se
N
N
NH2
OHO
HO
F
N
R
N
N
NH2
OHO
HO
F
N
R
H2N
HN
N
O
OHO
HO
F
N
R
HN
N
NH2
OHO
HO
F
NO
Fig (24).
N
N NH
Cl
H2NN
N
Cl
H2N27a
162 163
OBzO
BzO
F
Br
+
OBzO
BzO
F
N
N
N
R
H2N
OHO
HO
F
N
HN
N
X
H2N
OBzO
BzO
F
N
165b: X = S
c: X = Se
58%
b: thiourea, EtOH
c: selenourea, EtOH
HN
N
X
H2N
OHO
HO
F
N
159b: X = S
c: X = Se
NH3/MeOH
b: 92%
c: 91%
b: 93%
c: 75%
a: NH3/MeOH
b: NaOMe/MeOH
a: 93%
b: 73%
163
Me3SiI,
MeCN
92%
HN
N
O
H2N
OHO
HO
F
N
164a: R = Cl
b: R = OMe
159a
NaH,
MeCN
Scheme 43.
192 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
displaced by an amino group ( 158a). Selective deamination at
the 2-amino group yielded 2'-deoxy-2'-fluoro- -D-arabino-
furanosyl-7-deazaisoguanosine (161) [202].
Similarly, glycosylation of 162 with the 7-halogenated 7-deaza-
2-pivaloylaminopurines 41b-d [70] (MeCN/KOH/TDA-1) afforded
-D-nucleosides (Scheme 45). However, this reaction resulted in a
mixture of the N7-linked nucleosides 166b-d and N
2, N
7-
bisglycosylated products 167b-d [202]. This was not observed
when 41b-d were glycosylated with 2-deoxy-3,5-di-O-(p-toluoyl)-
-D-erythro-pentofuranosyl chloride (7) (see section 3.2.3). It was
noticed that the formation of 166b-d required an extended reaction
time due to the decreased reactivity of the fluoro sugar 162. Thus,
product formation occurred under thermodynamic control leading
to two regioisomeric series of reaction products (166b-d and 167b-
d). Compounds 166b-d were converted to the 2,4-diamino deriva-
tives 158b-d (25% aq. NH3/dioxane), and compounds 167b-d were
deblocked with NaOMe/MeOH to yield the 6-methoxy nucleosides
168b-d [202].
The solid-liquid glycosylation of 6-chloro-7-deaza-7-
fluoropurine (33e) with the halogenose 162 (TDA-1/KOH/MeCN)
resulted in the formation of the -D-nucleoside 169e. The latter was
deprotected in 25% aq. ammonia with the concomitant displace-
ment of 6-chloro substituent by an amino group affording the
fluorinated tubercidin derivative 157e (Scheme 46). Compound
169e was also transformed to the 6-methoxy compound 170 using
NaOMe/MeOH. Demethylation of 170 gave the inosine analog
160b (2 N NaOH) [67-69].
In a few cases, the formation of -D-nucleosides was accompa-
nied by small amounts of -anomers. This was the result of an in-
creased reaction temperature [203] or a low nucleophilicity of the
nucleobase allowing the sugar bromide to equilibrate [198]. As the
glycosylation reaction of 33a with the sugar bromide 162 required
20 h for completion, anomerization of the halogenose took place
with the formation of a mixture of the -nucleoside 169a and its -
anomer 171 (Scheme 47), which were separated by silica gel chro-
matography [198]. Debenzoylation of 169a with methanolic am-
monia (room temperature) afforded 172, which upon amination
(NH3/MeOH at 120°C) gave the tubercidin derivative 157a. Thia-
tion of 169a with thiourea in the presence of a catalytic amount of
formic acid followed by debenzoylation afforded 7-deaza-2’-deoxy-
2’-fluoroarabino-6-thioinosine (173). Oxidation of 173 (30%
H2O2/NH3·H2O) gave 7-deaza-2’-deoxy-2’-fluoroarabinoinosine
(160a).
N
N NH
Cl
H2N
27a
162
164a
161
(i) MeCN, TDA-1, KOH
(ii) NH3/MeOH, rt, 18h
aq. NH3,
dioxane
NaNO2,
AcOH/H2O
(i) 59%
(ii) 86%
90% 69%+
158a
N
N
NH2
H2N
OHO
HO
F
N
HN
N
NH2
O
OHO
HO
F
N
Scheme 44.
N
N NH
Cl
PivHN
R
N
N N
Cl
PivHN
N
N N
Cl
N
R
R
41b-d
b: R = Cl; c: R = Br; d: R = I
MeCN, TDA-1, KOH
NaOMe,
MeOH
aq. NH3/dioxane
90°C, 24h
+
OBzO
BzO
F
OBzO
BzO
F
OBzO
BzO
F
Piv
166b-d
167b-d
158b-d
168b-d
b: 87%
c: 90%
d: 86%
b: 89%
c: 90%
d: 82%
166b: 45%; 167b: 11%
166c: 45%; 167c: 10%
166d: 44%; 167d: 12%
N
N N
NH2
H2N
R
OHO
HO
F
N
N N
OMe
H2N
R
OHO
HO
F
162
OBzO
BzO
F
Br
Scheme 45.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 193
Similarly, glycosylation of the sodium salt of 174 [106, 110]
with 162 required a longer reaction time (18 h, rt) or high tempera-
ture (80°C, 4 h) which resulted in the formation of an anomeric
mixture with the -D-anomer 175 as the main product and its -
anomer 176 as minor product (Scheme 48) [198, 203]. Treatment of
175 with NH3/MeOH at room temperature led to ring closure with
concomitant removal of the benzoyl groups affording 8-bromo-7-
cyano-7-deazapurine nucleoside 177. Selective acetylation of 177
with acetic anhydride in the presence of 4-(dimethylamino)pyridine
(DMAP) in DMF gave 3’,5’-di-O-acetyl derivative 178a, which on
reductive debromination (5% Pd/C, MgO, H2), yielded the nucleo-
side 178b. Deacetylation of 178b (Na2CO3 in aq. 1,4-dioxane) fur-
nished the fluorinated 2’-deoxytoyocamycin 157b. Oxidative hy-
drolysis of the 7-carbonitrile group of 157b (30% H2O2/aq. NH3)
yielded 2’-deoxy-2’-fluoroarabinosangivamycin (157c). The corre-
sponding thiosangivamycin 157d was obtained from 157b using
H2S in dry pyridine.
The replacement of a hydroxy group by a fluorine atom in 7-
deazapurine 2’-deoxyribonucleosides can change the N/S-
conformational equilibrium of the pentofuranose moiety. This equi-
librium is driven by various stereoelectronic gauche and anomeric
effects. The sugar pucker is described relative to the exocyclic atom
C5’, and defined as endo if the puckered atom is at the same side of
N
N NH
Cl N
N
Cl
33e
162
TDA-1, KOH,
MeCN, rt, 10 min
+
OBzO
BzO
F
N
169e
F
F
67%
N
N
NH2
OHO
HO
F
N
157e
F
N
N
OMe
OHO
HO
F
N
170
F
HN
N
O
OHO
HO
F
N
160b
F
aq. NH3
72%
NaOMe,
MeOH
97%
2N NaOH
37%
169e
Scheme 46.
N
N NH
ClN
N
Cl
33a 162
NaH, MeCN,
rt, 20hO
BzO
BzO
F
Br+
OBzO
BzO
F
N
OBzO
BzO
F
N
N
Cl
N
HN
N
S
OHO
HO
F
N
+
169a (57%) 171 (9%)
172: R = Cl (72%)
157a: R = NH2 (80%)
(i) thiourea, EtOH
(ii) NaOMe/MeOH NH3/MeOH
84%
169a
N
N
R
OHO
HO
F
N
HN
N
O
OHO
HO
F
N
aq. NH3,
30% H2O2
74%
173160a
Scheme 47.
194 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
the plane as C5’, otherwise it is exo. The north (N) conformation is
centered around the C3’-endo conformer, while the south (S) con-
formation is centered around the C2’-endo species (Fig. 25). The
conformational analysis of a series of fluorinated 7-deazapurine
nucleosides has been studied using the PSEUROT program (Table
1) [67, 202, 204].
As shown in Table 1, the presence of the fluorine atom of the
sugar moiety drives the N S equilibrium of 7-deazapurin-2,6-
diamine 2’-deoxy-2’-fluoroarabinonucleosides 158a-d towards N
(34-37% N) in comparison to the corresponding 2’-
deoxyribonucleosides 18a-d (26-28% N) [70, 202]. The same ob-
servation is made for 7-deaza-2’-deoxy-2’-fluoroisoguanosine (161)
(35% N) or 2’-deoxy-2’-fluoroadenosine (FdA) (36% N) and their
corresponding 2’-deoxyribonucleosides 36a (27% N) or dA (28%
N) [70, 202, 204]. This means that a fluorine atom in the 2’-‘up’
position increases the population of the N conformers by around
10%. Only with nucleoside 157e, the change is less pronounced
(3%) [67]. From the view point of the gauche disposition of the 2’-
N
NC
174 162
NaH, MeCN, rt, 18h
or 80°C, 4h
OBzO
BzO
F
Br+
OBzO
BzO
F
N
N
N
NH2
OHO
HO
F
N
OBzO
BzO
F
N
N
N
NH2
OAcO
AcO
F
N
+
175 (50%) 176 (16%)
178a: X = Br (77%)
b: X = H (84%)
177
EtO
CN
Br
CN
Br
CN
X
N
NC
N
EtO
CN
Br
N
N
NH2
OHO
HO
F
N
CN
157c: R = CONH2 (69%)
d: R = CSNH2 (93%)
88%
NH3,
MeOH
Ac2O,
DMAP,
DMF
Na2CO3,
aq. dioxane
96%
Br
CNNC
NEtO
175
Pd/C, H2
157c: aq. NH3,
30% H2O2
157d: H2S, Et3N,
pyridine
N
N
NH2
OHO
HO
F
N
R
157b
Scheme 48.
O
B
5'
C2'-exo-C3'-endo
O
B
C2'-endo-C3'-exo
F
OH
F
HO
4'
3'
2'
1'4'
2'
1'3'
5'
B = Base
HO
HO
N S
Fig (25). N and S conformations of sugar rings of 7-deazapurine 2’-deoxy-2’- fluoroarabinonucleosides.
Table 1. Population of Sugar Conformers of 7-Deazapurine Fluoroarabinonucleosides and 7-Deazpurine 2’-Deoxyribonucleosides
Fluoro-compd Conformation Deoxy-compd Conformation
FdA 36% N dA 28% N [204]
158a 34% N 18a 26% N
158b 36% N 18b 28% N
158c 37% N 18c 29% N
158d 36% N 18d 28% N
161 35% N 36a 27% N
160b 40% N 15e 30% N
157e 33% N 14e 30% N
N
N
NH2
H2N
R
OHO
HO
N
HN
N
NH2
OHO
HO
NO
18a-d 36a
N
N
NH2 F
OHO
HO
N
14e
HN
N
OHO
HO
N
15e
O
N
N
NH2 F
N
N
NH2
H2N
R
OHO
HO
N
OHO
HO
F
N
HN
N
NH2
OHO
HO
F
NO
F
161158a-d 157e
HN
N
OF
OHO
HO
N
F
160b
a: R = H; b: R = Cl; c: R = Br; d: R = I; e: R = F
F
N
N
N
NH2
OHO
HO
N
dA
N
N
N
NH2
OHO
HO
F
N
FdA
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 195
‘up’ fluoro substituent with the furanose oxygen, the 2’-‘up’ fluoro
favors the S conformation. However, this puts the F and the 3’-OH
in the ap orientation. Because the 2’-‘up’ fluoro and 3’-OH also
seek to have a gauche orientation, which can be best achieved in the
north conformation, there is a small pull towards N as a conse-
quence of this force.
The fluorinated nucleosides 7-deaza-2’-deoxy-7-fluoro-
adenosine (14e), 7-bromo-2'-deoxy-2,6-diamino-2'-fluoro- -D-
arabinofuranosyl-7-deazapurine (158c) and 7-deaza-7-fluoro-2'-C-
methylinosine (179) were subjected to single-crystal X-ray analysis
by our laboratory [205-207]. As it can been seen from Fig. (26),
nucleoside 179 is a close derivative of the highly active anti-HCV
compound 180 [208].
In the crystal structure of compound 179 (Fig. 27A), the glyco-
sylic bond torsion angle is in the anti range [ = 140.78 (14)°],
while that of 14e (Fig. 27B) is situated between anti and high-anti
[ = 101.1 (3)°]. The 2’-C-methylribofuranosyl moiety of 179
adopts an N sugar conformation with an unsymmetrical twist (C3'-
endo-C2'-exo, 3T2) and P = 5.9 (2)° and m = 42.1 (1)°. On the other
hand, the 2’-deoxyribonucleoside 14e exhibits an S-type sugar con-
formation (2T3) with P = 164.7 (3)° and m = 40.1 (2)°, which is
consistent with the predominat S conformation (70%) observed in
solution (Table 1). In the crystal structure of 158c, two conforma-
tions of the exocyclic C4’-C5’ bond were found, corresponding to
conformer 1 (0.69 occupancy) (Fig. 27C) and conformer 2 (0.31
occupancy) (Fig. 27D). However, both conformers show the same
N
N
NH2
OHO
HO
N
F
N
N
NH2
OHO
HO
F
N
Br
H2N
HN
N
O
OHO
HO
Me
N
F
OH
N
N
NH2
OHO
HO
Me
N
F
OH
179 18014e 158c
Fig (26).
(A) (B)
(C) (D)
Fig (27). Perspective views of (A) 7-deaza-7-fluoro-2'-C-methylinosine (179) [207], (B) 7-deaza-2’-deoxy-7-fluoroadenosine (14e) [205], (C) conformer 1 of
7-bromo-2'-deoxy-2,6-diamino-2'-fluoro- -D-arabinofuranosyl-7-deazapurine (158c) and (D) conformer 2 of 158c [206]. Displacement ellipsoids are drawn at
the 50% probability level and H atoms are shown as spheres of arbitrary size.
196 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
anti glycosylic bond torsion angle [ = 114.8 (4)°] and N-type sugar
conformation (3T4) with P = 23.3 (4)° and m = 36.5 (2)°. In this
case, the N-type sugar conformation found for the crystalline state
of 158c is not consistent with the predominant sugar pucker of 158c
observed in solution (63% S, Table 1).
2’,3’-Dideoxy-3’-fluorothymidine (FLT) shows antiviral activ-
ity against HIV; it is even a more potent inhibitor of HIV replica-
tion than AZT. Unfortunately, FLT and related nucleosides were
found to be highly cytotoxic. This prompted us to synthesize 7-
deazapurine 2’,3’-dideoxy-3’-fluoronucleosides such as 181a [209]
and 182a [210] (Scheme 49). The syntheses started from the 2’-
deoxyribonucleosides 183 or 29a. The 5’-hydroxyl groups were
protected with a (tert-butyl)diphenylsilyl residue affording 184a or
184b [211], which were then oxidized with CrO3 yielding 3’-oxo
nucleosides 185a or 185b. The reduction of 185a or 185b with
NaBH4 in EtOH resulted in the formation of compounds 186a or
186b having the 3'-hydroxyl group in the arabino configuration.
After protection of the 2-amino group of 186b with a mono-
methoxytrityl (MMT) residue ( 186c), the fluorine substituent
was introduced using (diethylamino)sulfur trifluoride (DAST) with
inversion of configuration ( 187a or 187b). Desilylation of com-
pound 187a with TBAF ( 188a) followed by amination with aq.
NH3 afforded 2’,3’-dideoxy-3’-fluorotubercidin 181a. In the other
route, the trityl residue of 187b was removed with 80% AcOH to
give 187c, which was treated with TBAF ( 188b), followed by 2
M NaOH thereby yielding 7-deaza-2’,3’-dideoxy-3’-
fluoroguanosine 182a. The triphosphates 181b or 182b were pre-
pared from 181a or 182a with POCl3 ( monophosphate) followed
by condensation with tetrabutylammonium diphosphate according
to the one-pot protocol of Ludwig (see also Scheme 59, section
4.3.4) [212]. Both triphosphates (181b, 182b) were found to be
inhibitors of HIV-1 reverse transcriptase (see Table 2, section
4.3.4).
O
N
N N
HO
Cl
183: R = H
29a: R = NH2
R
HO
O
N
N N
BuPh2SiO
Cl
184a: R = H
b: R = NH2
R
HO
tO
N
N N
BuPh2SiO
Cl
185a: R = H
b: R = NH2
R
t
O
O
N
N N
BuPh2SiO
Cl
186a: R = H
b: R = NH2
R
tOH
O
N
N N
BuPh2SiO
Cl
187a: R = H
b: R = NHMMT (29%)
R
t
F
tBuPh2SiCl
imidazole, DMF
CrO3, pyridine
Ac2O, CH2Cl2
186c: R = NHMMT (94%)
MMT-Cl, pyridine
NaBH4, EtOH
80% AcOH
187c: R = NH2 (73%)
O
N
N N
HO
Cl
R
F
188a: R = H
b: R = NH2
O
N
N N
RO
NH2
F
DAST,
CH2Cl2 or toluene
TBAF, THF
181a,b 182a,b
or
181a: aq. NH3/dioxane
182a: 2M NaOH
O
HN
N N
RO
F
H2N
O
a: 76%
b: 68%
a: 57%
b: 35%
a: 76%
b: 92%
P O
O
OH
P O P OH
O O
OH OH
181a: 76%
182a: 36%
b: R =a: R = H
Scheme 49.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 197
4.3. 7-Deazapurine 2’,3’-Dideoxyribonucleosides
2’,3’-Dideoxyribonucleoside triphosphates can act as termina-
tors of DNA-polymerase as well as of reverse transcriptase [213-
216]. Therefore, several purine and pyrimidine 2’,3’-
dideoxyribonucleosides show antiviral activity against RNA and
DNA viruses such as human immunodeficiency virus (HIV) [217-
219] and the Hepatitis B virus (HBV). Their triphosphates are also
used in the Sanger dideoxy sequencing of DNA [21, 220] which is a
standard tool of modern molecular biology. This method has been
modified in several ways. The replacement of dGTP by 7-deaza-
dGTP has been shown to resolve band compression during electro-
phoretic separation [19]. 7-Deazapurine 2’,3’-dideoxyribo-
nucleoside triphosphates bearing a fluorescent dye at position-7
serve as terminators in most currently available automated DNA
sequencing machines [221].
4.3.1. 7-Deazapurine 2’,3’-Dideoxyribonucleosides Used in DNA-
Sequencing
In 1987, Prober and coworkers reported on an improved Sanger
protocol which makes use of modified dideoxyribonucleoside
triphosphates bearing fluorescent dyes in the side chains [21]. The
fluorescent chain terminators can carry different labels for the four
different triphosphates used in the fluorescence-based automated
DNA sequencers [21, 222-225]. Succinylfluorescein has been at-
tached via an aminopropargyl linker to the 5-position of pyrimidi-
nes and to the 7-position of 7-deazapurines. The syntheses of the
triphosphates 189 and 190 are outlined in Schemes 50 and 51. The
protected 2’,3’-dideoxyribonucleoside 191 or 198 served as starting
material [22]. Iodination of 191 and 198 was performed with N-
iodosuccinimide (NIS) in DMF or ICl in CH2Cl2 affording 192 or
199. Then, the methoxy group of 192 was cleaved with sodium
(i) 3 POCl3, H2O, (MeO)3PO
(ii) H2P2O72-(Bu3NH+)2, NBu3
(iii) Et3NHHCO3, H2O
O
HN
N N
O
197
O
POPOPHO
O O O
OOO
C CCH2NH2
Pd(0)(PPh3)4, CuI, Et3N, DMF
CCH2NHCOCF3,HC
N
O CH3
O
O OAcO
O
H2N
O
HN
N N
O
O N
N
O CH3
O
O OO
POPOPHO
O O O
OOO
4 Et3NH+
189
O
HN
N N
HO
CO
H2N
CCH2NHCOCF3
O
N
N N
TrO
OMe
O
N
N N
TrO
OMe
O
HN
N N
TrO
O
O
HN
N N
TrO
O
O
HN
N N
HO
O
NIS, DMF
191 192 193
194 195b
MeS MeS
I
Na thiocresolate,
HMPA, toluene
MeS
1) m-CPBA, CH2Cl2
2) NH3, dioxane
96%
I
97%
70%
AcOH
81%
II
H2N H2N
H2N
196
3 Et3NH+
N
O
O
H
Scheme 50.
198 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
thiocresolate ( 193), which was followed by oxidative conversion
of the methylthio group into an amino group to yield 194. The pro-
tected 194 as well as 199 were deblocked affording the 7-iodinated
dideoxyribonucleosides 195b or 200b as central intermediates [22].
Sonogashira cross coupling (CuI, CPd(0)(PPh3)4, Et3N, DMF) of
the 7-iodo nucleoside 195b with N-trifluoroacetylpropargylamine
gave 196 [226]. The latter was converted into its 5’-triphosphate
and deacylated to afford 7-(3-amino-1-propynyl)-7-deaza-2’,3’-
dideoxyguanosine 197. The amino side chain was reacted with O-
acetyl protected succinylfluorescein and deprotected to give 189. In
a similar way, 7-deaza-2’,3’-dideoxy-7-iodoadenosine (200b) was
transformed into the corresponding fluorescence-labeled Sanger
sequencing reagent 190 (Scheme 51).
Sequencing by synthesis (SBS) – as the Sanger sequencing – is
based on the polymerase chain reaction [129-133]. In SBS, a primer
is extended on a template by a single nucleotide, and the identity is
determined by the use of fluorescent reporter groups. Four nucleo-
tides are designed with dyes attached to the 5-position of
pyrimidine nucleotides or the 7-position of 7-deazapurine nucleo-
tides. The dyes are linked to the base via photocleavable linkers and
are cleaved by light (355 nm). Various photo-cleavable linkers have
been developed. The part of the linker which stays on the growing
oligonucleotide chain should be as small as possible. A small cap-
ping group is employed for the 3’-OH position. After nucleotide
identification, the 3’-OH protecting groups are removed chemically.
Allyl groups can be used for that purpose and are stepwise removed
under palladium assistance. As purine nucleotide surrogates, 7-
deazapurine nucleotides are used. 7-Deaza-2’-deoxyadenosine and
7-deaza-2’-deoxyguanosine conjugates 202 and 205 bearing fluo-
rescent dyes and their photochemical cleavage is outlined in
Scheme 52.
7-Deazapurine 2’,3’-dideoxyribonucleosides were also used in
recently developed DNA sequencing approaches [227], such as
MALDI-TOF mass spectrometry DNA sequencing [228]. This
approach makes use of biotinylated 7-deazapurine 2’,3’-
dideoxyribonucleosides 206 and 207 to generate Sanger sequencing
termination products (Fig. 28). The biotin-streptavidin interaction is
used to capture the biotinylated DNA fragments on a streptavidin-
coated solid support followed by direct detection with matrix-
O
N
N N
O
201
NH2
POPOPHO
O O O
OOO
C CCH2NH2
N
O CH3
O
O O
CH3
AcO
H3C
O
O
N
N N
NH2 N
N
O CH3
O
O OO
OPOPOPHO
O O O
OOO
4 Et3NH+
190
O
N
N N
PivO
Cl
O
N
N N
PivO
Cl
O
N
N N
HO
NH2
ICl, Na2CO3, CH2Cl2
198 199
200b
I
NH3, MeOH
70%
I
70%
3 Et3NH+
H3C CH3
N
O
O
H
Scheme 51.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 199
O
HN
N N
O NH
OPOPOPO
O O O
OOO
202
HN
O
O2N
O
O
H2N
N
B
N
F F
CH3
CH3
O
O
N
N N
NH2 NH
O
OPOPOPO
O O O
OOO
205
NN
COO
HN
O
O2N
O
O
O
O
HN
N N
O NH2
OPOPOPO
O O O
OOO
203
H2N
O
HN
O
O2N
CH3
N
B
N
F F
CH3
CH3
+ CO2
+
irradiation at 355 nm,
10 sec
O
O
HN
N N
O NH2
OPOPOPO
O O O
OOO
204
H2N
HO
Pd-deallylation,
30 sec
Scheme 52.
200 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
O
N
N N
O
NH2
POPOPHO
O O O
OOO
N
NH
O
OH
S
NHHN
HH
O
206
O
HN
N N
O
O
POPOPHO
O O O
OOO
N
NH
O
OH
S
NHHN
HH
O
207
H2N
Fig (28).
O
HO
N
N N
HO
Cl
O
HO
N
N N
DMTO
Cl
O
PhOCO
N
N N
DMTO
Cl
O
N
N N
DMTO
Cl
O
N
N N
HO
Cl
200a: R = NH2 (65%)
213: R = OMe (78%)
DMT-Cl,
pyridine
ClCOPh,
S
MeCN
n-Bu3SnH,
AIBN
80% AcOH
200a: aq. NH3
213: NaOMe/MeOH
208a: 2N NaOH
183 209 210
211 212a
78% 76% 75%
67%
O
HN
N N
HO
O
or
O
N
N N
HO
R
208a (80%)
S
Scheme 53.
assisted laser desorption ionization time-of-flight mass spectrome-
try (MALDI-TOF MS).
4.3.2. Synthesis of 7-Deazapurine 2’,3’-Dideoxyribonucleosides
7-Deazapurine 2’,3’-dideoxyribonucleosides [61, 211, 229-231]
have been prepared from the corresponding 2’- or 3’-
deoxyribonucleosides as described for the formation of 2’-
deoxyribonucleosides from ribonucleosides (see section 3.3), as
well as by elimination of a 3’-mesyloxy group yielding unsaturated
nucleosides [95], which were subjected to catalytic hydrogenation
[119-122].
2’,3’-Dideoxytubercidin (200a) and 7-deaza-2’,3’-
dideoxyinosine (208a) were prepared by Barton-McCombie deoxy-
genation of the corresponding 2’-deoxyribonucleosides (Scheme
53) [61]. The 5’-hydroxyl group of compound 183 was selectively
protected with a DMT residue (DMT-Cl) to give 209. The 3’-
hydroxyl group of compound 209 was derivatized with phe-
noxythiocarbonyl chloride to give 210 (see also section 3.3).
Treatment with tri-n-butylstannane in toluene in the presence of
AIBN afforded 2’,3’-dideoxyribonucleoside 211. Detritylation of
211 gave the nucleoside 212a, which was further converted to
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 201
2’,3’-dideoxytubercidin (200a), the 6-methoxy derivative 213, and
7-deaza-2’,3’-dideoxyinosine (208a).
The 7-deazapurine 2’,3’-dideoxyribonucleoside 200a was also
synthesized by deoxygenation of 3’-deoxyribonucleosides (Scheme
54) [230]. 7-Deazacordycepin (214) was employed as starting mate-
rial and was prepared as described earlier [119, 232]. Compound
214 was converted to 215 with monomethoxytrityl chloride (MMT-
Cl) in pyridine using silver nitrate as a catalyst. The phenoxythio-
carbonyl derivative 216 was prepared next. Barton-McCombie de-
oxygenation of 216 with tri-n-butylstannane in benzene in the pres-
ence of AIBN afforded the 2’,3’-dideoxyribonucleoside 217 which
was deprotected to give 2’,3’-dideoxytubercidin (200a).
The protocols discussed above require sufficient amounts of the
corresponding 7-deazapurine ribo-, 2’- or 3’-deoxyribonucleosides.
This problem was circumvented by the nucleobase anion glycosyla-
tion of various 7-deazapurines employing 2,3-dideoxy-5-O-[(1,1-
dimethylethyl)dimethylsilyl]-D-glycero-pentofuranosyl chloride
(218a) or 2,3-dideoxy-5-O-[(1,1-dimethylethyl)diphenylsilyl]-D-
glycero-pentofuranosyl chloride (218b) [233-236] as activated
sugar components. Compounds 218a,b were prepared in situ from
the corresponding lactol [233] by Appel chlorination [237, 238].
The nucleobase anions of compounds 33a or 27a were gener-
ated under solid-liquid phase transfer conditions (KOH/TDA-
1/MeCN) and were reacted with the anomeric mixture of the sugar
halide 218a resulting in the -anomers 219a,b and the correspond-
ing -nucleosides 220a,b [236] (Scheme 55). Deprotection of
219a,b or 220a,b was performed with TBAF in THF to afford the
free nucleosides 212a, 221a or 222a,b in a total yield of 67-85%
[235, 236]. Further conversion of 212a or 221a provided 7-
deazapurine 2’,3’-dideoxyribonucleosides 200a, 208a, 223a, 224a
and 195a [61, 211, 231] (Scheme 55).
4.3.3. Synthesis of 7-Deazapurine 2’,3’-Didehydro-2’,3’-Dide-
oxyribonucleosides
Since 2’,3’-unsaturated nucleosides, such as d4T and d4C show
antiviral activity [239, 240], the synthesis of modified purine 2’,3’-
didehydro-2’,3’-dideoxyribonucleosides and their corresponding
triphosphates became of interest [95, 121, 210]. In this context, the
synthesis of the 2’,3’-didehydro-2’,3’-dideoxy analogs of tubercidin
(227a), toyocamycin (227f) and sangivamycin (227j) was carried
out starting from their parent ribonucleosides 89a,f,j (Scheme 56)
[121]. Treatment of 89a,f,j with 2-acetoxy-2-methylpropionyl bro-
mide led to a mixture of 2’-O-acetyl-3’-bromo-3’-deoxy sugar
modified nucleosides and corresponding 5’-trimethyldioxolanone
derivatives in the case of 89f and 89j. The addition of methanol
resulted in a complete removal of the 5’-trimethyldioxolanone moi-
ety to yield the 5’-hydroxyl derivatives. Subsequent acetylation
gave the 2’,5’-diacetylated compounds 225a,f,j. Treatment of
225a,f,j with a zinc-copper couple in DMF furnished the 5’-
acetylated 2’,3’-unsatured nucleosides 226a,f,j. Compounds
226a,f,j were deprotected (NH3/MeOH) to give the target nucleo-
sides 2’,3’-dideoxy-2’,3’-didehydrotubercidin (227a), 2’,3’-
dideoxy-2’,3’-didehydrotoyocamycin (227f), and 2’,3’-dideoxy-
2’,3’-didehydrosangivamycin (227j).
The 7-deazapurine nucleosides 233a and 234a, related to 2-
amino-7-deazaadenosine and 7-deazaguanosine, were synthesized
as shown in Scheme 57. 2-Amino-6-chloro-7-deazapurine 2’-
deoxyribonucleoside 29a was utilized as starting material. The 5’-
hydroxyl group of 29a was protected with a (t-Bu)Ph2Si residue to
afford compound 184b. Protection of 184b with dimethylforma-
mide diethyl acetal gave the labile amidine 228. Compound 228
was subjected to hydrolysis (MeOH/H2O) affording the formyl
derivative 229 (75% yield). Treatment of 229 with methanesulfonyl
chloride gave 230. Elimination of the mesyl group and deprotection
of 230 occurred simultaneously with TBAF/THF to give the the
2’,3’-didehydro-2’,3’-dideoxyribonucleoside 231 in 60% yield.
Then, the formyl group was removed (aq. NH3, dioxane) to give
232a. Further displacement of the chloro substituent of 232a (2 M
NaOH) afforded 234a, and nucleophilic displacement with 25% aq.
ammonia yielded 233a.
The isoinosine analog 238 was prepared according to Scheme
58 using the 2-methoxy nucleoside 61 as starting material [95]. The
synthetic route was performed as described above yielding the in-
termediate 237. Treatment of 237 with 2 N NaOH furnished 2',3'-
dideoxy-2'-3'-didehydro isoinosine derivative 238 (81% yield).
O
N
N N
HO
NH2
O
N
N N
MMTO
NH2
O
N
N N
MMTO
NH2
O
N
N N
MMTO
NH2
O
N
N N
HO
NH2
MMT-Cl, AgNO3,
THF/pyridine
ClCOPh,
S
MeCN/DMAP
Bu3SnH,
AIBN, benzene 80% AcOH
214 215 216
217 200a
72% 75%
90% 88%
OH OCOPh
S
OH
Scheme 54.
202 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
O
N
N N
SiO
Cl
33a: R = H
27a: R = NH2
218a
N
N NH
Cl
+
219a: R = H (46%)
b: R = NH2 (22%)
N
NN
Cl
O
N
N N
HO
Cl
212a: R = H (67%)
221a: R = NH2 (85%)
OHO
N
NN
Cl
O
SiOO
SiOCl
222a: R = H (82%)
b: R = NH2 (79%)
TBAF/THF
TBAF/THF
KOH,TDA-1
MeCN
R
R
R
R
R
220a: R = H (18%)
b: R = NH2 (20%)
O
N
N N
HO
NH2
O
N
N N
HO
NH2
H2N
O
HN
N N
HO
O
H2N
O
HN
N N
HO
O
O
HN
N N
HO
S
200a 224a 195a208a 223a
( / )
Scheme 55.
O
HO
N
N N
HO
NH2
OH
O
N
N N
AcO
NH2
OAc
AcO
Br
O
MeCN
(i)
(ii) MeOH for 89f,j
(iii) Ac2O/pyridine
Br
O
N
N N
AcO
NH2
Zn-Cu,
DMF
89a,f,j 225a,f,j
O
N
N N
HO
NH2
226a,f,j 227a,f,j
NH3/MeOH
a: R = H; f: R = CN; j: R = CONH2
R R R R
Scheme 56.
4.3.4. 7-Deazapurine 2’,3’-Dideoxyribonucleosides as Inhibitors
of DNA Polymerases
2’,3’-Dideoxyribonucleoside triphosphates act as terminators of
various DNA-polymerases and of viral reverse transcriptase [213-
216]. Therefore, several purine and pyrimidine 2’,3’-
dideoxyribonucleoside show antiviral activity by inhibiting the
growth of the human immunodeficiency virus (HIV) or the Hepati-
tis B virus (HBV) [217-219, 241]. Nucleobase as well as sugar
modified purine and pyrimidine 2’,3’-dideoxyribonucleosides have
been synthesized to improve the toxicology profile. Thus, a number
of 7-deazapurine 2’,3’-dideoxyribonucleotides and their 2’,3’-
didehydro or 2’,3’-dideoxy-3’-fluoro triphosphate analogs have
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 203
O
N
N N
HO
Cl
H2N
HO
O
N
N N
BuPh2SiO
Cl
H2N
HO
t
tBuPh2SiCl,
imidazole, DMF
O
N
N N
BuPh2SiO
Cl
(H3C)2NCH=N
HO
t
(H3C)2NCH(OC2H5)2,
DMF
MeOH,
H2O
O
N
N N
BuPh2SiO
Cl
HCHN
HO
t
O
O
N
N N
BuPh2SiO
Cl
HCHN
MsO
O
O
N
N N
HO
Cl
R-HN
CH3SO2Cl,
pyridine
t
(i) TBAF, THF
(ii) 25% aq. NH3,
dioxane
O
HN
N N
HO
O
H2N
29a 184b 228
75%
229 230
93%
O
N
N N
HO
NH2
H2N 2M NaOH
55%
25% aq. NH3,
dioxane
40%
231: R = HCO
232a: R = H
234a233a
231: 60%
232a: 55%
232a
Scheme 57.
O
HO
N
N N
HO
MeO
O
HO
N
N N
BuPh2SiO
MeO
O
MsO
N
N N
BuPh2SiO
MeO
O
N
N N
HO
MeO
O
HN
N N
HO
O
BuPh2SiCl,
pyridine MsCl, pyridine
TBAF/THF 2N NaOH
61 235 236
237 238
89% 82%
96% 81%
t
t t
Scheme 58.
been prepared (Fig. 29), and their activity data against HIV-1 re-
verse transcriptase are given in Table 2.
Fig. (29) displays a series of nucleoside triphosphates (181b,
182b, 200b, 208b, 212b, 223b, 224b, 195b, 234b, 233b, 239, or
232b) which were prepared from the corresponding nucleosides
181a, 182a, 200a, 208a, 212a, 223a, 224a, 195a, 234a, 233a, 227a,
or 232a in an one-pot reaction following a protocol originally de-
veloped for the phosphorylation of purine and pyrimidine 2’-
deoxyribonucleosides by Ludwig (Scheme 59) [212, 235]. The
nucleosides were dissolved in PO(MeO)3 (0°C). Two equivalents of
POCl3 were added resulting in the formation of an activated dichlo-
rophosphate which was directly condensed with bis-
tetrabutylammonium pyrophosphate. Purification on a DEAE-
Sephadex column with aq. triethylammonium bicarbonate yielded
204 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
the triphosphates in the form of their triethylammonium salts (Fig.
29).
The 7-deazapurine triphosphates shown in Table 2 were found
to be strong inhibitors of HIV-1 reverse transcriptase. 7-
Deazaguanine, 7-deazaadenine, and 2-amino-7-deazaadenine 2’,3’-
dideoxyribonucleoside triphosphates (195b, 200b, and 224b), their
2’,3’-didehydro analogs (234b, 239, and 233b) and 3’-fluorinated
derivatives (181b and 182b) show similar inhibitory activity against
HIV-1 reverse transcriptase as the corresponding purine 2’,3’-
dideoxyribonucleotides. In the case of compounds 212b, 7-deaza-
2’,3’-dideoxyinosine (208b), and its thio analog 223b less activity
was observed.
O
N
N N
HO
(i) POCl3, PO(MeO)3
(ii) H2P2O72-(Bu3NH+)2
200a
NH2
O
N
N N
O
200b
NH2
POPOPHO
O O O
OHOHOH
30-50%
Scheme 59.
O
N
N N
RO
NH2
O
N
N N
RO
NH2
H2N
O
HN
N N
RO
O
H2N
O
HN
N N
RO
O
O
HN
N N
RO
S
200a,b
224a,b 195a,b
208a,b 223a,b
a: R = H; 239, or b: R = P O
O
OH
P O P OH
O O
OH OH
O
N
N N
RO
NH2
227a or 239
O
N
N N
RO
NH2
H2N
233a,b
O
HN
N N
RO
O
H2N
234a,b
O
N
N N
HO
Cl
232a,b
H2N
O
N
N N
RO
Cl
212a,b
O
N
N N
RO
NH2
181a,b
O
HN
N N
RO
O
H2N
182a,b
F F
Fig (29).
Table 2. Inhibition of HIV-1 Reverse Transcriptase (RT) by Pyrrolo[2,3-d]pyrimidine and Purine 2’,3’-Dideoxyribonucleoside Triphosphates a [210,
235]
IC50 [μM] IC50 [μM] IC50 [μM]
212b 102.0 195b 0.12 239 0.53
200b 0.39 181b 0.43 AZTTP b 0.5
208b 20.0 182b 0.27 ddATP b 0.45
223b 1680.0 234b 0.09 ddGTP b 0.2
224b 0.5 233b 0.39
a The RT inhibitory tests were performed in the laboratories of Boehringer Mannheim GmbH.
b AZTTP = 3’-azido-3’-deoxythymidine 5’-triphosphate; ddATP = 2’,3’-dideoxyadenosine 5’-triphosphate; ddGTP = 2’,3’-dideoxyguanosine 5’-triphosphate.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 205
The inhibitory experiments indicate that a number of 7-
deazapurine 2',3'-dideoxyribonucleoside triphosphates display
strong inhibitory activity against HIV-1 reverse transcriptase in
enzymatic inhibitor assays. However, none of them was a powerful
inhibitor of the HIV virus growth in cellular assays. Probably, 7-
deazapurine 2’,3’-dideoxyribonucleosides withstand phosphoryla-
tion by cellular kinases but immediately form the monophosphates
acting as kinase inhibitors as it was reported for 7-deazapurine ri-
bonucleosides [242, 243]. Pronucleotide derivatives of monophos-
phates might overcome this activity barrier.
5. SYNTHESIS OF 7-DEAZAPURINE -L 2'-DEOXYRIBO-
NUCLEOSIDES
L-Nucleosides, the enantiomers of the naturally occurring D-
compounds are not recognized by mammalian enzymes but are
accepted by some bacteria- or virus-encoded enzymes. This results
in minimal host toxicity. The first L-nucleoside ( -L-dT) was syn-
thesized by mejkal and orm in 1964 [42]; in the same year Ac-
ton, Ryan, and Goodman described -L-adenosine [244]. Related
ribonucleosides were reported by Shimizu in 1967 [245] and by
Hol and orm in 1969 [246]. The discovery of the antiviral activ-
ity of 3TC, which is more active and less toxic than its D-
counterpart, attracted considerable interest in the synthesis and the
pharmacological activity of L-nucleosides [247-253]. Their antivi-
ral activity is comparable and even greater than that of their D-
counterparts. Better toxicological profiles and a greater metabolic
stability were observed in several cases. L-Nucleosides and their
analogs have become effective drugs for the treatment of viral dis-
eases. A number of them, such as lamivudine (3TC) and FTC are
commercialized; others like L-dT and L-FMAU are expected to get
approval by the FDA [248-250]. In the following, the synthesis of a
number of 7-deazapurine -L-2’-deoxyribonucleosides is described.
Considering the identical chemical properties of L- and D-
nucleosides and their precursors in a non-chiral environment, the
protocols developed for the 7-deazapurine D-nucleoside synthesis
can be employed for the preparation of the corresponding L-
enantiomers. Instead of the corresponding halogenose 7, 2-deoxy-
3,5-di-O-(p-toluoyl)- -L-erythro-pentofuranosyl chloride (8) was
used as sugar component. The L-sugar 8 was prepared according to
the procedure described for the corresponding D-enantiomer 7 [41,
254]. Solid-liquid nucleobase anion glycosylation [33, 34, 38] of 7-
deazapurines 33a,d with the halogenose 8 (TDA-1/KOH/MeCN)
afforded the toluoyl-protected -L-nucleosides 240a,d stereoselec-
tively in 68-89% yield (Scheme 60). Deprotection of compound
240d under concomitant nucleophilic displacement of the chloro
substituent (aq. NH3/dioxane, 80°C) gave -L-2’-deoxytubercidin
(242a). Compound 240d was treated with methanolic ammonia to
yield the 6-chloro intermediate 241d, which was later converted to
-L-2’-deoxy-7-iodotubercidin (242d) in aq. NH3/dioxane (120°C)
[255].
Also, a number of 6,7-disubstituted 2-amino-7-deazapurine -
L-2’-deoxyribonucleosides were prepared. The glycosylation of the
2-amino-6-chloro-7-deazapurines 27a,b [53, 256] with L-
halogenose 8 performed under solid-liquid conditions gave the -L-
nucleosides 243a (85%) or 243b (66%) exclusively (Scheme 61).
The intermediates 243a,b were deprotected (methanolic ammonia)
affording nucleosides 244a,b.
The protected nucleosides 243a,b were also employed in vari-
ous nucleophilic displacement reactions [255] (Scheme 61). Re-
moval of the toluoyl protecting groups and displacement of the 6-
chloro substituent occurred upon treatment of 243a,b with 25% aq.
NH3 in a steel vessel furnishing the 2,6-diamino nucleosides
245a,b. Selective deamination of compounds 245a,b with sodium
nitrite in AcOH/H2O (V/V, 1:5) gave the 7-deazaisoguanine deriva-
tives 246a,b (60-70% yield). Compounds 243a,b were also con-
verted to the 6-methoxy nucleosides 247a,b with 0.5 M
NaOMe/MeOH (reflux conditions). The 7-deaza- -L-2’-
deoxyguanosines 248a,b were obtained from 247a,b by nucleo-
philic methoxy/hydroxy group displacement in refluxing 2 N
NaOH. The 6-thio compound 249 was prepared from 244a using
thiourea (Scheme 62).
Structural and conformational parameters of a few L-
nucleosides were obtained in the solid state from single-crystal X-
ray analyses. The X-ray structure of the L-enantiomer 2’-deoxy-7-
iodotubercidin (242d) is shown in Fig. (30) [255]. The conforma-
N
N NH
Cl
MeCN, KOH, TDA-1
N
N
Cl
NO
+ OTol
OTol
Cl
O
OTol
OTol
a: 68%
d: 89%
R
R
8
a: R = H; d: R = I
33a,d
240a: R = H
d: R = I
N
N
NH2
N
O
OH
OH
aq. NH3,
dioxane
NH3,
MeOH
87%
242a,d241d
R
N
N
Cl
N
O
OH
OH
I
240d 240a
aq. NH3,
dioxane
81%40%
Scheme 60.
206 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
tion around the glycosyl bond was found to be anti with the torsion
angle (O4’-C1’-N9-C4) of 147.1°, and the sugar moiety adopts
the S-conformation (3E; P = 197° and m = 32.7°).
The -L-configuration was deduced from the CD spectra meas-
ured for both, the 7-deazapurine L-nucleosides and the correspond-
ing D-enantiomers. Typical spectra are shown in Fig. (31a,b). By
comparing the CD spectra of the -L- and -D-nucleosides, mirror
images confirmed the enantiomeric character. The -L-nucleosides
show a positive lobe while a negative lobe is formed by the -D-
enantiomers.
6. SYNTHESIS OF PYRROLO[2,3-d]PYRIMIDINES WITH
UNUSUAL GLYCOSYLATION SITES
In the early days of 7-deazapurine 2’-deoxyribonucleoside syn-
thesis, the major research focus was directed towards the develop-
N
N NH
Cl
H2N
N
N
Cl
H2N N
O
+OTol
OTol
Cl
O
OTol
OTol
N
N
Cl
H2N N
O
OH
OH
R
R
R
8
a: R = H; b: R = Cl
27a,b
243a,b
244a,b
N
N
NH2
H2N N
O
OH
OH
245a,b 247a,b
R
246a,b
N
N
OMe
H2N N
O
OH
OH
R
HN
N
NH2
O N
O
OH
OH
R
248a,b
HN
N
O
H2N N
O
OH
OH
R
aq. NH3,
dioxane
NaOMe,
MeOH
AcOH/H2O,
NaNO2, rt2N NaOH,
reflux
TDA-1, KOH,
MeCN
NH3/MeOH, rt
a: 85%
b: 66%
a: 88%
b: 90%
a: 71%
b: 91%
a: 90%
b: 80%
a: 81%
b: 87%
a: 70%
b: 60%
Scheme 61.
HN
N
S
H2N N
O
OH
OH
N
N
Cl
H2N N
O
OH
OH
(NH2)2CS, EtOH,
reflux
244a 249
85%
Scheme 62.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 207
ment of a convenient glycosylation protocol allowing glycosylation
at the desired position-9 (pyrrole nitrogen) of the 7-deazapurine
moiety (see also section 2). However, 7-deazapurine 2’-deoxyribo-
nucleosides with unusual glycosylation sites, e.g. with nitrogen-1,
carbon-8 or carbon-7 as glycosylation position, show interesting
properties.
6.1. 7-Deazapurine Nucleosides with Carbon-8 as Glycosylation
Position
Because of the weak nucleophilicity of the pyrrole nitrogen of
7-deazapurines, nucleosides with unusual glycosylation positions
are readily formed (see also section 2). Even C-nucleosides can be
generated selectively in the presence of a protected sugar moiety
with Friedel-Crafts catalysts such as SnCl4 [257]. This protocol,
which is commonly named Vorbrüggen glycosylation, has been
applied to the synthesis of 7-deazaguanine ribonucleosides with
carbon-8 or carbon-7 as glycosylation position. Corresponding 2’-
deoxyribonucleosides were synthesized by Barton deoxygenation
(see also section 3.3).
Glycosylation of the unprotected 7-deazaguanine base (250a)
with 1-O-acetyl-2,3,5-tri-O-benzoyl-D-ribofuranose (251) in ni-
tromethane in the presence of SnCl4 afforded the benzoyl protected
C8-ribonucleoside 252 (Scheme 63) [257]. Compound 252 was
deprotected with saturated methanolic ammonia to give the C8-
glycosylated 7-deazaguanine ribonucleoside 253 (78% yield).
The same authors evaluated the reactivity of 8-methyl-7-
deazaguanine (250b) with the protected sugar 251 under the condi-
tions mentioned above (nitromethane, SnCl4). In this case position-
8 is blocked by the methyl group. Now, electrophilic C-
glycosylation occurred at position-7 ( 254); however the yield
was lower (47%) compared to glycosylation at position-8 (56%)
Fig (30). Perspective view showing the displacement ellipsoids obtained
from the single-crystal X-ray analyses of compound 242d [255].
Fig (31). CD spectra of L-7-deaza-2’-deoxyguanosines (248a,b), L-2’-deoxytubercidin (242a) and their corresponding D-counterparts (12a,b and 14a). Spec-
tra were measured in MeOH with nucleoside concentrations of 4.1 10-5
(a) and 9.5 10-5
mol/L (b) [255].
O
BzO
BzO
252
HN
NH2N N
O
H
O
BzO
BzO
OBz
OAc
NH
N
H2N
N
O
OBz
nitromethane, SnCl4,
60°C, 3h
O
HO
HO
253
NH
N
H2N
N
O
OH
H HNH3/MeOH
56%78%
250a
251
Scheme 63.
208 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
employing 7-deazaguanine (250a) as nucleobase. Deprotection
(NH3/MeOH) of the protected intermediate 254 afforded the free
C7-ribonucleoside 255 in 48% yield (Scheme 64). The authors con-
cluded that electrophilic C-glycosylation in nitromethan using
SnCl4 as catalyst occurs preferentially at position-8, unless that
position is blocked. In that case, glycosylation proceeds at position-
7 [257].
The synthesis of the corresponding isobutyrylated 2’-
deoxyribonucleoside 260 employing ribonucleoside 253 as starting
material was achieved by Barton deoxygenation [117, 258]. First,
the amino group of 253 was protected with an isobutyryl residue
employing the protocol of transient protection ( 256; 72% yield)
(Scheme 65) [259]. Compound 256 was treated with Markiewicz’s
reagent [118] to give the silyl derivative 257 (57% yield). Reaction
O
BzO
BzO
254
HN
NH2N N
O
H
HN
NH2N N
O
OBz
251, nitromethane,
SnCl4, 60°C, 3h
H
NH3/MeOH
47% 48%
250b
CH3
CH3
O
HO
HO
255
HN
NH2N N
O
OH
H
CH3
Scheme 64.
O
HO
HO
253
NH
N
H2N
N
O
OH
H
O
HO
HO
256
NH
N
i-BuHN
N
O
OH
H
(i-Bu)2O, pyridine,
rt, 3h
O
O
O
257
NH
N
i-BuHN
N
O
OH
H
Si
O
Si
i-Pr i-Pr
i-Pr
i-Pr
(i-Pr)2ClSiOSi(i-Pr)2Cl,
pyridine, rt, 12h
O
O
O
258a: R = H (48%)
b: R = C(S)OPh (10%)
NH
N
i-BuHN
N
O
OCOPh
Si
O
Si
i-Pr i-Pr
i-Pr
i-Pr
R
S
PhOC(S)Cl, MeCN,
rt, 12h
O
O
O
259
NH
N
i-BuHN
N
O
H
Si
O
Si
i-Pr i-Pr
i-Pr
i-Pr
258a: (n-Bu)3SnH,
AIBN, toluene,
60°C, 4h
257
O
HO
HO
260
NH
N
i-BuHN
N
O
H
0.1N TBAF/THF,
rt, 3h
O
HO
DMTO
261
NH
N
i-BuHN
N
O
H
DMT-Cl, pyridine,
rt, 3h
O
O
DMTO
262
NH
N
i-BuHN
N
O
H
P
NCCH2CH2O N(i-Pr)2
(i-Pr)2NP(Cl)O(CH2)2CN,
CH2Cl2, rt, 20 min
72% 57%
90% 82%
72% 78%
260
Scheme 65.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 209
of the silylated 257 with phenoxythiocarbonyl chloride in MeCN
furnished the 2’-O-phenoxythiocarbonyl derivative 258a (48%)
together with the bisphenoxythiocarbonyl compound 258b (10%) as
by-product. Reductive cleavage of 258a with tri-n-butyltin(IV)
hydride in toluene in the presence of AIBN yielded the 2’-deoxy
derivative 259 (90%). Desilylation with 0.1 N TBAF in anhydrous
THF gave the isobutyrylated 2’-deoxyribonucleoside 260 (82%).
However, the unprotected nucleoside was difficult to isolate due to
its unfavourable chromatographic properties. In order to prepare a
phosphoramidite building block of 260 which can be employed in
solid-phase oligonucleotide synthesis, the DMT group was intro-
duced at the 5’-hydroxyl group ( 261; 72%). The phosphoramid-
ite 262 was prepared under standard conditions from compound 261
(78% yield). For application of 262 in oligonucleotide synthesis and
properties of corresponding oligonucleotides, we refer to reference
[258].
6.2. 7-Deazapurine Nucleosides with Nitrogen-1 as Glycosyla-
tion Site
6.2.1. Fluorescent Pyrrolo-C 2’-Deoxyribonucleosides and
Analogs Thereof
Pyrrolo-dC (263a) contains the same 7-deazapurine (pyr-
rolo[2,3-d]pyrimidine) moiety as the fluorescent 7-deaza-2’-
deoxyisoinosine (37), but employing nitrogen-1 as glycosylation
position (Fig. 32). Also, pyrrolo-dC (263a), the corresponding ribo-
nucleoside pyrrolo-C (263b), and its 6-alkylated or alkynylated
derivatives (263c-j) develop significant fluorescence. This section
deals with the synthesis of pyrrolo-dC derivatives comprising sim-
ple alkyl or alkynyl side chains and studies of their fluorescence
properties, whereas pyrrolo-dC derivatives with complex side
chains that are prone to form additional hydrogen bonds in duplex
or triplex DNA are addressed in section 6.2.2. Due to the common
HN
NO
O
HO
N
HO
37
N
N
O
O
HO
HO
263a: R = H
b: R = OH
N
N
O
O
HO
HO
c: R = CH3
R
HN HN
R
d: R = (CH2)3
e: R = (CH2)4
f: R = (CH2)3 CH3
g: R = (CH2)5 CH3
h: R = CH2NHCOCF3
Phj: R =
i: R = CH2OCH3
263c-j
(3)
(1)
(7)(6)
(4a)
(systematic numbering)
purine numbering
1
3
98
5
Fig (32).
I
O
O
N
N
O
NH2
O
N
N
AcN
O
(Ph3P)PdCl2, CuI, Et3N,
DMF, N2, 60°C
265a: R = Si(CH3)3
R
f: R = (CH2)3
Si
O
Si
i-Pri-Pr
i-Pr
i-Pr
C
O
O
N
N
O
NH2
O
Si
O
Si
i-Pri-Pr
i-Pr
i-Pr
HC C R,C R
Ac2O
C
O
O
N
N
O
NHAc
O
Si
O
Si
i-Pri-Pr
i-Pr
i-Pr
C R
264
CH3
266a: R = Si(CH3)3
f: R = (CH2)3CH3
O
O
O
Si
O
Si
i-Pri-Pr
i-Pr
i-Pr
267a: R = H
f: R = (CH2)3CH3
N
N
HN
O
R
O
HO
HO
263a: R = H
f: R = (CH2)3CH3
CuI, DMF,
N2, 120°C
1) TBAF
2) NH4OH
266a,f
a: 90%
f: 83%
a: 78% over 2 steps
f: 81.5% over 2 steps
a: 75%
f: 75%
Scheme 66.
210 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
application of the systematic numbering in reports on pyrrolo-C
nucleosides, we employed this numbering throughout sections
6.2.1. and 6.2.2.
The synthesis of pyrrolo-dC (263a) using silylated 2’-deoxy-5-
iodocytidine (264) as starting material was first reported by
Ohtsuka and co-workers [260]. Introduction of the silylated ethynyl
side chain was achieved by cross-coupling in DMF in the presence
of (Ph3P)PdCl2, CuI and EtN3 to give the protected intermediate
265a in 90% yield. Next, the amino group was protected by acetyla-
tion ( 266a). Ring closure was performed in DMF with CuI, and
the silylated pyrrolo-dC derivative 267a was obtained in 78% over
two steps. Desilylation with TBAF followed by NH4OH treatment
afforded the unsubstituted pyrrolo-dC 2’-deoxyribonucleoside 263a
(75% yield). The 6-butyl derivative of pyrrolo-dC 263f was pre-
pared following the same procedure as shown in Scheme 66 [260].
Later, Gamper et al. obtained pyrrolo-dC (263a) via the
furano[2,3-d]pyrimidine intermediate 269a by oxygen nitrogen
exchange [261]. This route employs 2’-deoxy-5-ethynyluridine
(268a) as precursor [262, 263]. Cyclization of 268a was achieved
with copper iodide in the presence of Et3N in DMF to give the
furano[2,3-d]pyrimidine intermediate 269a in 83% yield. Treatment
of compound 269a with 30% aq. ammonia at room temperature
afforded the 7-deazapurine nucleoside 263a in high yield (95%)
(Scheme 67). On the contrary, it was demonstrated by others that no
oxygen nitrogen exchange takes place when a 5’,3’-di-acetylated
6-p-toluoyl furano[2,3-d]pyrimidine nucleoside was treated with
NH3 in methanol at room temperature for 20 h [264]. Under these
conditions, only the deprotected 6-p-toluoyl furano[2,3-
d]pyrimidine nucleoside was obtained.
Accordingly, a number of 6-alkylated or alkynylated pyrrolo-
dC analogs (263c-e) were prepared from their corresponding 5-
alkynylated 2’-deoxyuridine precursors (268c-e) as shown in
Scheme 68 [82, 265-267]. It should be noted that in some cases
already during Sonogashira cross-coupling, the formation of cy-
clized furano-pyrimidine nucleosides was reported [82, 267]. Al-
though in the context of pyrrolo-dC synthesis, furano-pyrimidine
nucleosides only function as intermediates, it is worthwhile to men-
tion that several analogs thereof show promising antiviral activities
[268, 269].
An alternative synthetic route makes use of the low nucleophil-
icity of the pyrrole nitrogen allowing the regioselective nucleobase
anion glycosylation at nitrogen-3 (pyrimidine moiety). This route
was exemplified on the ribonucleoside pyrrolo-C (263b) by our
laboratory and others (Scheme 69) [82, 270, 271], but was not ap-
plied to its 2’-deoxy derivative.
O
HO
HN
N
HO
O
O
O
HO
N
N
HO
O
O
O
HO
N
N
HO
HN
O
CuI, Et3N, DMF 30% aq. NH3, rt
83% 95%
268a 269a 263a
Scheme 67.
R
O
HO
HN
N
HO
O
O
O
HO
N
N
HO
O
O
O
HO
N
N
HO
HN
O
CuI, Et3N, MeOH,
reflux
25% aq. NH3,
rt or 50°C
268c: R =
R R
CH3
(CH2)3
(CH2)4
269c: R = CH3 (61% or 95%)
(CH2)3
(CH2)4
263c: R = CH3 (90% or 95%)
(CH2)3
(CH2)4
(90%)
(87%)
(92%)
(90%)
d: R =
e: R =
d: R =
e: R =
d: R =
e: R =
Scheme 68.
O
BzO
BzOO
BzO
N
N
BzO
HN
O
O
HO
N
N
HO
HN
O
BSA, TMSOTf,
80°C NaOMe/MeOH, rt
87% 81%
270
271 263b
OBz
OAc
HN
N NO
H
251
+
OBz OH
Scheme 69.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 211
6-Methylpyrrolo-dC (263c) was also converted into its corre-
sponding phosphoramidite building block (Scheme 70) and em-
ployed in solid-phase oligonucleotide synthesis [266]. However, as
reported by Berry and co-workers, building block synthesis used
the furano-pyrimidine intermediate 269c as precursor and introduc-
tion of the DMT residue was performed first ( 272c; 87% yield).
In the next step, conversion of the DMT protected furano-
pyrimidine intermediate 272c into the 7-deazapurine pyrrolo-dC
nucleoside 273c was accomplished using saturated ammonia in
MeOH in a pressure bottle at 50°C (49% yield). Phosphitylation ((i-
Pr2)2POCH2CH2CN, 1H-tetrazole, CH2Cl2, rt) of 273c gave com-
pound 274c in 53% yield as outlined in Scheme 70.
Similar routes were chosen for the phosphoramidite building
block synthesis of compounds 274h-j [272-274]. However, the
O
HO
N
N
DMTO
HN
O
O
O
N
N
DMTO
HN
O
sat. NH3 in MeOH,
pressure bottle, 50°C
(i-Pr2N)2POCH2CH2CN,
1H-tetrazole, CH2Cl2,
rt, 4.5h
P
NCCH2CH2O N(i-Pr)2
O
HO
N
N
DMTO
O
O
DMT-Cl, pyridine,
CH2Cl2
87% 49%
53%
272c
273c
274c
O
HO
N
N
HO
O
O
269c
273c
Scheme 70.
O
HO
N
N
DMTO
HN
O
O
O
N
N
DMTO
HN
O
aq. NH3, MeOH,
55°C
i-Pr2NP(Cl)OCH2CH2CN,
Et3N, CH2Cl2, rt
R R
P
NCCH2CH2O N(i-Pr)2
O
HO
N
N
DMTO
O
O
269h: DMT-Cl,
pyridine, DMAP
O
HO
HN
N
R2O
O
O
R1
268h: R1 = CH2NHCOCF3, R2 = H
Ph, R2 = DMT275j: R1 =
O
HO
N
N
R2O
O
O
R
CuI, Et3N, MeOH,
reflux
275i: R1 = CH2OCH3, R2 = DMT
269h: R = CH2NHCOCF3, R2 = H (77%)
Ph, R2 = DMT272j: R =
272i: R = CH2OCH3, R2 = DMT
272h (84%)
272h-j
273h: R = CH2NHCOCF3 (50%)
Ph (78%)j: R =
i: R = CH2OCH3 (85%)
274h: R = CH2NHCOCF3
Ph (80%)j: R =
i: R = CH2OCH3
NH
CF3
O
Scheme 71.
212 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
DMT residue was introduced at different stages for 274h and 274i,j
as demonstrated in Scheme 71. The first method employs the
trifluoroacetyl protected 6-propargyl derivative of 2’-deoxyuridine
268h as precursor [272]. First, ring closure towards the furano-
pyrimidine intermediate 269h was performed (77%), then the 5’-
hydroxyl group was protected with the DMT residue ( 272h;
84%). The second method utilizes the DMT-protected 5-
functionalized 2’-deoxyuridines 275i,j as precursors [273, 274].
Cyclization was performed on the DMT compounds 275i,j to afford
the furano[2,3-d]pyrimidine nucleosides 272i,j. Next, the oxygen to
nitrogen exchange was performed on 272h-j by ammonia treatment
yielding 273h-j. Conversion into the phosphoramidite building
blocks 274h-j was achieved under standard conditions (i-
Pr2P(Cl)OCH2CH2CN, Et3N, CH2Cl2, rt). Compounds 274h-j were
employed in solid-phase oligonucleotide synthesis, and the proper-
ties of 263i,j as constituents of oligonucleotides were investigated.
Oligonucleotides containing 263h were labelled with pyrene on the
oligonucleotide level after removal of the trifluoroacetyl protecting
group and utilized as molecular beacons [272].
A variation of the phosphoramidite building block synthesis of
the 6-phenyl derivative of pyrrolo-dC (274j) comprises acetylated
2’-deoxy-5-phenylethynyluridine (276) as starting material
(Scheme 72) [273]. Cyclization to the acetylated furano[2,3-
d]pyrimidine compound 277 was performed under Ag+-catalyzed
conditions (AgNO3, acetone, rt) as reported by Agrofoglio [275].
Deacetylation and oxygen to nitrogen exchange were performed in
one step followed by dimethoxytritylation to afford the DMT-
protected pyrrolo-dC compound 273j in 68% over two steps. As
described above, 273j was converted into phosphoramidite 274j.
Alternatively, conversion of furano[2,3-d]pyrimidine nucleo-
sides into their corresponding pyrrolo-dC derivatives can also be
achieved on oligonucleotide level by ammonia treatment (standard
oligonucleotide deprotection conditions) [261, 265, 266, 276, 277].
In this context, phosphoramidite building blocks of the furano[2,3-
d]pyrimidine nucleoside 278a,c,e,g,k,l were synthesized first as
shown in Scheme 73. This protocol was chosen for the synthesis of
oligonucleotides incorporating nucleosides 263a,c,e,g. Two routes
1) aq. NH3, MeOH,
55°C
2) DMT-Cl, pyridinei-Pr2NP(Cl)OCH2CH2CN,
Et3N, CH2Cl2, rt
O
AcO
N
N
AcO
O
O
Ph
O
AcO
HN
N
AcO
O
O
AgNO3, acetone,
rt, 24h
274j273j
Ph
75% 68% over 2 steps 80%
276 277
Scheme 72.
R
O
HO
HN
N
DMTO
O
O
O
HO
N
N
DMTO
O
O
O
O
N
N
DMTO
O
O
CuI, Et3N,
MeOH,
reflux
i-Pr2NP(Cl)OCH2CH2CN,
i-Pr2EtN, CH2Cl2, rt
R R
(CH2)4
(CH2)5
P
NCCH2CH2O N(i-Pr)2
CH3
CH3
O
HO
N
N
HO
O
O
R
269a: R = H
c: R = CH3
e: R = (CH2)4
275e: R =
g: R =
278a: R = H
c: R = CH3
e: R = (CH2)4
g: R = (CH2)5
CH3
272a: R = H
c: R = CH3
e: R = (CH2)4
g: R = (CH2)5
DMT-Cl,
pyridine
(CH2)2NPhthk: R =
(CH2)3NPhthl: R =
(CH2)2NPhthk: R =
(CH2)3NPhthl: R =
(CH2)2NPhthk: R =
(CH2)3NPhthl: R =
Scheme 73.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 213
were employed for the preparation of the DMT compounds
272a,c,e,g,k,l. The first method starts with the dimethoxytritylation
of the furano[2,3-d]pyrimidine nucleosides 269a,c,e yielding the
DMT protected compounds 272a,c,e. The second method begins
with the cyclization of the already DMT-protected alkylated or
alkynylated 2’-deoxyuridine analogs 275e,g,k,l affording the DMT-
protected furano[2,3-d]pyrimidine nucleosides 272e,g,k,l.
Phosphitylation was performed under standard conditions. The
phosphoramidites 278a,c,e,g,k,l were employed in solid-phase oli-
gonucleotides synthesis [261, 265, 266, 276, 277]. During deprotec-
tion (25% aq. NH3, 60°C, 14 h) of the oligonucleotides, the
furano[2,3-d]pyrimidine residues derived from 278a,c,e,g were
converted into their corresponding 7-deazapurine (pyrrolo[2,3-
d]pyrimidine) congeners as indicated in Scheme 74A [261, 265,
266, 277]. Deprotection of some oligonucleotides containing
furano[2,3-d]pyrimidine moieties derived from 278c,k,l was per-
formed in the presence of 30% aq. methylamine followed by recy-
clization with DOWEX ion-exchange resin (H+ form) (Scheme
74B) [276]. In this case, oxygen-7 of the furano[2,3-d]pyrimidine
moiety was replaced by an N-methyl group resulting in N7-
methylated 6-substitued pyrrolo-dC analogs as components of oli-
gonucleotides.
Pyrrolo-dC (263a), the corresponding ribonucleoside pyrrolo-C
(263b), and its 6-alkylated or alkynylated derivatives (263c-j) de-
velop significant fluorescence. Fluorescence spectra of 6-
hexynylpyrrolo-dC (263e) show an excitation maximum at 340 nm
with an emission at 466 nm (stoke shift of 126 nm), and the fluo-
rescence quantum yield ( ) of 263e in double distilled water was
determined to be 0.05 [265]. A representative emission and excita-
tion spectrum of 263e is shown in Fig. (33). Compared to the parent
ribonucleoside 263b, the introduction of a hexynyl side chain at
position-6 has almost no influence on the excitation maximum
(263b: 336 nm) and the quantum yield (263b: = 0.06). However,
the emission maximum of 263e is shifted to higher wavelengths
compared to 263b (450 nm).
Although pyrrolo-dC (263a) and its derivatives show signifi-
cant fluorescence, their extinction coefficients and quantum yields
are low when compared to fluorescent dyes like coumarin or
pyrene. In this context, our laboratory subjected 6-hexynylpyrrolo-
dC (263e) bearing a terminal triple bond to the copper(I)-catalyzed
Huisgen-Meldal-Sharpless “click” reaction (see also section 4.1.1)
and conjugated 263e to 3-azido-7-hydroxycoumarin (118) as shown
in Scheme 75 [265]. The pyrrolo-dC-coumarin conjugate 279 has
an excitation maximum at 350 nm with an emission at 476 nm
(stoke shift of 126 nm) at neutral pH (Fig. 34a) and an excitation
maximum at 392 nm with an emission at 476 nm at pH 8.5 (Fig.
34b). The fluorescence properties of coumarin derivatives are
strongly pH dependent, and at neutral pH the generation of the
O
O
N
N
O
O
O
O
O
N
N
O
HN
O
25% aq. NH3,
60°C, 14h
R R
DNA
DNA
DNA
DNA
CH3
R = H
R = (CH2)4
R = (CH2)5
R = CH3
O
O
N
N
O
O
O
O
O
N
N
O
N
O
R R
DNA
DNA
DNA
DNA
NH3R = (CH2)2
R = CH3
NH3R = (CH2)3
30% aq. MeNH2, Dowex
H+
A
B
Scheme 74.
Fig (33). The emission and excitation spectra of nucleoside 263e measured
at room temperature in double distilled water.
214 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
highly fluorescent phenolate anions is limited. At pH 7, the stoke
shifts of the 6-hexynylpyrrolo-dC nucleoside 263e and its click
derivative 279 are almost identical. However, the fluorescence
quantum yield of 279 was determined to be 0.18, which is signifi-
cantly higher than that of the parent nucleoside 263e ( = 0.05). On
the contrary, the closely related coumarin click conjugate of 2’-
deoxy-5-octa-1,7-diynyluridine (280; Scheme 75) reveals a quan-
tum yield of = 0.32 which is significantly higher than that of the
pyrrolo-dC click conjugate 279. From these findings, it was con-
cluded that the 7-deazapurine moiety of 279 quenches the fluores-
cence of the coumarin dye [265].
6.2.2. Pyrrolo-dC Analogs Forming Additional Hydrogen Bonds
Recent efforts have been dedicated to the development of novel
cytosine analogs which stabilize DNA duplexes through the forma-
tion of additional hydrogen bonds to the Hoogsteen binding sites of
guanine. These derivatives utilize either 9-aminoethoxy phenoxaz-
ine (“G-clamp”) [278, 279] or 6-substituted pyrrolo-C as their het-
erocyclic moiety [280, 281]. The work on phenoxazine nucleosides
goes back to a publication of Matteucci and co-workers reporting
for the first time on the synthesis of tricyclic phenoxazine deriva-
tives that are capable of base pairing [282]. Within the last years,
the extraordinary duplex stabilizing properties of the G-clamp com-
bined with its mismatch discrimination properties initialized the
syntheses of various closely related G-clamp analogs and their
building blocks for DNA, RNA or PNA [283-285]. Less work has
been dedicated to the development of 6-substituted pyrrolo-dC de-
rivatives which are also capable of forming additional hydrogen
bonds in duplex DNA. However, the extended hydrogen bonding
motif of 6-modified N-methylpyrrolo-dC nucleosides was used for
the recognition of the dC•dG base pair in triplex DNA [280, 281].
Two protocols have been used for the introduction of N-methyl
pyrrolo-dC nucleosides into oligonucleotides. The first one makes
use of furano[2,3-d]pyrimidine phosphoramidite building blocks as
demonstrated in Schemes 73 and 74, and the replacement of oxy-
gen-7 by the N-methyl group was accomplished on oligonucleotide
level. The second protocol utilizes N-methyl pyrrolo-dC phos-
phoramidite building blocks for solid-phase oligonucleotide synthe-
sis.
For the synthesis of the furano[2,3-d]pyrimidine phosphoramid-
ite building blocks 284a-c with m-trifluoroacetamido-, ureido-, and
O
HO
N
HO
N
O
HN
263e
N3
O OHO
118
O
O
N
N
N
OH
O
HO
N
HO
N
O
HN
279
CuSO4 5 H2O
Na-ascorbate
THF:H2O:t-BuOH
+85%
O
O
N
N
N
OH
O
HO
N
HO
HN
O
O
280
Scheme 75.
Fig (34). The emission and excitation spectra of the pyrrolo-dC coumarin conjugate 279 measured (a) in water (pH 7.0) and (b) in 0.1 M Tris-HCl buffer (pH
8.5).
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 215
acetamido-phenyl side chain modifications at position-6 of the het-
erocyclic moiety, the DMT protected 2’-deoxyuridine derivatives
281a-c were used as starting material. Compounds 281a-c were
constructed by Sonogashira cross-coupling [280] and were cyclized
using CuI and Et3N in MeOH to yield the DMT-protected
furano[2,3-d]pyrimidine nucleosides 282a-c in 69-96% yield. The
aniline moiety of 282a was further protected using ethyl
trifluoroacetate ( 283a) as indicated by Scheme 76. Phosphityla-
tion of 282b,c and 283a under standard conditions furnished the
phosphoramidites 284a-c in 64-69% yield. Conversion of the
furano[2,3-d]pyrimidine moieties to the N-methyl pyrrolo[2,3-
d]pyrimidine derivatives was achieved with aq. methylamine on the
oligonucleotide level as described in section 6.2.1 and illustrated in
Scheme 74B. However, this protocol was encountered with difficul-
ties as it was found that, the re-cyclization of the N-methyl 7-
deazapurine moiety was incomplete in some cases (identified by
HPLC analysis) [281].
The alternative strategy requires the synthesis of protected 2’-
deoxy-5-iodo-4N-methylcytidine (287) as starting material [281].
The synthesis route is outlined in Scheme 77 and uses 5’-DMT
protected 2’-deoxy-5-iodouridine (285) as precursor. Then, the 3’-
hydroxyl group was acetylated ( 286). For the conversion of 286
into the target compound 287, the 4-carbonyl position was activated
first using N-methylimidazole (NMI) and POCl3 in pyridine. After
the introduction of methylamine and deacetylation, the DMT com-
pound 2’-deoxy-5-iodo-4N-methylcytidine 287 was obtained in
88% yield. Several side chains were introduced via the Pd-
catalyzed Sonogashira cross-coupling reaction employing different
phenyl acetylene residues (288a-d). Cyclization was achieved with
CuI and Et3N in DMF to yield N-methyl pyrrolo-dC analogs with
phenyl (289a), acetamido (289b), ureido (289c) or phenylamino
(289d) modifications at position-6 (Scheme 78). The amino group
of 289d was further protected using ethyl trifluoroacetate ( 289e).
Conversion into the corresponding phosphoramidites was achieved
under standard conditions and gave 290a-c,e in 51-86% yield [281].
The 6-guanidinyl derivative 292 was obtained in 44% yield
from compound 289d using isothiourea (291) as guanidinylation
reagent (Scheme 79). Subsequent phosphitylation afforded phos-
phoramidite 293 in 47% yield. The building blocks 290a-c,e and
293 were incorporate into triplex forming oligonucleotides (TFOs)
and deprotected under standard conditions. The binding affinity and
selectivity for dC•dG was evaluated. For details, we refer to [281].
7. CONCLUSION AND OUTLOOK
As 7-deazapurines mimic the shape of purines ideally, the cor-
responding 2'-deoxyribonucleosides can replace canonical purine
constituents of DNA. Moreover, they are substrates or inhibitors of
various enzymes. A highly stable glycosylic bond and a rather inert
6-amino function make this class of modified nucleosides resistant
against nucleoside converting enzymes, such as nucleoside phos-
phorylase or adenosine deaminase.
O
HO
HN
N
DMTO
O
O
O
HO
N
N
DMTO
O
O
O
O
N
N
DMTO
O
O
CuI, Et3N,
MeOH, 80°C
i-Pr2NP(Cl)OCH2CH2CN,
i-Pr2EtN, THF, rt
P
NCCH2CH2ON(i-Pr)2
281a: R = H
b: R = COCH3
c: R = CONH2
NHR
NHR
NHR
282a: R = H (96%)
b: R = COCH3 (89%)
c: R = CONH2 (69%)
O
HO
N
N
DMTO
O
O
NHR
282a: CF3COOEt, DMAP,
Et3N, THF, 80-85°C
283a: R = COCF3 (35%)
a: R = COCF3 (64%)
b: R = COCH3 (69%)
c: R = CONH2 (69%)
284a-c
Scheme 76.
216 Current Organic Chemistry, 2012, Vol. 16, No. 2 Seela et al.
Initially, the preparation of 7-deazapurine 2’-deoxyribo-
nucleosides was encountered with difficulties, but after the devel-
opment of the nucleobase anion glycosylation protocol they became
easily accessible. The central key step of this protocol comprises
the formation of the nucleobase anion as highly reactive compo-
nent. Generation of the nucleobase anion can be achieved (i) under
liquid-liquid conditions, (ii) under solid-liquid conditions or (iii) by
the sodium salt glycosylation. The second important component of
the nucleobase anion glycosylation is the halogenose. By employ-
ing one among the different halogenose derivatives, the -D or -L
enantiomers of 7-deazapurine 2’-deoxyribonucleosides, 7-
deazapurine 2’-deoxy-2’-fluoroarabinonucleosides or 7-deazapurine
2’,3’-dideoxyribonucleosides can be prepared. Up to now, a broad
diversity of 7-deazapurine nucleosides has been synthesized using
one of the various glycosylation protocols. Alternatively, 7-
deazapurine 2’-deoxyribonucleosides can also be obtained from
their corresponding ribonucleosides by Barton-McCombie deoxy-
genation. Nucleophilic substitution reactions were carried out on
central intermidiates and led to various 7-deazapurine nucleosides
related to 2’-deoxyadensoine, 2’-deoxyguanosine or 2’-
deoxyinosine.
As position-7 of 7-deazapurine nucleosides points towards the
major groove of DNA, this site is an ideal position to introduce
functionalities into DNA. Regioselective halogenation reactions on
the various 7-deazapurine nucleosides have been performed, and it
was found that halogen substituents introduced at position-7 greatly
O
HO
HN
N
DMTO
O
OAc2O, pyridine,
0°C-rt, 3h
285
I
O
AcO
HN
N
DMTO
O
O
286
I
O
HO
N
N
DMTO
NH
O
287
I1) POCl3, NMI,
pyridine, 0°C-rt, 1h
2) aq. MeNH2,
0°C-rt, 16h
97% 88%
288a-d
alkyne, Pd(PPh3)4,
CuI, Et3N, DMF,
dark, rt
Scheme 77.
O
HO
N
N
DMTO
N
O
O
O
N
N
DMTO
N
O
CuI, Et3N, DMF,
dark, 125°C
289a-c,e:
i-Pr2NP(Cl)O(CH2)2CN,
i-Pr2EtN, CH2Cl2 or THF
or THF-DMF, rt
P
NCCH2CH2O N(i-Pr)2
R R
290a: R = H (66%)
b: R = NHCOCH3 (86%)
c: R = NHCONH2 (51%)
289a: R = H (71%)
b: R = NHCOCH3
c: R = NHCONH2
d: R = NH2 (80%)
e: R = NHCOCF3 (65%)
CF3CO2Et, DMAP, Et3N,
CH2Cl2
e: R = NHCOCF3 (70%)
O
HO
N
N
DMTO
NH
O
R
288a: R = H (90%)
b: R = NHCOCH3
c: R = NHCONH2
d: R = NH2 (95%)
Scheme 78.
O
HO
N
N
DMTO
N
O
O
O
N
N
DMTO
N
O
291, pyridine,
CH2Cl2, reflux, 2d
i-Pr2NP(Cl)O(CH2)2CN,
i-Pr2EtN, CH2Cl2, rt, 2h
P
NCCH2CH2O N(i-Pr)2
NH
293
289d 292
O
HO
N
N
DMTO
N
O
NH2
RN
NHR
R = CO2(CH2)2CN
NH
RN
NHR
44% 47%
NCO N
H
O
N O
S O
CN
291
Scheme 79.
7-Deazapurine (Pyrrolo[2,3-d]pyrimidine) 2’-Deoxyribonucleosides Current Organic Chemistry, 2012, Vol. 16, No. 2 217
increase duplex stability. An even better stabilization was observed
when the halogen substituents are replaced by alkynyl residues. The
strongly stabilizing effect on DNA makes 7-deazapurines good
candidates for primers or probes used in DNA diagnostics, sequenc-
ing or for antisense technology. Also, 7-deazapurine nucleosides
substituted with side chains carrying terminal triple bonds are ad-
vantageous for further functionalization employing the protocol of
the copper(I)-catalyzed Huisgen-Meldal-Sharpless “click” reaction.
“Click” reactions on 7-alkynylated 7-deazapurine 2’-
deoxyribonucleosides have been performed on nucleoside, nucleo-
tide as well as on oligonucleotide level and a broad range of azido
reporter groups have been utilized for labelling. Recently, the
“click” reaction has been used to crosslink 7-deazapurine 2’-
deoxyribonucleosides and oligonucleotides [149, 178].
The assembly of nucleosides and oligonucleotides to su-
pramolecular devices is another promising topic. Since 7-
deazapurines 2’-deoxyribonucleosides are very robust against alka-
line and acidic conditions, stable supramolecular structures can be
formed which might show advantages over the assemblies built up
by purine systems. As an example, aggregates of oligonucleotide
294 incorporating 7-bromo-7-deaza-2’-deoxyisoguanosine (36c;
Br7c
7iGd) have been shown to form a tetraplex in the presence of
Na+ (Fig. 35a) and Rb
+ (Fig. 35b), while a pentaplex is generated in
the presence of Cs+
(Fig. 35c) [286, 287]. Possible structures of
quadruplexes or pentaplexes are shown in Fig. (35d,e).
A variety of pyrrolo[2,3-d]pyrimidines develop antitumor activ-
ity. One of the recent prominent examples is Pemetrexed (Alimta®;
295), a folate antimetabolite which was developed by E. C. Taylor
[289] and brought into the market by the pharmaceutical company
Lilly (Fig. 36). Its antitumor activity is based on the inhibition of
enzymes acting in purine and pyrimidine synthesis. In that respect,
it can be expected that 7-dezapurine nucleosides possess a tremen-
dous potential to be used as anticancer or antiviral agents [208].
HN
N NH
O
H2N
295
NH
O
COOH
COOH
Pemetrexed, Alimta
Fig (36).
Moreover, modified pyrrolo[2,3-d]pyrimidine 2’-
deoxyribonucleosides and oligonucleotides will be applicable in
new areas of chemistry, physics, and biology, including nanotech-
nology and nanoelectronics. The use of DNA as a building block
for nanoelectronic sensors and devices due to its efficient hole-
conducting properties is currently under investigation. Recently, it
was reported that the charge-transfer efficiency of DNA can be
efficiently increased when 7-deaza-2’-deoxyadenosine is used as a
substitute of 2’-deoxyadenosine [290].
ACKNOWLEDGMENTS
We thank Mr. Ping Ding, Mr. Suresh S. Pujari, Mr. Sachin A.
Ingale and Dr. Peter Leonard for reading the manuscript and for
helpful comments. Financial support by the Bundesministerium für
Bildung und Forschung (BMBF) and ChemBiotech (Germany) is
gratefully acknowledged. We thank all people whose work is de-
scribed in this review.
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Received: 09 December, 2010 Revised: 02 May, 2011 Accepted: 05 August, 2011